Ronald J. Wong

All Publications

• Is it time for a precision health approach to the management of newborn hyperbilirubinemia? Journal of perinatology : official journal of the California Perinatal Association Stevenson, D. K., Wells, G. S., Wong, R. J. 2024

  Abstract

  Newborn hyperbilirubinemia during the first two weeks of life is one of most common problems requiring management decisions by a pediatrician. However, high bilirubin levels in the circulation have been associated with neurologic injury under a variety of conditions encountered in the newborn infant, such as hemolysis. The risk for developing dangerous hyperbilirubinemia is multifactorial and is determined by a complex set of factors related to a newborn infant's genetic capacities as well as intra- and extrauterine exposures. To this end, a precision health approach based on the integration of prenatal genetic and postnatal diagnostic measures might improve the management of neonatal hyperbilirubinemia.

  View details for DOI 10.1038/s41372-024-01941-3

  View details for PubMedID 38514741

  View details for PubMedCentralID 225223

• Large-scale proteomics in the first trimester of pregnancy predict psychopathology and temperament in preschool children: an exploratory study. Journal of child psychology and psychiatry, and allied disciplines Buthmann, J. L., Miller, J. G., Aghaeepour, N., King, L. S., Stevenson, D. K., Shaw, G. M., Wong, R. J., Gotlib, I. H. 2024

  Abstract

  Understanding the prenatal origins of children's psychopathology is a fundamental goal in developmental and clinical science. Recent research suggests that inflammation during pregnancy can trigger a cascade of fetal programming changes that contribute to vulnerability for the emergence of psychopathology. Most studies, however, have focused on a handful of proinflammatory cytokines and have not explored a range of prenatal biological pathways that may be involved in increasing postnatal risk for emotional and behavioral difficulties.Using extreme gradient boosted machine learning models, we explored large-scale proteomics, considering over 1,000 proteins from first trimester blood samples, to predict behavior in early childhood. Mothers reported on their 3- to 5-year-old children's (N = 89, 51% female) temperament (Child Behavior Questionnaire) and psychopathology (Child Behavior Checklist).We found that machine learning models of prenatal proteomics predict 5%-10% of the variance in children's sadness, perceptual sensitivity, attention problems, and emotional reactivity. Enrichment analyses identified immune function, nervous system development, and cell signaling pathways as being particularly important in predicting children's outcomes.Our findings, though exploratory, suggest processes in early pregnancy that are related to functioning in early childhood. Predictive features included far more proteins than have been considered in prior work. Specifically, proteins implicated in inflammation, in the development of the central nervous system, and in key cell-signaling pathways were enriched in relation to child temperament and psychopathology measures.

  View details for DOI 10.1111/jcpp.13948

  View details for PubMedID 38287782

• Integrative analysis of noncoding mutations identifies the druggable genome in preterm birth. Science advances Wang, C., Wang, Y. J., Ying, L., Wong, R. J., Quaintance, C. C., Hong, X., Neff, N., Wang, X., Biggio, J. R., Mesiano, S., Quake, S. R., Alvira, C. M., Cornfield, D. N., Stevenson, D. K., Shaw, G. M., Li, J. 2024; 10 (3): eadk1057

  Abstract

  Preterm birth affects ~10% of pregnancies in the US. Despite familial associations, identifying at-risk genetic loci has been challenging. We built deep learning and graphical models to score mutational effects at base resolution via integrating the pregnant myometrial epigenome and large-scale patient genomes with spontaneous preterm birth (sPTB) from European and African American cohorts. We uncovered previously unidentified sPTB genes that are involved in myometrial muscle relaxation and inflammatory responses and that are regulated by the progesterone receptor near labor onset. We studied genomic variants in these genes in our recruited pregnant women administered progestin prophylaxis. We observed that mutation burden in these genes was predictive of responses to progestin treatment for preterm birth. To advance therapeutic development, we screened ~4000 compounds, identified candidate molecules that affect our identified genes, and experimentally validated their therapeutic effects on regulating labor. Together, our integrative approach revealed the druggable genome in preterm birth and provided a generalizable framework for studying complex diseases.

  View details for DOI 10.1126/sciadv.adk1057

  View details for PubMedID 38241369

  View details for PubMedCentralID PMC10798565

• Microbiome preterm birth DREAM challenge: Crowdsourcing machine learning approaches to advance preterm birth research. Cell reports. Medicine Golob, J. L., Oskotsky, T. T., Tang, A. S., Roldan, A., Chung, V., Ha, C. W., Wong, R. J., Flynn, K. J., Parraga-Leo, A., Wibrand, C., Minot, S. S., Oskotsky, B., Andreoletti, G., Kosti, I., Bletz, J., Nelson, A., Gao, J., Wei, Z., Chen, G., Tang, Z. Z., Novielli, P., Romano, D., Pantaleo, E., Amoroso, N., Monaco, A., Vacca, M., De Angelis, M., Bellotti, R., Tangaro, S., Kuntzleman, A., Bigcraft, I., Techtmann, S., Bae, D., Kim, E., Jeon, J., Joe, S., Theis, K. R., Ng, S., Lee, Y. S., Diaz-Gimeno, P., Bennett, P. R., MacIntyre, D. A., Stolovitzky, G., Lynch, S. V., Albrecht, J., Gomez-Lopez, N., Romero, R., Stevenson, D. K., Aghaeepour, N., Tarca, A. L., Costello, J. C., Sirota, M. 2023: 101350

  Abstract

  Every year, 11% of infants are born preterm with significant health consequences, with the vaginal microbiome a risk factor for preterm birth. We crowdsource models to predict (1) preterm birth (PTB; <37 weeks) or (2) early preterm birth (ePTB; <32 weeks) from 9 vaginal microbiome studies representing 3,578 samples from 1,268 pregnant individuals, aggregated from public raw data via phylogenetic harmonization. The predictive models are validated on two independent unpublished datasets representing 331 samples from 148 pregnant individuals. The top-performing models (among 148 and 121 submissions from 318 teams) achieve area under the receiver operator characteristic (AUROC) curve scores of 0.69 and 0.87 predicting PTB and ePTB, respectively. Alpha diversity, VALENCIA community state types, and composition are important features in the top-performing models, most of which are tree-based methods. This work is a model for translation of microbiome data into clinically relevant predictive models and to better understand preterm birth.

  View details for DOI 10.1016/j.xcrm.2023.101350

  View details for PubMedID 38134931

• The placental vasculature is affected by changes in gene expression and glycogen-rich cells in a diet-induced obesity mouse model. PloS one Zhao, H., Wong, R. J., Stevenson, D. K. 2023; 18 (11): e0294185

  Abstract

  Maternal obesity is a risk factor for pregnancy complications. Obesity caused by a high-fat diet (HFD) may alter maternal glucose/glycogen metabolism. Here, our objective was to investigate whether the placental vasculature is altered via changes in gene expression and glycogen-rich cells using a preclinical mouse model of diet-induced obesity. We subjected female FVB/N mice to one of three feeding regimens: regular chow (RC) given at preconception and during pregnancy (Control); RC given at preconception and then a HFD during pregnancy (HFD-P); or HFD initiated 4 weeks preconception and during pregnancy (HFD-PreCP). Daily food consumption and weekly maternal weights were recorded. Maternal blood glucose levels were measured at preconception and 4 gestational epochs (E6.5-E9.5, E10.5-E12.5, E13.5-E15.5, E16.5-E19.5). At E8.5-E16.5, total RNA in placentas were isolated for gene expression analyses. Placentas were also collected for HE and periodic acid Schiff's (PAS) staining and glycogen content assays. Dams in the HFD-P and HFD-PreCP groups gained significantly more weight than controls. Pre- and antenatal glucose levels were also significantly higher (15%-30%) in HFD-PreCP dams. Expression of several placental genes were also altered in HFD dams compared with controls. Consumption of the HFD also led to phenotypic and morphologic changes in glycogen trophoblasts (GlyTs) and uterine natural killer (uNK) cells. Alterations in vascularity were also observed in the labyrinth of HFD-PreCP placentas, which correlated with decreased placental efficiency. Overall, we observed that a HFD induces gestational obesity in mice, alters expression of placental genes, affects glucose homeostasis, and alters glycogen-positive GlyTs and uNK cells. All these changes may lead to impaired placental vascular development, and thus heighten the risk for pregnancy complications.

  View details for DOI 10.1371/journal.pone.0294185

  View details for PubMedID 37948457

  View details for PubMedCentralID PMC10637699

• Challenges to the study of gender dysphoria. Acta paediatrica (Oslo, Norway : 1992) Stevenson, D. K., Murray, J. C., Muglia, L. J., Wong, R. J., Katz, M. 2023

  View details for DOI 10.1111/apa.16975

  View details for PubMedID 37724910

• Comparative predictive power of serum vs plasma proteomic signatures in feto-maternal medicine. AJOG global reports Espinosa, C., Ali, S. M., Khan, W., Khanam, R., Pervin, J., Price, J. T., Rahman, S., Hasan, T., Ahmed, S., Raqib, R., Rahman, M., Aktar, S., Nisar, M. I., Khalid, J., Dhingra, U., Dutta, A., Deb, S., Stringer, J. S., Wong, R. J., Shaw, G. M., Stevenson, D. K., Darmstadt, G. L., Gaudilliere, B., Baqui, A. H., Jehan, F., Rahman, A., Sazawal, S., Vwalika, B., Aghaeepour, N., Angst, M. S. 2023; 3 (3): 100244

  Abstract

  Blood proteins are frequently measured in serum or plasma, because they provide a wealth of information. Differences in the ex vivo processing of serum and plasma raise concerns that proteomic health and disease signatures derived from serum or plasma differ in content and quality. However, little is known about their respective power to predict feto-maternal health outcomes. Predictive power is a sentinel characteristic to determine the clinical use of biosignatures.This study aimed to compare the power of serum and plasma proteomic signatures to predict a physiological pregnancy outcome.Paired serum and plasma samples from 73 women were obtained from biorepositories of a multinational prospective cohort study on pregnancy outcomes. Gestational age at the time of sampling was the predicted outcome, because the proteomic signatures have been validated for such a prediction. Multivariate and cross-validated models were independently derived for serum and plasma proteins.A total of 1116 proteins were measured in 88 paired samples from 73 women with a highly multiplexed platform using proximity extension technology (Olink Proteomics Inc, Watertown, MA). The plasma proteomic signature showed a higher predictive power (R=0.64; confidence interval, 0.42-0.79; P=3.5×10-6) than the serum signature (R=0.45; confidence interval, 0.18-0.66; P=2.2×10-3). The serum signature was validated in plasma with a similar predictive power (R=0.58; confidence interval, 0.34-0.75; P=4.8×10-5), whereas the plasma signature was validated in serum with reduced predictive power (R=0.53; confidence interval, 0.27-0.72; P=2.6×10-4). Signature proteins largely overlapped in the serum and plasma, but the strength of association with gestational age was weaker for serum proteins.Findings suggest that serum proteomics are less informative than plasma proteomics. They are compatible with the view that the partial ex-vivo degradation and modification of serum proteins during sample processing are an underlying reason. The rationale for collecting and analyzing serum and plasma samples should be carefully considered when deriving proteomic biosignatures to ascertain that specimens of the highest scientific and clinical yield are processed. Findings suggest that plasma is the preferred matrix.

  View details for DOI 10.1016/j.xagr.2023.100244

  View details for PubMedID 37456144

  View details for PubMedCentralID PMC10339042

• A "Gold Standard" Test for Diagnosing and Quantifying Hemolysis in Neonates and Infants. Journal of perinatology : official journal of the California Perinatal Association Christensen, R. D., Bahr, T. M., Wong, R. J., Vreman, H. J., Bhutani, V. K., Stevenson, D. K. 2023

  Abstract

  Identifying "gold standard" diagnostic tests can promote evidence-based neonatology practice. Hemolysis is a pathological shortening of the erythrocyte lifespan, differing from erythrocyte senescence in responsible mechanisms and clinical implications. Diagnosing hemolysis goes beyond a binary (yes vs. no) determination. It is characterized according to magnitude, and as acute vs. chronic, and genetically based vs. not. For neonates with significant hyperbilirubinemia or anemia, detecting hemolysis and quantifying its magnitude provides diagnostic clarity. The 2022 American Academy of Pediatrics (AAP) Clinical Practice Guideline on management of hyperbilirubinemia in the newborn states that hemolysis is a risk factor for developing significant hyperbilirubinemia and neurotoxicity. The guideline recommends identifying hemolysis from any cause, but specific guidance is not provided. A spectrum of laboratory tests has been endorsed as diagnostic methods for hemolysis. Herein we examine these laboratory tests and recommend one as the "gold standard" for diagnosing and quantifying hemolysis in neonates and infants.

  View details for DOI 10.1038/s41372-023-01730-4

  View details for PubMedID 37468612

  View details for PubMedCentralID 4623764

• Abrupt perturbation and delayed recovery of the vaginal ecosystem following childbirth. Nature communications Costello, E. K., DiGiulio, D. B., Robaczewska, A., Symul, L., Wong, R. J., Shaw, G. M., Stevenson, D. K., Holmes, S. P., Kwon, D. S., Relman, D. A. 2023; 14 (1): 4141

  Abstract

  The vaginal ecosystem is closely tied to human health and reproductive outcomes, yet its dynamics in the wake of childbirth remain poorly characterized. Here, we profile the vaginal microbiota and cytokine milieu of participants sampled longitudinally throughout pregnancy and for at least one year postpartum. We show that delivery, regardless of mode, is associated with a vaginal pro-inflammatory cytokine response and the loss of Lactobacillus dominance. By contrast, neither the progression of gestation nor the approach of labor strongly altered the vaginal ecosystem. At 9.5-months postpartum-the latest timepoint at which cytokines were assessed-elevated inflammation coincided with vaginal bacterial communities that had remained perturbed (highly diverse) from the time of delivery. Time-to-event analysis indicated a one-year postpartum probability of transitioning to Lactobacillus dominance of 49.4%. As diversity and inflammation declined during the postpartum period, dominance by L. crispatus, the quintessential health-associated commensal, failed to return: its prevalence before, immediately after, and one year after delivery was 41%, 4%, and 9%, respectively. Revisiting our pre-delivery data, we found that a prior live birth was associated with a lower odds of L. crispatus dominance in pregnant participants-an outcome modestly tempered by a longer ( > 18-month) interpregnancy interval. Our results suggest that reproductive history and childbirth in particular remodel the vaginal ecosystem and that the timing and degree of recovery from delivery may help determine the subsequent health of the woman and of future pregnancies.

  View details for DOI 10.1038/s41467-023-39849-9

  View details for PubMedID 37438386

  View details for PubMedCentralID 4355684

• Breath: The Exhaust of Metabolism JOURNAL OF PEDIATRICS Stevenson, D. K., Maric, I., Wong, R. J. 2023; 257: 1-3

  View details for DOI 10.1018/j.jpeds.2023.03.002

  View details for Web of Science ID 001030453900001

• Development of a Urine Metabolomics Biomarker-Based Prediction Model for Preeclampsia during Early Pregnancy. Metabolites Zhang, Y., Sylvester, K. G., Jin, B., Wong, R. J., Schilling, J., Chou, C. J., Han, Z., Luo, R. Y., Tian, L., Ladella, S., Mo, L., Maric, I., Blumenfeld, Y. J., Darmstadt, G. L., Shaw, G. M., Stevenson, D. K., Whitin, J. C., Cohen, H. J., McElhinney, D. B., Ling, X. B. 2023; 13 (6)

  Abstract

  Preeclampsia (PE) is a condition that poses a significant risk of maternal mortality and multiple organ failure during pregnancy. Early prediction of PE can enable timely surveillance and interventions, such as low-dose aspirin administration. In this study, conducted at Stanford Health Care, we examined a cohort of 60 pregnant women and collected 478 urine samples between gestational weeks 8 and 20 for comprehensive metabolomic profiling. By employing liquid chromatography mass spectrometry (LCMS/MS), we identified the structures of seven out of 26 metabolomics biomarkers detected. Utilizing the XGBoost algorithm, we developed a predictive model based on these seven metabolomics biomarkers to identify individuals at risk of developing PE. The performance of the model was evaluated using 10-fold cross-validation, yielding an area under the receiver operating characteristic curve of 0.856. Our findings suggest that measuring urinary metabolomics biomarkers offers a noninvasive approach to assess the risk of PE prior to its onset.

  View details for DOI 10.3390/metabo13060715

  View details for PubMedID 37367874

• Multiomic signals associated with maternal epidemiological factors contributing to preterm birth in low- and middle-income countries. Science advances Espinosa, C. A., Khan, W., Khanam, R., Das, S., Khalid, J., Pervin, J., Kasaro, M. P., Contrepois, K., Chang, A. L., Phongpreecha, T., Michael, B., Ellenberger, M., Mehmood, U., Hotwani, A., Nizar, A., Kabir, F., Wong, R. J., Becker, M., Berson, E., Culos, A., De Francesco, D., Mataraso, S., Ravindra, N., Thuraiappah, M., Xenochristou, M., Stelzer, I. A., Marić, I., Dutta, A., Raqib, R., Ahmed, S., Rahman, S., Hasan, A. S., Ali, S. M., Juma, M. H., Rahman, M., Aktar, S., Deb, S., Price, J. T., Wise, P. H., Winn, V. D., Druzin, M. L., Gibbs, R. S., Darmstadt, G. L., Murray, J. C., Stringer, J. S., Gaudilliere, B., Snyder, M. P., Angst, M. S., Rahman, A., Baqui, A. H., Jehan, F., Nisar, M. I., Vwalika, B., Sazawal, S., Shaw, G. M., Stevenson, D. K., Aghaeepour, N. 2023; 9 (21): eade7692

  Abstract

  Preterm birth (PTB) is the leading cause of death in children under five, yet comprehensive studies are hindered by its multiple complex etiologies. Epidemiological associations between PTB and maternal characteristics have been previously described. This work used multiomic profiling and multivariate modeling to investigate the biological signatures of these characteristics. Maternal covariates were collected during pregnancy from 13,841 pregnant women across five sites. Plasma samples from 231 participants were analyzed to generate proteomic, metabolomic, and lipidomic datasets. Machine learning models showed robust performance for the prediction of PTB (AUROC = 0.70), time-to-delivery (r = 0.65), maternal age (r = 0.59), gravidity (r = 0.56), and BMI (r = 0.81). Time-to-delivery biological correlates included fetal-associated proteins (e.g., ALPP, AFP, and PGF) and immune proteins (e.g., PD-L1, CCL28, and LIFR). Maternal age negatively correlated with collagen COL9A1, gravidity with endothelial NOS and inflammatory chemokine CXCL13, and BMI with leptin and structural protein FABP4. These results provide an integrated view of epidemiological factors associated with PTB and identify biological signatures of clinical covariates affecting this disease.

  View details for DOI 10.1126/sciadv.ade7692

  View details for PubMedID 37224249

• Target-agnostic drug prediction integrated with medical record analysis uncovers differential associations of statins with increased survival in COVID-19 patients. PLoS computational biology Sperry, M. M., Oskotsky, T. T., Marić, I., Kaushal, S., Takeda, T., Horvath, V., Powers, R. K., Rodas, M., Furlong, B., Soong, M., Prabhala, P., Goyal, G., Carlson, K. E., Wong, R. J., Kosti, I., Le, B. L., Logue, J., Hammond, H., Frieman, M., Stevenson, D. K., Ingber, D. E., Sirota, M., Novak, R. 2023; 19 (5): e1011050

  Abstract

  Drug repurposing requires distinguishing established drug class targets from novel molecule-specific mechanisms and rapidly derisking their therapeutic potential in a time-critical manner, particularly in a pandemic scenario. In response to the challenge to rapidly identify treatment options for COVID-19, several studies reported that statins, as a drug class, reduce mortality in these patients. However, it is unknown if different statins exhibit consistent function or may have varying therapeutic benefit. A Bayesian network tool was used to predict drugs that shift the host transcriptomic response to SARS-CoV-2 infection towards a healthy state. Drugs were predicted using 14 RNA-sequencing datasets from 72 autopsy tissues and 465 COVID-19 patient samples or from cultured human cells and organoids infected with SARS-CoV-2. Top drug predictions included statins, which were then assessed using electronic medical records containing over 4,000 COVID-19 patients on statins to determine mortality risk in patients prescribed specific statins versus untreated matched controls. The same drugs were tested in Vero E6 cells infected with SARS-CoV-2 and human endothelial cells infected with a related OC43 coronavirus. Simvastatin was among the most highly predicted compounds (14/14 datasets) and five other statins, including atorvastatin, were predicted to be active in > 50% of analyses. Analysis of the clinical database revealed that reduced mortality risk was only observed in COVID-19 patients prescribed a subset of statins, including simvastatin and atorvastatin. In vitro testing of SARS-CoV-2 infected cells revealed simvastatin to be a potent direct inhibitor whereas most other statins were less effective. Simvastatin also inhibited OC43 infection and reduced cytokine production in endothelial cells. Statins may differ in their ability to sustain the lives of COVID-19 patients despite having a shared drug target and lipid-modifying mechanism of action. These findings highlight the value of target-agnostic drug prediction coupled with patient databases to identify and clinically evaluate non-obvious mechanisms and derisk and accelerate drug repurposing opportunities.

  View details for DOI 10.1371/journal.pcbi.1011050

  View details for PubMedID 37146076

• Breath: The Exhaust of Metabolism. The Journal of pediatrics Stevenson, D. K., Marić, I., Wong, R. J. 2023

  View details for DOI 10.1016/j.jpeds.2023.03.002

  View details for PubMedID 36925060

• Heme, Heme Oxygenase-1, Statins, and SARS-CoV-2. Antioxidants (Basel, Switzerland) Stevenson, D. K., Vreman, H. J., Wong, R. J. 2023; 12 (3)

  Abstract

  Heme, a metalloporphyrin, or more specifically, a tetrapyrrole containing ferrous iron, is an ancient molecule [...].

  View details for DOI 10.3390/antiox12030614

  View details for PubMedID 36978862

• Prefrontal activation in preschool children is associated with maternal adversity and child temperament: A preliminary fNIRS study of inhibitory control DEVELOPMENTAL PSYCHOBIOLOGY Miller, J. G., Hyat, M., Perlman, S. B., Wong, R. J., Shaw, G. M., Stevenson, D. K., Gotlib, I. H. 2023; 65 (1): e22351

  Abstract

  Exposure to adversity is a well-documented risk factor for cognitive, behavioral, and mental health problems. In fact, the consequences of adversity may be intergenerational. A growing body of research suggests that maternal exposures to adversity, including those prior to childbirth, are associated with offspring biobehavioral development. In a sample of 36 mothers and their preschool-age children (mean child age = 4.21 ± 0.92 years), we used functional near-infrared spectroscopy to replicate and extend this work to include brain activation during inhibitory control in young children. We found that measures of maternal exposure to adversity, including cumulative, childhood, and preconception exposures, were significantly and positively associated with activation in the right frontopolar prefrontal cortex (PFC) and in the left temporal and parietal clusters during inhibitory control. In addition, and consistent with previous findings, children's increased negative affect and decreased effortful control were associated with increased right PFC activation during inhibitory control. These findings provide preliminary evidence that maternal and dispositional risk factors are linked to alterations in PFC functioning during the preschool years. Children of mothers with a history of exposure to adversity, as well as children who are less temperamentally regulated, may require increased neural resources to meet the cognitive demands of inhibitory control.

  View details for DOI 10.1002/dev.22351

  View details for Web of Science ID 000895767900001

  View details for PubMedID 36567657

• Early prediction and longitudinal modeling of preeclampsia from multiomics. Patterns (New York, N.Y.) Maric, I., Contrepois, K., Moufarrej, M. N., Stelzer, I. A., Feyaerts, D., Han, X., Tang, A., Stanley, N., Wong, R. J., Traber, G. M., Ellenberger, M., Chang, A. L., Fallahzadeh, R., Nassar, H., Becker, M., Xenochristou, M., Espinosa, C., De Francesco, D., Ghaemi, M. S., Costello, E. K., Culos, A., Ling, X. B., Sylvester, K. G., Darmstadt, G. L., Winn, V. D., Shaw, G. M., Relman, D. A., Quake, S. R., Angst, M. S., Snyder, M. P., Stevenson, D. K., Gaudilliere, B., Aghaeepour, N. 2022; 3 (12): 100655

  Abstract

  Preeclampsia is a complex disease of pregnancy whose physiopathology remains unclear. We developed machine-learning models for early prediction of preeclampsia (first 16weeks of pregnancy) and over gestation by analyzing six omics datasets from a longitudinal cohort of pregnant women. For early pregnancy, a prediction model using nine urine metabolites had the highest accuracy and was validated on an independent cohort (area under the receiver-operating characteristic curve [AUC]= 0.88, 95% confidence interval [CI] [0.76, 0.99] cross-validated; AUC= 0.83, 95% CI [0.62,1] validated). Univariate analysis demonstrated statistical significance of identified metabolites. An integrated multiomics model further improved accuracy (AUC= 0.94). Several biological pathways were identified including tryptophan, caffeine, and arachidonic acid metabolisms. Integration with immune cytometry data suggested novel associations between immune and proteomic dynamics. While further validation in a larger population is necessary, these encouraging results can serve as a basis for a simple, early diagnostic test for preeclampsia.

  View details for DOI 10.1016/j.patter.2022.100655

  View details for PubMedID 36569558

• Omics approaches: interactions at the maternal-fetal interface and origins of child health and disease. Pediatric research Ozen, M., Aghaeepour, N., Maric, I., Wong, R. J., Stevenson, D. K., Jantzie, L. L. 2022

  Abstract

  Immunoperinatology is an emerging field. Transdisciplinary efforts by physicians, physician-scientists, basic science researchers, and computational biologists have made substantial advancements by identifying unique immunologic signatures of specific diseases, discovering innovative preventative or treatment strategies, and establishing foundations for individualized neonatal intensive care of the most vulnerable neonates. In this review, we summarize the immunobiology and immunopathology of pregnancy, highlight omics approaches to study the maternal-fetal interface, and their contributions to pregnancy health. We examined the importance of transdisciplinary, multiomic (such as genomics, transcriptomics, proteomics, metabolomics, and immunomics) and machine-learning strategies in unraveling the mechanisms of adverse pregnancy, neonatal, and childhood outcomes and how they can guide the development of novel therapies to improve maternal and neonatal health. IMPACT: Discuss immunoperinatology research from the lens of omics and machine-learning approaches. Identify opportunities for omics-based approaches to delineate infection/inflammation-associated maternal, neonatal, and later life adverse outcomes (e.g., histologic chorioamnionitis [HCA]).

  View details for DOI 10.1038/s41390-022-02335-x

  View details for PubMedID 36216868

• HMOX1 Genetic Polymorphisms Display Ancestral Diversity and May Be Linked to Hypertensive Disorders in Pregnancy. Reproductive sciences (Thousand Oaks, Calif.) Sun, T., Cruz, G. I., Mousavi, N., Maric, I., Brewer, A., Wong, R. J., Aghaeepour, N., Sayed, N., Wu, J. C., Stevenson, D. K., Leonard, S. A., Gymrek, M., Winn, V. D. 2022

  Abstract

  Racial disparity exists for hypertensive disorders in pregnancy (HDP), which leads to disparate morbidity and mortality worldwide. The enzyme heme oxygenase-1 (HO-1) is encoded by HMOX1, which has genetic polymorphisms in its regulatory region that impact its expression and activity and have been associated with various diseases. However, studies of these genetic variants in HDP have been limited. The objective of this study was to examine HMOX1 as a potential genetic contributor of ancestral disparity seen in HDP. First, the 1000 Genomes Project (1KG) phase 3 was utilized to compare the frequencies of alleles, genotypes, and estimated haplotypes of guanidine thymidine repeats (GTn; containing rs3074372) and A/T SNP (rs2071746) among females from five ancestral populations (Africa, theAmericas, Europe, East Asia, and South Asia, N=1271). Then, using genomic DNA from women with a history of HDP, we explored the possibility of HMOX1 variants predisposing women to HDP (N=178) compared with an equivalent ancestral group from 1KG (N=263). Both HMOX1 variants were distributed differently across ancestries, with African women having a distinct distribution and an overall higher prevalence of the variants previously associated with lower HO-1 expression. The two HMOX1 variants display linkage disequilibrium in all but the African group, and within EUR cohort, LL and AA individuals have a higher prevalence in HDP. HMOX1 variants demonstrate ancestral differences that may contribute to racial disparity in HDP. Understanding maternal genetic contribution to HDP will help improve prediction and facilitate personalized approaches to care for HDP.

  View details for DOI 10.1007/s43032-022-01001-1

  View details for PubMedID 35697922

• Bilirubin-induced neurotoxicity and visuocortical dysfunction. Journal of perinatology : official journal of the California Perinatal Association Good, W. V., Wong, R. J., Norcia, A. M., Stevenson, D. K., Slagel, T., Hou, C., Bhutani, V. K. 2022

  View details for DOI 10.1038/s41372-022-01417-2

  View details for PubMedID 35618749

• Prediction of gestational age using urinary metabolites in term and preterm pregnancies. Scientific reports Contrepois, K., Chen, S., Ghaemi, M. S., Wong, R. J., Alliance for Maternal and Newborn Health Improvement (AMANHI), Global Alliance to Prevent Prematurity and Stillbirth (GAPPS), Shaw, G., Stevenson, D. K., Aghaeepour, N., Snyder, M. P., Jehan, F., Sazawal, S., Baqui, A. H., Nisar, M. I., Dhingra, U., Khanam, R., Ilyas, M., Dutta, A., Mehmood, U., Deb, S., Hotwani, A., Ali, S. M., Rahman, S., Nizar, A., Ame, S. M., Muhammad, S., Chauhan, A., Khan, W., Raqib, R., Das, S., Ahmed, S., Hasan, T., Khalid, J., Juma, M. H., Chowdhury, N. H., Kabir, F., Aftab, F., Quaiyum, M. A., Manu, A., Yoshida, S., Bahl, R., Rahman, A., Pervin, J., Price, J. T., Rahman, M., Kasaro, M. P., Litch, J. A., Musonda, P., Vwalika, B., Stringer, J. S. 2022; 12 (1): 8033

  Abstract

  Assessment of gestational age (GA) is key to provide optimal care during pregnancy. However, its accurate determination remains challenging in low- and middle-income countries, where access to obstetric ultrasound is limited. Hence, there is an urgent need to develop clinical approaches that allow accurate and inexpensive estimations of GA. We investigated the ability of urinary metabolites to predict GA at time of collection in a diverse multi-site cohort of healthy and pathological pregnancies (n=99) using a broad-spectrum liquid chromatography coupled with mass spectrometry (LC-MS) platform. Our approach detected a myriad of steroid hormones and their derivatives including estrogens, progesterones, corticosteroids, and androgens which were associated with pregnancy progression. We developed a restricted model that predicted GA with high accuracy using three metabolites (rho=0.87, RMSE=1.58weeks) that was validated in an independent cohort (n=20). The predictions were more robust in pregnancies that went to term in comparison to pregnancies that ended prematurely. Overall, we demonstrated the feasibility of implementing urine metabolomics analysis in large-scale multi-site studies and report a predictive model of GA with a potential clinical value.

  View details for DOI 10.1038/s41598-022-11866-6

  View details for PubMedID 35577875

• Early prediction of preeclampsia in pregnancy with cell-free RNA. Nature Moufarrej, M. N., Vorperian, S. K., Wong, R. J., Campos, A. A., Quaintance, C. C., Sit, R. V., Tan, M., Detweiler, A. M., Mekonen, H., Neff, N. F., Baruch-Gravett, C., Litch, J. A., Druzin, M. L., Winn, V. D., Shaw, G. M., Stevenson, D. K., Quake, S. R. 2022

  Abstract

  Liquid biopsies that measure circulating cell-free RNA (cfRNA) offer an opportunity to study the development of pregnancy-related complications in a non-invasive manner and to bridge gaps in clinical care1-4. Here we used 404 blood samples from 199 pregnant mothers to identify and validate cfRNA transcriptomic changes that are associated with preeclampsia, a multi-organ syndrome that is the second largest cause of maternal death globally5. We find that changes in cfRNA gene expression between normotensive and preeclamptic mothers are marked and stable early in gestation, well before the onset of symptoms. These changes are enriched for genes specific to neuromuscular, endothelial and immune cell types and tissues that reflect key aspects of preeclampsia physiology6-9, suggest new hypotheses for disease progression and correlate with maternal organ health. This enabled the identification and independent validation of a panel of 18 genes that when measured between 5 and 16 weeks of gestation can form the basis of a liquid biopsy test that would identify mothers at risk of preeclampsia long before clinical symptoms manifest themselves. Tests based on these observations could help predict and manage who is at risk for preeclampsia-an important objective for obstetric care10,11.

  View details for DOI 10.1038/s41586-022-04410-z

  View details for PubMedID 35140405

• MULTIOMICS LONGITUDINAL MODELING OF PREECLAMPTIC PREGNANCIES Espinosa, C., Maric, I., Contrepois, K., Moufarrej, M., Stelzer, I. S., Feyaerts, D., Han, X., Tang, A., Wong, R. J., Darmstadt, G. L., Winn, V. D., Shaw, G. M., Relman, D. A., Quake, S. R., Angst, M. S., Snyder, M., Stevenson, D. K., Gaudilliere, B., Aghaeepour, N. BMJ PUBLISHING GROUP. 2022: 309

  View details for DOI 10.1136/jim-2022-WRMC.400

  View details for Web of Science ID 000737295900421

• Revealing the impact of lifestyle stressors on the risk of adverse pregnancy outcomes with multitask machine learning. Frontiers in pediatrics Becker, M., Dai, J., Chang, A. L., Feyaerts, D., Stelzer, I. A., Zhang, M., Berson, E., Saarunya, G., De Francesco, D., Espinosa, C., Kim, Y., Maric, I., Mataraso, S., Payrovnaziri, S. N., Phongpreecha, T., Ravindra, N. G., Shome, S., Tan, Y., Thuraiappah, M., Xue, L., Mayo, J. A., Quaintance, C. C., Laborde, A., King, L. S., Dhabhar, F. S., Gotlib, I. H., Wong, R. J., Angst, M. S., Shaw, G. M., Stevenson, D. K., Gaudilliere, B., Aghaeepour, N. 2022; 10: 933266

  Abstract

  Psychosocial and stress-related factors (PSFs), defined as internal or external stimuli that induce biological changes, are potentially modifiable factors and accessible targets for interventions that are associated with adverse pregnancy outcomes (APOs). Although individual APOs have been shown to be connected to PSFs, they are biologically interconnected, relatively infrequent, and therefore challenging to model. In this context, multi-task machine learning (MML) is an ideal tool for exploring the interconnectedness of APOs on the one hand and building on joint combinatorial outcomes to increase predictive power on the other hand. Additionally, by integrating single cell immunological profiling of underlying biological processes, the effects of stress-based therapeutics may be measurable, facilitating the development of precision medicine approaches.Objectives: The primary objectives were to jointly model multiple APOs and their connection to stress early in pregnancy, and to explore the underlying biology to guide development of accessible and measurable interventions.Materials and Methods: In a prospective cohort study, PSFs were assessed during the first trimester with an extensive self-filled questionnaire for 200 women. We used MML to simultaneously model, and predict APOs (severe preeclampsia, superimposed preeclampsia, gestational diabetes and early gestational age) as well as several risk factors (BMI, diabetes, hypertension) for these patients based on PSFs. Strongly interrelated stressors were categorized to identify potential therapeutic targets. Furthermore, for a subset of 14 women, we modeled the connection of PSFs to the maternal immune system to APOs by building corresponding ML models based on an extensive single cell immune dataset generated by mass cytometry time of flight (CyTOF).Results: Jointly modeling APOs in a MML setting significantly increased modeling capabilities and yielded a highly predictive integrated model of APOs underscoring their interconnectedness. Most APOs were associated with mental health, life stress, and perceived health risks. Biologically, stressors were associated with specific immune characteristics revolving around CD4/CD8 T cells. Immune characteristics predicted based on stress were in turn found to be associated with APOs.Conclusions: Elucidating connections among stress, multiple APOs simultaneously, and immune characteristics has the potential to facilitate the implementation of ML-based, individualized, integrative models of pregnancy in clinical decision making. The modifiable nature of stressors may enable the development of accessible interventions, with success tracked through immune characteristics.

  View details for DOI 10.3389/fped.2022.933266

  View details for PubMedID 36582513

• Maternal stress and its consequences - biological strain. American journal of perinatology Stevenson, D. K., Gotlib, I. H., Buthmann, J. L., Maric, I., Aghaeepour, N., Gaudilliere, B., Angst, M. S., Darmstadt, G. L., Druzin, M. L., Wong, R. J., Shaw, G. M., Katz, M. 2022

  Abstract

  Understanding the role of stress in pregnancy and its consequences is important, particularly given documented associations between maternal stress and preterm birth and other pathologic outcomes. Physical and psychological stressors can elicit the same biological responses, known as biological strain. Chronic stressors, like poverty and racism (race-based discriminatory treatment), may create a legacy or trajectory of biological strain that no amount of coping can relieve in the absence of larger-scale socio-behavioral or societal changes. An integrative approach that takes into consideration simultaneously social and biological determinants of stress may provide the best insights into risk for preterm birth. The most successful computational approaches and the most predictive machine-learning models are likely to be those that combine information about the stressors and the biological strain (for example, as measured by different omics) experienced during pregnancy.

  View details for DOI 10.1055/a-1798-1602

  View details for PubMedID 35292943

• Perinatal infection, inflammation, preterm birth, and brain injury: A review with proposals for future investigations. Experimental neurology Reiss, J. D., Peterson, L. S., Nesamoney, S. N., Chang, A. L., Pasca, A. M., Marić, I., Shaw, G. M., Gaudilliere, B., Wong, R. J., Sylvester, K. G., Bonifacio, S. L., Aghaeepour, N., Gibbs, R. S., Stevenson, D. K. 2022: 113988

  Abstract

  Preterm newborns are exposed to several risk factors for developing brain injury. Clinical studies have suggested that the presence of intrauterine infection is a consistent risk factor for preterm birth and white matter injury. Animal models have confirmed these associations by identifying inflammatory cascades originating at the maternofetal interface that penetrate the fetal blood-brain barrier and result in brain injury. Acquired diseases of prematurity further potentiate the risk for cerebral injury. Systems biology approaches incorporating ante- and post-natal risk factors and analyzing omic and multiomic data using machine learning are promising methodologies for further elucidating biologic mechanisms of fetal and neonatal brain injury.

  View details for DOI 10.1016/j.expneurol.2022.113988

  View details for PubMedID 35081400

• Black swans and ambitious overgeneralization in newborn intensive care. Pediatric research Stevenson, D. K., Wong, R. J., Shaw, G. M., Aghaeepour, N., Maric, I., Prince, L. S., Reiss, J. D., Katz, M. 2021

  View details for DOI 10.1038/s41390-021-01771-5

  View details for PubMedID 34601493

• Integrated trajectories of the maternal metabolome, proteome, and immunome predict labor onset. Science translational medicine Stelzer, I. A., Ghaemi, M. S., Han, X., Ando, K., Hedou, J. J., Feyaerts, D., Peterson, L. S., Rumer, K. K., Tsai, E. S., Ganio, E. A., Gaudilliere, D. K., Tsai, A. S., Choisy, B., Gaigne, L. P., Verdonk, F., Jacobsen, D., Gavasso, S., Traber, G. M., Ellenberger, M., Stanley, N., Becker, M., Culos, A., Fallahzadeh, R., Wong, R. J., Darmstadt, G. L., Druzin, M. L., Winn, V. D., Gibbs, R. S., Ling, X. B., Sylvester, K., Carvalho, B., Snyder, M. P., Shaw, G. M., Stevenson, D. K., Contrepois, K., Angst, M. S., Aghaeepour, N., Gaudilliere, B. 2021; 13 (592)

  Abstract

  Estimating the time of delivery is of high clinical importance because pre- and postterm deviations are associated with complications for the mother and her offspring. However, current estimations are inaccurate. As pregnancy progresses toward labor, major transitions occur in fetomaternal immune, metabolic, and endocrine systems that culminate in birth. The comprehensive characterization of maternal biology that precedes labor is key to understanding these physiological transitions and identifying predictive biomarkers of delivery. Here, a longitudinal study was conducted in 63 women who went into labor spontaneously. More than 7000 plasma analytes and peripheral immune cell responses were analyzed using untargeted mass spectrometry, aptamer-based proteomic technology, and single-cell mass cytometry in serial blood samples collected during the last 100 days of pregnancy. The high-dimensional dataset was integrated into a multiomic model that predicted the time to spontaneous labor [R = 0.85, 95% confidence interval (CI) [0.79 to 0.89], P = 1.2 * 10-40, N = 53, training set; R = 0.81, 95% CI [0.61 to 0.91], P = 3.9 * 10-7, N = 10, independent test set]. Coordinated alterations in maternal metabolome, proteome, and immunome marked a molecular shift from pregnancy maintenance to prelabor biology 2 to 4 weeks before delivery. A surge in steroid hormone metabolites and interleukin-1 receptor type 4 that preceded labor coincided with a switch from immune activation to regulation of inflammatory responses. Our study lays the groundwork for developing blood-based methods for predicting the day of labor, anchored in mechanisms shared in preterm and term pregnancies.

  View details for DOI 10.1126/scitranslmed.abd9898

  View details for PubMedID 33952678

• Understanding how biologic and social determinants affect disparities in preterm birth and outcomes of preterm infants in the NICU. Seminars in perinatology Stevenson, D. K., Aghaeepour, N., Maric, I., Angst, M. S., Darmstadt, G. L., Druzin, M. L., Gaudilliere, B., Ling, X. B., Moufarrej, M. N., Peterson, L. S., Quake, S. R., Relman, D. A., Snyder, M. P., Sylvester, K. G., Shaw, G. M., Wong, R. J. 2021: 151408

  Abstract

  To understand the disparities in spontaneous preterm birth (sPTB) and/or its outcomes, biologic and social determinants as well as healthcare practice (such as those in neonatal intensive care units) should be considered. They have been largely intractable and remain obscure in most cases, despite a myriad of identified risk factors for and causes of sPTB. We still do not know how they might actually affect and lead to the different outcomes at different gestational ages and if they are independent of NICU practices. Here we describe an integrated approach to study the interplay between the genome and exposome, which may drive biochemistry and physiology, with health disparities.

  View details for DOI 10.1016/j.semperi.2021.151408

  View details for PubMedID 33875265

• Proteomic signatures predict preeclampsia in individual cohorts but not across cohorts - implications for clinical biomarker studies. The journal of maternal-fetal & neonatal medicine : the official journal of the European Association of Perinatal Medicine, the Federation of Asia and Oceania Perinatal Societies, the International Society of Perinatal Obstetricians Ghaemi, M. S., Tarca, A. L., Romero, R. n., Stanley, N. n., Fallahzadeh, R. n., Tanada, A. n., Culos, A. n., Ando, K. n., Han, X. n., Blumenfeld, Y. J., Druzin, M. L., El-Sayed, Y. Y., Gibbs, R. S., Winn, V. D., Contrepois, K. n., Ling, X. B., Wong, R. J., Shaw, G. M., Stevenson, D. K., Gaudilliere, B. n., Aghaeepour, N. n., Angst, M. S. 2021: 1–8

  Abstract

  Early identification of pregnant women at risk for preeclampsia (PE) is important, as it will enable targeted interventions ahead of clinical manifestations. The quantitative analyses of plasma proteins feature prominently among molecular approaches used for risk prediction. However, derivation of protein signatures of sufficient predictive power has been challenging. The recent availability of platforms simultaneously assessing over 1000 plasma proteins offers broad examinations of the plasma proteome, which may enable the extraction of proteomic signatures with improved prognostic performance in prenatal care.The primary aim of this study was to examine the generalizability of proteomic signatures predictive of PE in two cohorts of pregnant women whose plasma proteome was interrogated with the same highly multiplexed platform. Establishing generalizability, or lack thereof, is critical to devise strategies facilitating the development of clinically useful predictive tests. A second aim was to examine the generalizability of protein signatures predictive of gestational age (GA) in uncomplicated pregnancies in the same cohorts to contrast physiological and pathological pregnancy outcomes.Serial blood samples were collected during the first, second, and third trimesters in 18 women who developed PE and 18 women with uncomplicated pregnancies (Stanford cohort). The second cohort (Detroit), used for comparative analysis, consisted of 76 women with PE and 90 women with uncomplicated pregnancies. Multivariate analyses were applied to infer predictive and cohort-specific proteomic models, which were then tested in the alternate cohort. Gene ontology (GO) analysis was performed to identify biological processes that were over-represented among top-ranked proteins associated with PE.The model derived in the Stanford cohort was highly significant (p = 3.9E-15) and predictive (AUC = 0.96), but failed validation in the Detroit cohort (p = 9.7E-01, AUC = 0.50). Similarly, the model derived in the Detroit cohort was highly significant (p = 1.0E-21, AUC = 0.73), but failed validation in the Stanford cohort (p = 7.3E-02, AUC = 0.60). By contrast, proteomic models predicting GA were readily validated across the Stanford (p = 1.1E-454, R = 0.92) and Detroit cohorts (p = 1.1.E-92, R = 0.92) indicating that the proteomic assay performed well enough to infer a generalizable model across studied cohorts, which makes it less likely that technical aspects of the assay, including batch effects, accounted for observed differences.Results point to a broader issue relevant for proteomic and other omic discovery studies in patient cohorts suffering from a clinical syndrome, such as PE, driven by heterogeneous pathophysiologies. While novel technologies including highly multiplex proteomic arrays and adapted computational algorithms allow for novel discoveries for a particular study cohort, they may not readily generalize across cohorts. A likely reason is that the prevalence of pathophysiologic processes leading up to the "same" clinical syndrome can be distributed differently in different and smaller-sized cohorts. Signatures derived in individual cohorts may simply capture different facets of the spectrum of pathophysiologic processes driving a syndrome. Our findings have important implications for the design of omic studies of a syndrome like PE. They highlight the need for performing such studies in diverse and well-phenotyped patient populations that are large enough to characterize subsets of patients with shared pathophysiologies to then derive subset-specific signatures of sufficient predictive power.

  View details for DOI 10.1080/14767058.2021.1888915

  View details for PubMedID 33653202

• Mortality Risk Among Patients With COVID-19 Prescribed Selective Serotonin Reuptake Inhibitor Antidepressants. JAMA network open Oskotsky, T., Maric, I., Tang, A., Oskotsky, B., Wong, R. J., Aghaeepour, N., Sirota, M., Stevenson, D. K. 2021; 4 (11): e2133090

  Abstract

  Antidepressant use may be associated with reduced levels of several proinflammatory cytokines suggested to be involved with the development of severe COVID-19. An association between the use of selective serotonin reuptake inhibitors (SSRIs)-specifically fluoxetine hydrochloride and fluvoxamine maleate-with decreased mortality among patients with COVID-19 has been reported in recent studies; however, these studies had limited power due to their small size.To investigate the association of SSRIs with outcomes in patients with COVID-19 by analyzing electronic health records (EHRs).This retrospective cohort study used propensity score matching by demographic characteristics, comorbidities, and medication indication to compare SSRI-treated patients with matched control patients not treated with SSRIs within a large EHR database representing a diverse population of 83 584 patients diagnosed with COVID-19 from January to September 2020 and with a duration of follow-up of as long as 8 months in 87 health care centers across the US.Selective serotonin reuptake inhibitors and specifically (1) fluoxetine, (2) fluoxetine or fluvoxamine, and (3) other SSRIs (ie, not fluoxetine or fluvoxamine).Death.A total of 3401 adult patients with COVID-19 prescribed SSRIs (2033 women [59.8%]; mean [SD] age, 63.8 [18.1] years) were identified, with 470 receiving fluoxetine only (280 women [59.6%]; mean [SD] age, 58.5 [18.1] years), 481 receiving fluoxetine or fluvoxamine (285 women [59.3%]; mean [SD] age, 58.7 [18.0] years), and 2898 receiving other SSRIs (1733 women [59.8%]; mean [SD] age, 64.7 [18.0] years) within a defined time frame. When compared with matched untreated control patients, relative risk (RR) of mortality was reduced among patients prescribed any SSRI (497 of 3401 [14.6%] vs 1130 of 6802 [16.6%]; RR, 0.92 [95% CI, 0.85-0.99]; adjusted P = .03); fluoxetine (46 of 470 [9.8%] vs 937 of 7050 [13.3%]; RR, 0.72 [95% CI, 0.54-0.97]; adjusted P = .03); and fluoxetine or fluvoxamine (48 of 481 [10.0%] vs 956 of 7215 [13.3%]; RR, 0.74 [95% CI, 0.55-0.99]; adjusted P = .04). The association between receiving any SSRI that is not fluoxetine or fluvoxamine and risk of death was not statistically significant (447 of 2898 [15.4%] vs 1474 of 8694 [17.0%]; RR, 0.92 [95% CI, 0.84-1.00]; adjusted P = .06).These results support evidence that SSRIs may be associated with reduced severity of COVID-19 reflected in the reduced RR of mortality. Further research and randomized clinical trials are needed to elucidate the effect of SSRIs generally, or more specifically of fluoxetine and fluvoxamine, on the severity of COVID-19 outcomes.

  View details for DOI 10.1001/jamanetworkopen.2021.33090

  View details for PubMedID 34779847

• The Impact of Hypoxia in Early Pregnancy on Placental Cells. International journal of molecular sciences Zhao, H., Wong, R. J., Stevenson, D. K. 2021; 22 (18)

  Abstract

  Oxygen levels in the placental microenvironment throughout gestation are not constant, with severe hypoxic conditions present during the first trimester. This hypoxic phase overlaps with the most critical stages of placental development, i.e., blastocyst implantation, cytotrophoblast invasion, and spiral artery remodeling initiation. Dysregulation of any of these steps in early gestation can result in pregnancy loss and/or adverse pregnancy outcomes. Hypoxia has been shown to regulate not only the self-renewal, proliferation, and differentiation of trophoblast stem cells and progenitor cells, but also the recruitment, phenotype, and function of maternal immune cells. In this review, we will summarize how oxygen levels in early placental development determine the survival, fate, and function of several important cell types, e.g., trophoblast stem cells, extravillous trophoblasts, syncytiotrophoblasts, uterine natural killer cells, Hofbauer cells, and decidual macrophages. We will also discuss the cellular mechanisms used to cope with low oxygen tensions, such as the induction of hypoxia-inducible factor (HIF) or mammalian target of rapamycin (mTOR) signals, regulation of the metabolic pathway, and adaptation to autophagy. Understanding the beneficial roles of hypoxia in early placental development will provide insights into the root cause(s) of some pregnancy disorders, such as spontaneous abortion, preeclampsia, and intrauterine growth restriction.

  View details for DOI 10.3390/ijms22189675

  View details for PubMedID 34575844

• A Peripheral Immune Signature of Labor Induction. Frontiers in immunology Ando, K., Hédou, J. J., Feyaerts, D., Han, X., Ganio, E. A., Tsai, E. S., Peterson, L. S., Verdonk, F., Tsai, A. S., Marić, I., Wong, R. J., Angst, M. S., Aghaeepour, N., Stevenson, D. K., Blumenfeld, Y. J., Sultan, P., Carvalho, B., Stelzer, I. A., Gaudillière, B. 2021; 12: 725989

  Abstract

  Approximately 1 in 4 pregnant women in the United States undergo labor induction. The onset and establishment of labor, particularly induced labor, is a complex and dynamic process influenced by multiple endocrine, inflammatory, and mechanical factors as well as obstetric and pharmacological interventions. The duration from labor induction to the onset of active labor remains unpredictable. Moreover, prolonged labor is associated with severe complications for the mother and her offspring, most importantly chorioamnionitis, uterine atony, and postpartum hemorrhage. While maternal immune system adaptations that are critical for the maintenance of a healthy pregnancy have been previously characterized, the role of the immune system during the establishment of labor is poorly understood. Understanding maternal immune adaptations during labor initiation can have important ramifications for predicting successful labor induction and labor complications in both induced and spontaneous types of labor. The aim of this study was to characterize labor-associated maternal immune system dynamics from labor induction to the start of active labor. Serial blood samples from fifteen participants were collected immediately prior to labor induction (baseline) and during the latent phase until the start of active labor. Using high-dimensional mass cytometry, a total of 1,059 single-cell immune features were extracted from each sample. A multivariate machine-learning method was employed to characterize the dynamic changes of the maternal immune system after labor induction until the establishment of active labor. A cross-validated linear sparse regression model (least absolute shrinkage and selection operator, LASSO) predicted the minutes since induction of labor with high accuracy (R = 0.86, p = 6.7e-15, RMSE = 277 min). Immune features most informative for the model included STAT5 signaling in central memory CD8+ T cells and pro-inflammatory STAT3 signaling responses across multiple adaptive and innate immune cell subsets. Our study reports a peripheral immune signature of labor induction, and provides important insights into biological mechanisms that may ultimately predict labor induction success as well as complications, thereby facilitating clinical decision-making to improve maternal and fetal well-being.

  View details for DOI 10.3389/fimmu.2021.725989

  View details for PubMedID 34566984

  View details for PubMedCentralID PMC8458888

• Decreased Mortality Rate Among COVID-19 Patients Prescribed Statins: Data From Electronic Health Records in the US. Frontiers in medicine Maric, I., Oskotsky, T., Kosti, I., Le, B., Wong, R. J., Shaw, G. M., Sirota, M., Stevenson, D. K. 2021; 8: 639804

  Abstract

  The severe respiratory illness due to SARS-CoV-2, the virus responsible for coronavirus disease 2019 (COVID-19), is triggered by an intense pro-inflammatory host response. Statins, prescribed primarily for lipid reduction, are known to have anti-inflammatory and immunomodulatory properties and have been associated with a reduced mortality rate among COVID-19 patients taking statins as reported in two recent retrospective studies. However, a meta-analysis that included nine studies showed that statin use did not improve in-hospital outcomes of those with COVID-19. In addition, concerns regarding the use of statins and an increase in COVID-19 infections have been raised, as statins may increase the expression of angiotensin-converting enzyme 2 (ACE2), the primary receptor for the SARS-CoV-2 virus. Our goal was to investigate the effect of statins in COVID-19 patients in a large, diverse patient population across the United States containing nearly 120,000 patients diagnosed with COVID-19. We used propensity score matching of demographics, comorbidities, and medication indication to compare statin-treated patients (N = 2,297) with matched controls (N = 4,594). We observed a small, but statistically significant, decrease in mortality among patients prescribed statins (16.1%) when compared with matched COVID-19-positive controls (18.0 to 20.6%). These results support previous evidence that statins do not increase COVID-19-related mortality and may, in fact, have a mitigating effect on severity of the disease reflected in a slight reduction in mortality. Mixed findings on effects of statins in COVID-19 patients reported in the literature should prompt prospective randomized controlled trials in order to define better who might be advantaged with respect to clinical outcomes.

  View details for DOI 10.3389/fmed.2021.639804

  View details for PubMedID 33614688

• Data-Driven Modeling of Pregnancy-Related Complications. Trends in molecular medicine Espinosa, C. n., Becker, M. n., Marić, I. n., Wong, R. J., Shaw, G. M., Gaudilliere, B. n., Aghaeepour, N. n., Stevenson, D. K. 2021

  Abstract

  A healthy pregnancy depends on complex interrelated biological adaptations involving placentation, maternal immune responses, and hormonal homeostasis. Recent advances in high-throughput technologies have provided access to multiomics biological data that, combined with clinical and social data, can provide a deeper understanding of normal and abnormal pregnancies. Integration of these heterogeneous datasets using state-of-the-art machine-learning methods can enable the prediction of short- and long-term health trajectories for a mother and offspring and the development of treatments to prevent or minimize complications. We review advanced machine-learning methods that could: provide deeper biological insights into a pregnancy not yet unveiled by current methodologies; clarify the etiologies and heterogeneity of pathologies that affect a pregnancy; and suggest the best approaches to address disparities in outcomes affecting vulnerable populations.

  View details for DOI 10.1016/j.molmed.2021.01.007

  View details for PubMedID 33573911

• Maternal metabolic profiling to assess fetal gestational age and predict preterm delivery: a two-centre retrospective cohort study in the US. BMJ open Sylvester, K. G., Hao, S., You, J., Zheng, L., Tian, L., Yao, X., Mo, L., Ladella, S., Wong, R. J., Shaw, G. M., Stevenson, D. K., Cohen, H. J., Whitin, J. C., McElhinney, D. B., Ling, X. B. 2020; 10 (12): e040647

  Abstract

  OBJECTIVES: The aim of this study was to develop a single blood test that could determine gestational age and estimate the risk of preterm birth by measuring serum metabolites. We hypothesised that serial metabolic modelling of serum analytes throughout pregnancy could be used to describe fetal gestational age and project preterm birth with a high degree of precision.STUDY DESIGN: A retrospective cohort study.SETTING: Two medical centres from the USA.PARTICIPANTS: Thirty-six patients (20 full-term, 16 preterm) enrolled at Stanford University were used to develop gestational age and preterm birth risk algorithms, 22 patients (9 full-term, 13 preterm) enrolled at the University of Alabama were used to validate the algorithms.OUTCOME MEASURES: Maternal blood was collected serially throughout pregnancy. Metabolic datasets were generated using mass spectrometry.RESULTS: A model to determine gestational age was developed (R2=0.98) and validated (R2=0.81). 66.7% of the estimates fell within ±1week of ultrasound results during model validation. Significant disruptions from full-term pregnancy metabolic patterns were observed in preterm pregnancies (R2=-0.68). A separate algorithm to predict preterm birth was developed using a set of 10 metabolic pathways that resulted in an area under the curve of 0.96 and 0.92, a sensitivity of 0.88 and 0.86, and a specificity of 0.96 and 0.92 during development and validation testing, respectively.CONCLUSIONS: In this study, metabolic profiling was used to develop and test a model for determining gestational age during full-term pregnancy progression, and to determine risk of preterm birth. With additional patient validation studies, these algorithms may be used to identify at-risk pregnancies prompting alterations in clinical care, and to gain biological insights into the pathophysiology of preterm birth. Metabolic pathway-based pregnancy modelling is a novel modality for investigation and clinical application development.

  View details for DOI 10.1136/bmjopen-2020-040647

  View details for PubMedID 33268420

• A novel point-of-care device for measuring glucose-6-phosphate dehydrogenase enzyme deficiency. Seminars in perinatology Wong, R. J., Montiel, C., Kunda, M., Stevenson, D. K., Bhutani, V. K. 2020: 151356

  Abstract

  Extreme hyperbilirubinemia can cause bilirubin neurotoxicity. Infants with glucose-6-phosphate dehydrogenase (G6PD) deficiency can develop hemolysis and thus are at high risk. We evaluated a device that quantitatively measures G6PD activity kinetically using digital microfluidics (DMF). Intra- and inter-instrument and -day imprecision (CVs) were first assessed. G6PD activity in 86 samples was then measured and compared between DMF and 2 reference methods. Overall DMF reproducibility was 3.8% over 5 days by 2 operators on 2 instruments. Mean intra- and inter-instrument variabilities were 3.6% and 3.9%, respectively (n=28), with a user variability of 4.3%. Mean G6PD activity was 6.40±4.62 and 6.37±4.62U/g hemoglobin for DMF and Reference Methods 1 (n=46) and 12.15±3.86 and 11.48±1.55 for DMF and 2 (n=40), respectively, and strongly correlated (r=0.95 and 0.95) with mean biases of +0.04±2.90 and +0.67±1.55 for methods 1 and 2, respectively. The novel device could be used for early newborn G6PD screening.

  View details for DOI 10.1016/j.semperi.2020.151356

  View details for PubMedID 33293060

• Dysregulation of hypoxia-inducible factor-1alpha (Hif1alpha) expression in the Hmox1-deficient placenta. Placenta Zhao, H., Narasimhan, P., Kalish, F., Wong, R. J., Stevenson, D. K. 2020; 99: 108–16

  Abstract

  INTRODUCTION: Severe hypoxia exists in placentas during early pregnancy, with reoxygenation during mid-gestation. Hypoxia-inducible factor-1alpha (Hif1alpha), an oxygen sensor, initiates placental vascular development. We have shown that the placental vasculature in Hmox1-deficient (Hmox1+/-, Het) pregnancies is impaired, with morphological defects similar to Hif1alpha-deficient placentas.MATERIALS AND METHODS: Whole wild-type (WT) and Het mouse placentas were collected at E8.5 (1%-3% O2) and E9.5-15.5 (8%-10% O2). mRNA levels were determined using real-time RT-PCR or PCR arrays and protein levels using Western blot. Bone marrow-derived macrophages (BMDMs) from WT, Het, and Hmox1 knockout (KO) mice, representing different Hmox1 cellular levels, were generated to study the role of Hmox1 on Hif1alpha 's response to hypoxia-reoxygenation and gestational age-specific placental lysates.RESULTS: Hif1alpha was expressed in WT and Het placentas throughout gestation, with protein levels peaking at E8.5 and mRNA levels significantly upregulated from E9.5-E13.5, but significantly lower in Het placentas. Genes associated with angiogenesis (Vegfa, Vegfr1, Mmp2, Cxcl12, Angpt1, Nos3), antioxidants (Sod1, Gpx1), and transcription factors (Ap2, Bach1, Nrf2) were significantly different in Het placentas. In response to in vitro hypoxia-reoxygenation and to WT or Het placental lysates, Hif1alpha transcription was lower in Het and Hmox1 KO BMDMs compared with WT BMDMs.DISCUSSION: These findings suggest that deficiencies in Hmox1 underlie the insufficient placental Hif1alpha response to hypoxia-reoxygenation during gestation and subsequently impair downstream placental vascular formation. Therefore, a dysregulation of Hif1alpha expression caused by any genetic defect or environmental influence in early pregnancy could be the root cause of pregnancy disorders.

  View details for DOI 10.1016/j.placenta.2020.07.015

  View details for PubMedID 32784053

• Molecular Physiology and Pathophysiology of Bilirubin Handling by the Blood, Liver, Intestine, and Brain in the Newborn. Physiological reviews Hansen, T. W., Wong, R. J., Stevenson, D. K. 2020; 100 (3): 1291–1346

  Abstract

  Bilirubin is the end product of heme catabolism formed during a process that involves oxidation-reduction reactions and conserves iron body stores. Unconjugated hyperbilirubinemia is common in newborn infants, but rare later in life. The basic physiology of bilirubin metabolism, such as production, transport, and excretion, has been well described. However, in the neonate, numerous variables related to nutrition, ethnicity, and genetic variants at several metabolic steps may be superimposed on the normal physiological hyperbilirubinemia that occurs in the first week of life and results in bilirubin levels that may be toxic to the brain. Bilirubin exists in several isomeric forms that differ in their polarities and is considered a physiologically important antioxidant. Here we review the chemistry of the bilirubin molecule and its metabolism in the body with a particular focus on the processes that impact the newborn infant, and how differences relative to older children and adults contribute to the risk of developing both acute and long-term neurological sequelae in the newborn infant. The final section deals with the interplay between the brain and bilirubin and its entry, clearance, and accumulation. We conclude with a discussion of the current state of knowledge regarding the mechanism(s) of bilirubin neurotoxicity.

  View details for DOI 10.1152/physrev.00004.2019

  View details for PubMedID 32401177

• Towards personalized medicine in maternal and child health: integrating biologic and social determinants. Pediatric research Stevenson, D. K., Wong, R. J., Aghaeepour, N., Maric, I., Angst, M. S., Contrepois, K., Darmstadt, G. L., Druzin, M. L., Eisenberg, M. L., Gaudilliere, B., Gibbs, R. S., Gotlib, I. H., Gould, J. B., Lee, H. C., Ling, X. B., Mayo, J. A., Moufarrej, M. N., Quaintance, C. C., Quake, S. R., Relman, D. A., Sirota, M., Snyder, M. P., Sylvester, K. G., Hao, S., Wise, P. H., Shaw, G. M., Katz, M. 2020

  View details for DOI 10.1038/s41390-020-0981-8

  View details for PubMedID 32454518

• Charles E. Ahlfors (1944-2020). Pediatric research Wennberg, R. P., Wong, R. J., Shapiro, S. M., Tiribelli, C. 2020

  View details for DOI 10.1038/s41390-020-0938-y

  View details for PubMedID 32375163

• Cycled Phototherapy Dose-Finding Study for Extremely Low-Birth-Weight Infants: A Randomized Clinical Trial. JAMA pediatrics Arnold, C., Tyson, J. E., Pedroza, C., Carlo, W. F., Stevenson, D. K., Wong, R., Dempsey, A., Khan, A., Fonseca, R., Wyckoff, M., Moreira, A., Lasky, R. 2020

  Abstract

  Importance: Cycled (intermittent) phototherapy (PT) might adequately control peak total serum bilirubin (TSB) level and avoid mortality associated with usual care (continuous PT) among extremely low-birth-weight (ELBW) infants (401-1000 g).Objective: To identify a cycled PT regimen that substantially reduces PT exposure, with an increase in mean peak TSB level lower than 1.5 mg/dL in ELBW infants.Design, Setting, and Participants: This dose-finding randomized clinical trial of cycled PT vs continuous PT among 305 ELBW infants in 6 US newborn intensive care units was conducted from March 12, 2014, to November 14, 2018.Interventions: Two cycled PT regimens (≥15 min/h and ≥30 min/h) were provided using a simple, commercially available timer to titrate PT minutes per hour against TSB level. The comparator arm was usual care (continuous PT).Main Outcomes and Measures: Mean peak TSB level and total PT hours through day 14 in all 6 centers and predischarge brainstem auditory-evoked response wave V latency in 1 center. Mortality and major morbidities were secondary outcomes despite limited power.Results: Consent was requested for 452 eligible infants and obtained for 305 (all enrolled) (mean [SD] birth weight, 749 [152] g; gestational age, 25.7 [1.9] weeks; 81 infants [27%] were multiple births; 137 infants [45%] were male; 112 [37%] were black infants; and 107 [35%] were Hispanic infants). Clinical and demographic characteristics of the groups were similar at baseline. After a preplanned interim analysis of 100 infants, the regimen of 30 min/h or more was discontinued, and the study proceeded with 2 arms. Comparing 128 infants receiving PT of 15 min/h or more with 128 infants receiving continuous PT among those surviving to 14 days, mean peak TSB levels were 7.1 vs 6.4 mg/dL (adjusted difference, 0.7; 95% CI, 0.4-1.1 mg/dL) and mean total PT hours were 34 vs 72 (adjusted difference, -39; 95% CI, -45 to -32). Wave V latency adjusted for postmenstrual age was similar in 37 infants receiving 15 min/h or more of PT and 33 infants receiving continuous PT: 7.42 vs 7.32 milliseconds (difference, 0.10; 95% CI, -0.11 to 0.30 millisecond). The relative risk for death was 0.79 (95% CI, 0.40-1.54), with a risk difference of -4.5% (95% CI, -10.9 to 2.0). Morbidities did not differ between groups.Conclusions and Relevance: Cycled PT can substantially reduce total PT with little increase in peak TSB level. A large, randomized trial is needed to assess whether cycled PT would increase survival and survival without impairment in small, preterm infants.Trial Registration: ClinicalTrials.gov Identifier: NCT01944696.

  View details for DOI 10.1001/jamapediatrics.2020.0559

  View details for PubMedID 32338720

• Cellular Aging in Pregnancy: Telomere Dynamics Across Gestation. Panelli, D. M., Leonard, S. A., Wong, R. J., Girsen, A. I., Baskovic, M., Stevenson, D. K., Bianco, K. SPRINGER HEIDELBERG. 2020: 127A–128A

  View details for Web of Science ID 000525432600181

• Dysregulation of HIF1a Impairs Placental Angiogenesisin Heme Oxygenase-1-Deficient Pregnancies. Zhao, H., Narasimhan, P., Kalish, F., Winn, V. D., Wong, R. J., Stevenson, D. K. SPRINGER HEIDELBERG. 2020: 256A

  View details for Web of Science ID 000525432601139

• Multi-Omic, Longitudinal Profile of Third-Trimester Pregnancies Identifies a Molecular Switch That Predicts the Onset of Labor. Stelzer, I., Ghaemi, M., Han, X., Ando, K., Peterson, L., Contrepois, K., Ganio, E., Tsai, A., Tsai, E., Rumer, K., Stanley, N., Fallazadeh, R., Becker, M., Culos, A., Gaudilliere, D., Wong, R., Winn, V., Shaw, G., Stevenson, D., Snyder, M., Angst, M., Aghaeepour, N., Gaudilliere, B. SPRINGER HEIDELBERG. 2020: 89A

  View details for Web of Science ID 000525432600082

• Multiomic immune clockworks of pregnancy. Seminars in immunopathology Peterson, L. S., Stelzer, I. A., Tsai, A. S., Ghaemi, M. S., Han, X. n., Ando, K. n., Winn, V. D., Martinez, N. R., Contrepois, K. n., Moufarrej, M. N., Quake, S. n., Relman, D. A., Snyder, M. P., Shaw, G. M., Stevenson, D. K., Wong, R. J., Arck, P. n., Angst, M. S., Aghaeepour, N. n., Gaudilliere, B. n. 2020

  Abstract

  Preterm birth is the leading cause of mortality in children under the age of five worldwide. Despite major efforts, we still lack the ability to accurately predict and effectively prevent preterm birth. While multiple factors contribute to preterm labor, dysregulations of immunological adaptations required for the maintenance of a healthy pregnancy is at its pathophysiological core. Consequently, a precise understanding of these chronologically paced immune adaptations and of the biological pacemakers that synchronize the pregnancy "immune clock" is a critical first step towards identifying deviations that are hallmarks of peterm birth. Here, we will review key elements of the fetal, placental, and maternal pacemakers that program the immune clock of pregnancy. We will then emphasize multiomic studies that enable a more integrated view of pregnancy-related immune adaptations. Such multiomic assessments can strengthen the biological plausibility of immunological findings and increase the power of biological signatures predictive of preterm birth.

  View details for DOI 10.1007/s00281-019-00772-1

  View details for PubMedID 32020337

• Multiomics Characterization of Preterm Birth in Low- and Middle-Income Countries. JAMA network open Jehan, F. n., Sazawal, S. n., Baqui, A. H., Nisar, M. I., Dhingra, U. n., Khanam, R. n., Ilyas, M. n., Dutta, A. n., Mitra, D. K., Mehmood, U. n., Deb, S. n., Mahmud, A. n., Hotwani, A. n., Ali, S. M., Rahman, S. n., Nizar, A. n., Ame, S. M., Moin, M. I., Muhammad, S. n., Chauhan, A. n., Begum, N. n., Khan, W. n., Das, S. n., Ahmed, S. n., Hasan, T. n., Khalid, J. n., Rizvi, S. J., Juma, M. H., Chowdhury, N. H., Kabir, F. n., Aftab, F. n., Quaiyum, A. n., Manu, A. n., Yoshida, S. n., Bahl, R. n., Rahman, A. n., Pervin, J. n., Winston, J. n., Musonda, P. n., Stringer, J. S., Litch, J. A., Ghaemi, M. S., Moufarrej, M. N., Contrepois, K. n., Chen, S. n., Stelzer, I. A., Stanley, N. n., Chang, A. L., Hammad, G. B., Wong, R. J., Liu, C. n., Quaintance, C. C., Culos, A. n., Espinosa, C. n., Xenochristou, M. n., Becker, M. n., Fallahzadeh, R. n., Ganio, E. n., Tsai, A. S., Gaudilliere, D. n., Tsai, E. S., Han, X. n., Ando, K. n., Tingle, M. n., Maric, I. n., Wise, P. H., Winn, V. D., Druzin, M. L., Gibbs, R. S., Darmstadt, G. L., Murray, J. C., Shaw, G. M., Stevenson, D. K., Snyder, M. P., Quake, S. R., Angst, M. S., Gaudilliere, B. n., Aghaeepour, N. n. 2020; 3 (12): e2029655

  Abstract

  Worldwide, preterm birth (PTB) is the single largest cause of deaths in the perinatal and neonatal period and is associated with increased morbidity in young children. The cause of PTB is multifactorial, and the development of generalizable biological models may enable early detection and guide therapeutic studies.To investigate the ability of transcriptomics and proteomics profiling of plasma and metabolomics analysis of urine to identify early biological measurements associated with PTB.This diagnostic/prognostic study analyzed plasma and urine samples collected from May 2014 to June 2017 from pregnant women in 5 biorepository cohorts in low- and middle-income countries (LMICs; ie, Matlab, Bangladesh; Lusaka, Zambia; Sylhet, Bangladesh; Karachi, Pakistan; and Pemba, Tanzania). These cohorts were established to study maternal and fetal outcomes and were supported by the Alliance for Maternal and Newborn Health Improvement and the Global Alliance to Prevent Prematurity and Stillbirth biorepositories. Data were analyzed from December 2018 to July 2019.Blood and urine specimens that were collected early during pregnancy (median sampling time of 13.6 weeks of gestation, according to ultrasonography) were processed, stored, and shipped to the laboratories under uniform protocols. Plasma samples were assayed for targeted measurement of proteins and untargeted cell-free ribonucleic acid profiling; urine samples were assayed for metabolites.The PTB phenotype was defined as the delivery of a live infant before completing 37 weeks of gestation.Of the 81 pregnant women included in this study, 39 had PTBs (48.1%) and 42 had term pregnancies (51.9%) (mean [SD] age of 24.8 [5.3] years). Univariate analysis demonstrated functional biological differences across the 5 cohorts. A cohort-adjusted machine learning algorithm was applied to each biological data set, and then a higher-level machine learning modeling combined the results into a final integrative model. The integrated model was more accurate, with an area under the receiver operating characteristic curve (AUROC) of 0.83 (95% CI, 0.72-0.91) compared with the models derived for each independent biological modality (transcriptomics AUROC, 0.73 [95% CI, 0.61-0.83]; metabolomics AUROC, 0.59 [95% CI, 0.47-0.72]; and proteomics AUROC, 0.75 [95% CI, 0.64-0.85]). Primary features associated with PTB included an inflammatory module as well as a metabolomic module measured in urine associated with the glutamine and glutamate metabolism and valine, leucine, and isoleucine biosynthesis pathways.This study found that, in LMICs and high PTB settings, major biological adaptations during term pregnancy follow a generalizable model and the predictive accuracy for PTB was augmented by combining various omics data sets, suggesting that PTB is a condition that manifests within multiple biological systems. These data sets, with machine learning partnerships, may be a key step in developing valuable predictive tests and intervention candidates for preventing PTB.

  View details for DOI 10.1001/jamanetworkopen.2020.29655

  View details for PubMedID 33337494

• Computational discovery of therapeutic candidates for preventing preterm birth. JCI insight Le, B. L., Iwatani, S. n., Wong, R. J., Stevenson, D. K., Sirota, M. n. 2020; 5 (3)

  Abstract

  Few therapeutic methods exist for preventing preterm birth (PTB), or delivery before completing 37 weeks of gestation. In the US, progesterone (P4) supplementation is the only FDA-approved drug for use in preventing recurrent spontaneous PTB. However, P4 has limited effectiveness, working in only approximately one-third of cases. Computational drug repositioning leverages data on existing drugs to discover novel therapeutic uses. We used a rank-based pattern-matching strategy to compare the differential gene expression signature for PTB to differential gene expression drug profiles in the Connectivity Map database and assigned a reversal score to each PTB-drug pair. Eighty-three drugs, including P4, had significantly reversed differential gene expression compared with that found for PTB. Many of these compounds have been evaluated in the context of pregnancy, with 13 belonging to pregnancy category A or B - indicating no known risk in human pregnancy. We focused our validation efforts on lansoprazole, a proton-pump inhibitor, which has a strong reversal score and a good safety profile. We tested lansoprazole in an animal inflammation model using LPS, which showed a significant increase in fetal viability compared with LPS treatment alone. These promising results demonstrate the effectiveness of the computational drug repositioning pipeline to identify compounds that could be effective in preventing PTB.

  View details for DOI 10.1172/jci.insight.133761

  View details for PubMedID 32051340

• Changes in pregnancy-related serum biomarkers early in gestation are associated with later development of preeclampsia. PloS one Hao, S. n., You, J. n., Chen, L. n., Zhao, H. n., Huang, Y. n., Zheng, L. n., Tian, L. n., Maric, I. n., Liu, X. n., Li, T. n., Bianco, Y. K., Winn, V. D., Aghaeepour, N. n., Gaudilliere, B. n., Angst, M. S., Zhou, X. n., Li, Y. M., Mo, L. n., Wong, R. J., Shaw, G. M., Stevenson, D. K., Cohen, H. J., Mcelhinney, D. B., Sylvester, K. G., Ling, X. B. 2020; 15 (3): e0230000

  Abstract

  Placental protein expression plays a crucial role during pregnancy. We hypothesized that: (1) circulating levels of pregnancy-associated, placenta-related proteins throughout gestation reflect the temporal progression of the uncomplicated, full-term pregnancy, and can effectively estimate gestational ages (GAs); and (2) preeclampsia (PE) is associated with disruptions in these protein levels early in gestation; and can identify impending PE. We also compared gestational profiles of proteins in the human and mouse, using pregnant heme oxygenase-1 (HO-1) heterozygote (Het) mice, a mouse model reflecting PE-like symptoms.Serum levels of placenta-related proteins-leptin (LEP), chorionic somatomammotropin hormone like 1 (CSHL1), elabela (ELA), activin A, soluble fms-like tyrosine kinase 1 (sFlt-1), and placental growth factor (PlGF)-were quantified by ELISA in blood serially collected throughout human pregnancies (20 normal subjects with 66 samples, and 20 subjects who developed PE with 61 samples). Multivariate analysis was performed to estimate the GA in normal pregnancy. Mean-squared errors of GA estimations were used to identify impending PE. The human protein profiles were then compared with those in the pregnant HO-1 Het mice.An elastic net-based gestational dating model was developed (R2 = 0.76) and validated (R2 = 0.61) using serum levels of the 6 proteins measured at various GAs from women with normal uncomplicated pregnancies. In women who developed PE, the model was not (R2 = -0.17) associated with GA. Deviations from the model estimations were observed in women who developed PE (P = 0.01). The model developed with 5 proteins (ELA excluded) performed similarly from sera from normal human (R2 = 0.68) and WT mouse (R2 = 0.85) pregnancies. Disruptions of this model were observed in both human PE-associated (R2 = 0.27) and mouse HO-1 Het (R2 = 0.30) pregnancies. LEP outperformed sFlt-1 and PlGF in differentiating impending PE at early human and late mouse GAs.Serum placenta-related protein profiles are temporally regulated throughout normal pregnancies and significantly disrupted in women who develop PE. LEP changes earlier than the well-established biomarkers (sFlt-1 and PlGF). There may be evidence of a causative action of HO-1 deficiency in LEP upregulation in a PE-like murine model.

  View details for DOI 10.1371/journal.pone.0230000

  View details for PubMedID 32126118

• Investigating Pregnancy and Its Complications Using Circulating Cell-Free RNA in Women's Blood During Gestation. Frontiers in pediatrics Moufarrej, M. N., Wong, R. J., Shaw, G. M., Stevenson, D. K., Quake, S. R. 2020; 8: 605219

  Abstract

  In recent years, there have been major advances in the application of non-invasive techniques to predict pregnancy-related complications, for example by measuring cell-free RNA (cfRNA) in maternal blood. In contrast to cell-free DNA (cfDNA), which is already in clinical use to diagnose fetal aneuploidy, circulating RNA levels can correspond with tissue-specific gene expression and provide a snapshot of prenatal health across gestation. Here, we review the physiologic origins of cfRNA and its novel applications and corresponding challenges to monitor fetal and maternal health and predict pregnancy-related complications.

  View details for DOI 10.3389/fped.2020.605219

  View details for PubMedID 33381480

  View details for PubMedCentralID PMC7767905

• VoPo leverages cellular heterogeneity for predictive modeling of single-cell data. Nature communications Stanley, N. n., Stelzer, I. A., Tsai, A. S., Fallahzadeh, R. n., Ganio, E. n., Becker, M. n., Phongpreecha, T. n., Nassar, H. n., Ghaemi, S. n., Maric, I. n., Culos, A. n., Chang, A. L., Xenochristou, M. n., Han, X. n., Espinosa, C. n., Rumer, K. n., Peterson, L. n., Verdonk, F. n., Gaudilliere, D. n., Tsai, E. n., Feyaerts, D. n., Einhaus, J. n., Ando, K. n., Wong, R. J., Obermoser, G. n., Shaw, G. M., Stevenson, D. K., Angst, M. S., Gaudilliere, B. n., Aghaeepour, N. n. 2020; 11 (1): 3738

  Abstract

  High-throughput single-cell analysis technologies produce an abundance of data that is critical for profiling the heterogeneity of cellular systems. We introduce VoPo (https://github.com/stanleyn/VoPo), a machine learning algorithm for predictive modeling and comprehensive visualization of the heterogeneity captured in large single-cell datasets. In three mass cytometry datasets, with the largest measuring hundreds of millions of cells over hundreds of samples, VoPo defines phenotypically and functionally homogeneous cell populations. VoPo further outperforms state-of-the-art machine learning algorithms in classification tasks, and identified immune-correlates of clinically-relevant parameters.

  View details for DOI 10.1038/s41467-020-17569-8

  View details for PubMedID 32719375

• Comments on the 20th Anniversary of NeoReviews. NeoReviews Stevenson, D. K., Wong, R. J., Hay, W. W. 2020; 21 (10): e643–e648

  View details for DOI 10.1542/neo.21-10-e643

  View details for PubMedID 33004557

• TREATMENT WITH LANSOPRAZOLE IMPROVES PREGNANCY OUTCOME IN MICE Iwatani, S., Le, B. L., Konecny, C. M., Vijayakumar, D., Wong, R. T., Sirota, M., Wong, R. J., Stevenson, D. K. BMJ PUBLISHING GROUP. 2020: A149–A150

  View details for DOI 10.1136/jim-2019-WMRC.345

  View details for Web of Science ID 000507513300345

• Increased Carbon Monoxide Washout Rates in Newborn Infants. Neonatology Stevenson, D. K., Wong, R. J., Ostrander, C. R., Maric, I., Vreman, H. J., Cohen, R. S. 2019: 1–5

  Abstract

  BACKGROUND: Endogenous carbon monoxide (CO) production is primarily due to heme degradation, which also results in the equimolar production of bilirubin. Thus, estimates of total body CO production can serve as indices of total body bilirubin formation. The elimination rate of CO from a person's body (CO washout rate) after exposure to an elevated ambient CO concentration is determined by a variety of factors, and is very different between babies and adults.OBJECTIVE: We determined CO washout rates for babies using a simplified technique to measure total body CO excretion rates (VeCO).METHODS: Using a simplified technique, we measured the times to reach an approximate steady state after a change in ambient CO concentration (decay time constant) and CO washout rates in normal newborn infants using non-linear least squares curve fitting.RESULTS: We found a mean CO washout time of 18.7 ± 4.2 min and a CO equilibration (decay time) constant of 0.12 ± 0.04/min (0.08-0.21) for newborn infants.CONCLUSIONS: We confirm that CO washout rates for babies are much faster than those for adults. Therefore, measurements of carboxyhemoglobin (COHb) or end-tidal CO (ETCO), corrected for ambient CO, (COHbc and ETCOc, respectively) can be used as surrogates for VeCO and can provide accurate estimates of endogenous CO (VCO) and bilirubin production rates under normal environmental conditions. Such measurements can be used to identify infants with severe hyperbilirubinemia due to hemolysis and thus at high risk for bilirubin neurotoxicity.

  View details for DOI 10.1159/000503635

  View details for PubMedID 31634890

• Sixty years of phototherapy for neonatal jaundice - from serendipitous observation to standardized treatment and rescue for millions. Journal of perinatology : official journal of the California Perinatal Association Hansen, T. W., Maisels, M. J., Ebbesen, F., Vreman, H. J., Stevenson, D. K., Wong, R. J., Bhutani, V. K. 2019

  Abstract

  A breakthrough discovery 60 years ago by Cremer et al. has since changed the way we treat infants with hyperbilirubinemia and saved the lives of millions from death and disabilities. "Photobiology" has evolved by inquiry of diverse light sources: fluorescent tubes (wavelength range of 400-520nm; halogen spotlights that emit circular footprints of light; fiberoptic pads/blankets (mostly, 400-550nm range) that can be placed in direct contact with skin; and the current narrow-band blue light-emitting diode (LED) light (450-470nm), which overlaps the peak absorption wavelength (458nm) for bilirubin photoisomerization. Excessive bombardment with photons has raised concerns for oxidative stress in very low birthweight versus term infants treated aggressively with phototherapy. Increased emphasis on prescribing phototherapy as a "drug" that is dosed cautiously and judiciously is needed. In this historical review, we chronicled the basic to the neurotoxic components of severe neonatal hyperbilirubinemia and the use of standardized interventions.

  View details for DOI 10.1038/s41372-019-0439-1

  View details for PubMedID 31420582

• Maternal Height and Risk of Preeclampsia among Race/Ethnic Groups AMERICAN JOURNAL OF PERINATOLOGY Maric, I., Mayo, J. A., Druzin, M. L., Wong, R. J., Winn, V. D., Stevenson, D. K., Shaw, G. M. 2019; 36 (8): 864–71

  View details for DOI 10.1055/s-0038-1675205

  View details for Web of Science ID 000474763600013

• Frequency-following response among neonates with progressive moderate hyperbilirubinemia. Journal of perinatology : official journal of the California Perinatal Association Musacchia, G., Hu, J., Bhutani, V. K., Wong, R. J., Tong, M., Han, S., Blevins, N. H., Fitzgerald, M. B. 2019

  Abstract

  OBJECTIVE: To evaluate the feasibility of auditory monitoring of neurophysiological status using frequency-following response (FFR) in neonates with progressive moderate hyperbilirubinemia, measured by transcutaneous (TcB) levels.STUDY DESIGN: ABR and FFR measures were compared and correlated with TcB levels across three groups. Group I was a healthy cohort (n=13). Group II (n=28) consisted of neonates with progressive, moderate hyperbilirubinemia and Group III consisted of the same neonates, post physician-ordered phototherapy.RESULT: FFR amplitudes in Group I controls (TcB=83.1±32.5mol/L; 4.9±1.9mg/dL) were greater than Group II (TcB=209.3±48.0mol/L; 12.1±2.8mg/dL). After TcB was lowered by phototherapy, FFR amplitudes in Group III were similar to controls. Lower TcB levels correlated with larger FFR amplitudes (r=-0.291, p=0.015), but not with ABR wave amplitude or latencies.CONCLUSION: The FFR is a promising measure of the dynamic neurophysiological status in neonates, and may be useful in tracking neurotoxicity in infants with hyperbilirubinemia.

  View details for DOI 10.1038/s41372-019-0421-y

  View details for PubMedID 31263204

• Differential Dynamics of the Maternal Immune System in Healthy Pregnancy and Preeclampsia. Frontiers in immunology Han, X., Ghaemi, M. S., Ando, K., Peterson, L. S., Ganio, E. A., Tsai, A. S., Gaudilliere, D. K., Stelzer, I. A., Einhaus, J., Bertrand, B., Stanley, N., Culos, A., Tanada, A., Hedou, J., Tsai, E. S., Fallahzadeh, R., Wong, R. J., Judy, A. E., Winn, V. D., Druzin, M. L., Blumenfeld, Y. J., Hlatky, M. A., Quaintance, C. C., Gibbs, R. S., Carvalho, B., Shaw, G. M., Stevenson, D. K., Angst, M. S., Aghaeepour, N., Gaudilliere, B. 2019; 10: 1305

  Abstract

  Preeclampsia is one of the most severe pregnancy complications and a leading cause of maternal death. However, early diagnosis of preeclampsia remains a clinical challenge. Alterations in the normal immune adaptations necessary for the maintenance of a healthy pregnancy are central features of preeclampsia. However, prior analyses primarily focused on the static assessment of select immune cell subsets have provided limited information for the prediction of preeclampsia. Here, we used a high-dimensional mass cytometry immunoassay to characterize the dynamic changes of over 370 immune cell features (including cell distribution and functional responses) in maternal blood during healthy and preeclamptic pregnancies. We found a set of eight cell-specific immune features that accurately identified patients well before the clinical diagnosis of preeclampsia (median area under the curve (AUC) 0.91, interquartile range [0.82-0.92]). Several features recapitulated previously known immune dysfunctions in preeclampsia, such as elevated pro-inflammatory innate immune responses early in pregnancy and impaired regulatory T (Treg) cell signaling. The analysis revealed additional novel immune responses that were strongly associated with, and preceded the onset of preeclampsia, notably abnormal STAT5ab signaling dynamics in CD4+T cell subsets (AUC 0.92, p = 8.0E-5). These results provide a global readout of the dynamics of the maternal immune system early in pregnancy and lay the groundwork for identifying clinically-relevant immune dysfunctions for the prediction and prevention of preeclampsia.

  View details for DOI 10.3389/fimmu.2019.01305

  View details for PubMedID 31263463

  View details for PubMedCentralID PMC6584811

• Differential Dynamics of the Maternal Immune System in Healthy Pregnancy and Preeclampsia FRONTIERS IN IMMUNOLOGY Han, X., Ghaemi, M. S., Ando, K., Peterson, L. S., Ganio, E. A., Tsai, A. S., Gaudilliere, D. K., Stelzer, I. A., Einhaus, J., Bertrand, B., Stanley, N., Culos, A., Tanada, A., Hedou, J., Tsai, E. S., Fallahzadeh, R., Wong, R. J., Judy, A. E., Winn, V. D., Druzins, M. L., Blumenfeld, Y. J., Hlatky, M. A., Quaintance, C. C., Gibbs, R. S., Carvalho, B., Shaw, G. M., Stevenson, D. K., Angst, M. S., Aghaeepour, N., Gaudilliere, B. 2019; 10

  View details for DOI 10.3389/fimmu.2019.01305

  View details for Web of Science ID 000470999000001

• A multi-center evaluation of a device for measurement of bilirubin binding capacity in neonates: the effects of gestational age, Intralipid exposure and illness severity JOURNAL OF PERINATOLOGY Schutzman, D. L., Bhutani, V. K., Cuadrado, M., Lamola, A. A., Frantz, I., Obregon, E., Wong, R. J. 2019; 39 (6): 883–88

  View details for DOI 10.1038/s41372-019-0365-2

  View details for Web of Science ID 000468895300016

• The effect of light wavelength on in vitro bilirubin photodegradation and photoisomer production (vol 85, pg 865, 2019) PEDIATRIC RESEARCH Vreman, H. J., Kourula, S., Jasprova, J., Ludvikova, L., Klan, P., Muchova, L., Vitek, L., Cline, B. K., Wong, R. J., Stevenson, D. K. 2019; 85 (6): 905

  View details for DOI 10.1038/s41390-019-0356-1

  View details for Web of Science ID 000466408300029

• The effect of light wavelength on in vitro bilirubin photodegradation and photoisomer production PEDIATRIC RESEARCH Vreman, H. J., Kourula, S., Jasprova, J., Ludvikova, L., Klan, P., Muchova, L., Vitek, L., Cline, B. K., Wong, R. J., Stevenson, D. K. 2019; 85 (6): 865–73

  View details for DOI 10.1038/s41390-019-0310-2

  View details for Web of Science ID 000466408300024

• The effect of light wavelength on in vitro bilirubin photodegradation and photoisomer production. Pediatric research Vreman, H. J., Kourula, S., Jasprova, J., Ludvikova, L., Klan, P., Muchova, L., Vitek, L., Cline, B. K., Wong, R. J., Stevenson, D. K. 2019

  Abstract

  BACKGROUND: The action spectrum for bilirubin photodegradation has been intensively studied. However, questions still remain regarding which light wavelength most efficiently photodegrades bilirubin. In this study, we determined the in vitro effects of different irradiation wavelength ranges on bilirubin photodegradation.METHODS: In our in vitro method, normalized absolute irradiance levels of 4.2*1015photons/cm2/s from light-emitting diodes (ranging from 390-530nm) and 10-nm band-pass filters were used to irradiate bilirubin solutions (25mg/dL in 4% human serum albumin). Bilirubin and its major photoisomer concentrations were determined; the half-life time of bilirubin (t1/2) was calculated for each wavelength range, and the spectral characteristics for bilirubin photodegradation products were obtained for key wavelengths.RESULTS: The in vitro photodegradation of bilirubin at 37°C decreased linearly as the wavelengthwas increased from 390 to 500nm with t1/2 decreasing from 63 to 17min, respectively. At 460±10nm, a significantly lower rate of photodegradation and thus higher t1/2 (31min) than that at 500nm (17min) was demonstrated.CONCLUSION: In our system, the optimum bilirubin photodegradation and lumirubin production rates occurred between 490 and 500nm. Spectra shapes were remarkably similar, suggesting that lumirubin production was the major process of bilirubin photodegradation.

  View details for PubMedID 30710116

• The contributions of genetics to premature birth. Pediatric research Stevenson, D. K., Wong, R. J., Shaw, G. M., Li, J., Wise, P. H., Davis, J. M. 2019

  View details for PubMedID 30644444

• Low-dose lipopolysaccharide exposure can increase in vivo bilirubin production rates in newborn mice. Acta paediatrica (Oslo, Norway : 1992) Iwatani, S. n., Jacobsen, D. R., Wong, R. J., Stevenson, D. K. 2019

  Abstract

  Neonatal hemolysis increases bilirubin production rates, which can then lead to severe hyperbilirubinemia and bilirubin neurotoxicity. During hemolysis, heme is degraded by heme oxygenase (HO), which can be induced under stress conditions. It is known that neonatal sepsis is a risk factor for hemolysis and severe hyperbilirubinemia. Here, we evaluated if an exposure to lipopolysaccharide (LPS) to induce sepsis can upregulate HO-1, further increasing in vivo bilirubin production rates in mouse pups under heme loads.3-day-old pups were given LPS (1250 μg/kg) or saline (controls). Liver HO enzyme activity, HO-1 mRNA, and HO-1 protein were measured post-LPS exposure. We then assessed the effects of LPS treatment on in vivo bilirubin production rates after heme loading.Liver HO activity significantly increased (142%) over controls 24 hours after treatment with LPS (1250 μg/kg) without mortality. Liver HO-1 mRNA was significantly upregulated 2.47-fold at 24 hours post-LPS administration, while liver HO-1 protein increased 1.29-fold 24 hours post-LPS treatment. After heme loading, pups exposed to LPS had significantly higher bilirubin production rates (1.30-fold) compared with age-matched, saline-treated controls.Low-dose LPS treatment can upregulate liver HO-1 expression and may underlie the severe hyperbilirubinemia seen in septic infants, particularly when undergoing hemolysis.

  View details for DOI 10.1111/apa.15143

  View details for PubMedID 31860732

• Data-driven queries between medications and spontaneous preterm birth among 2.5 million pregnancies. Birth defects research Marić, I. n., Winn, V. D., Borisenko, E. n., Weber, K. A., Wong, R. J., Aziz, N. n., Blumenfeld, Y. J., El-Sayed, Y. Y., Stevenson, D. K., Shaw, G. M. 2019

  Abstract

  Our goal was to develop an approach that can systematically identify potential associations between medication prescribed in pregnancy and spontaneous preterm birth (sPTB) by mining large administrative "claims" databases containing hundreds of medications. One such association that we illustrate emerged with antiviral medications used for herpes treatment.IBM MarketScan® databases (2007-2016) were used. A pregnancy cohort was established using International Classification of Diseases (ICD-9/10) codes. Multiple hypothesis testing and the Benjamini-Hochberg procedure that limited false discovery rate at 5% revealed, among 863 medications, five that showed odds ratios (ORs) <1. The statistically strongest was an association between antivirals and sPTB that we illustrate as a real example of our approach, specifically for treatment of genital herpes (GH). Three groups of women were identified based on diagnosis of GH and treatment during the first 36 weeks of pregnancy: (a) GH without treatment; (b) GH treated with antivirals; (c) no GH or treatment.We identified 2,538,255 deliveries. 0.98% women had a diagnosis of GH. Among them, 60.0% received antiviral treatment. Women with treated GH had OR < 1, (OR [95% CI] = 0.91 [0.85, 0.98]). In contrast, women with untreated GH had a small increased risk of sPTB (OR [95% CI] =1.22 [1.14, 1.32]).Data-driven approaches can effectively generate new hypotheses on associations between medications and sPTB. This analysis led us to examine the association with GH treatment. While unknown confounders may impact these findings, our results indicate that women with untreated GH have a modest increased risk of sPTB.

  View details for DOI 10.1002/bdr2.1580

  View details for PubMedID 31433567

• Multiomics modeling of the immunome, transcriptome, microbiome, proteome and metabolome adaptations during human pregnancy BIOINFORMATICS Ghaemi, M., DiGiulio, D. B., Contrepois, K., Callahan, B., Ngo, T. M., Lee-McMullen, B., Lehallier, B., Robaczewska, A., Mcilwain, D., Rosenberg-Hasson, Y., Wong, R. J., Quaintance, C., Culos, A., Stanley, N., Tanada, A., Tsai, A., Gaudilliere, D., Ganio, E., Han, X., Ando, K., McNeil, L., Tingle, M., Wise, P., Maric, I., Sirota, M., Wyss-Coray, T., Winn, V. D., Druzin, M. L., Gibbs, R., Darmstadt, G. L., Lewis, D. B., Nia, V., Agard, B., Tibshirani, R., Nolan, G., Snyder, M. P., Relman, D. A., Quake, S. R., Shaw, G. M., Stevenson, D. K., Angst, M. S., Gaudilliere, B., Aghaeepour, N. 2019; 35 (1): 95–103

  View details for DOI 10.1093/bioinformatics/bty537

  View details for Web of Science ID 000459313900012

• Pravastatin improves fetal survival in mice with a partial deficiency of heme oxygenase-1. Placenta Tsur, A. n., Kalish, F. n., Burgess, J. n., Nayak, N. R., Zhao, H. n., Casey, K. M., Druzin, M. L., Wong, R. J., Stevenson, D. K. 2019; 75: 1–8

  Abstract

  Statins induce heme oxygenase-1 (HO-1) expression in vitro and in vivo. Low HO-1 expression is associated with pregnancy complications, e.g. preeclampsia and recurrent miscarriages. Here, we investigated the effects of pravastatin on HO-1 expression, placental development, and fetal survival in mice with a partial HO-1 deficiency.At E14.5, untreated pregnant wild-type (WT, n=13-18), untreated HO-1+/- (Het, n=6-9), and Het mice treated with pravastatin (Het+Pravastatin, n=12-14) were sacrificed. Numbers of viable fetuses/resorbed concepti were recorded. Maternal livers and placentas were harvested for HO activity. Hematoxylin and eosin (H&E) and CD31 immunohistochemical staining were performed on whole placentas.Compared with WT, HO activity in Het livers (65±18%, P<0.001) and placentas (74±7%, P<0.001) were significantly decreased. Number of viable fetuses per dam was significantly lower in Untreated Het dams (6.0±2.2) compared with WT (9.1±1.4, P<0.01), accompanied by a higher relative risk (RR) for concepti resorption (17.1, 95% CI 4.0-73.2). In Hets treated with pravastatin, maternal liver and placental HO activity increased, approaching levels of WT controls (to 83±7% and 87±14%, respectively). The number of viable fetuses per dam increased to 7.7±2.5 with a decreased RR for concepti resorption (2.7, 95% CI 1.2-5.9). In some surviving Untreated Het placentas, there were focal losses of cellular architecture and changes suggestive of reduced blood flow in the labyrinth. These findings were absent in Het+Pravastatin placentas.Pravastatin induces maternal liver and placental HO activity, may affect placental function and improve fetal survival in the context of a partial deficiency of HO-1.

  View details for PubMedID 30712660

• Multiomics modeling of the immunome, transcriptome, microbiome, proteome and metabolome adaptations during human pregnancy. Bioinformatics (Oxford, England) Ghaemi, M. S., DiGiulio, D. B., Contrepois, K., Callahan, B., Ngo, T. T., Lee-McMullen, B., Lehallier, B., Robaczewska, A., Mcilwain, D., Rosenberg-Hasson, Y., Wong, R. J., Quaintance, C., Culos, A., Stanley, N., Tanada, A., Tsai, A., Gaudilliere, D., Ganio, E., Han, X., Ando, K., McNeil, L., Tingle, M., Wise, P., Maric, I., Sirota, M., Wyss-Coray, T., Winn, V. D., Druzin, M. L., Gibbs, R., Darmstadt, G. L., Lewis, D. B., Partovi Nia, V., Agard, B., Tibshirani, R., Nolan, G., Snyder, M. P., Relman, D. A., Quake, S. R., Shaw, G. M., Stevenson, D. K., Angst, M. S., Gaudilliere, B., Aghaeepour, N. 2019; 35 (1): 95–103

  Abstract

  Motivation: Multiple biological clocks govern a healthy pregnancy. These biological mechanisms produce immunologic, metabolomic, proteomic, genomic and microbiomic adaptations during the course of pregnancy. Modeling the chronology of these adaptations during full-term pregnancy provides the frameworks for future studies examining deviations implicated in pregnancy-related pathologies including preterm birth and preeclampsia.Results: We performed a multiomics analysis of 51 samples from 17 pregnant women, delivering at term. The datasets included measurements from the immunome, transcriptome, microbiome, proteome and metabolome of samples obtained simultaneously from the same patients. Multivariate predictive modeling using the Elastic Net (EN) algorithm was used to measure the ability of each dataset to predict gestational age. Using stacked generalization, these datasets were combined into a single model. This model not only significantly increased predictive power by combining all datasets, but also revealed novel interactions between different biological modalities. Future work includes expansion of the cohort to preterm-enriched populations and in vivo analysis of immune-modulating interventions based on the mechanisms identified.Availability and implementation: Datasets and scripts for reproduction of results are available through: https://nalab.stanford.edu/multiomics-pregnancy/.Supplementary information: Supplementary data are available at Bioinformatics online.

  View details for PubMedID 30561547

• Understanding health disparities. Journal of perinatology : official journal of the California Perinatal Association Stevenson, D. K., Wong, R. J., Aghaeepour, N., Angst, M. S., Darmstadt, G. L., DiGiulio, D. B., Druzin, M. L., Gaudilliere, B., Gibbs, R. S., B Gould, J., Katz, M., Li, J., Moufarrej, M. N., Quaintance, C. C., Quake, S. R., Relman, D. A., Shaw, G. M., Snyder, M. P., Wang, X., Wise, P. H. 2018

  Abstract

  Based upon our recent insights into the determinants of preterm birth, which is the leading cause of death in children under five years of age worldwide, we describe potential analytic frameworks that provides both a common understanding and, ultimately the basis for effective, ameliorative action. Our research on preterm birth serves as an example that the framing of any human health condition is a result of complex interactions between the genome and the exposome. New discoveries of the basic biology of pregnancy, such as the complex immunological and signaling processes that dictate the health and length of gestation, have revealed a complexity in the interactions (current and ancestral) between genetic and environmental forces. Understanding of these relationships may help reduce disparities in preterm birth and guide productive research endeavors and ultimately, effective clinical and public health interventions.

  View details for PubMedID 30560947

• Impact of post-collection freezing delay on the reliability of serum metabolomics in samples reflecting the California mid-term pregnancy biobank. Metabolomics : Official journal of the Metabolomic Society La Frano, M. R., Carmichael, S. L., Ma, C., Hardley, M., Shen, T., Wong, R., Rosales, L., Borkowski, K., Pedersen, T. L., Shaw, G. M., Stevenson, D. K., Fiehn, O., Newman, J. W. 2018; 14 (11): 151

  Abstract

  Population-based biorepositories are important resources, but sample handling can affect data quality.Identify metabolites of value for clinical investigations despite extended postcollection freezing delays, using protocols representing a California mid-term pregnancy biobank.Blood collected from non-pregnant healthy female volunteers (n = 20) underwent three handling protocols after 30 min clotting at room temperature: (1) ideal-samples frozen (- 80 °C) within 2 h of collection; (2) delayed freezing-samples held at room temperature for 3 days, then 4 °C for 9 days, the median times for biobank samples, and then frozen; (3) delayed freezing with freeze-thaw-the delayed freezing protocol with a freeze-thaw cycle simulating retrieved sample sub-aliquoting. Mass spectrometry-based untargeted metabolomic analyses of primary metabolism and complex lipids and targeted profiling of oxylipins, endocannabinoids, ceramides/sphingoid-bases, and bile acids were performed. Metabolite concentrations and intraclass correlation coefficients (ICC) were compared, with the ideal protocol as the reference.Sixty-two percent of 428 identified compounds had good to excellent ICCs, a metric of concordance between measurements of samples handled with the different protocols. Sphingomyelins, phosphatidylcholines, cholesteryl esters, triacylglycerols, bile acids and fatty acid diols were the least affected by non-ideal handling, while sugars, organic acids, amino acids, monoacylglycerols, lysophospholipids, N-acylethanolamides, polyunsaturated fatty acids, and numerous oxylipins were altered by delayed freezing. Freeze-thaw effects were assay-specific with lipids being most stable.Despite extended post-collection freezing delays characteristic of some biobanks of opportunistically collected clinical samples, numerous metabolomic compounds had both stable levels and good concordance.

  View details for DOI 10.1007/s11306-018-1450-9

  View details for PubMedID 30830400

• Enabling precision medicine in neonatology, an integrated repository for preterm birth research. Scientific data Sirota, M., Thomas, C. G., Liu, R., Zuhl, M., Banerjee, P., Wong, R. J., Quaintance, C. C., Leite, R., Chubiz, J., Anderson, R., Chappell, J., Kim, M., Grobman, W., Zhang, G., Rokas, A., England, S. K., Parry, S., Shaw, G. M., Simpson, J. L., Thomson, E., Butte, A. J., March of Dimes Prematurity Research Centers, Driscoll, D., Macones, G., Muglia, L. J., Ober, C., Stevenson, D. K. 2018; 5: 180219

  Abstract

  Preterm birth, or the delivery of an infant prior to 37 weeks of gestation, is a significant cause of infant morbidity and mortality. In the last decade, the advent and continued development of molecular profiling technologies has enabled researchers to generate vast amount of 'omics' data, which together with integrative computational approaches, can help refine the current knowledge about disease mechanisms, diagnostics, and therapeutics. Here we describe the March of Dimes' Database for Preterm Birth Research (http://www.immport.org/resources/mod), a unique resource that contains a variety of 'omics' datasets related to preterm birth. The database is open publicly, and as of January 2018, links 13 molecular studies with data across tens of thousands of patients from 6 measurement modalities. The data in the repository are highly diverse and include genomic, transcriptomic, immunological, and microbiome data. Relevant datasets are augmented with additional molecular characterizations of almost 25,000 biological samples from public databases. We believe our data-sharing efforts will lead to enhanced research collaborations and coordination accelerating the overall pace of discovery in preterm birth research.

  View details for PubMedID 30398470

• Maternal Height and Risk of Preeclampsia among Race/Ethnic Groups. American journal of perinatology Maric, I., Mayo, J. A., Druzin, M. L., Wong, R. J., Winn, V. D., Stevenson, D. K., Shaw, G. M. 2018

  Abstract

  OBJECTIVE: Shorter maternal height has been associated with preeclampsia risk in several populations. It has been less evident whether an independent contribution to the risk exists from maternal height consistently across different races/ethnicities. We investigated associations between maternal height and risk of preeclampsia for different races/ethnicities.STUDY DESIGN: California singleton live births from 2007 to 2011 were analyzed. Logistic regression was used to estimate adjusted odds ratios for the association between height and preeclampsia after stratification by race/ethnicity. To determine the contribution of height that is as independent of body composition as possible, we performed one analysis adjusted for body mass index (BMI) and the other for weight. Additional analyses were performed stratified by parity, and the presence of preexisting/gestational diabetes and autoimmune conditions.RESULTS: Among 2,138,012 deliveries, 3.1% preeclampsia/eclampsia cases were observed. The analysis, adjusted for prepregnancy weight, revealed an inverse relation between maternal height and risk of mild and severe preeclampsia/eclampsia. When the analysis was adjusted for BMI, an inverse relation between maternal height was observed for severe preeclampsia/eclampsia. These associations were observed for each race/ethnicity.CONCLUSION: Using a large and diverse cohort, we demonstrated that shorter height, irrespective of prepregnancy weight or BMI, is associated with an increased risk of severe preeclampsia/eclampsia across different races/ethnicities.

  View details for PubMedID 30396225

• Epigenetic immune cell counting in human blood samples for immunodiagnostics SCIENCE TRANSLATIONAL MEDICINE Baron, U., Werner, J., Schildknecht, K., Schulze, J. J., Mulu, A., Liebert, U., Sack, U., Speckmann, C., Gossen, M., Wong, R. J., Stevenson, D. K., Babel, N., Schuermann, D., Baldinger, T., Bacchetta, R., Gruetzkau, A., Borte, S., Olek, S. 2018; 10 (452)

  Abstract

  Immune cell profiles provide valuable diagnostic information for hematologic and immunologic diseases. Although it is the most widely applied analytical approach, flow cytometry is limited to liquid blood. Moreover, either analysis must be performed with fresh samples or cell integrity needs to be guaranteed during storage and transport. We developed epigenetic real-time quantitative polymerase chain reaction (qPCR) assays for analysis of human leukocyte subpopulations. After method establishment, whole blood from 25 healthy donors and 97 HIV+ patients as well as dried spots from 250 healthy newborns and 24 newborns with primary immunodeficiencies were analyzed. Concordance between flow cytometric and epigenetic data for neutrophils and B, natural killer, CD3+ T, CD8+ T, CD4+ T, and FOXP3+ regulatory T cells was evaluated, demonstrating substantial equivalence between epigenetic qPCR analysis and flow cytometry. Epigenetic qPCR achieves both relative and absolute quantifications. Applied to dried blood spots, epigenetic immune cell quantification was shown to identify newborns suffering from various primary immunodeficiencies. Using epigenetic qPCR not only provides a precise means for immune cell counting in fresh-frozen blood but also extends applicability to dried blood spots. This method could expand the ability for screening immune defects and facilitates diagnostics of unobservantly collected samples, for example, in underdeveloped areas, where logistics are major barriers to screening.

  View details for PubMedID 30068569

• Prethreshold retinopathy of prematurity: VEGF inhibition without VEGF inhibitors. Journal of perinatology : official journal of the California Perinatal Association Gaynon, M. W., Wong, R. J., Stevenson, D. K., Sunshine, P. 2018

  Abstract

  The risk of developing treatment-warranted Type 1 retinopathy of prematurity (ROP) might be reduced in preterm infants by modifying certain systemic factors. There are steps that can be taken both early and late in the course of retinal vascular maturation that may potentially reduce an infant's risk of developing Type 1 ROP. In prethreshold stage 2-3 ROP without plus disease, a combination of supplemental oxygen, correction of severe anemia, and light adaptation to reduce rod photoreceptor oxygen consumption helped us to reduce ROP severity, and encouraged a return to a more physiologic retinal vascular maturation pattern. Thus, it may be possible to reduce the risk of developing Type 1 ROP by making adjustments in certain systemic parameters aimed at reducing retinal hypoxia, thereby gently lowering pathologically elevated levels of vascular endothelial growth factor (VEGF) within the eye.

  View details for PubMedID 30046180

• Bilirubin binding in jaundiced newborns: from bench to bedside? Pediatric research Ahlfors, C. E., Bhutani, V. K., Wong, R. J., Stevenson, D. K. 2018

  Abstract

  BACKGROUND: Bilirubin-induced neurologic dysfunction (BIND) is a spectrum of preventable neurological sequelae in jaundiced newborns. Current total plasma bilirubin (BT) concentration thresholds for phototherapy and/or exchange transfusion poorly predict BIND.METHODS: The unbound (free) bilirubin (Bf) measured at these BT thresholds provides additional information about the risk for BIND. Bf can be readily adapted to clinical use by determining Bf population parameters at current BT thresholds. These parameters can be established using a plasma bilirubin binding panel (BBP) consisting of BT, Bf, and two empiric constants, the maximum BT (BTmax) and the corresponding equilibrium association bilirubin constant (K).RESULTS: BTmax and K provide the variables needed to accurately estimate Bf at BT

  View details for PubMedID 29967530

• Hypoxia regulates placental angiogenesis via alternatively activated macrophages. American journal of reproductive immunology (New York, N.Y. : 1989) Zhao, H., Kalish, F. S., Wong, R. J., Stevenson, D. K. 2018: e12989

  Abstract

  PROBLEM: Uterine and placental macrophages play critical roles in maintaining a normal pregnancy. The majority of these macrophages are believed to be alternatively activated macrophages (M2).METHOD OF STUDY: Mouse bone marrow cells were differentiated into macrophages and polarized to M2 in vitro by treatment with IL-4 [M2a] or IL-10 [M2c] and M1 with LPS/IFN-gamma as controls. Macrophage subtypes were confirmed by surface markers and characterized by gene expression profiles.RESULTS: Compared to M1, M2 showed strong pro-angiogenic activity by expressing higher mRNA for angiogenic-associated factors (Cxcl12, Vegfa, PlGF, Mmp2). M2c produced the highest levels of PlGF, Mmp2, and Cxcr4. To mimic the normal placental microenvironment, M2 were exposed to hypoxia and sex hormones (progesterone, estrogen). In both M2c and M2a, severe hypoxic (1%-3% O2 ) exposure significantly suppressed PlGF, Cxcl12, and Mmp2 mRNA, but not Vegfa, compared to normoxia (21% O2 ) or physiological hypoxia (5% O2 ). mRNA expression returned to normal when hypoxic cells were returned to normoxia. Hypoxia (1%) reduced antioxidant levels in M2 and re-exposure to normoxia significantly increased superoxide dismutase (Sod1, Sod2) and heme oxygenase-1 (HO-1) levels in M2a, and only glutathione peroxidase (Gpx1, Gpx3, Gpx4) in M2c. However, progesterone and estrogen treatment had minimal effects on angiogenic factor expression in M2.CONCLUSION: M2, particularly M2c, displayed strong pro-angiogenic potential, which decreased under severe hypoxia such as in early pregnancy. Thus, conditions that alter the placental oxygen microenvironment or macrophage response in early pregnancy might cause aberrant angiogenesis and vascular remodeling, and lead to abnormal placental vascular development.

  View details for PubMedID 29932269

• Noninvasive blood tests for fetal development predict gestational age and preterm delivery SCIENCE Ngo, T. M., Moufarrej, M. N., Rasmussen, M. H., Camunas-Soler, J., Pan, W., Okamoto, J., Neff, N. F., Liu, K., Wong, R. J., Downes, K., Tibshirani, R., Shaw, G. M., Skotte, L., Stevenson, D. K., Biggio, J. R., Elovitz, M. A., Melbye, M., Quake, S. R. 2018; 360 (6393): 1133–36

  Abstract

  Noninvasive blood tests that provide information about fetal development and gestational age could potentially improve prenatal care. Ultrasound, the current gold standard, is not always affordable in low-resource settings and does not predict spontaneous preterm birth, a leading cause of infant death. In a pilot study of 31 healthy pregnant women, we found that measurement of nine cell-free RNA (cfRNA) transcripts in maternal blood predicted gestational age with comparable accuracy to ultrasound but at substantially lower cost. In a related study of 38 women (23 full-term and 15 preterm deliveries), all at elevated risk of delivering preterm, we identified seven cfRNA transcripts that accurately classified women who delivered preterm up to 2 months in advance of labor. These tests hold promise for prenatal care in both the developed and developing worlds, although they require validation in larger, blinded clinical trials.

  View details for PubMedID 29880692

• Heme oxygenase-1 deficiency promotes severity of sepsis in a non-surgical preterm mouse model. Pediatric research Fujioka, K., Kalish, F., Zhao, H., Wong, R. J., Stevenson, D. K. 2018

  Abstract

  BACKGROUND: Sepsis in preterm infants is associated with systemic inflammatory responses. The stress-response protein heme oxygenase-1 (HO-1) has protective anti-inflammatory properties. Recently, we reported a protective role of HO-1 using our non-surgical cecal slurry (CS) model in wild-type (WT) mouse pups. Here, we extend these findings to investigate the association of HO-1 deficiency with sepsis severity.METHODS: Adapting the Wynn model, we induced sepsis in 4-day-old HO-1-deficient (HO-1+/-, Het) pups to determine if HO-1 deficiency affected survival rates at the LD40 (2.0mg/g) of WT pups. To see if HO-1 induction affected sepsis severity, we gave 30-mumol heme/kg subcutaneously to 3-day-old mice 24h prior to sepsis induction.RESULTS: Post-sepsis induction, Het pups had a mortality of 85.0% (n=20) and increased expression of the pro-inflammatory gene in the livers and affected hematologic profiles. Heme treatment 24h prior to sepsis induction significantly increased liver HO activity, reduced mortality to 24.5% (n=17), attenuated inflammatory responses, reduced spleen bacterial counts, and significantly increased peripheral neutrophils.CONCLUSIONS: A partial deficiency in HO-1 increased the progression and mortality in sepsis. Furthermore, induction of HO-1 significantly reduced the mortality even in Het pups. Thus, we conclude that HO-1 plays an important role in the protection against preterm sepsis.

  View details for PubMedID 29795214

• Neuro-inflammatory effects of photodegradative products of bilirubin SCIENTIFIC REPORTS Jasprova, J., Dal Ben, M., Hurny, D., Hwang, S., Zizalova, K., Kotek, J., Wong, R. J., Stevenson, D. K., Gazzin, S., Tiribelli, C., Vitek, L. 2018; 8: 7444

  Abstract

  Phototherapy was introduced in the early 1950's, and is the primary treatment of severe neonatal jaundice or Crigler-Najjar syndrome. Nevertheless, the potential biological effects of the products generated from the photodegradation of bilirubin during phototherapy remain unknown. This is very relevant in light of recent clinical observations demonstrating that the use of aggressive phototherapy can increase morbidity or even mortality, in extremely low birthweight (ELBW) infants. The aim of our study was to investigate the effects of bilirubin, lumirubin (LR, its major photo-oxidative product), and BOX A and B (its monopyrrolic oxidative products) on the central nervous system (CNS) using in vitro and ex vivo experimental models. The effects of bilirubin photoproducts on cell viability and expression of selected genes were tested in human fibroblasts, three human CNS cell lines (neuroblastoma SH-SY5Y, microglial HMC3, and glioblastoma U-87 cell lines), and organotypic rat hippocampal slices. Neither bilirubin nor its photo-oxidative products affected cell viability in any of our models. In contrast, LR in biologically-relevant concentrations (25 μM) significantly increased gene expression of several pro-inflammatory genes as well as production of TNF-α in organotypic rat hippocampal slices. These findings might underlie the adverse outcomes observed in ELBW infants undergoing aggressive phototherapy.

  View details for PubMedID 29748620

• Heme oxygenase-1 deficiency results in splenic T-cell dysregulation in offspring of mothers exposed to late gestational inflammation AMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY Ozen, M., Zhao, H., Kalish, F., Yang, Y., Folkins, A., Burd, I., Wong, R. J., Stevenson, D. K. 2018; 79 (5): e12829

  Abstract

  Infection during pregnancy can disrupt regulatory/effector immune system balance, resulting in adverse pregnancy and fetal-neonatal outcomes. Heme oxygenase-1 (HO-1) is a major regulatory enzyme in the immune system. We observed maternal immune response dysregulation during late gestational inflammation (LGI), which may be mediated by HO-1. Here, we extend these studies to examine the immune response of offspring.Pregnant wild-type (Wt) and HO-1 heterozygote (Het) dams were treated with lipopolysaccharide (LPS) or vehicle at E15.5. Pups' splenic immune cells were characterized using flow cytometry.CD3+ CD4+ CD25+ (Tregs) and CD3+ CD8+ (Teffs) T cells in Wt and Het were similar in control neonates and increased with age. We showed not only age- but also genotype-specific and long-lasting T-cell dysregulation in pups after maternal LGI. The persistent immune dysregulation, mediated by HO-1 deficiency, was reflected as a decrease in Treg FoxP3 and CD3+ CD8+ T cells, and an increase in CD4+ /CD8+ T-cell and Treg/Teff ratios in Hets compared with Wt juvenile mice after maternal exposure to LGI.Maternal exposure to LGI can result in dysregulation of splenic T cells in offspring, especially in those with HO-1 deficiency. We speculate that these immune alterations are the basis of adverse outcomes in neonates from mothers exposed to low-grade (subclinical) infections.

  View details for PubMedID 29484761

• Heme oxygenase-1 genetic variants and the conundrum of hyperbilirubinemia in African-American newborns JOURNAL OF PERINATOLOGY Schutzman, D. L., Gatien, E., Ajayi, S., Wong, R. J. 2018; 38 (4): 345–50

  Abstract

  African-American (AA) infants are known to have, overall, lower bilirubin levels than infants of other ethnicities during their birth hospitalization. However, they are known to have a higher incidence of severe hyperbilirubinemia and are over represented in the US Kernicterus Registry. Heme oxygenase-1 (HO) is the rate limiting enzyme in heme metabolism leading to the equimolar production of bilirubin, carbon monoxide (CO) and free iron (Fe). Short (S) (GT)n repeats (<25) in the promoter region of the gene encoding the inducible HO-1 isozyme augment its expression, while long (L) repeats (>33) lead to an attenuation, modulating the production of bilirubin and CO. The impact of HO-1 promoter polymorphisms on bilirubin levels has not been well studied in (AA) infants.The objectives of this study were to compare the distribution of (GT)n repeat lengths in the HO-1 promoter region in a cohort of AA infants to those found in other ethnicities and to evaluate the contribution of this polymorphism to the degree of hyperbilirubinemia and the level of COHbc in this cohort.We prospectively studied a cohort of term AA infants with O+ mothers. Per hospital routine, infants' blood type, Rh status, direct antiglobulin test (DAT), and total bilirubin (TB) levels were checked prior to discharge. After parental consent, blood was collected for DNA extraction and carboxyhemoglobin (COHbc) measurements at the same time as the infants' newborn screen. An infant's TB percentile risk based on the Bhutani nomogram was used to determine need for phototherapy or follow-up. (GT)n repeat length in the HO-1 promoter was determined for each allele using PCR after DNA extraction from dried bloodspots. Size of allele lengths were typed as short (S, <25), medium (M, 25-33) or long (L, >33).One hundred eighty infants were studied for a total of 360 separate alleles. 12.2% (44/360) of alleles were S which was significantly less than all other ethnicities reviewed. Carboxyhemoglobin (COHbc) levels and bilirubin percentiles were higher among infants who had at least one S allele when compared to those who had at least one L allele in the cohort as a whole: COHbc 0.92 ± 0.35 vs. 0.85 ± 0.37; p = 0.28 and Bilirubin percentile 48.6 ± 34.0 vs. 44.9 ± 31.6; p = 0.51. This relationship remained when only those infants who were DAT neg were examined: COHbc 0.81 ± 0.26 vs. 0.74 ± 0.21; p = 0.11 and Bilirubin percentile 43.6 ± 29.9 vs. 37.5 ± 28.7; p = 0.28.The presence of L alleles of this variant is significantly greater among infants who are either African or of African descent. There was a trend toward lower COHbc levels among infants with at least one L allele as opposed to at least one S allele, although this did not have a statistically significant impact on TB risk percentile.

  View details for PubMedID 29302043

• Identification of risk for neonatal haemolysis. Acta paediatrica (Oslo, Norway : 1992) Bhutani, V. K., Maisels, M. J., Schutzman, D. L., Castillo Cuadrado, M. E., Aby, J. L., Bogen, D. L., Christensen, R. D., Watchko, J. F., Wong, R. J., Stevenson, D. K. 2018

  Abstract

  AIM: To identify neonates at risk of haemolytic hyperbilirubinaemia through near-concurrent measurements of total serum/plasma bilirubin (TB) or transcutaneous bilirubin (TcB) and end-tidal breath carbon monoxide (CO), corrected for ambient CO (ETCOc), an index of bilirubin production and haemolysis.METHODS: Paired TB/TcB (mg/dL) and ETCOc (ppm) measurements were obtained in newborns (n = 283) at 20 to <60 hours of age in five nurseries. TB/TcB values were assigned TB/TcB percentile risk values using the Bhutani hour-specific nomogram. In infants having two serial TB/TcB measurements (n = 76), TB rate of rise (ROR, mg/dL/h) was calculated.RESULTS: For the entire cohort (n = 283), 67.1% and 32.9% had TB/TcB<75th and ≥75th percentile, respectively. TB/TcB (5.79 ± 1.84 vs 9.14 ± 2.25 mg/dL) and ETCOc (1.61 ± 0.45 vs 2.02 ± 1.35 ppm, p = 0.0002) were different between the groups. About 36.6% of infants with TB/TcB ≥75th percentile had ETCOc ≥ 2.0 ppm. In the subcohort of infants with serial TB/TcB measurements (n = 76), 44.7% and 55.3% had TB/TcB<75th and ≥75th percentile, respectively. TB/TcB (5.28 ± 1.97 vs 9.53 ± 2.78 mg/dL), ETCOc (1.72 ± 0.48 vs 2.38 ± 1.89 ppm, p = 0.05) and TB ROR (0.011 ± 0.440 vs 0.172 ± 0.471 mg/dL/h) were different between the groups.CONCLUSION: The combined use of TB/TcB percentile risk assessments and ETCOc measurements can identify infants with haemolytic hyperbilirubinaemia. The addition of TB ROR can identify those infants with elimination disorders.

  View details for PubMedID 29532503

• A genome-wide association study identifies only two ancestry specific variants associated with spontaneous preterm birth SCIENTIFIC REPORTS Rappoport, N., Toung, J., Hadley, D., Wong, R. J., Fujioka, K., Reuter, J., Abbott, C. W., Oh, S., Hu, D., Eng, C., Huntsman, S., Bodian, D. L., Niederhuber, J. E., Hong, X., Zhang, G., Sikora-Wohfeld, W., Gignoux, C. R., Wang, H., Oehlert, J., Jelliffe-Pawlowski, L. L., Gould, J. B., Darmstadt, G. L., Wang, X., Bustamante, C. D., Snyder, M. P., Ziv, E., Patsopoulos, N. A., Muglia, L. J., Burchard, E., Shaw, G. M., O'Brodovich, H. M., Stevenson, D. K., Butte, A. J., Sirota, M. 2018; 8: 226

  Abstract

  Preterm birth (PTB), or the delivery prior to 37 weeks of gestation, is a significant cause of infant morbidity and mortality. Although twin studies estimate that maternal genetic contributions account for approximately 30% of the incidence of PTB, and other studies reported fetal gene polymorphism association, to date no consistent associations have been identified. In this study, we performed the largest reported genome-wide association study analysis on 1,349 cases of PTB and 12,595 ancestry-matched controls from the focusing on genomic fetal signals. We tested over 2 million single nucleotide polymorphisms (SNPs) for associations with PTB across five subpopulations: African (AFR), the Americas (AMR), European, South Asian, and East Asian. We identified only two intergenic loci associated with PTB at a genome-wide level of significance: rs17591250 (P = 4.55E-09) on chromosome 1 in the AFR population and rs1979081 (P = 3.72E-08) on chromosome 8 in the AMR group. We have queried several existing replication cohorts and found no support of these associations. We conclude that the fetal genetic contribution to PTB is unlikely due to single common genetic variant, but could be explained by interactions of multiple common variants, or of rare variants affected by environmental influences, all not detectable using a GWAS alone.

  View details for PubMedID 29317701

• Hemolysis and Glucose-6-Phosphate Dehydrogenase Deficiency-Related Neonatal Hyperbilirubinemia NEONATOLOGY Kaplan, M., Wong, R. J., Stevenson, D. K. 2018; 114 (3): 223–25

  Abstract

  Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a common enzyme deficiency affecting more than 300 million individuals worldwide. Extreme neonatal hyperbilirubinemia, with its severe sequelae of bilirubin neurotoxicity and the potential of death, is the most devastating manifestation of G6PD deficiency. In a recent review of Favism, Luzzatto and Arese state that the pathophysiology of jaundice in G6PD-deficient neonates is different from that of favism, as there is little evidence of hemolysis in these infants.To explore the role of hemolysis in neonatal hyperbilirubinemia associated with G6PD deficiency.Previously published works including studies of endogenous production of carbon monoxide (CO), an index of heme catabolism, in hyperbilirubinemic G6PD-deficient neonates were reviewed to determine the role of hemolysis in this condition.Three studies demonstrated that endogenous CO production is elevated in G6PD-deficient neonates with extreme hyperbilirubinemia.Hemolysis is an important pathogenetic factor in G6PD deficiency-associated neonatal hyperbilirubinemia.

  View details for PubMedID 29940590

• Simultaneously Monitoring Immune Response and Microbial Infections during Pregnancy through Plasma cfRNA Sequencing CLINICAL CHEMISTRY Pan, W., Ngo, T. M., Camunas-Soler, J., Song, C., Kowarsky, M., Blumenfeld, Y. J., Wong, R. J., Shaw, G. M., Stevenson, D. K., Quake, S. R. 2017; 63 (11): 1695–1704

  Abstract

  Plasma cell-free RNA (cfRNA) encompasses a broad spectrum of RNA species that can be derived from both human cells and microbes. Because cfRNA is fragmented and of low concentration, it has been challenging to profile its transcriptome using standard RNA-seq methods.We assessed several recently developed RNA-seq methods on cfRNA samples. We then analyzed the dynamic changes of both the human transcriptome and the microbiome of plasma during pregnancy from 60 women.cfRNA reflects a well-orchestrated immune modulation during pregnancy: an up-regulation of antiinflammatory genes and an increased abundance of antimicrobial genes. We observed that the plasma microbiome remained relatively stable during pregnancy. The bacteria Ureaplasma shows an increased prevalence and increased abundance at postpartum, which is likely to be associated with postpartum infection. We demonstrated that cfRNA-seq can be used to monitor viral infections. We detected a number of human pathogens in our patients, including an undiagnosed patient with a high load of human parvovirus B19 virus (B19V), which is known to be a potential cause of complications in pregnancy.Plasma cfRNA-seq demonstrates the potential to simultaneously monitor immune response and microbial infections during pregnancy.

  View details for PubMedID 28904056

• Numerous uncharacterized and highly divergent microbes which colonize humans are revealed by circulating cell-free DNA PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Kowarsky, M., Camunas-Soler, J., Kertesz, M., De Vlaminck, I., Koh, W., Pan, W., Martin, L., Neff, N. F., Okamoto, J., Wong, R. J., Kharbanda, S., El-Sayed, Y., Blumenfeld, Y., Stevenson, D. K., Shaw, G. M., Wolfe, N. D., Quake, S. R. 2017; 114 (36): 9623–28

  Abstract

  Blood circulates throughout the human body and contains molecules drawn from virtually every tissue, including the microbes and viruses which colonize the body. Through massive shotgun sequencing of circulating cell-free DNA from the blood, we identified hundreds of new bacteria and viruses which represent previously unidentified members of the human microbiome. Analyzing cumulative sequence data from 1,351 blood samples collected from 188 patients enabled us to assemble 7,190 contiguous regions (contigs) larger than 1 kbp, of which 3,761 are novel with little or no sequence homology in any existing databases. The vast majority of these novel contigs possess coding sequences, and we have validated their existence both by finding their presence in independent experiments and by performing direct PCR amplification. When their nearest neighbors are located in the tree of life, many of the organisms represent entirely novel taxa, showing that microbial diversity within the human body is substantially broader than previously appreciated.

  View details for PubMedID 28830999

• Heme oxygenase-1 promoter polymorphisms: do they modulate neonatal hyperbilirubinemia? JOURNAL OF PERINATOLOGY Kaplan, M., Wong, R. J., Stevenson, D. K. 2017; 37 (8): 901–5

  Abstract

  The role of genetic factors in the modulation of serum bilirubin levels and the pathophysiology of neonatal hyperbilirubinemia is being increasingly recognized. Heme oxygenase-1 (HO-1) is the rate-limiting enzyme by which heme is catabolized to biliverdin and thence to bilirubin, with the simultaneous release of equimolar quantities of ferrous iron (Fe3+) and carbon monoxide. Polymorphisms of the HO-1 gene promoter may modulate transcriptional activity, thereby augmenting or attenuating HO-1 expression with resultant modulation of the production of bilirubin. Few studies have related these polymorphisms to neonatal bilirubin metabolism and have reported conflicting results. In this clinical review, we surveyed the role of HO-1 gene promoter polymorphisms in the control of bilirubin production and further considered their role, if any, in the pathophysiology of neonatal hyperbilirubinemia.

  View details for PubMedID 28206992

• Effect of genetic variants of bilirubin metabolism on the degree of hyperbilirubinemia in African-American newborns. Journal of perinatology : official journal of the California Perinatal Association Schutzman, D. L., Baudhuin, L. M., Gatien, E., Ajayi, S., Wong, R. J. 2017; 37 (4): 432-435

  Abstract

  The objective of our study was to measure the effect of genetic variants of these two enzymes, UGT1A1 and SLCO1B1, in the bilirubin metabolic pathway on the degree of hyperbilirubinemia in a cohort of African-American (AA) infants from our well-baby nursery. In addition, a second objective was to document the types and frequencies of genetic variations of these enzymes in our cohort.A prospective study of 180 AA infants from the Well Baby Nursery of an inner city community hospital, all of whose mothers were type O pos. Sixty infants were ABO-incompatible direct antiglobulin test (DAT) pos, 60 were ABO-incompatible DAT neg and 60 were type O+. Blood for carboxyhemoglobin (COHb) and variants of the enzymes uridine diphosphoglucuronosyltransferase 1A1 and hepatic solute carrier organic anion transporter 1B1 (SLCO1B1) was drawn at the time of the infants' initial bilirubin, and the infants' precise percentile on the Bhutani nomogram was calculated.Variants in the two enzymes studied were quite common. In total, 21.1% were positive for a Gilbert phenotype, whereas an additional 42.4% were heterozygous for the *28 or *37 variant of UGT1A1. In total, 67.2% were homozygous for the *60 variant of the phenobarbital responsive enhancer module. In total, 41.1% were homozygous for the *1b variant of SLCO1B1, whereas an additional 12.7% were positive for the *4 variant of this gene. In total, 20.6% of infants had variations in both genes. Using logistic regression when COHbc was assessed with each of the different variants, only COHbc (P<0.0001 to 0.0004) was significantly associated with the level of hyperbilirubinemia as defined by the Bhutani nomogram.Although we have found quite a large number of genetic variants of the UGT1A1 and SLCO1B1 genes in the AA population, it does not appear that they have a significant impact on the incidence of hyperbilirubinemia among this group of infants.

  View details for DOI 10.1038/jp.2016.232

  View details for PubMedID 27977017

• Effects of Remote Ischemic Preconditioning on Heme Oxygenase-1 Expression and Cutaneous Wound Repair INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Cremers, N. A., Wever, K. E., Wong, R. J., van Rheden, R. E., Vermeij, E. A., van Dam, G. M., Carels, C. E., Lundvig, D. M., Wagener, F. A. 2017; 18 (2)

  Abstract

  Skin wounds may lead to scar formation and impaired functionality. Remote ischemic preconditioning (RIPC) can induce the anti-inflammatory enzyme heme oxygenase-1 (HO-1) and protect against tissue injury. We aim to improve cutaneous wound repair by RIPC treatment via induction of HO-1. RIPC was applied to HO-1-luc transgenic mice and HO-1 promoter activity and mRNA expression in skin and several other organs were determined in real-time. In parallel, RIPC was applied directly or 24h prior to excisional wounding in mice to investigate the early and late protective effects of RIPC on cutaneous wound repair, respectively. HO-1 promoter activity was significantly induced on the dorsal side and locally in the kidneys following RIPC treatment. Next, we investigated the origin of this RIPC-induced HO-1 promoter activity and demonstrated increased mRNA in the ligated muscle, heart and kidneys, but not in the skin. RIPC did not change HO-1 mRNA and protein levels in the wound 7 days after cutaneous injury. Both early and late RIPC did not accelerate wound closure nor affect collagen deposition. RIPC induces HO-1 expression in several organs, but not the skin, and did not improve excisional wound repair, suggesting that the skin is insensitive to RIPC-mediated protection.

  View details for DOI 10.3390/ijms18020438

  View details for Web of Science ID 000395457700206

  View details for PubMedID 28218659

• INDUCTION OF HEME OXYGENASE-1 ATTENUATES THE SEVERITY OF SEPSIS IN A NON-SURGICAL PRETERM MOUSE MODEL SHOCK Fujioka, K., Kalish, F., Zhao, H., Lu, S., Wong, S., Wong, R. J., Stevenson, D. K. 2017; 47 (2): 242-250

  Abstract

  Preterm sepsis is characterized by systemic bacterial invasion and inflammatory response. Its pathogenesis is unclear due to lack of proper animal models. Heme oxygenase-1 (HO-1) can affect physiologic and pathologic conditions through its anti-inflammatory, antioxidative, and anti-apoptotic properties. Since HO-1 is developmentally regulated, it may play a role in the pathogenesis of preterm sepsis. For this study, sepsis was induced using the non-surgical "cecal slurry" (CS) model. CS was given intraperitoneally at various doses to 4-day-old newborn mice to determine dose-dependent effects. The LD40 was then given and changes in bodyweight, bacterial colonization of organs, hematology, serum biochemistry, and immunomodulatory gene expression were determined. We found a dose-dependent mortality with an LD40 of 2.0 mg/g. Significant bacterial colonization and hematological changes (leukocytopenia, thrombocytopenia, and lymphocytopenia) and increased gene expression of pro-inflammatory cytokines, pattern-recognition receptors, and other genes related to immune responses were also observed. Twenty-four hours post-sepsis induction, bodyweight loss was associated with mortality and organ damage. Finally, to elucidate a protective role of HO-1, 30-μmol heme/kg was given subcutaneously 24 h pre-sepsis induction. HO activity in livers and spleens significantly increased 64% and 50% over age-matched controls 24 h post-heme administration. Importantly, heme significantly reduced mortality from 40.9% to 6.3% (P <0.005) and gene expression of pro-inflammatory cytokines (Ccl5, Cxcl10, IL-1b, and Ifng). We conclude that the CS model can be used as a model to study preterm sepsis. Because induction of HO-1 significantly reduced mortality, we speculate that HO-1 may confer protection against sepsis in preterm infants.

  View details for DOI 10.1097/SHK.0000000000000689

  View details for PubMedID 27454382

• Infiltration of myeloid cells in the pregnant uterus is affected by heme oxygenase-1 JOURNAL OF LEUKOCYTE BIOLOGY Zhao, H., Kalish, F., Wong, R. J., Stevenson, D. K. 2017; 101 (1): 217-226

  Abstract

  Infiltrating myeloid cells in pregnant uteri play critical roles in the establishment of the placenta and maintenance of normal pregnancies. Their recruitment and proliferation are primarily mediated by the interactions of cytokines and chemokines secreted locally with their corresponding receptors. Heme oxygenase-1 (HO-1) has various physiologic properties that contribute to placental vascular development, with deficiencies in HO-1 associated with pregnancy disorders. Here, we investigated the effect of HO-1 on myeloid cell infiltration into pregnant uteri using a partial HO-1-deficient (Het, HO-1(+/-)) mouse model. With the use of flow cytometry, HO-1 was found predominantly expressed in circulating and uterine myeloid cells, specifically neutrophils and monocytes/macrophages. In pregnant Het uteri, the numbers of neutrophils and monocytes/macrophages were significantly reduced compared with pregnant wild-type (WT; HO-1(+/+)) uteri. With the use of BrdU in vivo assays, HO-1 deficiency did not affect cell proliferation or blood cell populations. With the use of PCR arrays, gene expression of cytokines (Csf1, Csf3), chemokines (Ccl1, Ccl2, Ccl6, Ccl8, Ccl11, Ccl12, Cxcl4, Cxcl9, Cxcl12), and their receptors (Ccr1, Ccr2, Ccr3, Ccr5) were also reduced significantly in Het compared with pregnant WT uteri. Moreover, with the use of flow cytometry, myeloid CSF1R and CCR2 expression in blood and uteri from both pregnant and nonpregnant mice was characterized, and a deficiency in HO-1 significantly reduced CCR2 expression in infiltrating uterine monocytes/macrophages and dendritic cells (DCs). These data reveal that HO-1 regulates not only cytokine/chemokine production in pregnant uteri but also myeloid cell receptor numbers, suggesting a role of HO-1 in the recruitment and maintenance of myeloid cells in pregnant uteri and subsequent effects on placental vascular formation.

  View details for DOI 10.1189/jlb.1A0116-020RR

  View details for PubMedID 27468759

• Replication and refinement of a vaginal microbial signature of preterm birth in two racially distinct cohorts of US women. Proceedings of the National Academy of Sciences of the United States of America Callahan, B. J., DiGiulio, D. B., Goltsman, D. S., Sun, C. L., Costello, E. K., Jeganathan, P. n., Biggio, J. R., Wong, R. J., Druzin, M. L., Shaw, G. M., Stevenson, D. K., Holmes, S. P., Relman, D. A. 2017

  Abstract

  Preterm birth (PTB) is the leading cause of neonatal morbidity and mortality. Previous studies have suggested that the maternal vaginal microbiota contributes to the pathophysiology of PTB, but conflicting results in recent years have raised doubts. We conducted a study of PTB compared with term birth in two cohorts of pregnant women: one predominantly Caucasian (n = 39) at low risk for PTB, the second predominantly African American and at high-risk (n = 96). We profiled the taxonomic composition of 2,179 vaginal swabs collected prospectively and weekly during gestation using 16S rRNA gene sequencing. Previously proposed associations between PTB and lower Lactobacillus and higher Gardnerella abundances replicated in the low-risk cohort, but not in the high-risk cohort. High-resolution bioinformatics enabled taxonomic assignment to the species and subspecies levels, revealing that Lactobacillus crispatus was associated with low risk of PTB in both cohorts, while Lactobacillus iners was not, and that a subspecies clade of Gardnerella vaginalis explained the genus association with PTB. Patterns of cooccurrence between L. crispatus and Gardnerella were highly exclusive, while Gardnerella and L. iners often coexisted at high frequencies. We argue that the vaginal microbiota is better represented by the quantitative frequencies of these key taxa than by classifying communities into five community state types. Our findings extend and corroborate the association between the vaginal microbiota and PTB, demonstrate the benefits of high-resolution statistical bioinformatics in clinical microbiome studies, and suggest that previous conflicting results may reflect the different risk profile of women of black race.

  View details for PubMedID 28847941

• A Proteomic Clock of Human Pregnancy. American journal of obstetrics and gynecology Aghaeepour, N. n., Lehallier, B. n., Baca, Q. n., Ganio, E. A., Wong, R. J., Ghaemi, M. S., Culos, A. n., El-Sayed, Y. Y., Blumenfeld, Y. J., Druzin, M. L., Winn, V. D., Gibbs, R. S., Tibshirani, R. n., Shaw, G. M., Stevenson, D. K., Gaudilliere, B. n., Angst, M. S. 2017

  Abstract

  Early detection of maladaptive processes underlying pregnancy-related pathologies is desirable, as it will enable targeted interventions ahead of clinical manifestations. The quantitative analysis of plasma proteins features prominently among molecular approaches used to detect deviations from normal pregnancy. However, derivation of proteomic signatures sufficiently predictive of pregnancy-related outcomes has been challenging. An important obstacle hindering such efforts were limitations in assay technology, which prevented the broad examination of the plasma proteome.The recent availability of a highly-multiplexed platform affording the simultaneous measurement of 1,310 plasma proteins opens the door for a more explorative approach. The major aim of this study was to examine whether analysis of plasma collected during gestation of term pregnancy would allow identifying a set of proteins that tightly track gestational age. Establishing precisely-timed plasma proteomic changes during term pregnancy is a critical step in identifying deviations from regular patterns due to fetal and maternal maladaptations. A second aim was to gain insight into functional attributes of identified proteins, and link such attributes to relevant immunological changes.Pregnant women participated in this longitudinal study. In two subsequent subsets of 21 (training cohort) and 10 (validation cohort) women, specific blood specimens were collected during the first (7-14 wks), second (15-20 wks), and third (24-32 wks) trimesters, and 6 wks post-partum for analysis with a highly-multiplexed aptamer-based platform. An elastic net algorithm was applied to infer a proteomic model predicting gestational age. A bootstrapping procedure and piece-wise regression analysis was used to extract the minimum number of proteins required for predicting gestational age without compromising predictive power. Gene ontology analysis was applied to infer enrichment of molecular functions among proteins included in the proteomic model. Changes in abundance of proteins with such functions were linked to immune features predictive of gestational age at the time of sampling in pregnancies delivering at term.An independently validated model consisting of 74 proteins strongly predicted gestational age (p = 3.8x10-14, R = 0.97). The model could be reduced to eight proteins without losing its predictive power (p = 1.7x10-3, R = 0.91). The three top ranked proteins were glypican 3, chorionic somatomammotropin hormone, and granulins. Proteins activating the Janus kinase (JAK) and signal transducer and activator of transcription (STAT) pathway were enriched in the proteomic model, chorionic somatomammotropin hormone being the top ranked protein. Abundance of chorionic somatomammotropin hormone strongly correlated with STAT5 signaling activity in CD4 T cells, the endogenous cell-signaling event most predictive of gestational age.Results indicate that precisely timed changes in the plasma proteome during term pregnancy mirror a "proteomic clock". Importantly, the combined use of several plasma proteins was required for accurate prediction. The exciting promise of such a "clock" is that deviations from its regular chronological profile may assist in the early diagnoses of pregnancy-relate pathologies and point to underlying pathophysiology. Functional analysis of the proteomic model generated the novel hypothesis that somatomammotropin hormone may critically regulate T-cell function during pregnancy.

  View details for PubMedID 29277631

• Heme Oxygenase Activity and Heme Binding in a Neonatal Mouse Model NEONATOLOGY Kourula, S., Ang, J., Zhao, H., Kalish, F., Vandenabeele, P., Sylvester, K. G., Wong, R. J., Stevenson, D. K. 2017; 112 (4): 376–83

  Abstract

  Severe hemolytic disease of the newborn leads to the release of pro-oxidative free heme (FH). Heme oxygenase (HO) is primarily responsible for detoxifying FH.To investigate the protective effects of HO in a model of heme overload.For in vitro studies, NIH3T3 HO-1-luc cells were incubated with 10, 30, or 60 µM FH or methemalbumin (MHA). HO-1 promoter activity was assessed 3, 6, and 24 h after treatment. Cell survival was indexed by viability assays. For in vivo studies, 1- and 5-week-old wild-type (Wt) or HO-1-heterozygous (Het, HO-1+/-) mice were given 60 µmol FH or MHA/kg intraperitoneally. After 24 h, plasma aspartate aminotransferease (AST)/alanine transaminase (ALT) and hemopexin, liver HO activity, and lipid peroxidation (LP) were determined.In HO-1-luc cells, HO-1 promoter activity peaked 6 h after incubation with 30 µM FH (1.6-fold) or 60 µM MHA (2.1-fold) over baseline. Twenty-four hours after exposure to 60 µM FH, a decrease in viability of 80% was found, compared with no decrease after exposure to 60 µM MHA. In 1-week-old Wt and HO-1 Het pups given 60 µmol FH/kg, HO activity significantly increased 3.5- and 3.1-fold, respectively. No changes in LP or AST/ALT levels were observed. In adult Wt and HO-1 Het mice, HO activity increased (3.0- and 2.6-fold, respectively). LP and AST levels significantly increased 28.4- and 2.7-fold, respectively, in adult HO-1 Het mice. Hemopexin levels at baseline were higher in adults compared with newborns for both Wt and Het mice. In addition, FH induced hemopexin levels in both adults and newborns, but to a lesser degree in newborns.FH is highly toxic in vitro, but its toxicity is abolished when bound to albumin. Newborns appear to be protected from the pro-oxidative effects of FH, which may be mediated by heme binding and a higher absolute HO activity at baseline and after FH-mediated induction.

  View details for PubMedID 28926834

• An immune clock of human pregnancy. Science immunology Aghaeepour, N. n., Ganio, E. A., Mcilwain, D. n., Tsai, A. S., Tingle, M. n., Van Gassen, S. n., Gaudilliere, D. K., Baca, Q. n., McNeil, L. n., Okada, R. n., Ghaemi, M. S., Furman, D. n., Wong, R. J., Winn, V. D., Druzin, M. L., El-Sayed, Y. Y., Quaintance, C. n., Gibbs, R. n., Darmstadt, G. L., Shaw, G. M., Stevenson, D. K., Tibshirani, R. n., Nolan, G. P., Lewis, D. B., Angst, M. S., Gaudilliere, B. n. 2017; 2 (15)

  Abstract

  The maintenance of pregnancy relies on finely tuned immune adaptations. We demonstrate that these adaptations are precisely timed, reflecting an immune clock of pregnancy in women delivering at term. Using mass cytometry, the abundance and functional responses of all major immune cell subsets were quantified in serial blood samples collected throughout pregnancy. Cell signaling-based Elastic Net, a regularized regression method adapted from the elastic net algorithm, was developed to infer and prospectively validate a predictive model of interrelated immune events that accurately captures the chronology of pregnancy. Model components highlighted existing knowledge and revealed previously unreported biology, including a critical role for the interleukin-2-dependent STAT5ab signaling pathway in modulating T cell function during pregnancy. These findings unravel the precise timing of immunological events occurring during a term pregnancy and provide the analytical framework to identify immunological deviations implicated in pregnancy-related pathologies.

  View details for PubMedID 28864494

• The Importance of Hemolysis and Its Clinical Detection in Neonates with Hyperbilirubinemia. Current pediatric reviews Wong, R. J., Bhutani, V. K., Stevenson, D. K. 2017; 13 (3): 193–98

  Abstract

  BACKGROUND: Hyperbilirubinemia is a benign transitional phenomenon that occurs in 60% to 80% of all term infants. The degree of hyperbilirubinemia and hence risk for developing bilirubin-induced neurologic dysfunction or BIND is dependent upon two major processes: (i) bilirubin production and its elimination.OBJECTIVE: The aim of this review is to address the importance of hemolysis and its clinical detection in neonates with hyperbilirubinemia.RESULTS: In newborns, an increased bilirubin production rate due to hemolysis is often the primary cause of hyperbilirubinemia during the first week of life. If undiagnosed or untreated, it may lead to an increased risk for BIND. Therefore, the ability to identify infants with hemolytic disease is important in assessing those at risk for developing BIND. In addition, an infant's genetic profile and bilirubin binding status can also affect their overall capacity to cope with the resultant tissue bilirubin load and affect risk and guide appropriate management strategies.CONCLUSION: Therefore, the determination of a newborn's bilirubin production rate is critical to the assessment of a newborn's risk for developing unpredictable extreme hyperbilirubinemia and preventing BIND.

  View details for PubMedID 28782473

• Risk profiles for haemolytic and nonhaemolytic neonatal jaundice ACTA PAEDIATRICA Bhutani, V. K., Wong, R. J. 2016; 105 (12): 1387–88

  View details for PubMedID 27870210

• Mapping the Fetomaternal Peripheral Immune System at Term Pregnancy. Journal of immunology Fragiadakis, G. K., Baca, Q. J., Gherardini, P. F., Ganio, E. A., Gaudilliere, D. K., Tingle, M., Lancero, H. L., McNeil, L. S., Spitzer, M. H., Wong, R. J., Shaw, G. M., Darmstadt, G. L., Sylvester, K. G., Winn, V. D., Carvalho, B., Lewis, D. B., Stevenson, D. K., Nolan, G. P., Aghaeepour, N., Angst, M. S., Gaudilliere, B. L. 2016

  Abstract

  Preterm labor and infections are the leading causes of neonatal deaths worldwide. During pregnancy, immunological cross talk between the mother and her fetus is critical for the maintenance of pregnancy and the delivery of an immunocompetent neonate. A precise understanding of healthy fetomaternal immunity is the important first step to identifying dysregulated immune mechanisms driving adverse maternal or neonatal outcomes. This study combined single-cell mass cytometry of paired peripheral and umbilical cord blood samples from mothers and their neonates with a graphical approach developed for the visualization of high-dimensional data to provide a high-resolution reference map of the cellular composition and functional organization of the healthy fetal and maternal immune systems at birth. The approach enabled mapping of known phenotypical and functional characteristics of fetal immunity (including the functional hyperresponsiveness of CD4(+) and CD8(+) T cells and the global blunting of innate immune responses). It also allowed discovery of new properties that distinguish the fetal and maternal immune systems. For example, examination of paired samples revealed differences in endogenous signaling tone that are unique to a mother and her offspring, including increased ERK1/2, MAPK-activated protein kinase 2, rpS6, and CREB phosphorylation in fetal Tbet(+)CD4(+) T cells, CD8(+) T cells, B cells, and CD56(lo)CD16(+) NK cells and decreased ERK1/2, MAPK-activated protein kinase 2, and STAT1 phosphorylation in fetal intermediate and nonclassical monocytes. This highly interactive functional map of healthy fetomaternal immunity builds the core reference for a growing data repository that will allow inferring deviations from normal associated with adverse maternal and neonatal outcomes.

  View details for PubMedID 27793998

• Extreme hyperbilirubinemia and rescue exchange transfusion in California from 2007 to 2012. Journal of perinatology Bhutani, V. K., Meng, N. F., Knauer, Y., Danielsen, B. H., Wong, R. J., Stevenson, D. K., Gould, J. B. 2016; 36 (10): 853-857

  Abstract

  To evaluate the impact of statewide learning collaboratives that used national guidelines to manage jaundice on the serial prevalence of extreme hyperbilirubinemia (EHB, total bilirubin ⩾25 mg dl(-1)) and exchange transfusions introduced in California Perinatal Quality Care Collaborative (CPQCC) hospitals in 2007.Adverse outcomes were retrieved from statewide databases on re-admissions for live births ⩾35 weeks' gestation (2007 to 2012) in diverse CPQCC hospitals. Individual and cumulative select perinatal risk factors and frequencies were the outcomes measures.For 3 172 762 babies (2007 to 2012), 92.5% were ⩾35 weeks' gestation. Statewide EHB and exchange rates decreased from 28.2 to 15.3 and 3.6 to 1.9 per 100 000 live births, respectively. From 2007 to 2012, the trends for TB>25 mg dl(-1) rates were -0.92 per 100 000 live births per year (95% CI: -3.71 to 1.87, P=0.41 and R(2)=0.17).National guidelines complemented by statewide learning collaboratives can decrease or modify outcomes among all birth facilities and impact clinical practice behavior.

  View details for DOI 10.1038/jp.2016.106

  View details for PubMedID 27442156

• Phototherapy and the Risk of Photo-Oxidative Injury in Extremely Low Birth Weight Infants CLINICS IN PERINATOLOGY Stevenson, D. K., Wong, R. J., Arnold, C. C., Pedroza, C., Tyson, J. E. 2016; 43 (2): 291-?

  Abstract

  Phototherapy has been used to treat newborns with jaundice for more than 50 years with the presumption that it is safe and effective for all infants. In fact, this presumption may not be true for all infants, especially the smallest and most immature. The safety and efficacy of phototherapy have never really been questioned or adequately tested in the latter, yet clinical applications of phototherapy have been further refined as its mechanisms of action have been better understood and alternative light sources have become available. This article addresses what is known about the possible risks of photo-oxidative injury in extremely low birth weight infants.

  View details for DOI 10.1016/j.clp.2016.01.005

  View details for PubMedID 27235208

• Hyperbilirubinemia in Preterm Neonates CLINICS IN PERINATOLOGY Bhutani, V. K., Wong, R. J., Stevenson, D. K. 2016; 43 (2): 215-?

  Abstract

  Preterm neonates with increased bilirubin production loads are more likely to sustain adverse outcomes due to either neurotoxicity or overtreatment with phototherapy and/or exchange transfusion. Clinicians should rely on expert consensus opinions to guide timely and effective interventions until there is better evidence to refine bilirubin-induced neurologic dysfunction or benefits of bilirubin. In this article, we review the evolving evidence for bilirubin-induced brain injury in preterm infants and highlight the clinical approaches that minimize the risk of bilirubin neurotoxicity.

  View details for DOI 10.1016/j.clp.2016.01.001

  View details for PubMedID 27235203

• Identification of neonatal haemolysis: an approach to predischarge management of neonatal hyperbilirubinemia. Acta paediatrica (Oslo, Norway : 1992) Bhutani, V. K., Srinivas, S., Castillo Cuadrado, M. E., Aby, J. L., Wong, R. J., Stevenson, D. K. 2016; 105 (5): e189-94

  Abstract

  Relative contributions of increased production [by end-tidal carbon monoxide concentrations (ETCOc)] and decreased elimination of bilirubin to predischarge hour-specific total bilirubin (TB) levels were assessed in healthy late-preterm and term newborns. Secondly, we report predischarge ETCOc ranges to guide clinical management of hyperbilirubinemia.TB and ETCOc (≤3 timepoints) determinations of newborns aged between six hours and <6 days (n = 79) were stratified by postnatal age epochs. Hyperbilirubinemia risk was assessed by plotting TB values as a function of ETCOc.Stratifications of ETCOc (in ppm, mean, median and interquartile ranges) by postnatal age epochs (0-24, 24-48 and 48-72) were as follows: 2.0, 1.9, 1.8-2.2 (n = 11); 1.6, 1.5, 1.1-2.0 (n = 58); and 2.0, 1.8, 1.6-2.3 (n = 9), respectively. Infants with ETCOc ≥ 2.5 were at high risk, between 1.5 and 2.5 at moderate risk and ≤1.5 were at low risk. Risk due to haemolysis alone was not independent (p < 0.01). For infants with TB >75th percentile (n = 31), 23% had ETCO ≤1.5, and 77% had ETCOc > 1.5 (p < 0.00003).Near-simultaneous ETCOc and TB measurements in infants with TB >75th percentile accurately identify haemolytic hyperbilirubinemia.

  View details for DOI 10.1111/apa.13341

  View details for PubMedID 26802319

• Carboxyhemoglobin levels as a predictor of risk for significant hyperbilirubinemia in African-American DAT(+) infants. Journal of perinatology : official journal of the California Perinatal Association Schutzman, D. L., Gatien, E., Ajayi, S., Wong, R. J. 2016; 36 (5): 386-8

  Abstract

  To compare the degree of hemolysis in a group of direct antiglobulin test (DAT) positive (pos) African-American (AA) infants as measured by carboxyhemoglobin corrected (COHbc) for carbon monoxide in ambient air to a similar group of DAT negative (neg) ABO incompatible infants and a group without blood group incompatibility. To determine if COHbc is a better predictor of significant hyperbilirubinemia than DAT status.A prospective study of 180 AA infants from the Well-Baby Nursery of an inner city community hospital, all of whose mothers were type O pos. Infants (60) were ABO incompatible DAT pos, 60 were ABO incompatible DAT neg and 60 were type O(+). Blood for COHbc was drawn at the time of the infants' initial bilirubin and the infants' precise percentile on the Bhutani nomogram was calculated.Mean COHbc of type O(+) infants was 0.76±0.21 and 0.78±0.24% for ABO incompatible DAT neg infants (P=0.63). Mean CoHbc for the ABO incompatible DAT pos infants was 1.03±0.41% (P<0.0001 compared with both type O and DAT neg infants). Optimal cutoff on the receiver operating characteristic curve for COHbc to determine the risk for being in the Bhutani curve high risk zone was COHbc >0.90% (area under the curve(AUC) 0.8113). This was similar to the AUC of the receiver operating characteristic curve using any titer strength of DAT pos as a cutoff (0.7960).Although not greatly superior to the titer strength of DAT pos, COHbc is useful in determining if the etiology of severe hyperbilirubinemia is a hemolytic process.

  View details for DOI 10.1038/jp.2015.206

  View details for PubMedID 26765551

• The effect of hematocrit on in vitro bilirubin photoalteration PEDIATRIC RESEARCH Linfield, D. T., Lamola, A. A., Mei, E., Hwang, A. Y., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2016; 79 (3): 387-390

  Abstract

  Phototherapy using light in the spectral range of 410-500 nm, which overlaps the absorption of bilirubin, is the common treatment for neonatal hyperbilirubinemia. Hemoglobin (Hb) absorbs light strongly throughout this same range and thus can compete with bilirubin for this light and consequently reduce the efficacy of phototherapy. Here, we determined the effect of hematocrit (Hct) on in vitro bilirubin photoalteration using narrow-band blue (450 nm) light-emitting diodes (LEDs).Suspensions with Hcts from 0 to 80% and 16 ± 1 mg/dl bilirubin were prepared by mixing red blood cells (RBCs), bilirubin (30 mg/dl) in 4% human serum albumin, and normal saline. Aliquots of each suspension were exposed to blue light at equal irradiances. Before and after 60 min of exposure, bilirubin levels in supernatants (n = 46) were measured using a diazo-dye method.Bilirubin photoalteration steeply decreased by ~60% as Hct increased from 0 to 10%. Over the clinically relevant range of 30-70% Hct, the decrease was significant, but less drastic, exhibiting a quasi-linear dependence on Hct.Bilirubin photoalteration under blue light in vitro is significantly reduced as Hct increases. Clinical studies are warranted to confirm these in vitro observations that Hct can affect the efficacy of phototherapy.

  View details for DOI 10.1038/pr.2015.240

  View details for PubMedID 26571225

• Inhibition of heme oxygenase activity using a microparticle formulation of zinc protoporphyrin in an acute hemolytic newborn mouse model. Pediatric research Fujioka, K., Kalish, F., Wong, R. J., Stevenson, D. K. 2016; 79 (2): 251-257

  Abstract

  Increased bilirubin production due to hemolysis can lead to neonatal hyperbilirubinemia. Inhibition of heme oxygenase (HO), the rate-limiting enzyme in heme catabolism, by metalloporphyrins (Mps) may be an ideal preventive strategy for neonatal hemolytic disease. Zinc protoporphyrin (ZnPP) is a naturally occurring Mp, potent, not phototoxic, with minimal HO-1 upregulation, but is not orally absorbed. Recently, we designed a lipid-based ZnPP formulation (ZnPP-Lipid), which is orally absorbed by newborn mice. Here, we evaluated the efficacy of ZnPP-Lipid in heme-loaded newborn mice, a model analogous to hemolytic infants.After 24 h of heme administration (30 µmol/kg s.c.), 4-d-old mice were given 30 µmol ZnPP-Lipid/kg via intragastric injections. After 3 h, liver and brain HO activity were measured. HO-1 upregulation was assessed by determinations of HO-1 protein, promoter activity, and mRNA by Western blot, in vivo bioluminescence imaging, and RT-PCR, respectively.After heme loading, liver HO activity significantly increased ~1.6-fold, which was inhibited in a dose-dependent manner by ZnPP-Lipid. A dose of 30 µmol/kg returned activity to control levels. Brain HO activity was not inhibited. No significant increases in liver and brain HO-1 protein, promoter activity, and mRNA were observed.ZnPP-Lipid is effective and thus has potential for treating neonatal hyperbilirubinemia due to hemolysis.

  View details for DOI 10.1038/pr.2015.207

  View details for PubMedID 26488552

• Inhibition of heme oxygenase activity using a microparticle formulation of zinc protoporphyrin in an acute hemolytic newborn mouse model PEDIATRIC RESEARCH Fujioka, K., Kalish, F., Wong, R. J., Stevenson, D. K. 2016; 79 (2): 251-257

  Abstract

  Increased bilirubin production due to hemolysis can lead to neonatal hyperbilirubinemia. Inhibition of heme oxygenase (HO), the rate-limiting enzyme in heme catabolism, by metalloporphyrins (Mps) may be an ideal preventive strategy for neonatal hemolytic disease. Zinc protoporphyrin (ZnPP) is a naturally occurring Mp, potent, not phototoxic, with minimal HO-1 upregulation, but is not orally absorbed. Recently, we designed a lipid-based ZnPP formulation (ZnPP-Lipid), which is orally absorbed by newborn mice. Here, we evaluated the efficacy of ZnPP-Lipid in heme-loaded newborn mice, a model analogous to hemolytic infants.After 24 h of heme administration (30 µmol/kg s.c.), 4-d-old mice were given 30 µmol ZnPP-Lipid/kg via intragastric injections. After 3 h, liver and brain HO activity were measured. HO-1 upregulation was assessed by determinations of HO-1 protein, promoter activity, and mRNA by Western blot, in vivo bioluminescence imaging, and RT-PCR, respectively.After heme loading, liver HO activity significantly increased ~1.6-fold, which was inhibited in a dose-dependent manner by ZnPP-Lipid. A dose of 30 µmol/kg returned activity to control levels. Brain HO activity was not inhibited. No significant increases in liver and brain HO-1 protein, promoter activity, and mRNA were observed.ZnPP-Lipid is effective and thus has potential for treating neonatal hyperbilirubinemia due to hemolysis.

  View details for DOI 10.1038/pr.2015.207

  View details for Web of Science ID 000372377200003

• Heme Oxygenase-1 Expression Affects Murine Abdominal Aortic Aneurysm Progression. PloS one Azuma, J., Wong, R. J., Morisawa, T., Hsu, M., Maegdefessel, L., Zhao, H., Kalish, F., Kayama, Y., Wallenstein, M. B., Deng, A. C., Spin, J. M., Stevenson, D. K., Dalman, R. L., Tsao, P. S. 2016; 11 (2)

  Abstract

  Heme oxygenase-1 (HO-1), the rate-limiting enzyme in heme degradation, is a cytoprotective enzyme upregulated in the vasculature by increased flow and inflammatory stimuli. Human genetic data suggest that a diminished HO-1 expression may predispose one to abdominal aortic aneurysm (AAA) development. In addition, heme is known to strongly induce HO-1 expression. Utilizing the porcine pancreatic elastase (PPE) model of AAA induction in HO-1 heterozygous (HO-1+/-, HO-1 Het) mice, we found that a deficiency in HO-1 leads to augmented AAA development. Peritoneal macrophages from HO-1+/- mice showed increased gene expression of pro-inflammatory cytokines, including MCP-1, TNF-alpha, IL-1-beta, and IL-6, but decreased expression of anti-inflammatory cytokines IL-10 and TGF-beta. Furthermore, treatment with heme returned AAA progression in HO-1 Het mice to a wild-type profile. Using a second murine AAA model (Ang II-ApoE-/-), we showed that low doses of the HMG-CoA reductase inhibitor rosuvastatin can induce HO-1 expression in aortic tissue and suppress AAA progression in the absence of lipid lowering. Our results support those studies that suggest that pleiotropic statin effects might be beneficial in AAA, possibly through the upregulation of HO-1. Specific targeted therapies designed to induce HO-1 could become an adjunctive therapeutic strategy for the prevention of AAA disease.

  View details for DOI 10.1371/journal.pone.0149288

  View details for PubMedID 26894432

• Editorial: Heme Oxygenases: Novel Regulators of Reproductive Processes. Frontiers in pharmacology Wong, R. J., Zenclussen, A. C. 2015; 6: 282

  View details for DOI 10.3389/fphar.2015.00282

  View details for PubMedID 26640438

  View details for PubMedCentralID PMC4661693

• Temporal and spatial variation of the human microbiota during pregnancy PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA DiGiulio, D. B., Callahan, B. J., McMurdie, P. J., Costello, E. K., Lyell, D. J., Robaczewska, A., Sun, C. L., Goltsman, D. S., Wong, R. J., Shaw, G., Stevenson, D. K., Holmes, S. P., Relman, D. A. 2015; 112 (35): 11060-11065

  Abstract

  Despite the critical role of the human microbiota in health, our understanding of microbiota compositional dynamics during and after pregnancy is incomplete. We conducted a case-control study of 49 pregnant women, 15 of whom delivered preterm. From 40 of these women, we analyzed bacterial taxonomic composition of 3,767 specimens collected prospectively and weekly during gestation and monthly after delivery from the vagina, distal gut, saliva, and tooth/gum. Linear mixed-effects modeling, medoid-based clustering, and Markov chain modeling were used to analyze community temporal trends, community structure, and vaginal community state transitions. Microbiota community taxonomic composition and diversity remained remarkably stable at all four body sites during pregnancy (P > 0.05 for trends over time). Prevalence of a Lactobacillus-poor vaginal community state type (CST 4) was inversely correlated with gestational age at delivery (P = 0.0039). Risk for preterm birth was more pronounced for subjects with CST 4 accompanied by elevated Gardnerella or Ureaplasma abundances. This finding was validated with a set of 246 vaginal specimens from nine women (four of whom delivered preterm). Most women experienced a postdelivery disturbance in the vaginal community characterized by a decrease in Lactobacillus species and an increase in diverse anaerobes such as Peptoniphilus, Prevotella, and Anaerococcus species. This disturbance was unrelated to gestational age at delivery and persisted for up to 1 y. These findings have important implications for predicting premature labor, a major global health problem, and for understanding the potential impact of a persistent, altered postpartum microbiota on maternal health, including outcomes of pregnancies following short interpregnancy intervals.

  View details for DOI 10.1073/pnas.1502875112

  View details for Web of Science ID 000360383200068

• Bilirubin production and hour-specific bilirubin levels. Journal of perinatology Bhutani, V. K., Wong, R. J., Vreman, H. J., Stevenson, D. K. 2015; 35 (9): 735-738

  Abstract

  We assessed the relative contributions of increased bilirubin production (indexed by end-tidal carbon monoxide (CO) concentrations, corrected for ambient CO (ETCOc)) to hour-specific total bilirubin (TB) levels in healthy late preterm and term newborns.Post hoc analyses of concurrent ETCOc and TB (at 30±6 h of age) and follow-up TB levels at age 96±12 h and up to 168 h after birth were performed in a cohort of 641 term and late preterm infants.Increased bilirubin production (hour-specific ETCOc ⩾1.7 p.p.m. at age 30±6 h) was noted in ~80%, 42% and 32% of infants in the high-, intermediate- and low-risk TB zones, respectively. One infant with TB <40th percentile and ETCOc <1.7 p.p.m. developed TB ⩾95th percentile at age 168 h, probably due to decreased bilirubin elimination.Infants in the high-risk quartile of the hour-specific bilirubin nomogram have a higher mean bilirubin production. Infants with TB levels ⩾95th percentile without increased bilirubin production have impaired bilirubin elimination.

  View details for DOI 10.1038/jp.2015.32

  View details for PubMedID 25880796

• Implementing Mass Cytometry at the Bedside to Study the Immunological Basis of Human Diseases: Distinctive Immune Features in Patients with a History of Term or Preterm Birth CYTOMETRY PART A Gaudilliere, B., Ganio, E. A., Tingle, M., Lancero, H. L., Fragiadakis, G. K., Baca, Q. J., Aghaeepour, N., Wong, R. J., Quaintance, C., El-Sayed, Y. Y., Shaw, G. M., Lewis, D. B., Stevenson, D. K., Nolan, G. P., Angst, M. S. 2015; 87A (9): 817-829

  Abstract

  Single-cell technologies have immense potential to shed light on molecular and biological processes that drive human diseases. Mass cytometry (or Cytometry by Time Of Flight mass spectrometry, CyTOF) has already been employed in clinical studies to comprehensively survey patients' circulating immune system. As interest in the "bedside" application of mass cytometry is growing, the delineation of relevant methodological issues is called for. This report uses a newly generated dataset to discuss important methodological considerations when mass cytometry is implemented in a clinical study. Specifically, the use of whole blood samples versus peripheral blood mononuclear cells (PBMCs), design of mass-tagged antibody panels, technical and analytical implications of sample barcoding, and application of traditional and unsupervised approaches to analyze high-dimensional mass cytometry datasets are discussed. A mass cytometry assay was implemented in a cross-sectional study of 19 women with a history of term or preterm birth to determine whether immune traits in peripheral blood differentiate the two groups in the absence of pregnancy. Twenty-seven phenotypic and 11 intracellular markers were simultaneously analyzed in whole blood samples stimulated with lipopolysaccharide (LPS at 0, 0.1, 1, 10, and 100 ng mL(-1) ) to examine dose-dependent signaling responses within the toll-like receptor 4 (TLR4) pathway. Complementary analyses, grounded in traditional or unsupervised gating strategies of immune cell subsets, indicated that the prpS6 and pMAPKAPK2 responses in classical monocytes are accentuated in women with a history of preterm birth (FDR<1%). The results suggest that women predisposed to preterm birth may be prone to mount an exacerbated TLR4 response during the course of pregnancy. This important hypothesis-generating finding points to the power of single-cell mass cytometry to detect biologically important differences in a relatively small patient cohort. © 2015 International Society for Advancement of Cytometry.

  View details for DOI 10.1002/cyto.a.22720

  View details for Web of Science ID 000360590500009

• Temporal and spatial variation of the human microbiota during pregnancy. Proceedings of the National Academy of Sciences of the United States of America DiGiulio, D. B., Callahan, B. J., McMurdie, P. J., Costello, E. K., Lyell, D. J., Robaczewska, A., Sun, C. L., Goltsman, D. S., Wong, R. J., Shaw, G., Stevenson, D. K., Holmes, S. P., Relman, D. A. 2015

  Abstract

  Despite the critical role of the human microbiota in health, our understanding of microbiota compositional dynamics during and after pregnancy is incomplete. We conducted a case-control study of 49 pregnant women, 15 of whom delivered preterm. From 40 of these women, we analyzed bacterial taxonomic composition of 3,767 specimens collected prospectively and weekly during gestation and monthly after delivery from the vagina, distal gut, saliva, and tooth/gum. Linear mixed-effects modeling, medoid-based clustering, and Markov chain modeling were used to analyze community temporal trends, community structure, and vaginal community state transitions. Microbiota community taxonomic composition and diversity remained remarkably stable at all four body sites during pregnancy (P > 0.05 for trends over time). Prevalence of a Lactobacillus-poor vaginal community state type (CST 4) was inversely correlated with gestational age at delivery (P = 0.0039). Risk for preterm birth was more pronounced for subjects with CST 4 accompanied by elevated Gardnerella or Ureaplasma abundances. This finding was validated with a set of 246 vaginal specimens from nine women (four of whom delivered preterm). Most women experienced a postdelivery disturbance in the vaginal community characterized by a decrease in Lactobacillus species and an increase in diverse anaerobes such as Peptoniphilus, Prevotella, and Anaerococcus species. This disturbance was unrelated to gestational age at delivery and persisted for up to 1 y. These findings have important implications for predicting premature labor, a major global health problem, and for understanding the potential impact of a persistent, altered postpartum microbiota on maternal health, including outcomes of pregnancies following short interpregnancy intervals.

  View details for DOI 10.1073/pnas.1502875112

  View details for PubMedID 26283357

• Association of HMOX1 gene promoter polymorphisms with hyperbilirubinemia in the early neonatal period. Pediatrics international : official journal of the Japan Pediatric Society Katayama, Y., Yokota, T., Zhao, H., Wong, R. J., Stevenson, D. K., Taniguchi-Ikeda, M., Nakamura, H., Iijima, K., Morioka, I. 2015; 57 (4): 645-9

  Abstract

  Heme oxygenase (HO) is the rate-limiting enzyme in the heme degradation pathway that produces bilirubin. The promoter region of human heme oxygenase-1 (HMOX1) contains a polymorphic (GT)n repeat that can regulate gene expression. Here, we investigated the association of (GT)n repeat length in the HMOX1 promoter region with neonatal hyperbilirubinemia in a population of Japanese term neonates.Using polymerase chain reaction and fragment analysis, we determined the number of (GT)n repeats in 149 Japanese neonates. To omit the effects of the G71R mutation in uridine diphosphoglucuronosyltransferase on hyperbilirubinemia, we excluded 41 neonates with the G71R mutation. As a result, 25 neonates with hyperbilirubinemia and 83 non-hyperbilirubinemic controls were included in this prospective case-control study. Allele and genotype frequencies of (GT)n repeats in the HMOX1 gene were compared between hyperbilirubinemic and non-hyperbilirubinemic control neonates.The prevalence of short alleles (< 22 (GT)n repeats) was significantly higher in hyperbilirubinemic than in control neonates (18% vs 7%, P = 0.015). Hyperbilirubinemia was more frequent in homozygous or heterozygous short allele carriers than control neonates (28% vs 11%, respectively, P = 0.03). Possession of short alleles was significantly associated with the development of neonatal hyperbilirubinemia (OR, 3.1; 95%CI: 1.03-9.53).Infants carrying short alleles (< 22 (GT)n repeats) in the HMOX1 gene promoter region appear to be at a higher risk for developing neonatal hyperbilirubinemia.

  View details for DOI 10.1111/ped.12591

  View details for PubMedID 25625535

• Evaluation of a new end-tidal carbon monoxide monitor from the bench to the bedside. Acta paediatrica (Oslo, Norway : 1992) Castillo Cuadrado, M. E., Bhutani, V. K., Aby, J. L., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2015; 104 (6): e279-82

  View details for DOI 10.1111/apa.12938

  View details for PubMedID 25640053

• Heme oxygenase-1 confers protection and alters T-cell populations in a mouse model of neonatal intestinal inflammation PEDIATRIC RESEARCH Schulz, S., Chisholm, K. M., Zhao, H., Kalish, F., Yang, Y., Wong, R. J., Stevenson, D. K. 2015; 77 (5): 640-648

  Abstract

  Necrotizing enterocolitis (NEC), an intestinal inflammatory disease affecting premature infants, is associated with low regulatory T (Treg) to effector T (Teff) cell ratios. We recently demonstrated that heme oxygenase-1 (HO-1) deficiency leads to increased NEC development. Here, we investigated the effects of HO-1 on T-cell proportions in a murine NEC-like injury model.Intestinal injury was induced in 7-d-old wild-type (WT) or HO-1 heterozygous (HO-1 Het) pups by formula-feeding every 4 h for 24-78 h by oral gavage and exposures to 5%O2. Controls remained breastfed. HO-1 was induced in WT pups by administering heme preinjury induction. Lamina propria T cells were identified by flow cytometry. For adoptive transfer studies, WT splenic/thymic Tregs were injected intraperitoneally into HO-1 Het pups 12-24 h preinduction.Het mice showed increased intestinal injury and decreased Treg/Teff ratios. Genes for pattern recognition (Toll-like receptor-4, C-reactive protein, MyD88) and neutrophil recruitment increased in Het pups after NEC induction. Inducing intestinal HO-1 decreased NEC scores and incidence, and increased Treg/Teff ratios. Moreover, adoptive transfer of Tregs from WT to HO-1 Het pups decreased NEC scores and incidence and restored Treg/Teff ratios.HO-1 can change Treg proportions in the lamina propria of young mice under inflammatory conditions, which might, in part, confer intestinal protection.

  View details for DOI 10.1038/pr.2015.22

  View details for PubMedID 25665053

• Neonatal bilirubin binding capacity discerns risk of neurological dysfunction. Pediatric research Lamola, A. A., Bhutani, V. K., Du, L., Castillo Cuadrado, M., Chen, L., Shen, Z., Wong, R. J., Stevenson, D. K. 2015; 77 (2): 334-339

  Abstract

  Bilirubin binding capacity (BBC) defines the dynamic relationship between an infant's level of unbound or "free" bilirubin and his/her ability to "tolerate" increasing bilirubin loads. BBC is not synonymous with albumin (Alb) levels because Alb binding of bilirubin is confounded by a variety of molecular, biologic, and metabolic factors.We utilized a novel modification of a previously developed hematofluorometric method to directly assay BBC in whole blood from preterm and term neonates and then combined these data with an archived database. Total bilirubin (TB) was also measured, and multiple regression modeling was used to determine whether BBC in combination with TB measurements can assess an infant's risk for developing bilirubin-induced neurotoxicity.TB and BBC levels ranged from 0.7-22.8 to 6.3-47.5 mg/dl, respectively. Gestational age (GA) correlated with BBC (r = 0.54; P < 0.0002) with a slope of 0.93 mg/dl/wk by logistic regression. Our calculations demonstrate that recently recommended GA-modulated TB thresholds for phototherapy and exchange transfusion correspond to 45 and 67% saturation of our observed regression line, respectively.We speculate that the spread of BBC levels around the regression line (±5.8 mg/dl) suggests that individualized BBC assays would provide a robust approach to gauge risk of bilirubin neurotoxicity compared with TB and GA.

  View details for DOI 10.1038/pr.2014.191

  View details for PubMedID 25420178

• Bilirubin-induced neurologic dysfunction (BIND) SEMINARS IN FETAL & NEONATAL MEDICINE Bhutani, V. K., Wong, R. 2015; 20 (1): 1

  View details for PubMedID 25577656

• Neonatal bilirubin binding capacity discerns risk of neurological dysfunction. Pediatric research Lamola, A. A., Bhutani, V. K., Du, L., Castillo Cuadrado, M., Chen, L., Shen, Z., Wong, R. J., Stevenson, D. K. 2015; 77 (2): 334-339

  View details for DOI 10.1038/pr.2014.191

  View details for PubMedID 25420178

• Neonatal hemolysis and risk of bilirubin-induced neurologic dysfunction. Seminars in fetal & neonatal medicine Wong, R. J., Stevenson, D. K. 2015; 20 (1): 26-30

  Abstract

  The pathologic phenotype of severe hyperbilirubinemia in the newborn infant is primarily due to excessive bilirubin production and/or impaired conjugation, resulting in an increased bilirubin load. This may, in turn, increase an infant's risk for the development of bilirubin-induced neurologic dysfunction (BIND). The highest-risk infants are those with increased bilirubin production rates due to hemolysis. Several immune and non-immune conditions have been found to cause severe hemolysis, and these are often exacerbated in those infants with perinatal sepsis and genetic predispositions. Therefore, identification of these infants, with novel technologies, is paramount in reducing the incidence of BIND and the long-term neurologic sequelae for these at-risk infants.

  View details for DOI 10.1016/j.siny.2014.12.005

  View details for PubMedID 25560401

• Heme oxygenase and the immune system in normal and pathological pregnancies. Frontiers in pharmacology Ozen, M., Zhao, H., Lewis, D. B., Wong, R. J., Stevenson, D. K. 2015; 6: 84-?

  Abstract

  Normal pregnancy is an immunotolerant state. Many factors, including environmental, socioeconomic, genetic, and immunologic changes by infection and/or other causes of inflammation, may contribute to inter-individual differences resulting in a normal or pathologic pregnancy. In particular, imbalances in the immune system can cause many pregnancy-related diseases, such as infertility, abortions, pre-eclampsia, and preterm labor, which result in maternal/fetal death, prematurity, or small-for-gestational age newborns. New findings imply that myeloid regulatory cells and regulatory T cells (Tregs) may mediate immunotolerance during normal pregnancy. Effector T cells (Teffs) have, in contrast, been implicated to cause adverse pregnancy outcomes. Furthermore, feto-maternal tolerance affects the developing fetus. It has been shown that the Treg/Teff balance affects litter size and adoptive transfer of pregnancy-induced Tregs can prevent fetal rejection in the mouse. Heme oxygenase-1 (HO-1) has a protective role in many conditions through its anti-inflammatory, anti-apoptotic, antioxidative, and anti-proliferative actions. HO-1 is highly expressed in the placenta and plays a role in angiogenesis and placental vascular development and in regulating vascular tone in pregnancy. In addition, HO-1 is a major regulator of immune homeostasis by mediating crosstalk between innate and adaptive immune systems. Moreover, HO-1 can inhibit inflammation-induced phenotypic maturation of immune effector cells and pro-inflammatory cytokine secretion and promote anti-inflammatory cytokine production. HO-1 may also be associated with T-cell activation and can limit immune-based tissue injury by promoting Treg suppression of effector responses. Thus, HO-1 and its byproducts may protect against pregnancy complications by its immunomodulatory effects, and the regulation of HO-1 or its downstream effects has the potential to prevent or treat pregnancy complications and prematurity.

  View details for DOI 10.3389/fphar.2015.00084

  View details for PubMedID 25964759

• Mechanical Stress Changes the Complex Interplay Between HO-1, Inflammation and Fibrosis, During Excisional Wound Repair. Frontiers in medicine Cremers, N. A., Suttorp, M., Gerritsen, M. M., Wong, R. J., van Run-van Breda, C., van Dam, G. M., Brouwer, K. M., Kuijpers-Jagtman, A. M., Carels, C. E., Lundvig, D. M., Wagener, F. A. 2015; 2: 86-?

  Abstract

  Mechanical stress following surgery or injury can promote pathological wound healing and fibrosis, and lead to functional loss and esthetic problems. Splinted excisional wounds can be used as a model for inducing mechanical stress. The cytoprotective enzyme heme oxygenase-1 (HO-1) is thought to orchestrate the defense against inflammatory and oxidative insults that drive fibrosis. Here, we investigated the activation of the HO-1 system in a splinted and non-splinted full-thickness excisional wound model using HO-1-luc transgenic mice. Effects of splinting on wound closure, HO-1 promoter activity, and markers of inflammation and fibrosis were assessed. After seven days, splinted wounds were more than three times larger than non-splinted wounds, demonstrating a delay in wound closure. HO-1 promoter activity rapidly decreased following removal of the (epi)dermis, but was induced in both splinted and non-splinted wounds during skin repair. Splinting induced more HO-1 gene expression in 7-day wounds; however, HO-1 protein expression remained lower in the epidermis, likely due to lower numbers of keratinocytes in the re-epithelialization tissue. Higher numbers of F4/80-positive macrophages, αSMA-positive myofibroblasts, and increased levels of the inflammatory genes IL-1β, TNF-α, and COX-2 were present in 7-day splinted wounds. Surprisingly, mRNA expression of newly formed collagen (type III) was lower in 7-day wounds after splinting, whereas, VEGF and MMP-9 were increased. In summary, these data demonstrate that splinting delays cutaneous wound closure and HO-1 protein induction. The pro-inflammatory environment following splinting may facilitate higher myofibroblast numbers and increase the risk of fibrosis and scar formation. Therefore, inducing HO-1 activity against mechanical stress-induced inflammation and fibrosis may be an interesting strategy to prevent negative effects of surgery on growth and function in patients with orofacial clefts or in patients with burns.

  View details for DOI 10.3389/fmed.2015.00086

  View details for PubMedID 26697429

  View details for PubMedCentralID PMC4678194

• Direct Antiglobulin Titer Strength and Hyperbilirubinemia PEDIATRICS Kaplan, M., Hammerman, C., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2014; 134 (5): E1340-E1344

  Abstract

  We recently demonstrated that direct antiglobulin titer (DAT) positive, blood group A or B newborns born to group O mothers had a high incidence of hyperbilirubinemia, attributable to increased hemolysis. We reanalyzed our data asking whether increasing DAT strength plays a modulating role in the pathophysiology of the hemolysis and hyperbilirubinemia.Data from previously published DAT-positive, ABO-heterospecific neonates were analyzed for hyperbilirubinemia and hemolysis according to strength of DAT. DAT was measured by using a gel agglutination technique and reported as values ranging from DAT ± to DAT ++++. Hemolysis was evaluated by blood carboxyhemoglobin corrected for inspired, ambient CO (COHbc), and expressed as percent total hemoglobin (tHb). Hyperbilirubinemia was defined as any plasma total bilirubin value >95th percentile on the hour-specific nomogram.Hyperbilirubinemia was more prevalent in those with DAT ++ readings (16 of 20, 80%) than those both DAT ± (37 of 87 [42.5%], relative risk: 1.88, 95% confidence interval: 1.35-2.61) and DAT + (32 of 56 [57.1%], relative risk: 1.40, 95% confidence interval: 1.02-1.92). COHbc values were higher for those with DAT ++ (1.45 ± 0.49% tHb [mean ± SD]) than those DAT ± (1.20 ± 0.37% tHb, P = .01) or DAT + (1.22 ± 0.37% tHb, P = .02).DAT ++ readings were associated with a higher incidence of hyperbilirubinemia and higher COHbc values than DAT ± or DAT + counterparts. Increasing DAT strength may be a modulator of hemolysis and hyperbilirubinemia in ABO-heterospecific neonates. DAT strength, and not merely DAT presence or absence, should be taken into consideration in the management of ABO-heterospecific newborns.

  View details for DOI 10.1542/peds.2014-1290

  View details for Web of Science ID 000344385900009

• End-tidal carbon monoxide and hemolysis JOURNAL OF PERINATOLOGY Tidmarsh, G. F., Wong, R. J., Stevenson, D. K. 2014; 34 (8): 577-581

  Abstract

  Hemolytic disease in newborns can result from a number of conditions, which can place such infants at an increased risk for the development of severe hyperbilirubinemia. Because the catabolism of heme produces equimolar amounts of carbon monoxide (CO) and bilirubin, measurements of end-tidal breath CO (corrected for ambient CO) or ETCOc can serve as an index of hemolysis as well as of bilirubin production from any cause. Elevated levels of ETCOc have been correlated with blood carboxyhemoglobin levels and thus hemolysis. However, the detection of hemolysis can be a clinically challenging problem in newborns. Here, we describe the importance of determining ETCOc levels and their application in identifying infants at risk for developing hyperbilirubinemia associated with hemolysis and other causes of increased bilirubin production.

  View details for DOI 10.1038/jp.2014.66

  View details for Web of Science ID 000339706400001

  View details for PubMedID 24743136

• Safety and Efficacy of Filtered Sunlight in Treatment of Jaundice in African Neonates PEDIATRICS Slusher, T. M., Vreman, H. J., Olusanya, B. O., Wong, R. J., Brearley, A. M., Vaucher, Y. E., Stevenson, D. K. 2014; 133 (6): E1568-E1574

  Abstract

  Evaluate safety and efficacy of filtered-sunlight phototherapy (FS-PT).Term/late preterm infants #14 days old with clinically significant jaundice, assessed by total bilirubin (TB) levels, were recruited from a maternity hospital in Lagos, Nigeria. Sunlight was filtered with commercial window-tinting films that remove most UV and significant levels of infrared light and transmit effective levels of therapeutic blue light. After placing infants under an FS-PT canopy, hourly measurements of axillary temperatures, monitoring for sunburn, dehydration, and irradiances of filtered sunlight were performed. Treatment was deemed safe and efficacious if infants were able to stay in FS-PT for $5 hours and rate of rise of TB was ,0.2 mg/dL/h for infants #72 hours of age or TB decreased for infants .72 hours of age.A total of 227 infants received 258 days of FS-PT. No infant developed sunburn or dehydration. On 85 (33%) of 258 treatment days, infants were removed briefly from FS-PT due to minor temperature-related adverse events. No infant met study exit criteria. FS-PT was efficacious in 92% (181/197) of evaluable treatment days. Mean 6 SD TB change was –0.06 6 0.19 mg/dL/h. The mean 6 SD (range) irradiance of FS-PT was 38 6 22 (2–115) mW/cm2/nm, measured by the BiliBlanket Meter II.With appropriate monitoring, filtered sunlight is a novel, practical, and inexpensive method of PT that potentially offers safe and efficacious treatment strategy for management of neonatal jaundice in tropical countries where conventional PT treatment is not available.

  View details for DOI 10.1542/peds.2013-3500

  View details for Web of Science ID 000337172600012

  View details for PubMedID 24864170

• Effects of light on metalloporphyrin-treated newborn mice. Acta paediatrica Wong, R. J., Schulz, S., Espadas, C., Vreman, H. J., Rajadas, J., Stevenson, D. K. 2014; 103 (5): 474-479

  Abstract

  Zinc protoporphyrin (ZnPP) is a promising metalloporphyrin with sufficient potency, but has poor solubility and is not absorbed well orally. Intragastric administration of ZnPP microparticles (30 μmol/kg) to 3-day-old mice resulted in a twofold increase in potency and no signs of phototoxicity.The use of polymeric particulate delivery systems can improve the stability and enhance intestinal absorption of ZnPP, while retaining HO inhibitory potency without photosensitising effects, and thus is potentially useful in treating neonatal hyperbilirubinemia.

  View details for DOI 10.1111/apa.12554

  View details for PubMedID 24417721

• Anti-cancer effects of blue-green alga Spirulina platensis, a natural source of bilirubin-like tetrapyrrolic compounds ANNALS OF HEPATOLOGY Konickova, R., Vankova, K., Vanikova, J., Vanova, K., Muchova, L., Subhanova, I., Zadinova, M., Zelenka, J., Dvorak, A., Kolar, M., Strnad, H., Rimpelova, S., Ruml, T., Wong, R. J., Vitek, L. 2014; 13 (2): 273-283

  View details for Web of Science ID 000341467100014

• Heme Oxygenase-1 Promoter Polymorphisms and Neonatal Jaundice NEONATOLOGY Kaplan, M., Renbaum, P., Hamrnernnan, C., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2014; 106 (4): 323-329

  Abstract

  Heme oxygenase (HO) is the initial, rate-limiting enzyme in the conversion of heme to bilirubin. Dinucleotide (GT)n repeat length in the promoter region of the encoding gene modulates transcription: shorter alleles, in contrast with longer allele counterparts, are associated with greater gene expression and should result in increased heme catabolism.We compared the rates of heme catabolism and plasma total bilirubin (TB) between HO-1 promoter genotypes of varying (GT)n repeat lengths in glucose-6-phosphate dehydrogenase (G6PD)-normal and -deficient neonates.HO-1 promoter length was determined from genomic DNA from previous studies by size discrimination of fluorescently-labeled PCR products with capillary electrophoresis. Sizing was confirmed by sequencing homozygote samples. Alleles were categorized as: short (≤24 GT repeats), medium (25-33 GT repeats), and long (≥34 GT repeats). Previously determined values for blood carboxyhemoglobin, corrected for inspired carbon monoxide (COHbc), and TB were used to determine the rate of heme catabolism and 3rd day TB values for each HO-1 promoter length genotype, respectively. G6PD Mediterranean was determined by PCR analysis.Neither COHbc nor TB values were significantly different between various HO-1 promoter genotypes for either G6PD-normal or -deficient neonates.In the steady state, HO-1 promoter genotypes, based on the length of (GT)n repeats, do not modulate heme catabolism or 3rd day TB values in either G6PD-normal or -deficient neonates.

  View details for DOI 10.1159/000365744

  View details for Web of Science ID 000344445400007

• Heme oxygenase-1 promoter polymorphisms and neonatal jaundice. Neonatology Kaplan, M., Renbaum, P., Hammerman, C., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2014; 106 (4): 323-329

  Abstract

  Heme oxygenase (HO) is the initial, rate-limiting enzyme in the conversion of heme to bilirubin. Dinucleotide (GT)n repeat length in the promoter region of the encoding gene modulates transcription: shorter alleles, in contrast with longer allele counterparts, are associated with greater gene expression and should result in increased heme catabolism.We compared the rates of heme catabolism and plasma total bilirubin (TB) between HO-1 promoter genotypes of varying (GT)n repeat lengths in glucose-6-phosphate dehydrogenase (G6PD)-normal and -deficient neonates.HO-1 promoter length was determined from genomic DNA from previous studies by size discrimination of fluorescently-labeled PCR products with capillary electrophoresis. Sizing was confirmed by sequencing homozygote samples. Alleles were categorized as: short (≤24 GT repeats), medium (25-33 GT repeats), and long (≥34 GT repeats). Previously determined values for blood carboxyhemoglobin, corrected for inspired carbon monoxide (COHbc), and TB were used to determine the rate of heme catabolism and 3rd day TB values for each HO-1 promoter length genotype, respectively. G6PD Mediterranean was determined by PCR analysis.Neither COHbc nor TB values were significantly different between various HO-1 promoter genotypes for either G6PD-normal or -deficient neonates.In the steady state, HO-1 promoter genotypes, based on the length of (GT)n repeats, do not modulate heme catabolism or 3rd day TB values in either G6PD-normal or -deficient neonates.

  View details for DOI 10.1159/000365744

  View details for PubMedID 25277974

• Heme oxygenase-1 in pregnancy and cancer: similarities in cellular invasion, cytoprotection, angiogenesis, and immunomodulation. Frontiers in pharmacology Zhao, H., Ozen, M., Wong, R. J., Stevenson, D. K. 2014; 5: 295-?

  Abstract

  Pregnancy can be defined as a "permissible" process, where a semi-allogeneic fetus and placenta are allowed to grow and survive within the mother. Similarly, in tumor growth, antigen-specific malignant cells proliferate and evade into normal tissues of the host. The microenvironments of the placenta and tumors are amazingly comparable, sharing similar mechanisms exploited by fetal or cancer cells with regard to surviving in a hypoxic microenvironment, invading tissues via degradation and vasculogenesis, and escaping host attack through immune privilege. Heme oxygease-1 (HO-1) is a stress-response protein that has antioxidative, anti-apoptotic, pro-angiogenic, and anti-inflammatory properties. Although a large volume of research has been published in recent years investigating the possible role(s) of HO-1 in pregnancy and in cancer development, the molecular mechanisms that regulate these "yin-yang" processes have still not been fully elucidated. Here, we summarize and compare pregnancy and cancer development, focusing primarily on the function of HO-1 in cellular invasion, cytoprotection, angiogenesis, and immunomodulation. Due to the similarities of both processes, a thorough understanding of the molecular mechanisms of each process may reveal and guide the development of new approaches to prevent not only pregnancy disorders; but also, to study cancer.

  View details for DOI 10.3389/fphar.2014.00295

  View details for PubMedID 25642189

• Spirulina platensis and phycocyanobilin activate atheroprotective heme oxygenase-1: a possible implication for atherogenesis. Food & function Strasky, Z., Zemankova, L., Nemeckova, I., Rathouska, J., Wong, R. J., Muchova, L., Subhanova, I., Vanikova, J., Vanova, K., Vitek, L., Nachtigal, P. 2013; 4 (11): 1586-1594

  Abstract

  Spirulina platensis, a water blue-green alga, has been associated with potent biological effects, which might have important relevance in atheroprotection. We investigated whether S. platensis or phycocyanobilin (PCB), its tetrapyrrolic chromophore, can activate atheroprotective heme oxygenase-1 (Hmox1), a key enzyme in the heme catabolic pathway responsible for generation of a potent antioxidant bilirubin, in endothelial cells and in a mouse model of atherosclerosis. In vitro experiments were performed on EA.hy926 endothelial cells exposed to extracts of S. platensis or PCB. In vivo studies were performed on ApoE-deficient mice fed a cholesterol diet and S. platensis. The effect of these treatments on Hmox1, as well as other markers of oxidative stress and endothelial dysfunction, was then investigated. Both S. platensis and PCB markedly upregulated Hmox1 in vitro, and a substantial overexpression of Hmox1 was found in aortic atherosclerotic lesions of ApoE-deficient mice fed S. platensis. In addition, S. platensis treatment led to a significant increase in Hmox1 promoter activity in the spleens of Hmox-luc transgenic mice. Furthermore, both S. platensis and PCB were able to modulate important markers of oxidative stress and endothelial dysfunction, such as eNOS, p22 NADPH oxidase subunit, and/or VCAM-1. Both S. platensis and PCB activate atheroprotective HMOX1 in endothelial cells and S. platensis increased the expression of Hmox1 in aortic atherosclerotic lesions in ApoE-deficient mice, and also in Hmox-luc transgenic mice beyond the lipid lowering effect. Therefore, activation of HMOX1 and the heme catabolic pathway may represent an important mechanism of this food supplement for the reduction of atherosclerotic disease.

  View details for DOI 10.1039/c3fo60230c

  View details for PubMedID 24056745

• Raltegravir in vitro effect on bilirubin binding. Pediatric infectious disease journal Clarke, D. F., Wong, R. J., Wenning, L., Stevenson, D. K., Mirochnick, M. 2013; 32 (9): 978-980

  Abstract

  Drugs that displace bilirubin from albumin may increase the risk of kernicterus in neonates. We evaluated the effect of raltegravir on bilirubin-albumin binding in pooled neonatal serum using the peroxidase method. Raltegravir had minimal effect on bilirubin-albumin binding at concentrations of 5 and 10 µM, caused a small but statistically significant increase in unbound bilirubin at 100 µM and caused potentially harmful increases at 500 and 1000 µM. Our data suggest that the effect of raltegravir on neonatal bilirubin binding is unlikely to be clinically significant at typical peak concentrations reached with usual dosing.

  View details for DOI 10.1097/INF.0b013e31829044a8

  View details for PubMedID 23470680

• The effect of hematocrit on the efficacy of phototherapy for neonatal jaundice PEDIATRIC RESEARCH Lamola, A. A., Bhutani, V. K., Wong, R. J., Stevenson, D. K., McDonagh, A. F. 2013; 74 (1): 54-60

  Abstract

  Background:The therapeutic phototherapy action spectrum ranges from 420 to 500nm. However, a recent report of improved efficacy offluorescent "turquoise"light (~490 nm) compared toblue light(~450 nm) underscores the need to define an optimal action spectrum for precision-targeted phototherapy using very narrow wavelength ranges.Methods:We used a current semi-empirical model of theoptical properties of skinfor robust calculations of the fraction of light absorbed by bilirubin at various wavelengths that could be confounded by hemoglobin, melanin and skin thickness. Applying assumptions regarding the wavelength dependence of bilirubin photochemistry, "action spectra"wereassembled from the calculated values.Results:All the calculated action spectra displayed a peak between 472 and 480 nm (most at 476 nm), which is a significant shift from the well-reported 460 nm absorption peak of bilirubin. Interestingly, the relative amplitudes of the action spectra showed an inverse relationship with hematocrit.Conclusion:We speculate that narrow range of light at 476 nmshould be 60% more effective than blue (broad-band) fluorescent lamps. Because hemoglobin serves as a major competitor of bilirubin for light absorption, the calculations also predict that the efficacy of phototherapy is dependent on the hematocrit.A high hematocrit could reduce therapeutic efficiency.Pediatric Research (2013); doi:10.1038/pr.2013.67.

  View details for DOI 10.1038/pr.2013.67

  View details for PubMedID 23604171

• Heme oxygenase-1 deficiency promotes the development of necrotizing enterocolitis-like intestinal injury in a newborn mouse model. American journal of physiology. Gastrointestinal and liver physiology Schulz, S., Wong, R. J., Jang, K. Y., Kalish, F., Chisholm, K. M., Zhao, H., Vreman, H. J., Sylvester, K. G., Stevenson, D. K. 2013; 304 (11): G991-G1001

  Abstract

  Necrotizing enterocolitis (NEC) is typified by mucosal destruction, which subsequently can lead to intestinal necrosis. Prematurity, enteral feeding, and bacterial colonization are the main risk factors and, combined with other stressors, can cause increased intestinal permeability, injury, and an exaggerated inflammatory response. Heme oxygenase-1 (HO-1) mediates intestinal protection due to anti-inflammatory, antioxidative, and antiapoptotic effects of its products carbon monoxide, biliverdin, and bilirubin. This study investigates a possible role of HO-1 in the pathogenesis of NEC using a newborn mouse model. We induced NEC-like intestinal injury in 7-day-old HO-1 heterozygous (HO-1 Het, Hmox1(+/-)) and wild-type (Wt, Hmox1(+/+)) mice by gavage feeding and hypoxic exposures. Control (Con) pups of both genotypes were dam-fed. Intestines of HO-1 Het Con pups appeared predisposed to injury, with higher histological damage scores, more TUNEL-positive cells, and a significant reduction in muscularis externa thickness compared with Wt Con pups. The increase in HO activity after HO-1 induction by the substrate heme or by hypoxic stress was significantly impaired in HO-1 Het pups. After induction of intestinal injury, HO-1 Het pups displayed significantly higher NEC incidence (78 vs. 43%), mortality (83 vs. 54%), and median scores (2.5 vs. 1.5) than Wt NEC pups. PCR array analyses revealed increased expressions of IL-1β, P-selectin, matrix metallopeptidase 2, collagen type XVIII-α1, serpine 1, and others in NEC-induced HO-1 Het ileal and jejunal tissues. We conclude that a partial HO-1 deficiency promotes experimental NEC-like intestinal injury, possibly mediated by exaggerated inflammation and disruption in tissue repair.

  View details for DOI 10.1152/ajpgi.00363.2012

  View details for PubMedID 23578787

• Bilirubin neurotoxicity in preterm infants: risk and prevention. Journal of clinical neonatology Bhutani, V. K., Wong, R. J. 2013; 2 (2): 61-69

  Abstract

  Hemolytic conditions in preterm neonates, including Rhesus (Rh) disease, can lead to mortality and long-term impairments due to bilirubin neurotoxicity. Universal access to Rh immunoprophylaxis, coordinated perinatal-neonatal care, and effective phototherapy has virtually eliminated the risk of kernicterus in many countries. In the absence of jaundice due to isoimmunization and without access to phototherapy or exchange transfusion (in 1955), kernicterus was reported at 10.1%, 5.5%, and 1.2% in babies <30, 31-32, and 33-34 wks gestational age, respectively. Phototherapy initiated at 24±12 hr effectively prevented hyperbilirubinemia in infants <2,000 g even in the presence of hemolysis. This approach (in 1985) reduced exchange transfusions from 23.9% to 4.8%. Now with 3 decades of experience in implementing effective phototherapy, the need for exchange transfusions has virtually been eliminated. However, bilirubin neurotoxicity continues to be associated with prematurity alone. The ability to better predict this risk, other than birthweight and gestation, has been elusive. Objective tests such as total bilirubin, unbound or free bilirubin, albumin levels, and albumin-bilirubin binding, together with observations of concurrent hemolysis, sepsis, and rapid rate of bilirubin rise have been considered, but their individual or combined predictive utility has yet to be refined. The disruptive effects of immaturity, concurrent neonatal disease, cholestasis, use of total parenteral nutrition or drugs that alter bilirubin-binding abilities augment the clinical risk of neurotoxicity. Current management options rely on the "fine-tuning" of each infant's exposure to beneficial antioxidants and avoidance of silent neurotoxic properties of bilirubin navigated within the safe spectrum of operational thresholds demarcated by experts.

  View details for DOI 10.4103/2249-4847.116402

  View details for PubMedID 24049745

  View details for PubMedCentralID PMC3775137

• Combining hand techniques with electric pumping increases the caloric content of milk in mothers of preterm infants JOURNAL OF PERINATOLOGY Morton, J., Wong, R. J., Hall, J. Y., Pang, W. W., Lai, C. T., Lui, J., Hartmann, P. E., Rhine, W. D. 2012; 32 (10): 791-796

  Abstract

  We previously reported that preterm mothers' milk production can exceed levels of term mothers by using early hand expression and hands-on pumping (HOP) with the highest production (955 ml per day) in frequent users of hand expression. In this study, we compared milk composition between mothers stratified by early hand expression frequency.A total of 67 mothers of infants <31 weeks gestation were instructed on hand expression and HOP. Subjects submitted expression records and 1-ml samples from each pumping session over 24 h once weekly for 8 weeks.78% (52/67) of mothers completed the study. But for Week 1, no compositional differences (despite production differences) were noted between the three groups. Protein and lactose tracked reported norms, but fat and energy of mature milk (Weeks 2-8) exceeded norms, 62.5 g l(-1) per fat and 892.7 cal l(-1) (26.4 cal oz(-1)), respectively.Mothers combining manual techniques with pumping express high levels of fat-rich, calorie-dense milk, unrelated to production differences.

  View details for DOI 10.1038/jp.2011.195

  View details for Web of Science ID 000309519800010

  View details for PubMedID 22222549

• A deficiency in haem oxygenase-1 induces foetal growth restriction by placental vasculature defects ACTA PAEDIATRICA Wong, R. J., Zhao, H., Stevenson, D. K. 2012; 101 (8): 827-834

  Abstract

  Haem oxygenase-1 (HO-1), the rate-limiting enzyme in haem degradation, plays a role in angiogenesis and vasculogenesis and is highly expressed in the placenta. Deficiencies in HO-1 are associated with several pregnancy disorders, such as recurrent miscarriages and pre-eclampsia. The unique combination of tissue protective, smooth muscle relaxing and angiogenesis regulatory properties makes HO-1 a key player in the maintenance of a healthy pregnancy through a direct effect on placental structural and vascular development, thus affecting foetal development. Conclusion:  Therefore, we conclude that HO-1 plays an important role in placental vasculature development and a deficiency in HO-1 may contribute to pregnancy complications, such as pre-eclampsia, spontaneous abortions and premature births.

  View details for DOI 10.1111/j.1651-2227.2012.02729.x

  View details for PubMedID 22594519

• Effect of light exposure on metalloporphyrin-treated newborn mice PEDIATRIC RESEARCH Schulz, S., Wong, R. J., Kalish, F. S., Zhao, H., Jang, K. Y., Vreman, H. J., Stevenson, D. K. 2012; 72 (2): 161-168

  Abstract

  Neonatal hyperbilirubinemia arises from increased bilirubin production and decreased bilirubin elimination. Although phototherapy safely and effectively reduces bilirubin levels, recent evidence shows that it has adverse effects. Therefore, alternative treatments are warranted. Metalloporphyrins, competitive inhibitors of heme oxygenase (HO), the rate-limiting enzyme in bilirubin production, effectively reduce bilirubin formation; however, many are photoreactive. Here, we investigated possible photosensitizing effects of chromium mesoporphyrin (CrMP) and zinc deuteroporphyrin bis-glycol (ZnBG).Administration of CrMP or ZnBG to 3-d-old mouse pups (3.75-30.0 μmol/kg intraperitoneally) and exposure to cool white (F20T12CW) and blue (TL20W/52) fluorescent lights (+L) for 3 h, resulted in a dose-dependent mortality (50% lethal dose (LD50) = 21.5 and 19.5 μmol/kg, respectively). In contrast to ZnBG, there was no significant difference in survival between the CrMP+L and CrMP groups. Following 30 μmol/kg ZnBG+L, we found significant weight loss, decreased liver antioxidant capacities, and increased aspartate aminotransaminase levels. At 6-d post-light exposure, ZnBG+L-treated pups showed gross and histologic skin changes at doses >7.5 μmol/kg. No lethality was observed following treatment with 30 μmol ZnBG/kg plus exposure to blue light-emitting diodes. Phototoxicity of ZnBG was dependent on light source, emission spectrum, and irradiance.Low doses of ZnBG (<3.75 μmol/kg) retained maximal HO inhibitory potency without photosensitizing effects, and therefore are potentially useful in treating neonatal hyperbilirubinemia.

  View details for DOI 10.1038/pr.2012.62

  View details for PubMedID 22580722

• Metalloporphyrins - an update. Frontiers in pharmacology Schulz, S., Wong, R. J., Vreman, H. J., Stevenson, D. K. 2012; 3: 68-?

  Abstract

  Metalloporphyrins are structural analogs of heme and their potential use in the management of neonatal hyperbilirubinemia has been the subject of considerable research for more than three decades. The pharmacological basis for using this class of compounds to control bilirubin levels is the targeted blockade of bilirubin production through the competitive inhibition of heme oxygenase (HO), the rate-limiting enzyme in the bilirubin production pathway. Ongoing research continues in the pursuit of identifying ideal metalloporphyrins, which are safe and effective, by defining therapeutic windows and targeted interventions for the treatment of excessive neonatal hyperbilirubinemia.

  View details for DOI 10.3389/fphar.2012.00068

  View details for PubMedID 22557967

• Metalloporphyrins - an update FRONTIERS IN PHARMACOLOGY Schulz, S., Wong, R. J., Vreman, H. J., Stevenson, D. K. 2012; 3

  Abstract

  Metalloporphyrins are structural analogs of heme and their potential use in the management of neonatal hyperbilirubinemia has been the subject of considerable research for more than three decades. The pharmacological basis for using this class of compounds to control bilirubin levels is the targeted blockade of bilirubin production through the competitive inhibition of heme oxygenase (HO), the rate-limiting enzyme in the bilirubin production pathway. Ongoing research continues in the pursuit of identifying ideal metalloporphyrins, which are safe and effective, by defining therapeutic windows and targeted interventions for the treatment of excessive neonatal hyperbilirubinemia.

  View details for DOI 10.3389/fphar.2012.00068

  View details for Web of Science ID 000209177700066

  View details for PubMedCentralID PMC3337460

• Effects of Zinc Deuteroporphyrin Bis Glycol on Newborn Mice After Heme Loading PEDIATRIC RESEARCH He, C. X., Campbell, C. M., Zhao, H., Kalish, F. S., Schulz, S., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2011; 70 (5): 467-472

  Abstract

  Infants with hemolytic diseases frequently develop hyperbilirubinemia and are treated with phototherapy, which only eliminates bilirubin after its production. A better strategy might be to directly inhibit heme oxygenase (HO), the rate-limiting enzyme in bilirubin production. Metalloporphyrins (Mps) are heme analogs that competitively inhibit HO activity in vitro and in vivo and suppress plasma bilirubin levels in vivo. A promising Mp, zinc deuteroporphyrin bis glycol (ZnBG), is orally absorbed and effectively inhibits HO activity at relatively low doses. We determined the I(50) (the dose needed to inhibit HO activity by 50%) of orally administered ZnBG in vivo and then evaluated ZnBG's effects on in vivo bilirubin production, HO activity, HO protein levels, and HO-1 gene expression in newborn mice after heme loading, a model analogous to a hemolytic infant. The I(50) of ZnBG was found to be 4.0 μmol/kg body weight (BW). At a dose of 15 μmol/kg BW, ZnBG reduced in vivo bilirubin production, inhibited heme-induced liver HO activity and spleen HO activity to and below baseline, respectively, transiently induced liver and spleen HO-1 gene transcription, and induced liver and spleen HO-1 protein levels. We conclude that ZnBG may be an attractive compound for treating severe neonatal hyperbilirubinemia caused by hemolytic disease.

  View details for PubMedID 21785387

• Maternal Heme Oxygenase 1 Regulates Placental Vasculature Development via Angiogenic Factors in Mice BIOLOGY OF REPRODUCTION Zhao, H., Azuma, J., Kalish, F., Wong, R. J., Stevenson, D. K. 2011; 85 (5): 1005-1012

  Abstract

  The placental vasculature is critical for nutrient, gas, and waste exchange between the maternal and fetal systems. Its development depends on the proper expression and interaction of angiogenesis and associated growth factors. Heme oxygenase (HMOX), the enzyme for heme degradation, plays a role in angiogenesis and is highly expressed in the placenta. To evaluate the role of maternal HMOX1, the inducible HMOX isozyme, on placental vasculature formation, mice with a partial deficiency in Hmox1 (Hmox1(+/-)) were used. Three-dimensional images of placental vasculatures as well as spiral arteries from Hmox1(+/+) or Hmox1(+/-) placentas were created by vascular corrosion casting technique and imaged by micro-computerized tomography (microCT). The structures and morphologies of fetomaternal interfaces were observed by histological staining and the ultrastructure of uterine natural killer (uNK) cells, a major regulator in spiral artery remodeling, was analyzed by transmission electron microscopy. A group of growth factors and angiogenic factors from the decidua/mesometrial lymphoid aggregate of pregnancy (MLAp) as well as labyrinth regions were quantified using an angiogenesis PCR array kit and compared between Hmox1(+/+) or Hmox1(+/-) placentas. In conclusion, a partial deficiency of maternal Hmox1 resulted in the malformation of fetomaternal interface, insufficiency of spiral artery remodeling, and alteration of uNK cell differentiation and maturation. These changes were independent of the fetal genotype, but relied on the maternal HMOX1 level, which determined the balance of expression levels of pro- and antiangiogenic factors in the decidua/MLAp region. These results implied that Hmox1 polymorphisms among the human population might contribute to some unexplained cases of pregnancy disorders, such as fetal growth retardation and preeclampsia.

  View details for DOI 10.1095/biolreprod.111.093039

  View details for PubMedID 21778140

• Panhematin provides a therapeutic benefit in experimental pancreatitis GUT Habtezion, A., Kwan, R., Akhtar, E., Wanaski, S. P., Collins, S. D., Wong, R. J., Stevenson, D. K., Butcher, E. C., Omary, M. B. 2011; 60 (5): 671-679

  Abstract

  Acute pancreatitis (AP) can result in pancreatic necrosis and inflammation, with subsequent multi-organ failure. AP is associated with increased neutrophil recruitment and a rise in pro-inflammatory cytokines such as TNFα. Pretreatment with haemin, results in recruitment of haem-oxygenase-1 (HO-1)(+) macrophages and protects against experimental pancreatitis. It is not clear whether modulation of HO-1 after onset of disease has a protective role. In this study, we tested the utility of Panhematin, a water-soluble haemin formulation, in activating and inducing pancreatic HO-1, and as a therapeutic agent in treating mouse acute pancreatitis.We defined the distribution of radiolabelled haemin, then used in vivo HO-1-luciferase bioluminescence imaging and the CO-release assay to test Panhematin-induced upregulation of HO-1 transcription and activity, respectively. Using two well-defined AP murine models, we tested the therapeutic benefit of Panhematin, and quantified cytokine release using a luminex assay.Intravenously administered Panhematin induces rapid recruitment of HO-1(+) cells to the pancreas within 2 h and de novo splenic HO-1 transcription by 12 h. Despite high baseline spleen HO-1 activity, the pancreas is particularly responsive to Panhematin-mediated HO-1 induction. Panhematin-treated mice, at various time points after AP induction had significant reduction in mortality, pancreatic injury, together with upregulation of HO-1 and downregulation of pro-inflammatory cytokines and CXCL1, a potent neutrophil chemoattractant.Despite AP-associated mortality and morbidity, no effective treatment other than supportive care exists. We demonstrate that Panhematin leads to: (i) rapid induction and activation of pancreatic HO-1 with recruitment of HO-1(+) cells to the pancreas, (ii) amelioration of AP even when given late during the course of disease, and (iii) a decrease in leucocyte infiltration and pro-inflammatory cytokines including CXCL1. The utility of Panhematin at modest doses as a therapeutic in experimental pancreatitis, coupled with its current use and safety in humans, raises the potential of its applicability to human pancreatitis.

  View details for DOI 10.1136/gut.2010.217208

  View details for Web of Science ID 000289076700015

  View details for PubMedID 21159893

  View details for PubMedCentralID PMC3580958

• In vitro inhibition of heme oxygenase isoenzymes by metalloporphyrins JOURNAL OF PERINATOLOGY Wong, R. J., Vreman, H. J., Schulz, S., Kalish, F. S., Pierce, N. W., Stevenson, D. K. 2011; 31: S35-S41

  Abstract

  Neonatal jaundice results from an increased bilirubin production and decreased hepatic bilirubin conjugation and excretion. Severe hyperbilirubinemia is currently treated with phototherapy or exchange transfusion; however, its prevention by inhibiting bilirubin formation is a more logical strategy. Heme oxygenase (HO), with inducible (HO-1) and constitutive (HO-2) isoenzymes, is the rate-limiting enzyme in heme catabolism, producing equimolar amounts of bilirubin and carbon monoxide (CO). Metalloporphyrins (Mps) are heme derivatives that competitively inhibit HO and thereby suppress hyperbilirubinemia. No systematic studies have been reported evaluating whether the HO isoenzymes are inhibited differentially by various Mps. Identification of Mps that selectively inhibit the inducible HO-1 without affecting the 'housekeeping' HO-2 isoenzyme might be desirable in the clinical setting of hemolytic disease, in which the Hmox1 gene is greatly induced. Although bilirubin production is due to the activity of both HO-1 and HO-2, the inhibition of HO-1 with a relative sparing of HO-2 activity might provide the most selective approach for the treatment of hemolytic disease.We determined for the deutero-, proto-, meso- and bis-glycol porphyrins with zinc, tin and chromium as central atoms, respectively, the concentration needed for 50% inhibition (I(50)) of HO-1 and HO-2 activities in rat spleen and brain tissue.For a given Mp, HO-1 activity was less inhibited than that of HO-2. The order of inhibitor potency of each Mp was nearly identical for both isoenzymes. Tin mesoporphyrin was the most potent inhibitor for both isoenzymes. HO-2 selectivity was greatest for tin protoporphyrin. Conversely, the Zn compounds were least inhibitory toward HO-2. No Mp preferentially inhibited HO-1.Mps that produce a less inhibitory effect on HO-2, while limiting the response of the inducible HO-1, such as ZnPP, may be a useful clinical tool.

  View details for DOI 10.1038/jp.2010.173

  View details for Web of Science ID 000289236900006

  View details for PubMedID 21448202

• Metalloporphyrins in the management of neonatal hyperbilirubinemia SEMINARS IN FETAL & NEONATAL MEDICINE Stevenson, D. K., Wong, R. J. 2010; 15 (3): 164-168

  Abstract

  Neonatal jaundice in the first week of life is a common problem in newborns. It is due to an imbalance of bilirubin production and its elimination, which can lead to significantly elevated levels of circulating bilirubin or hyperbilirubinemia. Use of phototherapy and/or exchange transfusion are the current modes for treating neonatal hyperbilirubinemia and preventing any neurologic damage. These strategies, however, only remove bilirubin that has already been formed. Preventing the production of excess bilirubin may be a more logical approach. Synthetic heme analogs, metalloporphyrins, are competitive inhibitors of heme oxygenase, the rate-limiting enzyme in bilirubin production, and their use has been proposed as an attractive alternative strategy for preventing or treating severe hyperbilirubinemia.

  View details for DOI 10.1016/j.siny.2009.11.004

  View details for PubMedID 20006567

• Combining hand techniques with electric pumping increases milk production in mothers of preterm infants JOURNAL OF PERINATOLOGY Morton, J., Hall, J. Y., Wong, R. J., Thairu, L., Benitz, W. E., Rhine, W. D. 2009; 29 (11): 757-764

  Abstract

  Pump-dependent mothers of preterm infants commonly experience insufficient production. We observed additional milk could be expressed following pumping using hand techniques. We explored the effect on production of hand expression of colostrum and hands-on pumping (HOP) of mature milk.A total of 67 mothers of infants <31 weeks gestation were enrolled and instructed on pumping, hand expression of colostrum and HOP. Expression records for 8 weeks and medical records were used to assess production variables.Seventy-eight percent of the mothers completed the study. Mean daily volumes (MDV) rose to 820 ml per day by week 8 and 955 ml per day in mothers who hand expressed >5 per day in the first 3 days. Week 2 and/or week 8 MDV related to hand expression (P<0.005), maternal age, gestational age, pumping frequency, duration, longest interval between pumpings and HOP (P<0.003). Mothers taught HOP increased MDV (48%) despite pumping less.Mothers of preterm infants may avoid insufficient production by combining hand techniques with pumping.

  View details for DOI 10.1038/jp.2009.87

  View details for Web of Science ID 000271187300009

  View details for PubMedID 19571815

• Effect of Heme Oxygenase-1 Deficiency on Placental Development PLACENTA Zhao, H., Wong, R. J., Kalish, F. S., Nayak, N. R., Stevenson, D. K. 2009; 30 (10): 861-868

  Abstract

  Heme oxygenase (HO) is the rate-limiting enzyme in the heme catabolic pathway and highly expressed in the placenta. Deficiencies in HO-1, the inducible isoform, have been associated with pregnancy disorders, such as recurrent miscarriages, intrauterine growth retardation, and pre-eclampsia. The aim of this study was to identify if a deficiency in HO-1 affects placental development using a mouse model. When HO-1 heterozygote (Het, HO-1(+/-)) mice were cross-bred, an extremely low birth rate in homozygote (Mut, HO-1(-/-)) offspring (2.4%) and small litter sizes were observed. Placentas and fetuses from Het cross-breedings were relatively smaller and weighed less than those from wild-type (WT) cross-breedings at E12.5 and E15.5. Furthermore, Het placentas had significantly less HO-1 mRNA and protein levels than WT placentas, but no significant differences in placental HO activity. Interestingly, HO-2, the constituitive HO isoform, as well as iNOS and eNOS expression were significantly upregulated in Het placentas. Histological examination showed that the junctional zone (JZ) of Het placentas were markedly thinner than those of WT placentas and appeared to be due to an increase in apoptosis. Immunohistochemistry revealed that HO-1-expressing cells were located primarily in the JZ of Het placentas, specifically in the spongiotrophoblast layer. In addition, diastolic blood pressures and plasma soluble VEGFR-1 (sFlt-1) levels were significantly elevated in pregnant Het mice. We conclude that a partial deficiency in HO-1 is associated with morphological changes in the placenta and elevations in maternal diastolic blood pressure and plasma sFlt-1 levels, despite a compensatory increase in HO-2 expression.

  View details for DOI 10.1016/j.placenta.2009.07.012

  View details for Web of Science ID 000270706300006

  View details for PubMedID 19699520

  View details for PubMedCentralID PMC2771543

• Dermal Carbon Monoxide Excretion in Neonatal Rats During Light Exposure PEDIATRIC RESEARCH Vreman, H. J., Knauer, Y., Wong, R. J., Chan, M., Stevenson, D. K. 2009; 66 (1): 66-69

  Abstract

  Total body, head, and trunk carbon monoxide (CO) excretion rates were measured separately by gas chromatography in 1- to 7-d-old Wistar rat pups exposed to the dark and to mixed blue (one Special Blue-F20T12/BB) and white (two Cool White-F20T12/CW) fluorescent light or blue light emitting diode (LED) sources. During 48-min cycled exposures to the dark and to either light source, total body CO excretion rapidly increased 1.9- and 1.4-fold, respectively, over dark control levels. When CO excretion rates from the head and trunk were measured separately during exposure to either light source, CO excretion from the head did not change significantly; however, a large mean 4.4-fold increase in CO excretion from the trunk was observed. When light intensity delivered by the blue LED source was varied, we found that trunk CO excretion increased with increasing light intensities. In the presence of riboflavin (10 micromol/kg), total body CO excretion increased 2.8- and 2.1-fold during exposure to the mixed fluorescent light and blue LED sources, respectively. We conclude that light-induced elevations in total body CO excretion may be caused by transdermally excreted CO, which is most likely produced through endogenous photosensitizer-mediated photooxidation of dermal biomolecules.

  View details for PubMedID 19342986

• Heme oxygenase-1 deficiency leads to disrupted response to acute stress in stem cells and progenitors BLOOD Cao, Y., Wagers, A. J., Karsunky, H., Zhao, H., Reeves, R., Wong, R. J., Stevenson, D. K., Weissman, I. L., Contag, C. H. 2008; 112 (12): 4494-4502

  Abstract

  An effective response to extreme hematopoietic stress requires an extreme elevation in hematopoiesis and preservation of hematopoietic stem cells (HSCs). These diametrically opposed processes are likely to be regulated by genes that mediate cellular adaptation to physiologic stress. Herein, we show that heme oxygenase-1 (HO-1), the inducible isozyme of heme degradation, is a key regulator of these processes. Mice lacking one allele of HO-1 (HO-1(+/-)) showed accelerated hematopoietic recovery from myelotoxic injury, and HO-1(+/-) HSCs repopulated lethally irradiated recipients with more rapid kinetics. However, HO-1(+/-) HSCs were ineffective in radioprotection and serial repopulation of myeloablated recipients. Perturbations in key stem cell regulators were observed in HO-1(+/-) HSCs and hematopoietic progenitors (HPCs), which may explain the disrupted response of HO-1(+/-) HPCs and HPCs to acute stress. Control of stem cell stress response by HO-1 presents opportunities for metabolic manipulation of stem cell-based therapies.

  View details for DOI 10.1182/blood-2007-12-127621

  View details for PubMedID 18509090

• Inhibition of heme oxygenase activity in newborn mice by azalanstat CANADIAN JOURNAL OF PHYSIOLOGY AND PHARMACOLOGY Morisawa, T., Wong, R. J., Bhutani, V. K., Vreman, H. J., Stevenson, D. K. 2008; 86 (10): 651-659

  Abstract

  Inhibition of heme oxygenase (HO), the rate-limiting enzyme in heme catabolism, may be an ideal strategy for preventing neonatal jaundice. Although natural and synthetic heme analogs, called metalloporphyrins (Mps), have been extensively investigated for this purpose, some Mps are phototoxic, affect the activity of other enzymes, or induce HO-1 transcription-properties that may limit their clinical use. Another class of compounds, imidazole-dioxolanes, has been shown to selectively inhibit the inducible isozyme HO-1. Therefore, we investigated the efficacy of azalanstat (AZA), an imidazole-dioxolane, towards inhibiting HO activity in 7-day-old mice. We found that a single dose of AZA at 500 micromol.kg(-1) body mass (BM) administered i.p. significantly inhibited HO activity and reduced in vivo bilirubin production. In the spleen, HO inhibition (>50%) was observed within 0.25-3 h after administration. After 24 h, however, spleen HO activity, HO-1 protein, and HO-1 mRNA levels significantly increased 1.2-, 2.4-, and 4.0-fold, respectively. We conclude that AZA effectively inhibits in vivo HO activity only at a high dose and that it also induces spleen HO-1 gene transcription. Therefore, other imidazole-dioxolanes should be evaluated to determine whether they are more potent than AZA for use in treating neonatal jaundice.

  View details for DOI 10.1139/Y08-069

  View details for PubMedID 18841169

• Regulation of maternal and fetal hemodynamics by heme oxygenase in mice BIOLOGY OF REPRODUCTION Zhao, H., Wong, R. J., Doyle, T. C., Nayak, N., Vreman, H. J., Contag, C. H., Stevenson, D. K. 2008; 78 (4): 744-751

  Abstract

  Heme oxygenase (HMOX) regulates vascular tone and blood pressure through the production of carbon monoxide (CO), a vasodilator derived from the heme degradation pathway. During pregnancy, the maternal circulation undergoes significant adaptations to accommodate the hemodynamic demands of the developing fetus. Our objective was to investigate the role of HMOX on maternal and fetal hemodynamics during pregnancy in a mouse model. We measured and compared maternal tissue and placental HMOX activity and endogenous CO production, represented by excreted CO and carboxyhemoglobin levels, during pregnancy (Embryonic Days 12.5-15.5) to nonpregnant controls. Micro-ultrasound was used to monitor maternal abdominal aorta diameters as well as blood flow velocities and diameters of fetal umbilical arteries. Tin mesoporphyrin, a potent HMOX inhibitor, was used to inhibit HMOX activity. Changes in maternal vascular tone were monitored by tail cuff blood pressure measurements. Effects of HMOX inhibition on placental structures were assessed by histology. We showed that maternal tissue and placental HMOX activity and CO production were significantly elevated during pregnancy. When HMOX in the placenta was inhibited, maternal and fetal hemodynamics underwent significant changes, with maternal blood pressures increasing. We concluded that increases in maternal tissue and placental HMOX activity contribute to the regulation of peripheral vascular resistance and therefore are important for the maintenance of normal maternal vascular tone and fetal hemodynamic functions during pregnancy.

  View details for DOI 10.1095/biolreprod.107.064899

  View details for PubMedID 18094356

• Standardized bench method for evaluating the efficacy of phototherapy devices 4th International Bilirubin Workshop Vreman, H. J., Wong, R. J., Murdock, J. R., Stevenson, D. K. WILEY-BLACKWELL. 2008: 308–16

  Abstract

  As phototherapy (PT) devices employ a variety of broadband light sources, we developed and tested a standardized bench method for evaluating the efficacy of some devices.To evaluate efficacy, we quantified the in vitro photodegradation rate (expressed as t1/2) of unconjugated bilirubin in solution at 37 degrees C during exposure to a given light source at its mean delivered irradiance to the 2D body surface area (BSA) of newborn models. Reproducibility (between-day variation) of the method was determined at irradiance levels from 10 to 70 microW/cm2/nm on three different days.Between-day t1/2 measurements had coefficients of variation from 3% to 10%. When t1/2 values were normalized to the exposable 2D horizontal BSA, halogen lamp devices, without and with fiberoptics, were least effective (t1/2=60-108 min and 100-126 min for preterm and term models, respectively). Fluorescent tube devices had t1/2=19-78 min and 25-78 min, for preterm and term models, respectively. Light-emitting diode (LED)-based devices yielded the shortest t1/2 values (16-24 min) for preterm and term newborn models.We demonstrated the applicability of the method through the determination of the efficacy of several commercially available PT devices. This standardized method is reproducible and effectively evaluates the relative in vitro efficacy of various devices and may guide further in vitro and in vivo evaluations of devices.

  View details for DOI 10.1111/j.1651-2227.2007.00631.x

  View details for Web of Science ID 000253504300012

  View details for PubMedID 18241292

• Statin treatment increases formation of carbon monoxide and bilirubin in mice: a novel mechanism of in vivo antioxidant protection CANADIAN JOURNAL OF PHYSIOLOGY AND PHARMACOLOGY Muchova, L., Wong, R. J., Hsu, M., Morioka, O., Vitek, L., Zelenka, J., Schroeder, H., Stevenson, D. K. 2007; 85 (8): 800-810

  Abstract

  Heme oxygenase (HO) has a central role in cellular antioxidant defences and vascular protection, and it may mediate pleiotropic actions of drugs used in cardiovascular therapy. We investigated whether long-term use of statins upregulates HO activity and increases carbon monoxide (CO) and bilirubin levels in vivo. Adult FvB mice were given atorvastatin or rosuvastatin (5 mg/kg) daily by i.p. injections for 1, 2, or 3 weeks. HO activity, tissue CO, bilirubin, and antioxidant levels, total plasma bilirubin, and carboxyhemoglobin (COHb) were measured. Fold changes in heart HO activity significantly increased after 1, 2, and 3 weeks of atorvastatin (1.24 +/- 0.06 (p < or = 0.05); 1.29 +/- 0.26 (p < or = 0.03); 1.33 +/- 0.08 (p < 0.01), respectively) and 2 and 3 weeks of rosuvastatin (1.23 +/- 0.20 (p < or = 0.03); 1.63 +/- 0.42 (p < 0.01), respectively). Heart tissue CO and COHb levels also increased after 3 weeks with atorvastatin (1.30 +/- 0.24 (p < or = 0.05); 1.92 +/- 0.17 (p < or = 0.001), respectively) and rosuvastatin (1.47 +/- 0.13 (p < or = 0.004); 1.63 +/- 0.12 (p < or = 0.001), respectively). Significant increases in heart antioxidant levels were observed after statin treatment and corroborated by heart bilirubin content elevations. Antioxidant level increases were abolished by treatment with an HO inhibitor. These findings suggest that the induction of HO and the production of its products, CO and bilirubin, may be a mechanism by which statins exert antioxidant actions and confer cardioprotection in vivo.

  View details for DOI 10.1139/Y07-077

  View details for PubMedID 17901890

• The role of Bach1 in the induction of heme oxygenase by tin mesoporphyrin BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Abate, A., Zhao, H., Wong, R. J., Stevenson, D. K. 2007; 354 (3): 757-763

  Abstract

  Tin mesoporphyrin (SnMP), a competitive heme oxygenase (HO) inhibitor, also induces HO-1 mRNA and protein expression by a mechanism that is not fully understood. We examined whether the induction by SnMP is mediated by a de-repression of Bach1, a transcription factor that suppresses the HO-1 gene. Incubation of NIH3T3-HO-1-luc cells with SnMP attenuated HO activity with a concomitant increase in HO-1 mRNA and protein and a decrease in Bach1 and HO-2 proteins, which was not due to transcriptional down-regulation, but accelerated protein decay. Similarly, HO-1 protein degradation was increased by SnMP, despite of an elevation in HO-1 transcription. Transfection of Bach1 shRNA in Hepa cells raised basal HO-1 expression significantly, and SnMP treatment further increased HO-1 mRNA. In conclusion, SnMP induces HO-1 expression not only by de-repressing the HO-1 promoter by binding Bach1, but also by accelerating Bach1 degradation.

  View details for DOI 10.1016/j.bbrc.2007.01.050

  View details for PubMedID 17257585

• Expression and regulation of heme oxygenase isozymes in the developing mouse cortex PEDIATRIC RESEARCH Zhao, H., Wong, R. J., Nguyen, X., Kalish, F., Mizobuchi, M., Vreman, H. J., Stevenson, D. K., Contag, C. H. 2006; 60 (5): 518-523

  Abstract

  Heme oxygenase (HO), the rate-limiting enzyme in heme degradation, plays a role in neonatal jaundice. Understanding the regulation of the developmental expression patterns of the two HO isozymes, HO-1 and HO-2, is essential for targeting HO to control pathologic jaundice, and uncovering the fundamental role that they play in mammalian development. Here we characterized the ontogeny of HO-1 and HO-2 expression in the developing mouse cortex by in vivo bioluminescence imaging, quantitative RT-PCR, and Western blot. HO-2, the predominant isoform in the adult cortex, was relatively stable throughout all ages. HO-1 was observed to be progressively down-regulated in an age-related manner. HO-1 expression in the adult cortex was also the lowest among the eight adult tissues analyzed. Because there is a 283-bp CpG island region in the HO-1 promoter, we hypothesized that methylation of the island is responsible for the age-related HO-1 down-regulation in the cortex. Methylation status was assessed using regular and quantitative methylation-specific PCR and the CpG island was found to be hypomethylated at all ages. Therefore, we conclude that HO-1 gene expression in the cortex is developmentally-regulated and that methylation of the HO-1 CpG island is not associated with the down-regulation of the gene.

  View details for DOI 10.1203/01.PDR.0000242374.21415.f5

  View details for PubMedID 16966352

• Tissue-specific effects of statins on the expression of heme oxygenase-1 in vivo BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS HSU, M., Muchova, L., Morioka, I., Wong, R. J., Schroder, H., Stevenson, D. K. 2006; 343 (3): 738-744

  Abstract

  Heme oxygenase-1 (HO-1) plays a central role in antioxidant and anti-inflammatory actions, which may be mediated through its formation of biliverdin/bilirubin and carbon monoxide. HMG-CoA reductase inhibitors (statins) induce in vitro HO-1 expression and are reported to have pleiotropic benefits that reduce oxidative stress in the vasculature. We characterized the effects of statins on in vivo HO-1 expression in various extravascular tissues: liver, lung, brain, and heart. Adult mice were orally administered simvastatin, lovastatin, atorvastatin, or rosuvastatin. HO activity significantly increased in a statin- and tissue-specific manner, with all statins increasing heart and lung activity within 24 h. Significant elevations of HO-1 protein and mRNA were also observed in heart and lung after atorvastatin treatment. We conclude that in vivo HO-1 induction is statin- and tissue-specific. Through this pathway, statins may confer antioxidant and anti-inflammatory actions in the vasculature and extravascular systems.

  View details for DOI 10.1016/j.bbrc.2006.03.036

  View details for PubMedID 16563347

• Systemic effects of orally-administered zinc and tin (IV) metalloporphyrins on heme oxygenase expression in mice PEDIATRIC RESEARCH Morioka, I., Wong, R. J., Abate, A., Vreman, H. J., Contag, C. H., Stevenson, D. K. 2006; 59 (5): 667-672

  Abstract

  Some metalloporphyrins (Mps) inhibit heme oxygenase (HO), the rate-limiting enzyme in the production of bilirubin, and are potential compounds for the treatment of neonatal jaundice. We studied the safety and efficacy of Mps following oral administration. Adult HO-1-luc reporter mice were administered 30 micromol/kg body weight of tin mesoporphyrin (SnMP), zinc bis glycol deuteroporphyrin (ZnBG), or zinc protoporphyrin (ZnPP), or vehicle by oral gavage. Bilirubin production was measured as total body carbon monoxide (CO) excretion (VeCO). HO activity was quantitated via CO measurements by gas chromatography. HO-1 protein was determined by Western blot. HO-1 transcription levels were assessed by in vivo bioluminescence imaging. A significant 28% decrease in bilirubin production occurred within 3 h of SnMP treatment and persisted beyond 48 h. Bilirubin production decreased 15% and 9% by 3 h after administration of ZnBG and ZnPP, respectively, but returned to baseline within 48 h. Maximal inhibition of liver, spleen, and intestine HO activity was seen at 3 h with inhibitory effects decreasing in the order: SnMP > or = ZnBG > or = ZnPP. After SnMP treatment, HO-1 transcription increased 5.7-fold after 24 h. Furthermore, liver and spleen HO-1 protein significantly increased 3.7- and 2.0-fold, respectively, after 24 h. HO-1 transcription and protein were not affected in ZnBG- or ZnPP-treated mice. We conclude that the three Mps are absorbed at different rates in the mouse and affect bilirubin production and HO-1 expression in a tissue- and time-dependent manner.

  View details for DOI 10.1203/01.pdr.0000215088.71481.a6

  View details for PubMedID 16627879

• Monitoring age-related susceptibility of young mice to oral Salmonella enterica serovar typhimurium infection using an in vivo murine model PEDIATRIC RESEARCH Burns-Guydish, S. M., Olomu, I. N., Zhao, H., Wong, R. J., Stevenson, D. K., Contag, C. H. 2005; 58 (1): 153-158

  Abstract

  Neonates and young children are acutely susceptible to infections by gastrointestinal bacterial pathogens, such as Salmonella enterica serovar Typhimurium (S. typhimurium). To reveal age-related differences in susceptibility to this pathogen, we used in vivo bioluminescence imaging (BLI) to monitor the progression of infection in neonatal (1-wk-old), suckling (2-wk-old), juvenile (4-wk-old), and adult (6-wk-old) BALB/c mice. Mice were orally infected with various doses of a bioluminescent-labeled wild-type or mutant S. typhimurium strain, and progression of infection was monitored by BLI for 2 wks. We found that neonatal and suckling mice were more susceptible to the wild-type strain at inoculum sizes 4 and 2 log(10)'s lower for neonatal and suckling mice, respectively, than those for adult mice. At the lower inocula, newborn mice showed disseminated systemic infection as indicated by the pattern of photon emission assessed by BLI, whereas no bioluminescent signals were detectable in adult mice. In addition, an orgA(-) mutant strain of S. typhimurium with reduced virulence in adult mice produced systemic infection in newborn, suckling, and juvenile mice. Furthermore, as low as 3 log(10) CFU could be detected by BLI in tissue. The present study demonstrates that susceptibility to S. typhimurium infection decreases with age. Also, we established that BLI can be used to monitor the progression of infection in mice. Thus, this model of age-related susceptibility to S. typhimurium using BLI can be used to advance our understanding of the mechanisms involved in newborn susceptibility to infection.

  View details for DOI 10.1203/01.PDR.0000157725.44213.C4

  View details for PubMedID 15774831

• Prediction of hyperbilirubinemia in near-term and term infants. Journal of perinatology Stevenson, D. K., Fanaroff, A. A., Maisels, M. J., Young, B. W., Wong, R. J., Vreman, H. J., MacMahon, J. R., Yeung, C. Y., Seidman, D. S., Gale, R., Oh, W., Bhutani, V. K., Johnson, L. H., Kaplan, M., Hammerman, C., Nakamura, H. 2001; 21: S63-72

  Abstract

  The purpose of this study was to determine whether end-tidal carbon monoxide (CO) corrected for ambient CO (ETCOc), as a single measurement or in combination with serum total bilirubin (STB) measurements, can predict the development of hyperbilirubinemia during the first 7 days of life.From nine multinational clinical sites, 1370 neonates completed this cohort study from February 20, 1998 through February 22, 1999. Measurements of both ETCOc and STB were performed at 30+/-6 hours of life; STB also was measured at 96+/-12 hours and subsequently following a flow diagram based on a table of hours of age-specific STB. An infant was defined as hyperbilirubinemic if the hours of age-specific STB was greater than or equal to the 95th percentile as defined by the table at any time during the study.A total of 120 (8.8%) of the enrolled infants became hyperbilirubinemic. Mean STB in breast-fed infants was 8.92+/-4.37 mg/dl at 96 hours versus 7.63+/-3.58 mg/dl in those fed formula only. The mean ETCOc at 30+/-6 hours for the total population was 1.48+/-0.49 ppm, whereas those of nonhyperbilirubinemic and hyperbilirubinemic infants were 1.45+/-0.47 and 1.81+/-0.59 ppm, respectively. Seventy-six percent (92 of 120) of hyperbilirubinemic infants had ETCOc greater than the population mean. An ETCOc greater than the population mean at 30+/-6 hours yielded a 13.0% positive predictive value (PPV) and a 95.8% negative predictive value (NPV) for STB > or =95th percentile. When infants with STB > or =95th percentile at <36 hours of age were excluded, the STB at 30+/-6 hours yielded a 16.7% PPV and a 98.1% NPV for STB >75th percentile. The combination of these two measurements at 30+/-6 hours (either ETCOc more than the population mean or STB >75th percentile) had a 6.4% PPV with a 99.0% NPV.This prospective cohort study supports previous observations that measuring STB before discharge may provide some assistance in predicting an infant's risk for developing hyperbilirubinemia. The addition of an ETCOc measurement provides insight into the processes that contribute to the condition but does not materially improve the predictive ability of an hours of age-specific STB in this study population. The combination of STB and ETCOc as early as 30+/-6 hours may identify infants with increased bilirubin production (eg, hemolysis) or decreased elimination (conjugation defects) as well as infants who require early follow-up after discharge for jaundice or other clinical problems such as late anemia. Depending on the incidence of hyperbilirubinemia within an institution, the criteria for decision making should vary according to its unique population.

  View details for PubMedID 11803421

• Simultaneous production of carbon monoxide and thiobarbituric acid reactive substances in rat tissue preparations by an iron-ascorbate system CANADIAN JOURNAL OF PHYSIOLOGY AND PHARMACOLOGY Vreman, H. J., Wong, R. J., Sanesi, C. A., Dennery, P. A., Stevenson, D. K. 1998; 76 (12): 1057-1065

  Abstract

  Most of the carbon monoxide (CO) produced by mammals is a product of the heme oxygenase (HO) reaction, the rate-limiting step in the heme degradation pathway leading to the generation of bilirubin in man. However, some CO is derived from other sources. We studied the association of CO production with lipid peroxidation in tissue preparations from adult male Wistar rats. Supernatants, from 20% tissue homogenates in potassium phosphate buffer, centrifuged for 1 min at 13,000 x g, were incubated for 30 min at 37 degrees C in septum-sealed vials in the dark with ascorbate (100 microM) and Fe(II) (6 microM) and (or) Fe(III) (60 microM). Butylated hydroxytoluene (BHT, 100 microM) was added for the blank reaction. CO produced into the headspace was quantitated by gas chromatography. Thiobarbituric acid reactive substances (TBARS), conjugated dienes (CD), and lipid hydroperoxides (LOOH) in the reaction medium were quantitated by spectrophotometry. Of the tissues studied, CO and TBARS formation was greatest for brain, followed by kidney, lung, spleen, and blood, but no CO or TBARS formation was detected for testes, intestine, liver, and heart. Cell fractionation studies indicated that these differences might be due to the presence of endogenous soluble antioxidants in the latter tissues. Furthermore, these studies demonstrated that CO was exclusively generated by subcellular fractions that contained membranes. The magnitude of the rate of product formation in brain supernatants depended on the concentration of Fe(II) and (or) Fe(III). The formation of CO, TBARS, CD, and LOOH increased linearly with time for up to 30 min, but the rates of product formation were different. Product formation was completely inhibited by BHT (100 microM), biliverdin (50 microM), bilirubin (50 microM), citrate (100 microM), and the Fe(II) chelators, desferrioxamine mesylate (100 microM) and diethylenetriaminepentaacetate, but not by 10 microM of the HO inhibitor, zinc deuteroporphyrin bis glycol. We conclude that CO generation is associated with the process of in vitro lipid peroxidation in tissues with limited antioxidant reserves.

  View details for Web of Science ID 000079780100001

  View details for PubMedID 10326826

• Light-emitting diodes: A novel light source for phototherapy PEDIATRIC RESEARCH Vreman, H. J., Wong, R. J., Stevenson, D. K., Route, R. K., Reader, S. D., Fejer, M. M., Gale, R., Seidman, D. S. 1998; 44 (5): 804-809

  Abstract

  High intensity light-emitting diodes (LEDs) are being studied as possible light sources for the phototherapy of hyperbilirubinemic neonates. These power-efficient, low heat-producing light sources have the potential to deliver high intensity light of narrow wavelength band in the blue-green portion of the visible light spectrum, which overlaps the absorption spectrum of bilirubin (BR). We compared the efficacy between single LEDs of different color and then constructed a prototype phototherapy device using 300 blue LEDs. The efficacy of this device was compared with that of conventional phototherapy devices by measuring the in vitro photodegradation of BR in human serum albumin. When blue, blue-green, green, and white LEDs were compared, the blue light was the most effective in degrading BR by 28% of dark control, followed by blue-green (18% of control), and then white light (14% of control). Green light was the least effective (11% of control). The prototype device with three focused arrays, each with 100 blue LEDs, generated greater irradiance (> 200 microW.cm-2.nm-1) than any of the conventional devices tested. It also supported the greatest rate of BR photodegradation. We conclude that light from LEDs should be considered a more effective treatment for hyperbilirubinemia than light from presently used phototherapy devices. Furthermore, the unique characteristics of this light source may make it especially suitable for use in safe and lightweight home phototherapy devices.

  View details for PubMedID 9803466

• Author Correction: Abrupt perturbation and delayed recovery of the vaginal ecosystem following childbirth. Nature communications Costello, E. K., DiGiulio, D. B., Robaczewska, A., Symul, L., Wong, R. J., Shaw, G. M., Stevenson, D. K., Holmes, S. P., Kwon, D. S., Relman, D. A. 2024; 15 (1): 1744

  View details for DOI 10.1038/s41467-024-46160-8

  View details for PubMedID 38409135

• Correction: Advances to diminish global newborn kernicterus mortality. Journal of perinatology : official journal of the California Perinatal Association Bhutani, V. K., Vidavalur, R., Wong, R. J. 2024

  View details for DOI 10.1038/s41372-024-01903-9

  View details for PubMedID 38383576

• Advances to diminish global newborn kernicterus mortality. Journal of perinatology : official journal of the California Perinatal Association Bhutani, V. K., Vidavalur, R., Wong, R. J. 2023

  Abstract

  Initiatives, "Every Newborn Action Plans" and "Sustainable Developmental Goals," are profoundly shaping global infant mortality trends. Concurrently, professional organizations recommended curricula to prevent extreme hyperbilirubinemia (EHB) sequelae. Therefore we assessed if these efforts have successfully decreased EHB-related mortality over time.We used the Global Burden of Diseases 2019 database to determine neonatal and infant mortality and the burden of kernicterus from 1990-2019.Globally, kernicterus accounted for 2.8 million infant deaths and trended downwards significantly from 1990 to 2019. By 2019, kernicterus-related mortality was 4 and 293 per million livebirths in high (HICs) and low income countries (LICs), respectively. 82% of deaths occurred in LICs and lower-middle income-countries. Average declines of mortality rates were 6.2% and 3.0% for HICs and LICs, respectively.Kernicterus-related mortality has been effectively reduced to <5 per million in HICs. Skills and knowledge transfer can potentially transform frontline services to bridge discordant kernicteric outcomes worldwide.

  View details for DOI 10.1038/s41372-023-01862-7

  View details for PubMedID 38151598

  View details for PubMedCentralID 3873706

• Microbiome Preterm Birth DREAM Challenge: Crowdsourcing Machine Learning Approaches to Advance Preterm Birth Research. medRxiv : the preprint server for health sciences Golob, J. L., Oskotsky, T. T., Tang, A. S., Roldan, A., Chung, V., Ha, C. W., Wong, R. J., Flynn, K. J., Parraga-Leo, A., Wibrand, C., Minot, S. S., Andreoletti, G., Kosti, I., Bletz, J., Nelson, A., Gao, J., Wei, Z., Chen, G., Tang, Z. Z., Novielli, P., Romano, D., Pantaleo, E., Amoroso, N., Monaco, A., Vacca, M., De Angelis, M., Bellotti, R., Tangaro, S., Kuntzleman, A., Bigcraft, I., Techtmann, S., Bae, D., Kim, E., Jeon, J., Joe, S., Theis, K. R., Ng, S., Lee Li, Y. S., Diaz-Gimeno, P., Bennett, P. R., MacIntyre, D. A., Stolovitzky, G., Lynch, S. V., Albrecht, J., Gomez-Lopez, N., Romero, R., Stevenson, D. K., Aghaeepour, N., Tarca, A. L., Costello, J. C., Sirota, M. 2023

  Abstract

  Globally, every year about 11% of infants are born preterm, defined as a birth prior to 37 weeks of gestation, with significant and lingering health consequences. Multiple studies have related the vaginal microbiome to preterm birth. We present a crowdsourcing approach to predict: (a) preterm or (b) early preterm birth from 9 publicly available vaginal microbiome studies representing 3,578 samples from 1,268 pregnant individuals, aggregated from raw sequences via an open-source tool, MaLiAmPi. We validated the crowdsourced models on novel datasets representing 331 samples from 148 pregnant individuals. From 318 DREAM challenge participants we received 148 and 121 submissions for our two separate prediction sub-challenges with top-ranking submissions achieving bootstrapped AUROC scores of 0.69 and 0.87, respectively. Alpha diversity, VALENCIA community state types, and composition (via phylotype relative abundance) were important features in the top performing models, most of which were tree based methods. This work serves as the foundation for subsequent efforts to translate predictive tests into clinical practice, and to better understand and prevent preterm birth.

  View details for DOI 10.1101/2023.03.07.23286920

  View details for PubMedID 36945505

  View details for PubMedCentralID PMC10029035

• VMAP: Vaginal Microbiome Atlas During Pregnancy. medRxiv : the preprint server for health sciences Parraga-Leo, A., Oskotsky, T. T., Oskotsky, B., Wibrand, C., Roldan, A., Tang, A., Ha, C. W., Wong, R. J., Minot, S. S., Andreoletti, G., Kosti, I., Theis, K. R., Ng, S., Lee, Y. S., Diaz-Gimeno, P., Bennett, P. R., MacIntyre, D. A., Lynch, S. V., Romero, R., Tarca, A. L., Stevenson, D. K., Aghaeepour, N., Golob, J., Sirota, M. 2023

  Abstract

  The vaginal microbiome has been shown to be associated with pregnancy outcomes including preterm birth (PTB) risk. Here we present VMAP: Vaginal Microbiome Atlas during Pregnancy (http://vmapapp.org), an application to visualize features of 3,909 vaginal microbiome samples of 1,416 pregnant individuals from 11 studies, aggregated from raw public and newly generated sequences via an open-source tool, MaLiAmPi. Our visualization tool (http://vmapapp.org) includes microbial features such as various measures of diversity, VALENCIA community state types (CST), and composition (via phylotypes and taxonomy). This work serves as a resource for the research community to further analyze and visualize vaginal microbiome data in order to better understand both healthy term pregnancies and those associated with adverse outcomes.

  View details for DOI 10.1101/2023.03.21.23286947

  View details for PubMedID 36993193

  View details for PubMedCentralID PMC10055588

• Molecular Mechanisms of Pregnancy-Related Vascular Remodeling and Pregnancy Complications. International journal of molecular sciences Stevenson, D. K., Wong, R. J., Nayak, N. R. 2023; 24 (4)

  Abstract

  The purpose of this editorial is to highlight the various observations made in this Special Issue in the International Journal of Molecular Sciences [...].

  View details for DOI 10.3390/ijms24043712

  View details for PubMedID 36835124

• Shorter maternal leukocyte telomere length following cesarean birth: Implications for future research Panelli, D. M., Mayo, J. A., Wong, R. J., Becker, M., Maric, I., Wu, E., Gotlib, I. H., Aghaeepour, N., Druzin, M. L., Stevenson, D. K., Shaw, G. M., Bianco, K. MOSBY-ELSEVIER. 2023: S456-S457

  View details for Web of Science ID 000909337401260

• ALTERATIONS IN PLACENTAL GENE EXPRESSION AND GLYCOGEN-POSITIVE CELLS IN OBESE PREGNANT MICE Zhao, H., Wong, R. J., Stevenson, D. K. SAGE PUBLICATIONS LTD. 2023: NP66

  View details for Web of Science ID 000972423800064

• Author Correction: Prediction of gestational age using urinary metabolites in term and preterm pregnancies. Scientific reports Contrepois, K., Chen, S., Ghaemi, M. S., Wong, R. J., Jehan, F., Sazawal, S., Baqui, A. H., Stringer, J. S., Rahman, A., Nisar, M. I., Dhingra, U., Khanam, R., Ilyas, M., Dutta, A., Mehmood, U., Deb, S., Hotwani, A., Ali, S. M., Rahman, S., Nizar, A., Ame, S. M., Muhammad, S., Chauhan, A., Khan, W., Raqib, R., Das, S., Ahmed, S., Hasan, T., Khalid, J., Juma, M. H., Chowdhury, N. H., Kabir, F., Aftab, F., Quaiyum, A., Manu, A., Yoshida, S., Bahl, R., Pervin, J., Price, J. T., Rahman, M., Kasaro, M. P., Litch, J. A., Musonda, P., Vwalika, B., Shaw, G., Stevenson, D. K., Aghaeepour, N., Snyder, M. P. 2022; 12 (1): 19753

  View details for DOI 10.1038/s41598-022-23715-7

  View details for PubMedID 36396676

• Leukocyte telomere dynamics across gestation in uncomplicated pregnancies and associations with stress. BMC pregnancy and childbirth Panelli, D. M., Leonard, S. A., Wong, R. J., Becker, M., Mayo, J. A., Wu, E., Girsen, A. I., Gotlib, I. H., Aghaeepour, N., Druzin, M. L., Shaw, G. M., Stevenson, D. K., Bianco, K. 2022; 22 (1): 381

  Abstract

  Short leukocyte telomere length is a biomarker associated with stress and morbidity in non-pregnant adults. Little is known, however, about maternal telomere dynamics in pregnancy. To address this, we examined changes in maternal leukocyte telomere length (LTL) during uncomplicated pregnancies and explored correlations with perceived stress.In this pilot study, maternal LTL was measured in blood collected from nulliparas who delivered live, term, singleton infants between 2012 and 2018 at a single institution. Participants were excluded if they had diabetes or hypertensive disease. Samples were collected over the course of pregnancy and divided into three time periods: < 200/7 weeks (Timepoint 1); 201/7 to 366/7 weeks (Timepoint 2); and 370/7 to 9-weeks postpartum (Timepoint 3). All participants also completed a survey assessing a multivariate profile of perceived stress at the time of enrollment in the first trimester. LTL was measured using quantitative polymerase chain reaction (PCR). Wilcoxon signed-rank tests were used to compare LTL differences within participants across all timepoint intervals. To determine whether mode of delivery affected LTL, we compared postpartum Timepoint 3 LTLs between participants who had vaginal versus cesarean birth. Secondarily, we evaluated the association of the assessed multivariate stress profile and LTL using machine learning analysis.A total of 115 samples from 46 patients were analyzed. LTL (mean ± SD), expressed as telomere to single copy gene (T/S) ratios, were: 1.15 ± 0.26, 1.13 ± 0.23, and 1.07 ± 0.21 for Timepoints 1, 2, and 3, respectively. There were no significant differences in LTL between Timepoints 1 and 2 (LTL T/S change - 0.03 ± 0.26, p = 0.39); 2 and 3 (- 0.07 ± 0.29, p = 0.38) or Timepoints 1 and 3 (- 0.07 ± 0.21, p = 0.06). Participants who underwent cesareans had significantly shorter postpartum LTLs than those who delivered vaginally (T/S ratio: 0.94 ± 0.12 cesarean versus 1.12 ± 0.21 vaginal, p = 0.01). In secondary analysis, poor sleep quality was the main stress construct associated with shorter Timepoint 1 LTLs (p = 0.02) and shorter mean LTLs (p = 0.03).In this cohort of healthy pregnancies, maternal LTLs did not significantly change across gestation and postpartum LTLs were shorter after cesarean than after vaginal birth. Significant associations between sleep quality and short LTLs warrant further investigation.

  View details for DOI 10.1186/s12884-022-04693-0

  View details for PubMedID 35501726

• Alterations of Placental Gene Expression in Obese Pregnant Mice Zhao, H., Wong, R. J., Stevenson, D. K. SPRINGER HEIDELBERG. 2022: 217-218

  View details for Web of Science ID 000762765300444

• Cellular aging and pregnancy complications: Examining maternal leukocyte telomere length in two diverse cohorts. Panelli, D. M., Wang, X., Wong, R. J., Cruz, G., Hong, X., Aghaeepour, N., Druzin, M. L., Shaw, G. M., Zuckerman, B. S., Stevenson, D. K., Bianco, K. MOSBY-ELSEVIER. 2022: S646

  View details for Web of Science ID 000737459401371

• Early-pregnancy prediction of risk for pre-eclampsia using maternal blood leptin/ceramide ratio: discovery and confirmation. BMJ open Huang, Q., Hao, S., You, J., Yao, X., Li, Z., Schilling, J., Thyparambil, S., Liao, W., Zhou, X., Mo, L., Ladella, S., Davies-Balch, S. R., Zhao, H., Fan, D., Whitin, J. C., Cohen, H. J., McElhinney, D. B., Wong, R. J., Shaw, G. M., Stevenson, D. K., Sylvester, K. G., Ling, X. B. 2021; 11 (11): e050963

  Abstract

  OBJECTIVE: This study aimed to develop a blood test for the prediction of pre-eclampsia (PE) early in gestation. We hypothesised that the longitudinal measurements of circulating adipokines and sphingolipids in maternal serum over the course of pregnancy could identify novel prognostic biomarkers that are predictive of impending event of PE early in gestation.STUDY DESIGN: Retrospective discovery and longitudinal confirmation.SETTING: Maternity units from two US hospitals.PARTICIPANTS: Six previously published studies of placental tissue (78 PE and 95 non-PE) were compiled for genomic discovery, maternal sera from 15 women (7 non-PE and 8 PE) enrolled at ProMedDx were used for sphingolipidomic discovery, and maternal sera from 40 women (20 non-PE and 20 PE) enrolled at Stanford University were used for longitudinal observation.OUTCOME MEASURES: Biomarker candidates from discovery were longitudinally confirmed and compared in parallel to the ratio of placental growth factor (PlGF) and soluble fms-like tyrosine kinase (sFlt-1) using the same cohort. The datasets were generated by enzyme-linked immunosorbent and liquid chromatography-tandem mass spectrometric assays.RESULTS: Our discovery integrating genomic and sphingolipidomic analysis identified leptin (Lep) and ceramide (Cer) (d18:1/25:0) as novel biomarkers for early gestational assessment of PE. Our longitudinal observation revealed a marked elevation of Lep/Cer (d18:1/25:0) ratio in maternal serum at a median of 23 weeks' gestation among women with impending PE as compared with women with uncomplicated pregnancy. The Lep/Cer (d18:1/25:0) ratio significantly outperformed the established sFlt-1/PlGF ratio in predicting impending event of PE with superior sensitivity (85% vs 20%) and area under curve (0.92 vs 0.52) from 5 to 25 weeks of gestation.CONCLUSIONS: Our study demonstrated the longitudinal measurement of maternal Lep/Cer (d18:1/25:0) ratio allows the non-invasive assessment of PE to identify pregnancy at high risk in early gestation, outperforming the established sFlt-1/PlGF ratio test.

  View details for DOI 10.1136/bmjopen-2021-050963

  View details for PubMedID 34824115

• Noninvasive Prediction of Preeclampsia in Pregnancy with Circulating RNA. Moufarrej, M. N., Wong, R. J., Campos, A. A., Quaintance, C. C., Sit, R. V., Tan, M., Neff, N. F., Druzin, M. L., Winn, V. D., Shaw, G. M., Stevenson, D. K., Quake, S. R. SPRINGER HEIDELBERG. 2021: 75A

  View details for Web of Science ID 000675441000068

• Multi-Omic, Longitudinal Profile of Third-Trimester Pregnancies Identifies a Molecular Switch That Predicts the Onset of Labor. Stelzer, I., Ghaemi, M., Han, X., Ando, K., Hedou, J., Feyaerts, D., Peterson, L., Ganio, E., Tsai, A., Tsai, E., Rumer, K., Stanley, N., Fallazadeh, R., Becker, M., Culos, A., Gaudilliere, D., Wong, R., Winn, V., Shaw, G., Snyder, M., Stevenson, D., Contrepois, K., Angst, M., Aghaeepour, N., Gaudilliere, B. SPRINGER HEIDELBERG. 2021: 233A-234A

  View details for Web of Science ID 000675441000486

• HO-1 Genetic Variants Display Racial Diversity and May Impact Hypertensive Disorders in Pregnancy. Sun, T., Mousavi, N., Wong, R. J., Sayed, N., Wu, J. C., Stevenson, D. K., Gymrek, M., Winn, V. D. SPRINGER HEIDELBERG. 2021: 105A

  View details for Web of Science ID 000675441000144

• Bilirubin/albumin (B/A) ratios correlate with unbound bilirubin levels in preterm infants. Pediatric research Abe, S. n., Fujioka, K. n., Nakasone, R. n., Suga, S. n., Ashina, M. n., Nishida, K. n., Wong, R. J., Iijima, K. n. 2021

  Abstract

  A strong correlation between the bilirubin/albumin (B/A) ratio and unbound bilirubin (UB) levels in newborns ≥35 weeks of gestation has been reported. However, in preterm infants, the usefulness of B/A ratios remains unclear.We obtained serum from 381 newborns <35 weeks of gestation. UB levels were measured using the glucose oxidase-peroxidase method. Total serum bilirubin (TB) and albumin (Alb) concentrations were measured spectrophotometrically. Samples were then stratified into two groups based on the infant's phototherapy use. B/A ratios were calculated and correlated with UB levels. Samples taken from infants prior to or never receiving phototherapy (No PTx) were then stratified by gestational age (GA) epochs: 22-27, 28-29, 30-31, and 32-34 weeks and B/A ratios correlated with UB levels.B/A ratios significantly correlated with UB levels in samples from the No PTx cohort (n = 1250; y = 1.83x - 0.15, r2 = 0.93) when compared with samples from infants post-phototherapy (Post-PTx, n = 2039; y = 1.05x + 0.09, r2 = 0.69). Even when stratified by GA, the correlation remained.In preterm infants <35 weeks of gestation, B/A ratios correlated with UB levels better in infants prior to or never receiving phototherapy than in those infants receiving phototherapy.The bilirubin/albumin (B/A) ratio significantly correlates with unbound bilirubin (UB) levels in preterm infants <35 weeks of gestation. The B/A ratio can be used as an index of UB levels in preterm infants <35 weeks of gestation. The B/A ratio is useful, especially when UB measurements are not available, for managing hyperbilirubinemia in preterm infants.

  View details for DOI 10.1038/s41390-020-01351-z

  View details for PubMedID 33469181

• Inflammation-induced alterations in maternal-fetal Heme Oxygenase (HO) are associated with sustained innate immune cell dysregulation in mouse offspring. PloS one Ozen, M., Zhao, H., Kalish, F., Yang, Y., Jantzie, L. L., Wong, R. J., Stevenson, D. K. 2021; 16 (6): e0252642

  Abstract

  Heme oxygenase-1 (HO-1) is an evolutionarily conserved stress response enzyme and important in pregnancy maintenance, fetal and neonatal outcomes, and a variety of pathologic conditions. Here, we investigated the effects of an exposure to systemic inflammation late in gestation [embryonic day (E)15.5] on wild-type (Wt) and HO-1 heterozygous (Het, HO-1+/-) mothers, fetuses, and offspring. We show that alterations in fetal liver and spleen HO homeostasis during inflammation late in gestation can lead to a sustained dysregulation of innate immune cell populations and intracellular myeloid HO-1 expression in the spleen through young adolescence [postnatal day 25] in mice.

  View details for DOI 10.1371/journal.pone.0252642

  View details for PubMedID 34086785

• Corrigendum: Investigating Pregnancy and Its Complications Using Circulating Cell-Free RNA in Women's Blood During Gestation. Frontiers in pediatrics Moufarrej, M. N., Wong, R. J., Shaw, G. M., Stevenson, D. K., Quake, S. R. 2021; 9: 680201

  Abstract

  [This corrects the article DOI: 10.3389/fped.2020.605219.].

  View details for DOI 10.3389/fped.2021.680201

  View details for PubMedID 33903851

  View details for PubMedCentralID PMC8065098

• DO BILIRUBIN/ALBUMIN (B/A) RATIOS CORRELATE WITH UNBOUND BILIRUBIN LEVELS IN PRETERM INFANTS? Abe, S., Fujioka, K., Wong, R. J., Iijima, K. BMJ PUBLISHING GROUP. 2021: 126–27

  View details for DOI 10.1136/jim-2021-WRMC.56

  View details for Web of Science ID 000608729100074

• DECREASED MORTALITY RATE AMONG COVID-19 PATIENTS USING STATINS: DATA FROM US ELECTRONIC HEALTH RECORDS Oskotsky, T., Maric, I., Kosti, I., Le, B. L., Wong, R. J., Shaw, G. M., Sirota, M., Stevenson, D. K. BMJ PUBLISHING GROUP. 2021: 219–20

  View details for DOI 10.1136/jim-2021-WRMC.262

  View details for Web of Science ID 000608729100280

• IN VITRO INHIBITORY POTENCY OF D-PENICILLAMINE ON HEME OXYGENASE ISOZYME ACTIVITY Fuwa, K., Konecny, C. M., Stevenson, D. K., Wong, R. J. BMJ PUBLISHING GROUP. 2021: 111–12

  View details for DOI 10.1136/jim-2021-WRMC.22

  View details for Web of Science ID 000608729100040

• High-throughput quantitation of serological ceramides/dihydroceramides by LC/MS/MS: Pregnancy baseline biomarkers and potential metabolic messengers. Journal of pharmaceutical and biomedical analysis Huang, Q. n., Hao, S. n., Yao, X. n., You, J. n., Li, X. n., Lai, D. n., Han, C. n., Schilling, J. n., Hwa, K. Y., Thyparambil, S. n., Whitin, J. n., Cohen, H. J., Chubb, H. n., Ceresnak, S. R., McElhinney, D. B., Wong, R. J., Shaw, G. M., Stevenson, D. K., Sylvester, K. G., Ling, X. B. 2020; 192: 113639

  Abstract

  Ceramides and dihydroceramides are sphingolipids that present in abundance at the cellular membrane of eukaryotes. Although their metabolic dysregulation has been implicated in many diseases, our knowledge about circulating ceramide changes during the pregnancy remains limited. In this study, we present the development and validation of a high-throughput liquid chromatography-tandem mass spectrometric method for simultaneous quantification of 16 ceramides and 10 dihydroceramides in human serum within 5 min. by using stable isotope-labeled ceramides as internal standards. This method employs a protein precipitation method for high throughput sample preparation, reverse phase isocratic elusion for chromatographic separation, and Multiple Reaction Monitoring for mass spectrometric detection. To qualify for clinical applications, our assay has been validated against the FDA guidelines for Lower Limit of Quantitation (1 nM), linearity (R2>0.99), precision (imprecision<15 %), accuracy (inaccuracy<15 %), extraction recovery (>90 %), stability (>85 %), and carryover (<0.01 %). With enhanced sensitivity and specificity from this method, we have, for the first time, determined the serological levels of ceramides and dihydroceramides to reveal unique temporal gestational patterns. Our approach could have value in providing insights into disorders of pregnancy.

  View details for DOI 10.1016/j.jpba.2020.113639

  View details for PubMedID 33017796

• A global approach to prevent kernicterus: A dedication to Lois Johnson-Hamerman (Aug. 1, 1927 to Aug. 11, 2019). Seminars in perinatology Bhutani, V. K., Wong, R. J. 2020: 151350

  View details for DOI 10.1016/j.semperi.2020.151350

  View details for PubMedID 33339605

• Sixty years of phototherapy for neonatal jaundice: from serendipitous observation to standardized treatment and rescue for millions. Journal of perinatology : official journal of the California Perinatal Association Hansen, T. W., Jeffrey Maisels, M. n., Ebbesen, F. n., Vreman, H. J., Stevenson, D. K., Wong, R. J., Bhutani, V. K. 2020

  View details for DOI 10.1038/s41372-020-0712-3

  View details for PubMedID 32561834

• A multi-center evaluation of a device for measurement of bilirubin binding capacity in neonates: the effects of gestational age, Intralipid exposure and illness severity. Journal of perinatology : official journal of the California Perinatal Association Schutzman, D. L., Bhutani, V. K., Castillo Cuadrado, M. E., Lamola, A. A., Frantz, I., Obregon, E., Wong, R. J. 2019

  Abstract

  OBJECTIVE: Measure daily bilirubin-binding capacity (BBC) variation using an automated, not as-yet FDA approved, Point-of-Care hematofluorometer. Measure the effects of prematurity, clinical instability and exposure to Intralipid on BBC.SUBJECTS: Convenience sample of 109 infants from well-baby and intensive care nurseries. Gestational ages 28-41 weeks. 261 specimens obtained from postnatal ages 1-4 days. Unstable neonates were defined by need for at least noninvasive respiratory support and FiO2≥0.25.RESULTS: Median interday variation was 2.9±5.1mg/dL. BBC (0.254mg/dL/wk) and albumin (0.037g/dL/wk) increased for each week of gestation. BBC was lower in unstable compared to well infants (26.1±7.6mg/dL v 28.6±6.3mg/dL). BBC was not significantly different in infants receiving or not receiving IL.CONCLUSIONS: BBC measurements using the device had acceptable intraspecimen reproducibility and interday variability. BBC may be helpful in guiding the assessment of aggressive versus conservative management decisions in preterm and sick infants with hyperbilirubinemia.

  View details for PubMedID 30918342

• Data-Driven Queries between Medications and Spontaneous Preterm Birth among 2.5 Million Pregnancies: Association with Genital Herpes and Antiviral Drugs. Maric, I., Borisenko, E., Winn, V. D., Weber, K. A., Wong, R. J., Aziz, N., Blumenfeld, Y. J., El-Sayed, Y. Y., Stevenson, D. K., Shaw, G. M. SAGE PUBLICATIONS INC. 2019: 389A

  View details for Web of Science ID 000459610400965

• Soluble Factors in the Murine Placenta Regulate Gene Expression in Alternatively-Activated Macrophages. Zhao, H., Kalish, F., Wong, R. J., Stevenson, D. K. SAGE PUBLICATIONS INC. 2019: 293A

  View details for Web of Science ID 000459610400682

• Differential Dynamics of the Maternal Immune System in Healthy Pregnancy and Preeclampsia. Han, X., Ghaemi, M. S., Ando, K., Peterson, L., Ganio, E. A., Tsai, A. S., Gaudilliere, D., Einhaus, J., Tsai, E. S., Stanley, N. M., Culos, A., Taneda, A. H., Fallahzadeh, R., Wong, R. J., Winn, V. D., Stevenson, D. K., Angst, M. S., Aghaeepour, N., Gaudilliere, B. SAGE PUBLICATIONS INC. 2019: 271A

  View details for Web of Science ID 000459610400616

• Correction: The effect of light wavelength on in vitro bilirubin photodegradation and photoisomer production. Pediatric research Vreman, H. J., Kourula, S., Jasprova, J., Ludvikova, L., Klan, P., Muchova, L., Vitek, L., Cline, B. K., Wong, R. J., Stevenson, D. K. 2019

  Abstract

  Following publication of this article, the authors noticed that an incorrect affiliation was assigned to the author "Lucie Muchova". The original article has now been updated so that the author "Lucie Muchova" is associated with the "Institute of Medical Biochemistry and Laboratory Diagnostics, 1st Faculty of Medicine, Charles University, Katerinska 32, 120 00 Prague, Czech Republic". This has been corrected in both the PDF and HTML versions of the article.

  View details for PubMedID 30814644

• "Following through": addressing the racial inequality for preterm infants and their families. Pediatric research Stevenson, D. K., Wong, R. J., Profit, J. n., Shaw, G. M., Jason Wang, C. n., Lee, H. C. 2019

  View details for DOI 10.1038/s41390-019-0602-6

  View details for PubMedID 31581171

• IN VITRO INHIBITORY POTENCY OF ZINC PROTOPORPHYRIN MICROSPHERES ON HEME OXYGENASE ISOZYME ACTIVITY Jacobsen, D., Iwatani, S., Onderdonk, Z., Wong, R. T., Wong, R. J., Stevenson, D. K. BMJ PUBLISHING GROUP. 2019: 117

  View details for DOI 10.1136/jim-2018-000939.115

  View details for Web of Science ID 000457712500125

• A NOVEL POINT-OF-CARE DEVICE FOR MEASURING GLUCOSE-6-PHOSPHATE DEHYDROGENASE ENZYME DEFICIENCY Montiel, C., Kunda, M., Stevenson, D. K., Wong, R. J., Bhutani, V. K. BMJ PUBLISHING GROUP. 2019: 117

  View details for DOI 10.1136/jim-2018-000939.116

  View details for Web of Science ID 000457712500126

• BILIRUBIN PRODUCTION IS INCREASED IN NEWBORN MICE TREATED WITH LIPOPOLYSACCHARIDE Iwatani, S., Jacobsen, D., Wong, R. T., Onderdonk, Z., Wong, R. J., Stevenson, D. K. BMJ PUBLISHING GROUP. 2019: 117–18

  View details for DOI 10.1136/jim-2018-000939.117

  View details for Web of Science ID 000457712500127

• Author Correction: Enabling precision medicine in neonatology, an integrated repository for preterm birth research. Scientific data Sirota, M., Thomas, C. G., Liu, R., Zuhl, M., Banerjee, P., Wong, R. J., Quaintance, C. C., Leite, R., Chubiz, J., Anderson, R., Chappell, J., Kim, M., Grobman, W., Zhang, G., Rokas, A., Muglia, L. J., Ober, C., England, S. K., Macones, G., Driscoll, D., Parry, S., Shaw, G. M., Stevenson, D. K., Simpson, J. L., Thomson, E., Butte, A. J., March of Dimes Prematurity Research Centers, Driscoll, D., Macones, G., Muglia, L. J., Ober, C., Stevenson, D. K. 2018; 5 (1): 3

  Abstract

  The original version of the Data Descriptor contained errors in the author list and affiliations. Rita Leite's first name was misspelled as "Rite" and affiliations 4 and 5 were incorrectly swapped. In addition, members of the March of Dimes Prematurity Research Center consortium were not listed in the agreed positions within the author list. These errors have now been corrected in the HTML and PDF versions.

  View details for PubMedID 30563979

• Enabling precision medicine in neonatology, an integrated repository for preterm birth research (vol 5, 180219, 2018) SCIENTIFIC DATA Sirota, M., Thomas, C. G., Liu, R., Zuhl, M., Banerjee, P., Wong, R. J., Quaintance, C. C., Leite, R., Chubiz, J., Anderson, R., Chappell, J., Kim, M., Grobman, W., Zhang, G., Rokas, A., Muglia, L. J., Ober, C., England, S. K., Macones, G., Driscoll, D., Parry, S., Shaw, G. M., Stevenson, D. K., Simpson, J., Thomson, E., Butte, A. J., March Dimes Prematurity Res Ctrs 2018; 5

  View details for DOI 10.1038/s41597-018-0004-3

  View details for Web of Science ID 000453586800001

• Enabling precision medicine in neonatology, an integrated repository for preterm birth research SCIENTIFIC DATA Sirota, M., Thomas, C. G., Liu, R., Zuhl, M., Banerjee, P., Wong, R. J., Quaintance, C. C., Leite, R., Chubiz, J., Anderson, R., Chappell, J., Kim, M., Grobman, W., Zhang, G., Rokas, A., England, S. K., Parry, S., Shaw, G. M., Simpson, J., Thomson, E., Butte, A. J., March Dimes Prematurity Res Ctr 2018; 5

  View details for DOI 10.1038/sdata.2018.219

  View details for Web of Science ID 000449252700001

• Impact of post-collection freezing delay on the reliability of serum metabolomics in samples reflecting the California mid-term pregnancy biobank METABOLOMICS La Frano, M. R., Carmichael, S. L., Ma, C., Hardley, M., Shen, T., Wong, R., Rosales, L., Borkowski, K., Pedersen, T. L., Shaw, G. M., Stevenson, D. K., Fiehn, O., Newman, J. W. 2018; 14 (11)

  View details for DOI 10.1007/s11306-018-1450-9

  View details for Web of Science ID 000450525400001

• Filtered sunlight versus intensive electric powered phototherapy in moderate-to-severe neonatal hyperbilirubinaemia: a randomised controlled non-inferiority trial LANCET GLOBAL HEALTH Slusher, T. M., Vreman, H. J., Brearley, A. M., Vaucher, Y. E., Wong, R. J., Stevenson, D. K., Adeleke, O. T., Ojo, I. P., Edowhorhu, G., Lund, T. C., Gbadero, D. A. 2018; 6 (10): E1122–E1131

  Abstract

  Kernicterus resulting from severe neonatal hyperbilirubinaemia is a leading cause of preventable deaths and disabilities in low-income and middle-income countries, partly because high-quality intensive phototherapy is unavailable. Previously, we showed that filtered-sunlight phototherapy (FSPT) was efficacious and safe for treatment of mild-to-moderate neonatal hyperbilirubinaemia. We aimed to extend these studies to infants with moderate-to-severe hyperbilirubinaemia.We did a prospective, randomised controlled non-inferiority trial in Ogbomoso, Nigeria-a simulated rural setting. Near-term or term infants aged 14 days or younger who were of 35 weeks or more gestational age and with total serum bilirubin concentrations at or above the recommended age-dependent treatment levels for high-risk neonates were randomly assigned (1:1) to either FSPT or intensive electric phototherapy (IEPT). Randomisation was computer-generated, and neither clinicians nor the parents or guardians of participants were masked to group allocation. FSPT was delivered in a transparent polycarbonate room lined with commercial tinting films that transmitted effective phototherapeutic light, blocked ultraviolet light, and reduced infrared radiation. The primary outcome was efficacy, which was based on assessable treatment days only (ie, those on which at least 4 h of phototherapy was delivered) and defined as a rate of increase in total serum bilirubin concentrations of less than 3·4 μmol/L/h in infants aged 72 h or younger, or a decrease in total serum bilirubin concentrations in those older than 72 h. Safety was defined as no sustained hypothermia, hyperthermia, dehydration, or sunburn and was based on all treatment days. Analysis was by intention to treat with a non-inferiority margin of 10%.Between July 31, 2015, and April 30, 2017, 174 neonates were enrolled and randomly assigned: 87 to FSPT and 87 to IEPT. Neonates in the FSPT group received 215 days of phototherapy, 82 (38%) of which were not assessable. Neonates in the IEPT group received 219 treatment days of phototherapy, 67 (31%) of which were not assessable. Median irradiance was 37·3 μW/cm2/nm (IQR 21·4-56·4) in the FSPT group and 50·4 μW/cm2/nm (44·5-66·2) in the IEPT group. FSPT was efficacious on 116 (87·2%) of 133 treatment days; IEPT was efficacious on 135 (88·8%) of 152 treatment days (mean difference -1·6%, 95% CI -9·9 to 6·7; p=0·8165). Because the CI did not extend below -10%, we concluded that FSPT was not inferior to IEPT. Treatment was safe for all neonates.FSPT is safe and no less efficacious than IEPT for treatment of moderate-to-severe neonatal hyperbilirubinaemia in near-term and term infants.Thrasher Research Fund and National Center for Advancing Translational Sciences.

  View details for PubMedID 30170894

• In Vitro Study on the Effect of Maraviroc or Dolutegravir on Bilirubin to Albumin Binding PEDIATRIC INFECTIOUS DISEASE JOURNAL Schreiner, C. N., Ahlfors, C. E., Wong, R. J., Stevenson, D. K., Clarke, D. F., Mirochnick, M. 2018; 37 (9): 908–9

  Abstract

  We performed an in vitro evaluation of the effect of maraviroc or dolutegravir on bilirubin to albumin binding. At typical treatment and low albumin concentrations, maraviroc had no impact, while dolutegravir affected bilirubin to albumin binding to an equivalent extent as sulfisoxazole. However in vivo, neither is likely to significantly impact bilirubin to albumin binding because of their low concentrations relative to albumin.

  View details for PubMedID 29561509

• Maternal Height and Risk of Preeclampsia. Maric, I., Mayo, J. A., Druzin, M. L., Wong, R. J., Winn, V. D., Stevenson, D. K., Shaw, G. M. SAGE PUBLICATIONS INC. 2018: 207A–208A

  View details for Web of Science ID 000429928200458

• Heme Oxygenase-1 Deficiency Results in T-Cell Dysregulation in Offspring of Mothers Exposed to Late Gestational Inflammation Ozen, M., Zhao, H., Kalish, F., Yang, Y., Wong, R. J., Stevenson, D. K. SAGE PUBLICATIONS INC. 2018: 248A–249A

  View details for Web of Science ID 000429928200583

• Pravastatin Induces Heme Oxygenase Activity and Enhances Placental Development in a Murine Model. Tsur, A., Kalish, F., Burgess, J., Zhao, H., Casey, K. M., Druzin, M. L., Wong, R. J., Stevenson, D. K. SAGE PUBLICATIONS INC. 2018: 155A–156A

  View details for Web of Science ID 000429928200299

• The Effects of Hypoxia and Progesterone/Estrogen on Polarized Macrophages In Vitro. Zhao, H., Kalish, F., Wong, R. J., Stevenson, D. K. SAGE PUBLICATIONS INC. 2018: 301A–302A

  View details for Web of Science ID 000429928200747

• Mass Cytometry and Proteomic Based Prediction of the Onset of Labor. Ando, K., Han, X., Ghaemi, S., Tsai, A., Ganio, E., Gaudilliere, D., Culos, T., Shaw, G., Wong, R., Stevenson, D., Carvalho, B., Tingle, M., Angst, M., Aghaeepor, N., Gaudilliere, B., Stanford March Dimes Prematurity SAGE PUBLICATIONS INC. 2018: 153A

  View details for Web of Science ID 000429928200292

• Pravastatin improves pregnancy outcome and placental development in heme oxygenase-1 deficient mice Tsur, A., Kalish, F., Burgess, J., Zhao, H., Casey, K., Druzin, M. L., Wong, R. J., Stevenson, D. K. MOSBY-ELSEVIER. 2018: S202

  View details for Web of Science ID 000422946900324

• Bilirubin Production Is Increased in Newborn Mice Exposed to Isoflurane. Neonatology Iwatani, S., Burgess, J., Kalish, F., Wong, R. J., Stevenson, D. K. 2018; 115 (1): 21-27

  Abstract

  Increased bilirubin production due to hemolysis can lead to severe neonatal hyperbilirubinemia and, if left untreated, to bilirubin neurotoxicity. Post-cardiac surgery newborns have been shown to be at an increased risk for developing hyperbilirubinemia and also hemolysis. Isoflurane (ISO), a volatile anesthetic agent routinely used in newborn surgery, has been reported to upregulate heme oxygenase 1 (HO-1) expression. HO is the rate-limiting enzyme in the bilirubin production pathway.Here, we evaluated whether ISO exposure induces HO-1 and further increases bilirubin production in a hemolytic newborn mouse model.Three-day-old newborn mice were exposed to 2% ISO for 18 min or air. Liver HO activity and HO-1 protein were measured after exposure to ISO. Next, we evaluated the effect of ISO exposure on bilirubin production as indexed by the total body excretion rate of carbon monoxide following heme loading.ISO significantly increased liver HO activity 120% and 116% at 24 and 48 h, respectively, after exposure. HO-1 protein levels also similarly increased after ISO exposure, but the increases were not statistically significant compared with controls. After heme loading, ISO-exposed pups had significantly higher bilirubin production rates (1.24-fold), and also peaked earlier, than age-matched nonexposed pups.ISO exposure can induce HO-1 expression in the liver and may explain the development of severe hyperbilirubinemia in postsurgical infants, especially in those undergoing hemolysis.

  View details for DOI 10.1159/000492421

• NEONATAL HYPERBILIRUBINEMIA AFTER MECHANICAL CIRCULATORY SUPPORT Bhombal, S., Dasani, R., Davis, A., Axelrod, D. M., Wong, R. J., Bhutani, V. K. BMJ PUBLISHING GROUP. 2018: 164–65

  View details for DOI 10.1136/jim-2017-000663.236

  View details for Web of Science ID 000432007400247

• TREATMENT WITH PRAVASTATIN IMPROVES PREGNANCY OUTCOME AND PLACENTAL DEVELOPMENT IN HEME OXYGENASE-1-DEFICIENT MICE Tsur, A., Kalish, F., Burgess, J., Zhao, H., Casey, K. M., Druzin, M. L., Wong, R. J., Stevenson, D. K. BMJ PUBLISHING GROUP. 2018: 241–42

  View details for DOI 10.1136/jim-2017-000663.422

  View details for Web of Science ID 000432007400432

• CYCLED PHOTOTHERAPY IS A SAFE AND EFFECTIVE TREATMENT FOR SMALL PREMATURE INFANTS WITH HYPERBILIRUBINEMIA Arnold, C. C., Tyson, J. E., Cuadrado, C. E., Dempsey, A. G., Khan, A. M., Pedroza, C., Bhutani, V. K., Wong, R. J., Stevenson, D. K. BMJ PUBLISHING GROUP. 2017: 150–51

  View details for DOI 10.1136/jim-2016-000365.120

  View details for Web of Science ID 000433055100121

• 'The obesity paradox': a reconsideration of obesity and the risk of preterm birth. Journal of perinatology : official journal of the California Perinatal Association Tsur, A. n., Mayo, J. A., Wong, R. J., Shaw, G. M., Stevenson, D. K., Gould, J. B. 2017

  Abstract

  The association between obesity and spontaneous preterm births (sPTBs) has been shown to be influenced by obesity-attendant comorbidities. Our objective was to better understand the complex relationship of obesity and its attendant comorbidities with sPTBs.A retrospective analysis utilizing maternally linked hospital and birth certificate records of 2 049 196 singleton California deliveries from 2007 to 2011. Adjusted relative risks (aRRs) for sPTBs were estimated using multivariate Poisson regression modeling.Obese women had higher aRRs for sPTBs than their normal body mass index (BMI) controls. aRRs (95% confidence interval) increased with increasing BMI category: Obese I=1.10 (1.08 to 1.12); Obese II=1.15 (1.12 to 1.18); and Obese III=1.26 (1.22 to 1.30). When comparing only obese women without comorbidities to their normal BMI controls, aRRs reversed, that is, obese women had lower aRRs of sPTBs: Obese I=0.96 (0.94 to 0.98), Obese II=0.95 (0.91 to 0.98); and Obese III=0.98 (0.94 to 1.03). This same reversal of aRR direction was also observed among women with comorbidities: 0.92 (0.89 to 0.96); 0.89 (0.85 to 0.93); and 0.89 (0.85 to 0.93), respectively. Increasing BMI increased the aRRs for sPTBs among patients with gestational diabetes (P<0.05), while decreasing the risk among patients with chronic hypertension and pregnancy-related hypertensive disease (P<0.05).The obesity and preterm birth paradox is an example of what has been described as 'Simpson's Paradox'. Unmeasured confounding factors mediated by comorbidities may explain the observed protective effect of obesity upon conditioning on the presence or absence of comorbidities and thus resolve the paradox.Journal of Perinatology advance online publication, 27 July 2017; doi:10.1038/jp.2017.104.

  View details for PubMedID 28749482

• HEME OXYGENASE-1 DEFICIENCY INCREASES THE SEVERITY OF SEPSIS IN A PRETERM MOUSE MODEL Fujioka, K., Kalish, F., Zhao, H., Wong, R. J., Stevenson, D. K. BMJ PUBLISHING GROUP. 2017: 114–15

  View details for DOI 10.1136/jim-2016-000365.39

  View details for Web of Science ID 000433055100040

• IDENTIFICATION OF RISK FOR NEONATAL HEMOLYSIS: A MULTI-CENTER STUDY Bhutani, V. K., Cuadrado, C. E., Schutzman, D. L., Aby, J. L., Bogen, D. L., Christensen, R. D., Watchko, J. F., Maisels, M., Wong, R. J., Stevenson, D. K. BMJ PUBLISHING GROUP. 2017: 123

  View details for DOI 10.1136/jim-2016-000365.57

  View details for Web of Science ID 000433055100058

• Identification of neonatal haemolysis: an approach to predischarge management of neonatal hyperbilirubinemia ACTA PAEDIATRICA Bhutani, V. K., Srinivas, S., Cuadrado, M. E., Aby, J. L., Wong, R. J., Stevenson, D. K. 2016; 105 (5): E189-E194

  Abstract

  Relative contributions of increased production [by end-tidal carbon monoxide concentrations (ETCOc)] and decreased elimination of bilirubin to predischarge hour-specific total bilirubin (TB) levels were assessed in healthy late-preterm and term newborns. Secondly, we report predischarge ETCOc ranges to guide clinical management of hyperbilirubinemia.TB and ETCOc (≤3 timepoints) determinations of newborns aged between six hours and <6 days (n = 79) were stratified by postnatal age epochs. Hyperbilirubinemia risk was assessed by plotting TB values as a function of ETCOc.Stratifications of ETCOc (in ppm, mean, median and interquartile ranges) by postnatal age epochs (0-24, 24-48 and 48-72) were as follows: 2.0, 1.9, 1.8-2.2 (n = 11); 1.6, 1.5, 1.1-2.0 (n = 58); and 2.0, 1.8, 1.6-2.3 (n = 9), respectively. Infants with ETCOc ≥ 2.5 were at high risk, between 1.5 and 2.5 at moderate risk and ≤1.5 were at low risk. Risk due to haemolysis alone was not independent (p < 0.01). For infants with TB >75th percentile (n = 31), 23% had ETCO ≤1.5, and 77% had ETCOc > 1.5 (p < 0.00003).Near-simultaneous ETCOc and TB measurements in infants with TB >75th percentile accurately identify haemolytic hyperbilirubinemia.

  View details for DOI 10.1111/apa.13341

  View details for Web of Science ID 000373921200001

• A Randomized Trial of Phototherapy with Filtered Sunlight in African Neonates. New England journal of medicine Slusher, T. M., Olusanya, B. O., Vreman, H. J., Brearley, A. M., Vaucher, Y. E., Lund, T. C., Wong, R. J., Emokpae, A. A., Stevenson, D. K. 2015; 373 (12): 1115-1124

  Abstract

  Sequelae of severe neonatal hyperbilirubinemia constitute a substantial disease burden in areas where effective conventional phototherapy is unavailable. We previously found that the use of filtered sunlight for the purpose of phototherapy is a safe and efficacious method for reducing total bilirubin. However, its relative safety and efficacy as compared with conventional phototherapy are unknown.We conducted a randomized, controlled noninferiority trial in which filtered sunlight was compared with conventional phototherapy for the treatment of hyperbilirubinemia in term and late-preterm neonates in a large, urban Nigerian maternity hospital. The primary end point was efficacy, which was defined as a rate of increase in total serum bilirubin of less than 0.2 mg per deciliter per hour for infants up to 72 hours of age or a decrease in total serum bilirubin for infants older than 72 hours of age who received at least 5 hours of phototherapy; we prespecified a noninferiority margin of 10% for the difference in efficacy rates between groups. The need for an exchange transfusion was a secondary end point. We also assessed safety, which was defined as the absence of the need to withdraw therapy because of hyperthermia, hypothermia, dehydration, or sunburn.We enrolled 447 infants and randomly assigned 224 to filtered sunlight and 223 to conventional phototherapy. Filtered sunlight was efficacious on 93% of treatment days that could be evaluated, as compared with 90% for conventional phototherapy, and had a higher mean level of irradiance (40 vs. 17 μW per square centimeter per nanometer, P<0.001). Temperatures higher than 38.0°C occurred in 5% of the infants receiving filtered sunlight and in 1% of those receiving conventional phototherapy (P<0.001), but no infant met the criteria for withdrawal from the study for reasons of safety or required an exchange transfusion.Filtered sunlight was noninferior to conventional phototherapy for the treatment of neonatal hyperbilirubinemia and did not result in any study withdrawals for reasons of safety. (Funded by the Thrasher Research Fund, Salt Lake City, and the National Center for Advancing Translational Sciences of the National Institutes of Health; Clinical Trials.gov number, NCT01434810.).

  View details for DOI 10.1056/NEJMoa1501074

  View details for PubMedID 26376136

• Heme oxygenase-1 promoter polymorphisms and risk of spina bifida. Birth defects research. Part A, Clinical and molecular teratology Fujioka, K., Yang, W., Wallenstein, M. B., Zhao, H., Wong, R. J., Stevenson, D. K., Shaw, G. M. 2015; 103 (9): 741-746

  Abstract

  Spina bifida is the most common form of neural tube defects (NTDs). Etiologies of NTDs are multifactorial, and oxidative stress is believed to play a key role in NTD development. Heme oxygenase (HO), the rate-limiting enzyme in heme degradation, has multiple protective properties including mediating antioxidant processes, making it an ideal candidate for study. The inducible HO isoform (HO-1) has two functional genetic polymorphisms: (GT)n dinucleotide repeats and A(-413)T SNP (rs2071746), both of which can affect its promoter activity. However, no study has investigated a possible association between HO-1 genetic polymorphisms and risk of NTDs.This case-control study included 152 spina bifida cases (all myelomeningoceles) and 148 nonmalformed controls obtained from the California Birth Defects Monitoring Program reflecting births during 1990 to 1999. Genetic polymorphisms were determined by polymerase chain reaction and amplified fragment length polymorphisms/restriction fragment length polymorphisms using genomic DNA extracted from archived newborn blood spots. Genotype and haplotype frequencies of two HO-1 promoter polymorphisms between cases and controls were compared.For (GT)n dinucleotide repeat lengths and the A(-413)T SNP, no significant differences in allele frequencies or genotypes were found. Linkage disequilibrium was observed between the HO-1 polymorphisms (D': 0.833); however, haplotype analyses did not show increased risk of spina bifida overall or by race/ethnicity.Although, an association was not found between HO-1 polymorphisms and risk of spina bifida, we speculate that the combined effect of low HO-1 expression and exposures to known environmental oxidative stressors (low folate status or diabetes), may overwhelm antioxidant defenses and increase risk of NTDs and warrants further study. Birth Defects Research (Part A), 2014. © 2014 Wiley Periodicals, Inc.

  View details for DOI 10.1002/bdra.23343

  View details for PubMedID 26173399

• Bilirubin production and hour-specific bilirubin levels JOURNAL OF PERINATOLOGY Bhutani, V. K., Wong, R. J., Vreman, H. J., Stevenson, D. K. 2015; 35 (9): 735-738

  Abstract

  We assessed the relative contributions of increased bilirubin production (indexed by end-tidal carbon monoxide (CO) concentrations, corrected for ambient CO (ETCOc)) to hour-specific total bilirubin (TB) levels in healthy late preterm and term newborns.Post hoc analyses of concurrent ETCOc and TB (at 30±6 h of age) and follow-up TB levels at age 96±12 h and up to 168 h after birth were performed in a cohort of 641 term and late preterm infants.Increased bilirubin production (hour-specific ETCOc ⩾1.7 p.p.m. at age 30±6 h) was noted in ~80%, 42% and 32% of infants in the high-, intermediate- and low-risk TB zones, respectively. One infant with TB <40th percentile and ETCOc <1.7 p.p.m. developed TB ⩾95th percentile at age 168 h, probably due to decreased bilirubin elimination.Infants in the high-risk quartile of the hour-specific bilirubin nomogram have a higher mean bilirubin production. Infants with TB levels ⩾95th percentile without increased bilirubin production have impaired bilirubin elimination.

  View details for DOI 10.1038/jp.2015.32

  View details for Web of Science ID 000360408600015

• Implementing Mass Cytometry at the Bedside to Study the Immunological Basis of Human Diseases: Distinctive Immune Features in Patients with a History of Term or Preterm Birth. Cytometry. Part A : the journal of the International Society for Analytical Cytology Gaudillière, B., Ganio, E. A., Tingle, M., Lancero, H. L., Fragiadakis, G. K., Baca, Q. J., Aghaeepour, N., Wong, R. J., Quaintance, C., El-Sayed, Y. Y., Shaw, G. M., Lewis, D. B., Stevenson, D. K., Nolan, G. P., Angst, M. S. 2015; 87 (9): 817-829

  Abstract

  Single-cell technologies have immense potential to shed light on molecular and biological processes that drive human diseases. Mass cytometry (or Cytometry by Time Of Flight mass spectrometry, CyTOF) has already been employed in clinical studies to comprehensively survey patients' circulating immune system. As interest in the "bedside" application of mass cytometry is growing, the delineation of relevant methodological issues is called for. This report uses a newly generated dataset to discuss important methodological considerations when mass cytometry is implemented in a clinical study. Specifically, the use of whole blood samples versus peripheral blood mononuclear cells (PBMCs), design of mass-tagged antibody panels, technical and analytical implications of sample barcoding, and application of traditional and unsupervised approaches to analyze high-dimensional mass cytometry datasets are discussed. A mass cytometry assay was implemented in a cross-sectional study of 19 women with a history of term or preterm birth to determine whether immune traits in peripheral blood differentiate the two groups in the absence of pregnancy. Twenty-seven phenotypic and 11 intracellular markers were simultaneously analyzed in whole blood samples stimulated with lipopolysaccharide (LPS at 0, 0.1, 1, 10, and 100 ng mL(-1) ) to examine dose-dependent signaling responses within the toll-like receptor 4 (TLR4) pathway. Complementary analyses, grounded in traditional or unsupervised gating strategies of immune cell subsets, indicated that the prpS6 and pMAPKAPK2 responses in classical monocytes are accentuated in women with a history of preterm birth (FDR<1%). The results suggest that women predisposed to preterm birth may be prone to mount an exacerbated TLR4 response during the course of pregnancy. This important hypothesis-generating finding points to the power of single-cell mass cytometry to detect biologically important differences in a relatively small patient cohort. © 2015 International Society for Advancement of Cytometry.

  View details for DOI 10.1002/cyto.a.22720

  View details for PubMedID 26190063

• Association of HMOX1 gene promoter polymorphisms with hyperbilirubinemia in the early neonatal period PEDIATRICS INTERNATIONAL Katayama, Y., Yokota, T., Zhao, H., Wong, R. J., Stevenson, D. K., Taniguchi-Ikeda, M., Nakamura, H., Iijima, K., Morioka, I. 2015; 57 (4): 645-649

  Abstract

  Heme oxygenase (HO) is the rate-limiting enzyme in the heme degradation pathway that produces bilirubin. The promoter region of human heme oxygenase-1 (HMOX1) contains a polymorphic (GT)n repeat that can regulate gene expression. Here, we investigated the association of (GT)n repeat length in the HMOX1 promoter region with neonatal hyperbilirubinemia in a population of Japanese term neonates.Using polymerase chain reaction and fragment analysis, we determined the number of (GT)n repeats in 149 Japanese neonates. To omit the effects of the G71R mutation in uridine diphosphoglucuronosyltransferase on hyperbilirubinemia, we excluded 41 neonates with the G71R mutation. As a result, 25 neonates with hyperbilirubinemia and 83 non-hyperbilirubinemic controls were included in this prospective case-control study. Allele and genotype frequencies of (GT)n repeats in the HMOX1 gene were compared between hyperbilirubinemic and non-hyperbilirubinemic control neonates.The prevalence of short alleles (< 22 (GT)n repeats) was significantly higher in hyperbilirubinemic than in control neonates (18% vs 7%, P = 0.015). Hyperbilirubinemia was more frequent in homozygous or heterozygous short allele carriers than control neonates (28% vs 11%, respectively, P = 0.03). Possession of short alleles was significantly associated with the development of neonatal hyperbilirubinemia (OR, 3.1; 95%CI: 1.03-9.53).Infants carrying short alleles (< 22 (GT)n repeats) in the HMOX1 gene promoter region appear to be at a higher risk for developing neonatal hyperbilirubinemia.

  View details for DOI 10.1111/ped.12591

  View details for Web of Science ID 000360501500024

• Escherichia coli heme oxygenase modulates host innate immune responses MICROBIOLOGY AND IMMUNOLOGY Maharshak, N., Ryu, H. S., Fan, T., Onyiah, J. C., Schulz, S., Otterbein, S. L., Wong, R., Hansen, J. J., Otterbein, L. E., Carroll, I. M., Plevy, S. E. 2015; 59 (8): 452-465

  Abstract

  Induction of mammalian heme oxygenase (HO)-1 and exposure of animals to carbon monoxide (CO) ameliorates experimental colitis. When enteric bacteria, including Escherichia coli, are exposed to low iron conditions, they express an HO-like enzyme, chuS, and metabolize heme into iron, biliverdin and CO. Given the abundance of enteric bacteria residing in the intestinal lumen, our postulate was that commensal intestinal bacteria may be a significant source of CO and those that express chuS and other Ho-like molecules suppress inflammatory immune responses through release of CO. According to real-time PCR, exposure of mice to CO results in changes in enteric bacterial composition and increases E. coli 16S and chuS DNA. Moreover, the severity of experimental colitis correlates positively with E. coli chuS expression in IL-10 deficient mice. To explore functional roles, E. coli were genetically modified to overexpress chuS or the chuS gene was deleted. Co-culture of chuS-overexpressing E. coli with bone marrow-derived macrophages resulted in less IL-12p40 and greater IL-10 secretion than in wild-type or chuS-deficient E. coli. Mice infected with chuS-overexpressing E. coli have more hepatic CO and less serum IL-12 p40 than mice infected with chuS-deficient E. coli. Thus, CO alters the composition of the commensal intestinal microbiota and expands populations of E. coli that harbor the chuS gene. These bacteria are capable of attenuating innate immune responses through expression of chuS. Bacterial HO-like molecules and bacteria-derived CO may represent novel targets for therapeutic intervention in inflammatory conditions.

  View details for DOI 10.1111/1348-0421.12282

  View details for Web of Science ID 000359787000003

  View details for PubMedID 26146866

  View details for PubMedCentralID PMC4582649

• Evaluation of a new end-tidal carbon monoxide monitor from the bench tothe bedside ACTA PAEDIATRICA Cuadrado, M. E., Bhutani, V. K., Aby, J. L., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2015; 104 (6): E279-E282

  View details for DOI 10.1111/apa.12938

  View details for Web of Science ID 000354528100008

• Protective effect of heme oxygenase induction in ethinylestradiol-induced cholestasis JOURNAL OF CELLULAR AND MOLECULAR MEDICINE Muchova, L., Vanova, K., Suk, J., Micuda, S., Dolezelova, E., Fuksa, L., Cerny, D., Farghali, H., Zelenkova, M., Lenicek, M., Wong, R. J., Vreman, H. J., Vitek, L. 2015; 19 (5): 924-933

  Abstract

  Estrogen-induced cholestasis is characterized by impaired hepatic uptake and biliary bile acids secretion because of changes in hepatocyte transporter expression. The induction of heme oxygenase-1 (HMOX1), the inducible isozyme in heme catabolism, is mediated via the Bach1/Nrf2 pathway, and protects livers from toxic, oxidative and inflammatory insults. However, its role in cholestasis remains unknown. Here, we investigated the effects of HMOX1 induction by heme on ethinylestradiol-induced cholestasis and possible underlying mechanisms. Wistar rats were given ethinylestradiol (5 mg/kg s.c.) for 5 days. HMOX1 was induced by heme (15 μmol/kg i.p.) 24 hrs prior to ethinylestradiol. Serum cholestatic markers, hepatocyte and renal membrane transporter expression, and biliary and urinary bile acids excretion were quantified. Ethinylestradiol significantly increased cholestatic markers (P ≤ 0.01), decreased biliary bile acid excretion (39%, P = 0.01), down-regulated hepatocyte transporters (Ntcp/Oatp1b2/Oatp1a4/Mrp2, P ≤ 0.05), and up-regulated Mrp3 (348%, P ≤ 0.05). Heme pre-treatment normalized cholestatic markers, increased biliary bile acid excretion (167%, P ≤ 0.05) and up-regulated hepatocyte transporter expression. Moreover, heme induced Mrp3 expression in control (319%, P ≤ 0.05) and ethinylestradiol-treated rats (512%, P ≤ 0.05). In primary rat hepatocytes, Nrf2 silencing completely abolished heme-induced Mrp3 expression. Additionally, heme significantly increased urinary bile acid clearance via up-regulation (Mrp2/Mrp4) or down-regulation (Mrp3) of renal transporters (P ≤ 0.05). We conclude that HMOX1 induction by heme increases hepatocyte transporter expression, subsequently stimulating bile flow in cholestasis. Also, heme stimulates hepatic Mrp3 expression via a Nrf2-dependent mechanism. Bile acids transported by Mrp3 to the plasma are highly cleared into the urine, resulting in normal plasma bile acid levels. Thus, HMOX1 induction may be a potential therapeutic strategy for the treatment of ethinylestradiol-induced cholestasis.

  View details for DOI 10.1111/jcmm.12401

  View details for Web of Science ID 000353991100003

  View details for PubMedID 25683492

• Unique Roles of Infiltrating Myeloid Cells in the Murine Uterus during Early to Midpregnancy. Journal of immunology Zhao, H., Kalish, F., Schulz, S., Yang, Y., Wong, R. J., Stevenson, D. K. 2015; 194 (8): 3713-3722

  Abstract

  Leukocyte infiltration into the uterus is a characteristic feature in early to midpregnancy, but the composition and function of these leukocytes are not well understood. Using a pregnant murine model, we showed that myeloid cells and uterine NK (uNK) cells were the predominant populations in uteri during early to midgestation, whereas T and B cells were constrained. Uterine myeloid populations included cells that infiltrated from the circulation (myeloid-derived suppressor cells [MDSCs], monocyte-derived macrophages [Mφs], and dendritic cells [DCs]) or proliferated from resident precursors (resident Mφs [Re-Mφs] and DCs). CD11b(hi)Ly6-G(hi) cells, representing neutrophils in both blood and uterine MDSCs, significantly increased from embryonic days 8.5 to 9.5. To understand their putative functions, we used anti-Gr-1 Ab to deplete circulating neutrophils and uterine MDSCs. In the absence of MDSC suppression, uterine DCs, T cells, and regulatory T cells expanded. Conversely, uterine MDSCs responded to LPS-induced inflammation and transformed into CD14(+)-activated neutrophils, resulting in an upregulation of tolerogenic DCs. A high dose of LPS (2.5 μg/mouse) significantly increased the influx of neutrophils and production of proinflammatory cytokines, such as IL-1β and TNF-α, resulting in the reduction of Re-Mφs and uNK cells, and led to placental hemorrhages and fetal deaths. In summary, uterine MDSCs are important in early to midpregnancy by responding to the maternal immunologic milieu and protecting uNK cells and Re-Mφs via MDSC's suppressive and anti-inflammatory functions. Upsetting this delicate immune balance by factors leading to either insufficient MDSCs or excessive neutrophil infiltration in the fetomaternal interface may contribute to pregnancy failure.

  View details for DOI 10.4049/jimmunol.1401930

  View details for PubMedID 25780045

• Heme oxygenase-1 in pregnancy and cancer: similarities in cellular invasion, cytoprotection, angiogenesis, and immunomodulation FRONTIERS IN PHARMACOLOGY Zhao, H., Ozen, M., Wong, R. J., Stevenson, D. K. 2015; 5

  View details for DOI 10.3389/fphar.2014.00295

  View details for Web of Science ID 000348209600001

• Heme oxygenase and the immune system in normal and pathological pregnancies. Frontiers in pharmacology Ozen, M., Zhao, H., Lewis, D. B., Wong, R. J., Stevenson, D. K. 2015; 6: 84-?

  View details for DOI 10.3389/fphar.2015.00084

  View details for PubMedID 25964759

• Direct antiglobulin titer strength and hyperbilirubinemia. Pediatrics Kaplan, M., Hammerman, C., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2014; 134 (5): e1340-4

  Abstract

  We recently demonstrated that direct antiglobulin titer (DAT) positive, blood group A or B newborns born to group O mothers had a high incidence of hyperbilirubinemia, attributable to increased hemolysis. We reanalyzed our data asking whether increasing DAT strength plays a modulating role in the pathophysiology of the hemolysis and hyperbilirubinemia.Data from previously published DAT-positive, ABO-heterospecific neonates were analyzed for hyperbilirubinemia and hemolysis according to strength of DAT. DAT was measured by using a gel agglutination technique and reported as values ranging from DAT ± to DAT ++++. Hemolysis was evaluated by blood carboxyhemoglobin corrected for inspired, ambient CO (COHbc), and expressed as percent total hemoglobin (tHb). Hyperbilirubinemia was defined as any plasma total bilirubin value >95th percentile on the hour-specific nomogram.Hyperbilirubinemia was more prevalent in those with DAT ++ readings (16 of 20, 80%) than those both DAT ± (37 of 87 [42.5%], relative risk: 1.88, 95% confidence interval: 1.35-2.61) and DAT + (32 of 56 [57.1%], relative risk: 1.40, 95% confidence interval: 1.02-1.92). COHbc values were higher for those with DAT ++ (1.45 ± 0.49% tHb [mean ± SD]) than those DAT ± (1.20 ± 0.37% tHb, P = .01) or DAT + (1.22 ± 0.37% tHb, P = .02).DAT ++ readings were associated with a higher incidence of hyperbilirubinemia and higher COHbc values than DAT ± or DAT + counterparts. Increasing DAT strength may be a modulator of hemolysis and hyperbilirubinemia in ABO-heterospecific neonates. DAT strength, and not merely DAT presence or absence, should be taken into consideration in the management of ABO-heterospecific newborns.

  View details for DOI 10.1542/peds.2014-1290

  View details for PubMedID 25332496

• Protective effects of inhaled carbon monoxide in endotoxin-induced cholestasis is dependent on its kinetics. Biochimie Vanova, K., Suk, J., Petr, T., Cerny, D., Slanar, O., Vreman, H. J., Wong, R. J., Zima, T., Vitek, L., Muchova, L. 2014; 97: 173-180

  Abstract

  Carbon monoxide (CO), a product of heme oxygenase (HMOX), has many beneficial biological functions and is a promising therapeutic agent for many pathological conditions. However, the kinetics of inhaled CO and its protective role in endotoxin-induced cholestasis is not fully known. Thus, our objective was to characterize the kinetics of inhaled CO and then investigate its use in early phase experimental endotoxin-induced cholestasis. Female Wistar rats were randomly divided into 4 groups: CON (control), LPS (lipopolysaccharide, 6 mg/kg), CO (250 ppm COx1h), and CO + LPS. Rats were sacrificed at 0-12 h after LPS administration. Tissues and blood were collected for liver injury markers and tissue CO distribution measurements. Livers were harvested for measurements of Hmox activity, Hmox1 mRNA expression, cytokines (IL10, IL6, TNF), and bile lipid and pigment transporters. Half-lives of CO in spleen, blood, heart, brain, kidney, liver, and lungs were 2.4 ± 1.5, 2.3 ± 0.8, 1.8 ± 1.6, 1.5 ± 1.2, 1.1 ± 1.1, 0.6 ± 0.3, 0.6 ± 0.2 h, respectively. CO treatment increased liver IL10 mRNA and decreased TNF expression 1 h after LPS treatment and prevented the down-regulation of bile acid and bilirubin hepatic transporters (Slc10a1, Abcb11, and Abcc2, p < 0.05), an effect closely related to the kinetics. The protective effect of CO against cholestatic liver injury persisted even 12 h after CO exposure, as shown by attenuation of serum cholestatic markers in CO-treated animals. CO exposure substantially attenuated endotoxin-induced cholestatic liver injury and was directly related to the kinetics of inhaled CO. This data underscores the importance of the kinetics of inhaled CO for the proper design of experimental and clinical studies of using CO as a treatment strategy.

  View details for DOI 10.1016/j.biochi.2013.10.009

  View details for PubMedID 24148277

• Risks and benefits of comparative effectiveness research in preterm infants: SUPPORT. Journal of comparative effectiveness research Stevenson, D. K., Wong, R. J., Tyson, J. E. 2014; 3 (1): 17-21

  View details for DOI 10.2217/cer.13.85

  View details for PubMedID 24345253

• Treatment of neonatal jaundice with filtered sunlight in Nigerian neonates: study protocol of a non-inferiority, randomized controlled trial TRIALS Slusher, T. M., Olusanya, B. O., Vreman, H. J., Wong, R. J., Brearley, A. M., Vaucher, Y. E., Stevenson, D. K. 2013; 14

  Abstract

  Severe neonatal jaundice and its progression to kernicterus is a leading cause of death and disability among newborns in poorly-resourced countries, particularly in sub-Saharan Africa. The standard treatment for jaundice using conventional phototherapy (CPT) with electric artificial blue light sources is often hampered by the lack of (functional) CPT devices due either to financial constraints or erratic electrical power. In an attempt to make phototherapy (PT) more readily available for the treatment of pathologic jaundice in underserved tropical regions, we set out to test the hypothesis that filtered sunlight phototherapy (FS-PT), in which potentially harmful ultraviolet and infrared rays are appropriately screened, will be as efficacious as CPT.This prospective, non-blinded randomized controlled non-inferiority trial seeks to enroll infants with elevated total serum/plasma bilirubin (TSB, defined as 3 mg/dl below the level recommended by the American Academy of Pediatrics for high-risk infants requiring PT) who will be randomly and equally assigned to receive FS-PT or CPT for a total of 616 days at an inner-city maternity hospital in Lagos, Nigeria. Two FS-PT canopies with pre-tested films will be used. One canopy with a film that transmits roughly 33% blue light (wavelength range: 400 to 520 nm) will be used during sunny periods of a day. Another canopy with a film that transmits about 79% blue light will be used during overcast periods of the day. The infants will be moved from one canopy to the other as needed during the day with the goal of keeping the blue light irradiance level above 8 μW/cm²/nm.Primary outcome: FS-PT will be as efficacious as CPT in reducing the rate of rise in bilirubin levels. Secondary outcome: The number of infants requiring exchange transfusion under FS-PT will not be more than those under CPT.This novel study offers the prospect of an effective treatment for infants at risk of severe neonatal jaundice and avoidable exchange transfusion in poorly-resourced settings without access to (reliable) CPT in the tropics.ClinicalTrials.gov Identifier: NCT01434810.

  View details for DOI 10.1186/1745-6215-14-446

  View details for Web of Science ID 000329516800001

  View details for PubMedID 24373547

  View details for PubMedCentralID PMC3879162

• Evaluation of window-tinting films for sunlight phototherapy. Journal of tropical pediatrics Vreman, H. J., Slusher, T. M., Wong, R. J., Schulz, S., Olusanya, B. O., Stevenson, D. K. 2013; 59 (6): 496-501

  Abstract

  We evaluated nine semi-transparent plastic window-tinting films for their ability to block ultraviolet A (UVA) and infrared (IR) radiation and transmit therapeutic blue light (400-520 nm) for treating jaundiced newborns. For indoor testing, three light sources (TL/52 special blue fluorescent, Black Light UVA and IR heat lamps) were positioned above each film and measured successively using a thermocouple thermometer, UVA radiometer and blue light irradiance meter, placed below each film. For outdoor testing, the same setup was used with the sun at zenith and a cloudless sky. Compared with unfiltered radiation, blue light transmission through films ranged from 24 to 83%, UVA transmission was 0.1-7.1% and reductions in IR heat were 6-12°C and 5-10°C for heat lamp and sun, respectively. The data suggest that most of the relatively low-cost window-tinting films tested can effectively reduce sunlight UV and IR and offer a range of significant attenuations of therapeutic blue light.

  View details for DOI 10.1093/tropej/fmt062

  View details for PubMedID 23880667

• HEME OXYGENASE ACTIVITY AND HEME BINDING ARE CRITICAL IN EARLY LIFE Western Regional Meeting of the American-Federation-for-Medical-Research Ang, J., Schulz, S., Wong, R. J., Sevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2013: 190–90

  View details for Web of Science ID 000312657900287

• HEME OXYGENASE ACTIVITY AND HEME BINDING ARE CRITICAL IN EARLY LIFE Western Regional Meeting of the American-Federation-for-Medical-Research Ang, J., Schulz, S., Wong, R. J., Sevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2013: 153–53

  View details for Web of Science ID 000312657900157

• Intracellular accumulation of bilirubin as a defense mechanism against increased oxidative stress BIOCHIMIE Zelenka, J., Muchova, L., Zelenkova, M., Vanova, K., Vreman, H. J., Wong, R. J., Vitek, L. 2012; 94 (8): 1821-1827

  Abstract

  Antioxidant, anti-inflammatory and anti-atherogenic effects have been associated with elevations of unconjugated bilirubin (UCB) in serum and with the induction of heme oxygenase-1 (HO-1), the rate-limiting enzyme in UCB synthesis. The aim of this study was to investigate the intracellular metabolism and antioxidant properties of UCB in human hepatoblastoma HepG2 cells and tissues of Wistar rats exposed to oxidative stressors and lipopolysaccharide (LPS), respectively. Intracellular UCB concentrations in HepG2 cells correlated with its levels in culture media (p < 0.001) and diminished lipid peroxidation in a dose-dependent manner (p < 0.001). Moreover, induction of HO-1 with sodium arsenite led to 2.4-fold (p = 0.01) accumulation of intracellular UCB over basal level while sodium azide-derived oxidative stress resulted in a 60% drop (p < 0.001). This decrease was ameliorated by UCB elevation in media or by simultaneous induction of HO-1. In addition, hyperbilirubinemia and liver HO-1 induction in LPS-treated rats resulted in a 2-fold accumulation of tissue UCB (p = 0.01) associated with enhanced protection against lipid peroxidation (p = 0.02). In conclusion, hyperbilirubinemia and HO-1 induction associated with inflammation and oxidative stress increase intracellular concentrations of UCB, thus enhancing the protection of cellular lipids against peroxidation. Therefore, the previously reported protective effects of hyperbilirubinemia and HO-1 induction are at least in part due to intracellular accumulation of UCB.

  View details for DOI 10.1016/j.biochi.2012.04.026

  View details for Web of Science ID 000306209000022

  View details for PubMedID 22580386

• Neonatal phototherapy: choice of device and outcome ACTA PAEDIATRICA Bhutani, V. K., Wong, R. J. 2012; 101 (5): 441-443

  View details for DOI 10.1111/j.1651-2227.2012.02605.x

  View details for Web of Science ID 000302349200016

  View details for PubMedID 22251098

• IN VITRO BILIRUBIN PHOTODESTRUCTION ACTION SPECTRUM REVISITED Western Regional Meeting of the American-Federation-for-Medical-Research Schulz, S., Vreman, H. J., Wong, R. J., Cline, B., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2012: 210–10

  View details for Web of Science ID 000298634401302

• HEME OXYGENASE-1 DEFICIENCY PROMOTES NECROTIZING ENTEROCOLITIS DEVELOPMENT IN A MURINE MOUSE MODEL Western Regional Meeting of the American-Federation-for-Medical-Research Schulz, S., Jang, K., Kalish, F. S., Zhao, H., Vreman, H. J., Sylvester, K. S., Wong, R. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2012: 158–58

  View details for Web of Science ID 000298634401114

• Is phototherapy exposure associated with better or worse outcomes in 501-to 1000-g-birth-weight infants? ACTA PAEDIATRICA Hintz, S. R., Stevenson, D. K., Yao, Q., Wong, R. J., Das, A., Van Meurs, K. P., Morris, B. H., Tyson, J. E., Oh, W., Poole, W. K., Phelps, D. L., McDavid, G. E., Grisby, C., Higgins, R. D. 2011; 100 (7): 960-965

  Abstract

   To compare risk-adjusted outcomes at 18- to 22-month-corrected age for extremely low birth weight (ELBW) infants who never received phototherapy (NoPTx) to those who received any phototherapy (PTx) in the NICHD Neonatal Research Network randomized trial of Aggressive vs. Conservative Phototherapy.Outcomes at 18 to 22-month-corrected age included death, neurodevelopmental impairment (NDI) and Bayley Scales Mental Developmental Index (MDI). Regression models evaluated the independent association of PTx with adverse outcomes controlling for centre and other potentially confounding variables.Of 1972 infants, 216 were NoPTx and 1756 were PTx. For the entire 501- to 1000-g-BW cohort, PTx was not independently associated with death or NDI (OR 0.85, 95% CI: 0.60-1.20), death or adverse neurodevelopmental endpoints. However, among infants 501-750 g BW, the rate of significant developmental impairment with MDI < 50 was significantly higher for NoPTx (29%) than PTx (12%) (p = 0.004).Phototherapy did not appear to be independently associated with death or NDI for the overall ELBW group. Whether PTx increases mortality could not be excluded because of bias from deaths before reaching conservative treatment threshold. The higher rate of MDI < 50 in the 501- to 750-g-BW NoPTx group is concerning and consistent with NRN Trial results.

  View details for DOI 10.1111/j.1651-2227.2011.02175.x

  View details for PubMedID 21272067

• Bilirubin Production and the Risk of Bilirubin Neurotoxicity SEMINARS IN PERINATOLOGY Stevenson, D. K., Vreman, H. J., Wong, R. J. 2011; 35 (3): 121-126

  Abstract

  Neonatal jaundice usually occurs in the transitional period after birth, presenting as an elevation of circulating bilirubin. Bilirubin neurotoxicity can occur if the levels of bilirubin become excessive (hyperbilirubinemia). This pathologic phenotype of newborn jaundice can develop because of excessive bilirubin production or impaired conjugation, with the risk for developing bilirubin-induced neurologic dysfunction, depending on the degree of the resultant bilirubin load. The plasma bilirubin level thus can be used to assess an infant's risk for developing bilirubin neurotoxicity relative to an infant's age in hours. Because all infants have an impaired conjugation ability, infants at greatest risk are those who have increased bilirubin production rates, because of hemolysis, for example. Therefore, developing potential preventive strategies as well as noninvasive technologies to treat and to identify infants with increased bilirubin production rates, respectively, are tantamount to reducing the incidence of bilirubin-induced neurologic dysfunction.

  View details for DOI 10.1053/j.semperi.2011.02.005

  View details for PubMedID 21641484

• Heme Oxygenase-1 Deletion Affects Stress Erythropoiesis PLOS ONE Cao, Y., Kusy, S., Luong, R., Wong, R. J., Stevenson, D. K., Contag, C. H. 2011; 6 (5)

  Abstract

  Homeostatic erythropoiesis leads to the formation of mature red blood cells under non-stress conditions, and the production of new erythrocytes occurs as the need arises. In response to environmental stimuli, such as bone marrow transplantation, myelosuppression, or anemia, erythroid progenitors proliferate rapidly in a process referred to as stress erythropoiesis. We have previously demonstrated that heme oxygenase-1 (HO-1) deficiency leads to disrupted stress hematopoiesis. Here, we describe the specific effects of HO-1 deficiency on stress erythropoiesis.We used a transplant model to induce stress conditions. In irradiated recipients that received hmox(+/-) or hmox(+/+) bone marrow cells, we evaluated (i) the erythrocyte parameters in the peripheral blood; (ii) the staining intensity of CD71-, Ter119-, and CD49d-specific surface markers during erythroblast differentiation; (iii) the patterns of histological iron staining; and (iv) the number of Mac-1(+)-cells expressing TNF-α. In the spleens of mice that received hmox(+/-) cells, we show (i) decreases in the proerythroblast, basophilic, and polychromatophilic erythroblast populations; (ii) increases in the insoluble iron levels and decreases in the soluble iron levels; (iii) increased numbers of Mac-1(+)-cells expressing TNF-α; and (iv) decreased levels of CD49d expression in the basophilic and polychromatophilic erythroblast populations.As reflected by effects on secreted and cell surface proteins, HO-1 deletion likely affects stress erythropoiesis through the retention of erythroblasts in the erythroblastic islands of the spleen. Thus, HO-1 may serve as a therapeutic target for controlling erythropoiesis, and the dysregulation of HO-1 may be a predisposing condition for hematologic diseases.

  View details for DOI 10.1371/journal.pone.0020634

  View details for PubMedID 21655188

• Bile acids decrease intracellular bilirubin levels in the cholestatic liver: implications for bile acid-mediated oxidative stress JOURNAL OF CELLULAR AND MOLECULAR MEDICINE Muchova, L., Vanova, K., Zelenka, J., Lenicek, M., Petr, T., Vejrazka, M., Sticova, E., Vreman, H. J., Wong, R. J., Vitek, L. 2011; 15 (5): 1156-1165

  Abstract

  High plasma concentrations of bile acids (BA) and bilirubin are hallmarks of cholestasis. BA are implicated in the pathogenesis of cholestatic liver damage through mechanisms involving oxidative stress, whereas bilirubin is a strong antioxidant. We evaluated the roles of bilirubin and BA on mediating oxidative stress in rats following bile duct ligation (BDL). Adult female Wistar and Gunn rats intraperitoneally anaesthetized with ketamine and xylazine underwent BDL or sham operation. Cholestatic markers, antioxidant capacity, lipid peroxidation and heme oxygenase (HO) activity were determined in plasma and/or liver tissue 5 days after surgery. HepG2-rNtcp cells were used for in vitro experiments. Plasma bilirubin levels in control and BDL animals positively correlated with plasma antioxidant capacity. Peroxyl radical scavenging capacity was significantly higher in the plasma of BDL Wistar rats (210 ± 12%, P < 0.0001) compared to controls, but not in the liver tissues. Furthermore after BDL, lipid peroxidation in the livers increased (179 ± 37%, P < 0.01), whereas liver HO activity significantly decreased to 61% of control levels (P < 0.001). Addition of taurocholic acid (TCA, ≥ 50 μmol/l) to liver homogenates increased lipid peroxidation (P < 0.01) in Wistar, but not in Gunn rats or after the addition of bilirubin. In HepG2-rNtcp cells, TCA decreased both HO activity and intracellular bilirubin levels. We conclude that even though plasma bilirubin is a marker of cholestasis and hepatocyte dysfunction, it is also an endogenous antioxidant, which may counteract the pro-oxidative effects of BA in circulation. However, in an animal model of obstructive cholestasis, we found that BA compromise intracellular bilirubin levels making hepatocytes more susceptible to oxidative damage.

  View details for DOI 10.1111/j.1582-4934.2010.01098.x

  View details for Web of Science ID 000291045800015

  View details for PubMedID 20518850

• Spontaneous hepatic repopulation in transgenic mice expressing mutant human alpha 1-antitrypsin by wild-type donor hepatocytes JOURNAL OF CLINICAL INVESTIGATION Ding, J., Yannam, G. R., Roy-Chowdhury, N., Hidvegi, T., Basma, H., Rennard, S. I., Wong, R. J., Avsar, Y., Guha, C., Perlmutter, D. H., Fox, I. J., Roy-Chowdhury, J. 2011; 121 (5): 1930-1934

  Abstract

  α1-Antitrypsin deficiency is an inherited condition that causes liver disease and emphysema. The normal function of this protein, which is synthesized by the liver, is to inhibit neutrophil elastase, a protease that degrades connective tissue of the lung. In the classical form of the disease, inefficient secretion of a mutant α1-antitrypsin protein (AAT-Z) results in its accumulation within hepatocytes and reduced protease inhibitor activity, resulting in liver injury and pulmonary emphysema. Because mutant protein accumulation increases hepatocyte cell stress, we investigated whether transplanted hepatocytes expressing wild-type AAT might have a competitive advantage relative to AAT-Z-expressing hepatocytes, using transgenic mice expressing human AAT-Z. Wild-type donor hepatocytes replaced 20%-98% of mutant host hepatocytes, and repopulation was accelerated by injection of an adenovector expressing hepatocyte growth factor. Spontaneous hepatic repopulation with engrafted hepatocytes occurred in the AAT-Z-expressing mice even in the absence of severe liver injury. Donor cells replaced both globule-containing and globule-devoid cells, indicating that both types of host hepatocytes display impaired proliferation relative to wild-type hepatocytes. These results suggest that wild-type hepatocyte transplantation may be therapeutic for AAT-Z liver disease and may provide an alternative to protein replacement for treating emphysema in AAT-ZZ individuals.

  View details for DOI 10.1172/JCI45260

  View details for Web of Science ID 000290246800027

  View details for PubMedID 21505264

  View details for PubMedCentralID PMC3083768

• Quantitating carbon monoxide production from heme by vascular plant preparations in vitro PLANT PHYSIOLOGY AND BIOCHEMISTRY Vreman, H. J., Wong, R. J., Stevenson, D. K. 2011; 49 (1): 61-68

  Abstract

  Heme in animals is mainly degraded enzymatically, producing a predictable amount of carbon monoxide (CO). Under some conditions, alternative sources of CO production are important, such as lipid peroxidation and photo-oxidation. Less is known about CO production in plants as a reflection of enzymatic activity or coupled oxidation, but a sensitive assay for CO production in plants would be a valuable tool to explore the various sources in plants as the conditions of the reactions and mechanisms are defined. Using gas chromatography, we determined the requirements for heme-supported in vitro CO generation by exogenous reactants (NADPH, tissue supernatant, oxygen), optimum reaction conditions (time, temperature, pH, light), and effects of various cofactors and substrates using supernatants from Spinacia oleracea (spinach) leaf and Solanum tuberosa (potato) tuber homogenates. We then determined the CO production rate distribution between organ (root, stem, leaf, flower, fruit) supernatants in a number of commercially available plant species. CO production ranged from 4-65 nmol CO/h/g fresh weight and occurred in all vascular plant tissues examined, with the highest rates in chloroplast-containing tissues. In spinach leaves, CO production was concentrated (>2-fold) in the particulate fraction, whereas in potato tubers, the particulate fraction accounted for <50% of the rates in homogenates. We conclude that gas chromatography is uniquely suited for the determination of CO production in pigmented, heterogeneous plant tissue preparations.

  View details for DOI 10.1016/j.plaphy.2010.09.021

  View details for PubMedID 21055958

• THE INHIBITORY POTENCY OF LOW DOSES OF ZINC DEUTEROPORPHYRIN BIS GLYCOL ON HEME OXYGENASE ACTIVITY IN 3-DAY-OLD MICE Western Regional Meeting of the American-Federation-for-Medical-Research Katayama, Y., Shaw, N., Yaffe, Z. A., He, C. X., Kalish, F. S., Schulz-Geske, S., Zhao, H., Wong, R. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2011: 170–70

  View details for Web of Science ID 000285542500310

• ROLE OF IIEME OXYGENASE IN A MURINE MODEL OF EARLY NECROTIZING ENTEROCOLITIS Western Regional Meeting of the American-Federation-for-Medical-Research Schulz-Geske, S., Kalish, F. S., Jang, K. Y., Zhao, H., Huey, M., Vreman, H. J., Sylvester, K. S., Wong, R. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2011: 126–27

  View details for Web of Science ID 000285542500153

• HEME OXYGENASE-1 EXPRESSION IS DIFFERENTIALLY REGULATED IN VITRO BY METALLOPORPHYRINS Western Regional Meeting of the American-Federation-for-Medical-Research Schulz-Geske, S., Zhao, H., Kalish, F. S., McCarthy, E., Vreman, H. J., Wong, R. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2011: 196–96

  View details for Web of Science ID 000285542500399

• EFFECT OF FLUORESCENT LIGHT EXPOSURE ON METALLOPORHYRIN-TREATED NEWBORN MICE Western Regional Meeting of the American-Federation-for-Medical-Research Schulz-Geske, S., Kalish, F. S., Zhao, H., Katayama, Y., Champion, K. A., Vreman, H. J., Wong, R. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2011: 126–26

  View details for Web of Science ID 000285542500152

• Hemolysis and Hyperbilirubinemia in Antiglobulin Positive, Direct ABO Blood Group Heterospecific Neonates JOURNAL OF PEDIATRICS Kaplan, M., Hammerman, C., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2010; 157 (5): 772-777

  Abstract

  We quantified hemolysis and determined the incidence of hyperbilirubinemia in neonates who were direct antiglobulin titer (DAT)-positive, ABO heterospecific, and compared variables among O-A and O-B subgroups.Plasma total bilirubin (PTB) was determined before the neonates were discharged from the hospital and more frequently when clinically warranted, in neonates who were DAT positive with blood group A or B and with mothers who had blood group O. Heme catabolism (and therefore bilirubin production) was indexed by blood carboxyhemoglobin corrected for inspired carbon monoxide (COHbc). Hyperbilirubinemia was defined as any PTB concentration >95th percentile on the hour-of-life-specific bilirubin nomogram.Of 164 neonates, 111 were O-A and 53 O-B. Overall, hyperbilirubinemia developed 85 neonates (51.8%), and it tended to be more prevalent in the O-B neonates than O-A neonates (62.3% versus 46.8%; P = .053). Hyperbilirubinemia developed in more O-B newborns than O-A newborns at <24 hours (93.9% versus 48.1%; P< .0001). COHbc values were globally higher than our previously published newborn values. Babies in whom hyperbilirubinemia developed had higher COHbc values than the already high values of babies who were non-hyperbilirubinemic, and O-B newborns tended to have higher values than their O-A counterparts.DAT-positive, ABO heterospecificity is associated with increased hemolysis and a high incidence of neonatal hyperbilirubinemia. O-B heterospecificity tends to confer even higher risk than O-A counterparts.

  View details for DOI 10.1016/j.jpeds.2010.05.024

  View details for Web of Science ID 000283045900020

  View details for PubMedID 20598320

  View details for PubMedCentralID PMC2951500

• Understanding Neonatal Jaundice: A Perspective on Causation PEDIATRICS AND NEONATOLOGY Cohen, R. S., Wong, R. J., Stevenson, D. K. 2010; 51 (3): 143-148

  Abstract

  Neonatal jaundice can be best understood as a balance between the production and elimination of bilirubin, with a multitude of factors and conditions affecting each of these processes. When an imbalance results because of an increase in circulating bilirubin (or the bilirubin load) to significantly high levels (severe hyperbilirubinemia), it may cause permanent neurologic sequelae (kernicterus). In most infants, an increase in bilirubin production (e.g., due to hemolysis) is the primary cause of severe hyperbilirubinemia, and thus reducing bilirubin production is a rational approach for its management. The situation can become critical in infants with an associated impaired bilirubin elimination mechanism as a result of a genetic deficiency and/or polymorphism. Combining information about bilirubin production and genetic information about bilirubin elimination with the tracking of bilirubin levels means that a relative assessment of jaundice risk might be feasible. Information on the level of bilirubin production and its rate of elimination may help to guide the clinical management of neonatal jaundice.

  View details for PubMedID 20675237

• Influence of clinical status on the association between plasma total and unbound bilirubin and death or adverse neurodevelopmental outcomes in extremely low birth weight infants ACTA PAEDIATRICA Oh, W., Stevenson, D. K., TYSON, J. E., Morris, B. H., Ahlfors, C. E., Bender, G. J., Wong, R. J., Perritt, R., Vohr, B. R., Van Meurs, K. P., Vreman, H. J., Das, A., Phelps, D. L., O'Shea, T. M., Higgins, R. D. 2010; 99 (5): 673-678

  Abstract

  To assess the influence of clinical status on the association between total plasma bilirubin and unbound bilirubin on death or adverse neurodevelopmental outcomes at 18-22 months corrected age in extremely low birth weight infants.Total plasma bilirubin and unbound bilirubin were measured in 1101 extremely low birth weight infants at 5 +/- 1 days of age. Clinical criteria were used to classify infants as clinically stable or unstable. Survivors were examined at 18-22 months corrected age by certified examiners. Outcome variables were death or neurodevelopmental impairment, death or cerebral palsy, death or hearing loss, and death prior to follow-up. For all outcomes, the interaction between bilirubin variables and clinical status was assessed in logistic regression analyses adjusted for multiple risk factors.Regardless of clinical status, an increasing level of unbound bilirubin was associated with higher rates of death or neurodevelopmental impairment, death or cerebral palsy, death or hearing loss and death before follow-up. Total plasma bilirubin values were directly associated with death or neurodevelopmental impairment, death or cerebral palsy, death or hearing loss, and death before follow-up in unstable infants, but not in stable infants. An inverse association between total plasma bilirubin and death or cerebral palsy was found in stable infants. CONCLUSIONs: In extremely low birth weight infants, clinical status at 5 days of age affects the association between total plasma bilirubin and death or adverse neurodevelopmental outcomes at 18-22 months of corrected age. An increasing level of UB is associated a higher risk of death or adverse neurodevelopmental outcomes regardless of clinical status. Increasing levels of total plasma bilirubin are directly associated with increasing risk of death or adverse neurodevelopmental outcomes in unstable, but not in stable infants.

  View details for DOI 10.1111/j.1651-2227.2010.01688.x

  View details for Web of Science ID 000276034800011

  View details for PubMedID 20105142

  View details for PubMedCentralID PMC2875328

• ELEVATED EXHALED CARBON MONOXIDE CONCENTRATION IN HEMOGLOBINOPATHIES AND ITS RELATION TO RED BLOOD CELL TRANSFUSION THERAPY PEDIATRIC HEMATOLOGY AND ONCOLOGY James, E. B., Vreman, H. J., Wong, R. J., Stevenson, D. K., Vichinsky, E., Schumacher, L., Hall, J. Y., Simon, J., Golden, D. W., Harmatz, P. 2010; 27 (2): 112-121

  Abstract

  In this study, the authors examined a possible role of measurements of end-tidal carbon monoxide (CO), corrected for inhaled CO (ETCOc), as a noninvasive screening tool for hemoglobinopathies and as an indicator for when transfusions would be required in patients receiving chronic transfusions. ETCOc measurements were obtained in subjects with sickle cell disease (n = 18), thalassemia (n = 21), and healthy controls (n = 62). ETCOc values less than 3 parts per million (ppm) yielded a positive predictive value of 93% and negative predictive value of 94% in identifying hemoglobinopathies. Subsequently, 7 subjects with thalassemia had laboratory parameters and ETCOc measured over 2 transfusion cycles. ETCOc values were 4.90 +/- 0.32 ppm (mean +/- SD), with 89% of values being above normal (>or=3 ppm). Pretransfusion ETCOc levels significantly correlated with pretransfusion reticulocyte count (r = .96, P <.001), but not with pretransfusion hemoglobin (r = .44, P = .16) or pretransfusion soluble transferrin receptors (sTfR, r = .52, P = .10). In conclusion, we found that patients with hemoglobinopathies have ETCOc values above the range for healthy controls and ETCOc measurements can be used as an adjunct to hemoglobin measurements to determine the proper timing of transfusions.

  View details for DOI 10.3109/08880010903536227

  View details for Web of Science ID 000275286100004

  View details for PubMedID 20201692

• MOLECULAR MECHANISM OF HEME OXYGENASE-1 INDUCTION BY STATINS Western Regional Meeting of the American-Federation-for-Medical-Research Schulz-Geske, S., Kalish, F. S., Zhao, H., Vreman, H. J., Wong, R. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2010: 208–

  View details for Web of Science ID 000273638400380

• AGE-DEPENDENT EXPRESSION OF HEME OXYGENASE-1 IN MICE FOLLOWING ORAL ADMINISTRATION OF ZING BIS GLYCOL PORPHYRIN Western Regional Meeting of the American-Federation-for-Medical-Research He, C. X., Zhao, H., Kalish, F., Wong, R. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2010: 162–63

  View details for Web of Science ID 000273638400223

• Molecular mechanism and functional consequences of lansoprazole-mediated heme oxygenase-1 induction WORLD JOURNAL OF GASTROENTEROLOGY Schulz-Geske, S., Erdmann, K., Wong, R. J., Stevenson, D. K., Schroeder, H., Grosser, N. 2009; 15 (35): 4392-4401

  Abstract

  To investigate the molecular mechanism and functional consequences of heme oxygenase-1 (HO-1) activation by lansoprazole in endothelial cells and macrophages.Expression of HO-1 mRNA was analyzed by Northern blotting. Western blotting was used to determine the HO-1 and ferritin protein levels. NADPH-dependent reactive oxygen species (ROS) formation was measured with lucigenin-enhanced chemiluminescence. HO-1 promoter activity in mouse fibroblasts, stably transfected with a 15-kb HO-1 gene that drives expression of the reporter gene luciferase, was assessed using in vivo bioluminescence imaging.Lansoprazole increased HO-1 mRNA levels in endothelial cells and HO-1 protein levels in macrophages. In addition, lansoprazole-induced ferritin protein levels in both cell systems. Moreover, induction of the antioxidant proteins HO-1 and ferritin by lansoprazole was followed by a decrease in NADPH-mediated ROS formation. The radical scavenging properties of lansoprazole were diminished in the presence of the HO inhibitor, chromium mesoporphyrin IX. Induction of HO-1 gene expression by lansoprazole was not related to oxidative stress or to the activation of the mitogen-activated protein kinase pathway. However, the phosphatidylinositol 3-kinase inhibitor LY294002 showed a concentration-dependent inhibition of HO-1 mRNA and promoter activity.Activation of HO-1 and ferritin may account for the gastric protection of lansoprazole and is dependent on a pathway blocked by LY294002.

  View details for DOI 10.3748/wjg.15.4392

  View details for Web of Science ID 000270080500007

  View details for PubMedID 19764090

  View details for PubMedCentralID PMC2747059

• Differentiation and Transplantation of Human Embryonic Stem Cell-Derived Hepatocytes GASTROENTEROLOGY Basma, H., Soto-Gutierrez, A., Yannam, G. R., Liu, L., Ito, R., Yamamoto, T., Ellis, E., Carson, S. D., Sato, S., Chen, Y., Muirhead, D., Navarro-Alvarez, N., Wong, R. J., Roy-Chowdhury, J., Platt, J. L., Mercer, D. F., Miller, J. D., Strom, S. C., Kobayashi, N., Fox, I. J. 2009; 136 (3): 990-999

  Abstract

  The ability to obtain unlimited numbers of human hepatocytes would improve the development of cell-based therapies for liver diseases, facilitate the study of liver biology, and improve the early stages of drug discovery. Embryonic stem cells are pluripotent, potentially can differentiate into any cell type, and therefore could be developed as a source of human hepatocytes.To generate human hepatocytes, human embryonic stem cells were differentiated by sequential culture in fibroblast growth factor 2 and human activin-A, hepatocyte growth factor, and dexamethasone. Functional hepatocytes were isolated by sorting for surface asialoglycoprotein-receptor expression. Characterization was performed by real-time polymerase chain reaction, immunohistochemistry, immunoblot, functional assays, and transplantation.Embryonic stem cell-derived hepatocytes expressed liver-specific genes, but not genes representing other lineages, secreted functional human liver-specific proteins similar to those of primary human hepatocytes, and showed human hepatocyte cytochrome P450 metabolic activity. Serum from rodents given injections of embryonic stem cell-derived hepatocytes contained significant amounts of human albumin and alpha1-antitrypsin. Colonies of cytokeratin-18 and human albumin-expressing cells were present in the livers of recipient animals.Human embryonic stem cells can be differentiated into cells with many characteristics of primary human hepatocytes. Hepatocyte-like cells can be enriched and recovered based on asialoglycoprotein-receptor expression and potentially could be used in drug discovery research and developed as therapeutics.

  View details for DOI 10.1053/j.gastro.2008.10.047

  View details for Web of Science ID 000263751400037

  View details for PubMedID 19026649

  View details for PubMedCentralID PMC2732349

• Photoisomers: Obfuscating Factors in Clinical Peroxidase Measurements of Unbound Bilirubin? PEDIATRICS McDonagh, A. F., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2009; 123 (1): 67-76

  Abstract

  The objectives of the study were to measure the effect of 4Z,15E-bilirubin on peroxidase free bilirubin measurements and to review the literature on this topic.4Z,15E-Bilirubin was generated in situ in serum or serum albumin solution through controlled irradiation of isomerically pure 4Z,15Z-bilirubin IXalpha, under conditions in which the total amount of bilirubin remained constant. Reactions were monitored by difference spectroscopy, to ensure that solutions were not irradiated beyond the initial photostationary state and that concentrations of other isomers were kept to a minimum. Prepared in this way, 10% to 25% of the total bilirubin in the final solutions was in the form of the 4Z,15E-isomer. Free bilirubin in the solutions was measured with a peroxidase method, before and after irradiation. The use of bovine serum albumin as a surrogate for human albumin in in vitro studies also was investigated.The findings of previous studies are not altogether consistent, with a common flaw in several being the failure to measure photoisomer concentrations. For bilirubin in serum albumin solution, conversion of approximately 25% of the 4Z,15Z-isomer to 4Z,15E-bilirubin led to a much smaller decrease (<20%) in the apparent free bilirubin concentration; for bilirubin in serum, conversion of approximately 15% of the 4Z,15Z-isomer to photoisomers resulted in a much larger increase ( approximately 40%). Irradiation of bilirubin in bovine serum albumin solution generated a very different array of photoisomers than that observed in human albumin solutions.The effect of photoisomers on the accuracy and specificity of free 4Z,15Z-bilirubin measurements remains uncertain. In a clinical setting, free bilirubin measurements need to be interpreted with caution when samples contain photoisomers. Irradiated bovine albumin solutions of isomerically impure bilirubin used in previous studies are poor models for investigating the effects of phototherapy in humans and the albumin binding of photoisomers.

  View details for DOI 10.1542/peds.2008-0492

  View details for Web of Science ID 000262046400010

  View details for PubMedID 19117862

• A NEW MOUSE MODEL OF NEONATAL BRAIN INJURY Western Regional Meeting of the American-Federation-for-Medical-Research Knauer, Y., Wong, R. J., Zhao, H., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2009: 129–30

  View details for Web of Science ID 000270092400124

• AGE-DEPENDENT EXPRESSION OF HEME OXYGENASE-1 IN MICE FOLLOWING ORAL ADMINISTRATION OF CHROMIUM MESOPORPHYRIN Western Regional Meeting of the American-Federation-for-Medical-Research He, C. X., Morisawa, T., Zhao, H., Wong, R. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2009: 177–77

  View details for Web of Science ID 000270092400293

• Failure to Predict Hemolysis and Hyperbilirubinemia by IgG Subclass in Blood Group A or B Infants Born to Group O Mothers PEDIATRICS Kaplan, M., Na'amad, M., Kenan, A., Rudensky, B., Hammerman, C., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2009; 123 (1): E132-E137

  Abstract

  Direct antibody titer-positive, blood group A or B neonates who are born to group O mothers may be at risk for hemolysis and hyperbilirubinemia. Immunoglobulin G1 and immunoglobulin G3 subclasses are associated with increased hemolysis relative to immunoglobulin G2 and immunoglobulin G4. We investigated whether identification of immunoglobulin G subclass 1 or 3 may be predictive of hemolysis and hyperbilirubinemia.Direct antibody titer-positive, blood group A and B neonates born to group O mothers were tested for the presence of immunoglobulin G subclasses 1 and 3 in umbilical cord blood by using a commercially available gel testing technology. By inference, neonates in whom neither immunoglobulin G1 nor immunoglobulin G3 were detected were designated immunoglobulin G2 and/or 4. Mandatory plasma total bilirubin was measured at discharge, and additional measurements performed as clinically indicated. Hyperbilirubinemia was defined as any plasma total bilirubin value >95th percentile for hour of life. Blood carboxyhemoglobin and total hemoglobin concentrations were also measured on the predischarge sample. Measured carboxyhemoglobin, expressed as percentage of total hemoglobin, was corrected for ambient carbon monoxide to derive "corrected carboxyhemoglobin," a sensitive index of heme catabolism. The corrected carboxyhemoglobin/total hemoglobin ratio was calculated to correct for any differences in total hemoglobin mass between groups.Eighty-two infants were studied, 18 of whom were designated as immunoglobulin G1, 0 as immunoglobulin G3, and 64 as immunoglobulin G2 and/or 4. The incidence of plasma total bilirubin >95th percentile was similar between the subgroupings. Corrected carboxyhemoglobin values and corrected carboxyhemoglobin/total hemoglobin ratio were also similar between the subgroupings.Immunoglobulin G1 was found in 22% of direct antibody titer-positive, group A and B neonates who were born to group O mothers, whereas immunoglobulin G3 was rare. Hemolysis and hyperbilirubinemia could not be predicted by this gel technique that enabled identification of these immunoglobulin G subclasses.

  View details for DOI 10.1542/peds.2008-2617

  View details for Web of Science ID 000262046400077

  View details for PubMedID 19114458

• Highly sensitive method for quantitative determination of bilirubin in biological fluids and tissues JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES Zelenka, J., Lenicek, M., Muchova, L., Jirsa, M., Kudla, M., Balaz, P., Zadinova, M., Ostrow, J. D., Wong, R. J., Vitek, L. 2008; 867 (1): 37-42

  Abstract

  Unconjugated bilirubin (UCB) exhibits potent antioxidant and cytoprotective properties, but causes apoptosis and cytotoxicity at pathologically elevated concentrations. Accurate measurement of UCB concentrations in cells, fluids and tissues is needed to evaluate its role in redox regulation, prevention of atherosclerotic and malignant diseases, and bilirubin encephalopathy. In the present study, we developed and validated a highly sensitive method for tissue UCB determinations. UCB was extracted from rat organs with chloroform/methanol/hexane at pH 6.2 and then partitioned into a minute volume of alkaline buffer that was subjected to HPLC using an octyl reverse phase (RP) column. Addition of mesobilirubin as an internal standard corrected for losses of UCB during extraction. Recoveries averaged 75+/-5%. The detection limit was 10pmol UCB/g wet tissue. Variance was +/-2.5%. When used to measure UCB concentrations in tissues of jaundiced Gunn rats, this procedure yielded UCB levels directly comparable to published methods, and accurately determined very low tissue bilirubin concentrations (

  View details for DOI 10.1016/j.jchromb.2008.03.005

  View details for Web of Science ID 000256200400007

  View details for PubMedID 18373963

• Severe hemolysis with normal blood count in a glucose-6-phosphate dehydrogenase deficient neonate JOURNAL OF PERINATOLOGY Kaplan, M., Hammerman, C., Vreman, H. J., Wong, R. J., Stevenson, D. K. 2008; 28 (4): 306-309

  Abstract

  A premature glucose-6-phosphate dehydrogenase (G-6-PD) deficient neonate was readmitted for exponential rise in the plasma bilirubin concentration to 33.0 mg dl(-1). Blood carboxyhemoglobin (2.8% of total hemoglobin, >threefold normal value) confirmed the presence of hemolysis; however, hematological indices were unchanged from the birth hospitalization. Serum unbound bilirubin, although present, was probably at a concentration insufficient to cause bilirubin encephalopathy. In G-6-PD deficient neonates, severe hemolysis may occur in the absence of hematological changes typical of a hemolytic process.

  View details for DOI 10.1038/sj.jp.7211919

  View details for Web of Science ID 000254782000012

  View details for PubMedID 18379570

• Upregulation of the HO-1 gene by proton pump inhibitors - molecular mechanisms and functinal consequences 49th Annual Meeting of the German-Society-for-Experimental-and-Clinical-Pharmacology-and-Toxicology Schulz, S., Grosser, N., Erdmann, K., Hinkelmann, U., Wong, R. J., Stevenson, D. K., Schroeder, H. SPRINGER. 2008: 60–60

  View details for Web of Science ID 000254204900291

• Heme oxygenase-1 and simvastatin: molecular mechanisms and in vivo effects 49th Annual Meeting of the German-Society-for-Experimental-and-Clinical-Pharmacology-and-Toxicology Hinkelmann, U., Schulz, S., Grosser, N., Wong, R. J., Morisawa, T., Stevenson, D. K., Muchova, L., Schroeder, H. SPRINGER. 2008: 58–58

  View details for Web of Science ID 000254204900282

• Efficacy of chromium mesoporphyrin in inhibiting heme oxygenase activity in heme-loaded newborn mice Western Regional Meeting of the American-Federation-for-Medical-Research Morisawa, T., Xiao, H. H., Wong, R. J., Bhutani, V. K., Vreman, H. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2008: 198–98

  View details for Web of Science ID 000252793300297

• Time-dependent effects of chromium mesoporphyrin on the inhibition of heme oxygenase activity in newborn mice Western Regional Meeting of the American-Federation-for-Medical-Research Xiao, H. H., Morisawa, T., Wong, R. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2008: 260–61

  View details for Web of Science ID 000252793301130

• (TA)(n) UGT 1A1 promoter polymorphism: A crucial factor in the pathophysiology of jaundice in G-6-PD deficient neonates PEDIATRIC RESEARCH Kaplan, M., Renbaum, P., Vreman, H. J., Wong, R. J., Levy-Lahad, E., Hammerman, C., Stevenson, D. K. 2007; 61 (6): 727-731

  Abstract

  Increased heme catabolism has been reported in glucose-6-phosphate dehydrogenase (G-6-PD)-normal neonates who were also homozygous for (TA)7/(TA)7 (UGT1A1*28) uridine diphosphoglucuronate-glucuronosyltransferase 1A1 (UGT) promoter polymorphism (Gilbert syndrome). As G-6-PD deficiency is associated with increased hemolysis, we hypothesized that in G-6-PD-deficient neonates who also have the (TA)7/(TA)7 UGT promoter genotype, steady-state hemolysis would be even further increased. Male G-6-PD-deficient neonates were sampled for plasma total bilirubin (PTB), blood carboxyhemoglobin corrected for inhaled carbon monoxide in ambient air (COHbc) (an index of heme catabolism), and UGT (TA)n promoter genotype determination and compared with previously published G-6-PD-normal neonates. Although COHbc values were higher in the G-6-PD-deficient than in the G-6-PD-normal cohorts (0.97 +/- 0.32% of total Hb (tHb) versus 0.76 +/- 0.19% of tHb, p < 0.001), PTB values were similar (9.2 +/- 3.4 mg/dL versus 8.9 +/- 3.0 mg/dL, respectively, p = 0.3). Within the G-6-PD-deficient group, although COHbc values were alike between the three UGT promoter genotypes, PTB was higher in the (TA)7/(TA)7 homozygotes (11.1 +/- 4.0 mg/dL) compared with (TA)6/(TA)7 heterozygotes (9.1 +/- 3.2 mg/dL, p = 0.03) and wild-type (TA)6/(TA)6 homozygotes (8.8 +/- 3.4 mg/dL, p = 0.02). In the steady state, similar rates of hemolysis, but increased PTB in the G-6-PD- deficient, (TA)7/(TA)7 homozygotes, imply that (TA)7/(TA)7, homozygosity is central to increased PTB.

  View details for DOI 10.1203/pdr.0b013e31805365c5

  View details for Web of Science ID 000246787300019

  View details for PubMedID 17426648

• Chronic administration of statins increases bilirubin and carbon monoxide formation in the heart - A novel mechanism of antioxidant protection 76th Congress of the European-Atherosclerosis-Society/15th Paavo Nurmi Symposium Muchova, L., Wong, R. J., HSU, M., Morioka, I., Zelenka, J., Vitek, L., Schroder, H., Stevenson, D. K. ELSEVIER IRELAND LTD. 2007: 206–

  View details for Web of Science ID 000247869100830

• Heme oxygenase-1 stabilizes the blood-spinal cord barrier and limits oxidative stress and white matter damage in the acutely injured murine spinal cord JOURNAL OF CEREBRAL BLOOD FLOW AND METABOLISM Lin, Y., Vreman, H. J., Wong, R. J., Tjoa, T., Yamauchi, T., Noble-Haeusslein, L. J. 2007; 27 (5): 1010-1021

  Abstract

  We hypothesized that heme oxygenase (HO)-1, the inducible form of HO, represents an important defense against early oxidative injury in the traumatized spinal cord by stabilizing the blood-spinal cord barrier and limiting the infiltration of leukocytes. To test this hypothesis, we first examined the immunoexpression of HO-1 and compared barrier permeability and leukocyte infiltration in spinal cord-injured HO-1-deficient (+/-) and wild-type (WT, +/+) mice. Heme oxygenase was expressed in both endothelial cells and glia of the injured cord. Barrier disruption to luciferase and infiltration of neutrophils were significantly greater in the HO-1+/- than WT mice at 24 h postinjury (P

  View details for DOI 10.1038/sj.jcbfm.9600412

  View details for Web of Science ID 000245999300012

  View details for PubMedID 17047682

• Effects of sample dilution, peroxidase concentration, and chloride ion on the measurement of unbound bilirubin in premature newborns CLINICAL BIOCHEMISTRY Ahlfors, C. E., Vreman, H. J., Wong, R. J., Bender, G. J., Oh, W., Morris, B. H., Stevenson, D. K. 2007; 40 (3-4): 261-267

  Abstract

  To assess the effects of sample dilution, peroxidase concentration, and chloride ion (Cl(-)) on plasma unbound bilirubin (B(f)) measurements made using a commercial peroxidase methodology (UB Analyzer) in a study population of ill, premature newborns.B(f) was measured with a UB Analyzer in 74 samples at the standard 42-fold sample dilution and compared with B(f) measured at a 2-fold sample dilution using a FloPro Analyzer. B(f) was measured at two peroxidase concentrations to determine whether the peroxidase steady state B(f) (B(fss)) measurements were significantly less than the equilibrium B(f) (B(feq)), in which case it was necessary to calculate B(feq) from the two B(fss) measurements. B(f) was also measured before and after adding 100 mmol/L Cl(-) to the UB Analyzer assay buffer.B(feq) at the 42-fold dilution was nearly 10-fold less than but it correlated significantly with B(feq) at the 2-fold dilution (mean 8.2+/-5.2 nmol/L versus 73.5+/-70 nmol/L, respectively, p<0.0001; correlation r=0.6). The two UB Analyzer B(fss) measurements were significantly less than B(feq) in 42 of 74 (57%) samples, and Cl(-) increased B(feq) in 66 of 74 (89%) samples by a mean of 82+/-67%.B(fss) measured by the UB Analyzer at the standard 42-fold sample dilution using assay buffer without Cl(-) and a single peroxidase concentration is significantly less than the B(feq) in undiluted plasma. Accurate B(f) measurements can be made only in minimally diluted serum or plasma.

  View details for DOI 10.1016/j.clinbiochem.2006.09.006

  View details for Web of Science ID 000244193000020

  View details for PubMedID 17069786

  View details for PubMedCentralID PMC1945224

• Inhibition of heme oxygenase activity following repeated heme loads by tin mesoporphyrin in newborn mice. 8th Conference of the Western Student Medical Research Forum/Western Section of the American-Federation-for-Medical-Research/Western Association-of-Physicians/Western-Society-for-Pediatric-Research/Western-Society-for-Clinical-Investigation Wong, R. J., Morioka, I., Muchova, L., Vreman, H. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2007: S88–S88

  View details for Web of Science ID 000247692400083

• Effect of early hand expression on milk production in pump-dependent mothers of preterm infants. 8th Conference of the Western Student Medical Research Forum/Western Section of the American-Federation-for-Medical-Research/Western Association-of-Physicians/Western-Society-for-Pediatric-Research/Western-Society-for-Clinical-Investigation Morton, J., Hall, J. Y., Thairu, L., Nomanbhoy, S., Bhutani, R., Carlson, S., Wong, R. J., Rhine, W. D. LIPPINCOTT WILLIAMS & WILKINS. 2007: S116–S116

  View details for Web of Science ID 000247692400255

• Maximizing milk volumes of pump-dependent mothers with breast massage. 8th Conference of the Western Student Medical Research Forum/Western Section of the American-Federation-for-Medical-Research/Western Association-of-Physicians/Western-Society-for-Pediatric-Research/Western-Society-for-Clinical-Investigation Morton, J., Hall, J. Y., Thairu, L., Nomanbhoy, S., Bhutani, R., Carlson, S., Wong, R. J., Rhine, W. D. LIPPINCOTT WILLIAMS & WILKINS. 2007: S117–S117

  View details for Web of Science ID 000247692400259

• Efficacy of azalanstat in inhibiting heme oxygenase activity in newborn mice. 8th Conference of the Western Student Medical Research Forum/Western Section of the American-Federation-for-Medical-Research/Western Association-of-Physicians/Western-Society-for-Pediatric-Research/Western-Society-for-Clinical-Investigation Morisawa, T., Wong, R. J., Bhutani, V. K., Vreman, H. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2007: S89–S89

  View details for Web of Science ID 000247692400087

• Comparison of heme oxygenase activity in gut-associated lymphoid tissues and the intestine. 8th Conference of the Western Student Medical Research Forum/Western Section of the American-Federation-for-Medical-Research/Western Association-of-Physicians/Western-Society-for-Pediatric-Research/Western-Society-for-Clinical-Investigation Burns-Guydisli, S. M., Wong, R. J., Kafsh, F., Contag, C. H., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2007: S117–S118

  View details for Web of Science ID 000247692400262

• Determination of in vivo carbon monoxide production in laboratory animals via exhaled air. Journal of pharmacological and toxicological methods Dercho, R. A., Nakatsu, K., Wong, R. J., Stevenson, D. K., Vreman, H. J. 2006; 54 (3): 288-295

  Abstract

  In vitro assays play an important role in the understanding of the heme oxygenase (HO)/carbon monoxide (CO) pathway. However, because physiological roles for the products of this pathway are hypothesized, it is becoming increasingly important to perform in vivo studies. Since CO production is primarily mediated by HO and is excreted mainly by the lungs, measurements of total body CO excretion (VeCO) via the breath allow continuous, noninvasive monitoring of heme degradation and CO and bilirubin production. Here, we describe a modified flow-through method for the collection and quantitation of CO from small laboratory animals.Mice and rats were studied in gas-tight chambers supplied with a continuous flow of CO-free air. CO in the exhaust air was measured by gas chromatography with a reduction gas analyzer. After establishing baseline VeCO levels, animals were administered various xenobiotics known to alter HO activity and further monitored for changes in CO production for up to 12 h without observable distress.Administration of heme (substrate for HO) resulted in reproducible increases in CO production; whereas, prior administration of zinc protoporphyrin (ZnPP, HO inhibitor) or cobalt protoporphyrin (CoPP, HO inducer) resulted in respective dose-dependent decreases and increases in the heme-induced CO production.We have demonstrated that this noninvasive method of CO quantitation reliably estimates heme degradation with sensitivity to distinguish between different types of HO-manipulating xenobiotics in a dose-dependant manner in both mouse and rat models. Furthermore, VeCO measurements allow nearly real-time determinations of CO and bilirubin formation, which helps to illustrate the time course of drug action.

  View details for PubMedID 16540352

• Concentration of carbon monoxide (CO) in postmortem human tissues: Effect of environmental CO exposure Annual Meeting of the Society-of-Pediatric-Research Vreman, H. J., Wong, R. J., Stevenson, D. K., Smialek, J. E., Fowler, D. R., Li, n. L., Vigorito, R. D., Zielke, H. R. WILEY-BLACKWELL PUBLISHING, INC. 2006: 1182–90

  Abstract

  We studied how carbon monoxide (CO) is distributed within the human body through quantitation of CO concentrations in postmortem tissue samples from fatalities including possible CO exposure. Stored, frozen tissues were diced, sonicated in water, and 0.01-8.0 mg wet weight (ww) tissues were incubated with sulfosalicylic acid in CO-purged, septum-sealed vials. CO released into the headspace was quantitated by reduction gas chromatography. Mean tissue CO concentrations (pmol/mg ww) from subjects diagnosed to have no known CO exposure (control, N=14), died from fire (N=13), and CO asphyxiation (N=7), respectively, were: adipose (2;13;9), brain (3;13;65), muscle (15;97;297), heart (30;99;371), kidney (22;432;709, lung (54;690;2638), spleen (73;1366;3548), and blood (162;2238;5070). Carboxyhemoglobin concentrations were 1.4%, 25.2%, and 69.1% of total hemoglobin, respectively. We conclude that measurements of CO concentration in a variety of tissues can be used as markers for the degree of exogenous CO exposure and the identification of possible causes of death.

  View details for DOI 10.1111/j.1556-4029.2006.00212.x

  View details for PubMedID 17018107

• End-tidal carbon monoxide measurements in infant respiratory distress syndrome ACTA PAEDIATRICA Krediet, T. G., Cirkel, G. A., Vreman, H. J., Wong, R. J., Stevenson, D. K., Groenendaal, F., Egberts, J., van Bel, F. 2006; 95 (9): 1075-1082

  Abstract

  RDS involving inflammatory and oxidative processes may lead to increased production of carbon monoxide (CO).The relationship between end-tidal CO, corrected for inhaled CO (ETCOc), and RDS severity was investigated in preterm infants as well as the value of early ETCOc measurements to predict chronic lung disease.78 infants (30 no RDS, 32 moderate RDS, 16 severe RDS) were included. ETCOc was measured using the CO-Stat End Tidal Breath Analyzer.ETCOc was significantly higher in RDS compared to no RDS during the first week (p<0.05). Severity of RDS was the most significant independent variable in a stepwise regression model related to ETCOc (F-test: 18.17). Negative predictive value of early (within first 12 h of life) ETCOc measurement (<2.5 ppm) for development of chronic lung disease was excellent (100%).During severe RDS, inflammation may contribute to increased lipid peroxidation leading to increased local CO production in the lung, indicated by increased ETCOc. Early ETCOc determinations may be helpful to exclude occurrence of chronic lung disease.

  View details for DOI 10.1080/08035250500537017

  View details for Web of Science ID 000240122500009

  View details for PubMedID 16938753

• The effect of tin-mesoporphyrin on Bach 1 expression Experimental Biology 2006 Annual Meeting Abate, A., Zhao, H., Wong, R. J., Stevenson, D. K. FEDERATION AMER SOC EXP BIOL. 2006: A1344–A1344

  View details for Web of Science ID 000236326204408

• Transcutaneous bilirubinometry: A noninvasive tool for studying newborn jaundiced rats before and after exposure to light PEDIATRIC RESEARCH Vreman, H. J., Wong, R. J., Chan, M. L., Young, B. W., Stevenson, D. K. 2006; 59 (2): 203-209

  Abstract

  The homozygous Gunn rat is the most frequently used animal model for the study of neonatal jaundice. We evaluated the applicability of noninvasive transcutaneous bilirubin (TcB) measurements as an index of serum total bilirubin (STB) levels in neonatal rats by comparison to invasive STB measurements. TcB measurements were made during the first 96 h of life with the Model 101 Minolta/Air-Shields Jaundice Meter (JM) and SpectRx BiliCheck System (BC). Measurements with both devices displayed parallel TcB profiles, rapidly rising within 24 h, increasing during the next 6 h, then leveling off after 30 h. Linear regressions for the JM (n = 60) were as follows: STB (mg/dL) = 0.79 (JM) - 0.01 (units, r = 0.95, head); STB (mg/dL) = 0.82 (JM) + 1.51 (units, r = 0.95, upper back); and STB (mg/dL) = 0.74 (JM) + 1.60 (units, r = 0.91, lower back). Mean bias +/- imprecision were as follows: -0.02 +/- 3.99 mg/dL, -0.01 +/- 3.90, and 0.01 +/- 4.28 at the head, upper back, and lower back, respectively. For the BC, only lower back measurements were taken, and the regression was as follows: STB (mg/dL) = 0.77 (BC) + 1.65 mg/dL, (r = 0.93, n = 29) with a mean bias +/- imprecision of -1.08 +/- 3.08 mg/dL. When pups were exposed to light, correlations remained strong but intercepts increased. These results demonstrate that noninvasive TcB measurements correlate highly with STB in the Gunn rat during the first 96 h of life and after exposure to light. We conclude that JM measurements at the head and BC at the lower back reflect STB most reliably and consistently. Thus, in addition to being a useful tool for evaluating jaundice in human neonates, TcB methodology can be used successfully for the noninvasive monitoring of jaundice in neonatal Gunn rats pre- and postlight exposure.

  View details for DOI 10.1203/01.pdr.000196737.73851.8a

  View details for PubMedID 16439579

• The effectiveness of oral tin mesoporphyrin prophylaxis in reducing bilirubin production after an oral heme load in a transgenic mouse model BIOLOGY OF THE NEONATE DeSandre, G. H., Wong, R. J., Morioka, I., Contag, C. H., Stevenson, D. K. 2006; 89 (3): 139-146

  Abstract

  Neonatal jaundice is commonly encountered and rarely associated with morbidity and mortality. Nonetheless, infants with glucose-6-phosphate dehydrogenase deficiency often have hemolysis (a heme load) caused by an environmental oxidant trigger, thus increasing their risk for serious morbidity. The use of tin mesoporphyrin (SnMP) has been proposed for interdicting the development of severe hyperbilirubinemia in a variety of conditions.We studied the in vivo effects of prophylactic oral SnMP on heme oxygenase (HO) activity and bilirubin production, as indexed by the excretion rate of carbon monoxide (VeCO), following a subsequent oral heme load.Adult mice were exposed serially to heme and assessed for in vivo bilirubin production rates, HO-1 transcription and protein, and HO activity. The effect of prophylaxis with a single oral dose of SnMP prior to an oral heme load was assessed by measuring VeCOand tissue HO activities.After serial heme exposures, VeCO, HO-1 transcription and protein, and liver and spleen HO activities increased incrementally. After pretreatment with oral SnMP, bilirubin production decreased in response to an oral heme load. Also, heme-mediated increases in liver, spleen, and intestine HO activities were significantly dampened.A single oral dose of SnMP results in durable inhibition of bilirubin production and HO activity for at least 24 h in a mouse model of oral heme loading. Further studies are needed to fully elucidate the duration of this protection against hyperbilirubinemia due to a delayed heme load and any long-term consequences of prophylaxis with SnMP on HO-1 transcription and HO-1 protein.

  View details for DOI 10.1159/000088717

  View details for PubMedID 16205054

• Metalloporphyrin inhibition of in vitro mouse heme oxygenase isozyme activity. Western Regional Meeting of the American-Federation-for-Medical-Research Pierce, N. W., Wong, R. J., Morioka, I., Vreman, H. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2006: S120–S121

  View details for Web of Science ID 000235301500251

• Statins as tissue-specific inducers of in vivo heme oxygenase expression. Western Regional Meeting of the American-Federation-for-Medical-Research HSU, M., Muchova, L., Morioka, I., Wong, R. J., Schroder, H., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2006: S121–S121

  View details for Web of Science ID 000235301500252

• Statins as tissue-specific inducers of in vivo heme oxygenase expression Western Regional Meeting of the American-Federation-for-Medical-Research HSU, M., Muchova, L., Morioka, I., Wong, R. J., Schroder, H., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2006: S86–S86

  View details for Web of Science ID 000235301500051

• Dose-dependent effects of tin mesoporphyrin on heme oxygenase activity inhibition in newborn mice. Western Regional Meeting of the American-Federation-for-Medical-Research Morioka, I., Wong, R. J., Vreman, H. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2006: S128–S128

  View details for Web of Science ID 000235301500293

• Heme regulation in traumatic brain injury: relevance to the adult and developing brain JOURNAL OF CEREBRAL BLOOD FLOW AND METABOLISM Chang, E. F., Claus, C. P., Vreman, H. J., Wong, R. J., Noble-Haeusslein, L. J. 2005; 25 (11): 1401-1417

  Abstract

  Intracranial bleeding is one of the most prominent aspects in the clinical diagnosis and prognosis of traumatic brain injury (TBI). Substantial amounts of blood products, such as heme, are released because of traumatic subarachnoid hemorrhages, intraparenchymal contusions, and hematomas. Despite this, surprisingly few studies have directly addressed the role of blood products, in particular heme, in the setting of TBI. Heme is degraded by heme oxygenase (HO) into three highly bioactive products: iron, bilirubin, and carbon monoxide. The HO isozymes, in particular HO-1 and HO-2, exhibit significantly different expression patterns and appear to have specific roles after injury. Developmentally, differences between the adult and immature brain have implications for endogenous protection from oxidative stress. The aim of this paper is to review recent advances in the understanding of heme regulation and metabolism after brain injury and its specific relevance to the developing brain. These findings suggest novel clinical therapeutic options for further translational study.

  View details for Web of Science ID 000232815000001

  View details for PubMedID 15917748

• Apigenin decreases hemin-mediated heme oxygenase-1 induction FREE RADICAL BIOLOGY AND MEDICINE Abate, A., Yang, G., Wong, R. J., Schroder, H., Stevenson, D. K., Dennery, P. A. 2005; 39 (6): 711-718

  Abstract

  Hemin is a strong inducer of heme oxygenase-1 (HO-1) expression in vitro and in vivo. Whereas moderate overexpression of HO-1 is protective against oxidative stress, uncontrolled levels of HO-1 can be detrimental. Therefore, we evaluated the effects of apigenin (APG), a flavonoid involved in a number of phosphorylation pathways and also known to inhibit inducible genes, such as iNOS and COX-2, on HO-1 expression. Incubation of mouse embryonic fibroblasts with APG (5--40 microM) decreased hemin-induced HO-1 protein and mRNA expression. APG also reduced the induction of HO-1 promoter activity, as assessed by bioluminescence imaging, in NIH3T3 cells transfected with the 15-kb HO-1 promoter fused with the reporter gene luciferase (HO-1-luc). Furthermore, through the use of specific inhibitors, APG's effect was found to be unrelated to its PKC, CK 2, PI 3 K, p38, or ERK inhibitory activities. Quercetin (10--40 microM), also a flavonoid, also inhibited hemin-induced HO-1 expression. Additionally, in vivo studies using HO-1-luc transgenic mice showed that APG (50 mg/kg) decreased hemin-induced HO activity and HO-1 protein expression in the liver. These results suggest that hemin-induced HO-1 expression can be attenuated by flavonoids, such as APG.

  View details for DOI 10.1016/j.freeradbiomed.2005.01.020

  View details for PubMedID 16109301

• Determination of carbon monoxide (CO) in rodent tissue: Effect of heme administration and environmental CO exposure ANALYTICAL BIOCHEMISTRY Vreman, H. J., Wong, R. J., Kadotani, T., Stevenson, D. K. 2005; 341 (2): 280-289

  Abstract

  Carbon monoxide (CO), produced endogenously during heme degradation, is considered a messenger molecule in vascular and neurologic tissues. To study this role, it is important to determine CO concentration in target tissues pre- and post-perturbations. Here, we describe a sensitive and reproducible method, which is linear and accurate, and provide some examples of its application for quantitation of CO concentrations in tissues pre- and post-perturbations. Tissues from adult rats and mice were sonicated (20% w/w), and volumes representing 0.04-8 mg fresh weight (FW) were incubated at 0 degrees C for 30 min with sulfosalicylic acid. CO liberated into the headspace was quantitated by gas chromatography. Tissue CO concentrations (mean+/-SD, pmol CO/mg FW) were as follows: blood (47+/-10, 45+/-5), muscle (4+/-4, 10+/-1), kidney (5+/-2, 7+/-2), heart (6+/-3, 6+/-1), spleen (11+/-3, 6+/-1), liver (4+/-1, 5+/-1), intestine (2+/-1, 4+/-2), lung (2+/-1, 3+/-1), testes (1+/-1, 2+/-1), and brain (2+/-1, 2+/-0) in untreated rat (n=3) and mouse (n=5), respectively. Between the rat and the mouse, only CO concentrations in the muscle and spleen were significantly different (p0.05). Endogenous CO generation, after administration of heme arginate to mice (n=3), increased CO concentrations by 0-43 pmol/mg FW. Exposure of mice (n=3) to 500 ppm CO for 30 min yielded significantly elevated CO concentrations by 4-2603 pmol/mg FW in all tissues over the native state. While blood had the highest CO concentration for all conditions, muscle, kidney, heart, spleen, and liver, all rich in hemoglobin and/or other CO-binding hemoproteins, also contained substantial CO concentrations. Intestine, lung, testes, and brain contained the lowest CO concentrations.

  View details for DOI 10.1016/j.ab.2005.03.019

  View details for PubMedID 15907874

• Reduction in hospital readmission rates for hyperbilirubinemia is associated with use of transcutaneous bilirubin measurements CLINICAL CHEMISTRY Stevenson, D. K., Wong, R. J., Vreman, H. J. 2005; 51 (3): 481-482

  View details for DOI 10.1373/clinchem.2004.046789

  View details for Web of Science ID 000227230600001

  View details for PubMedID 15738511

• Is carbon monoxide-mediated cyclic guanosine monophosphate production responsible for low blood pressure in neonatal respiratory distress syndrome? JOURNAL OF APPLIED PHYSIOLOGY van Bel, F., Latour, V., Vreman, H. J., Wong, R. J., Stevenson, D. K., Steendijk, P., Egberts, J., Krediet, T. G. 2005; 98 (3): 1044-1049

  Abstract

  Infant respiratory distress syndrome (RDS) involves inflammatory processes, causing an increased expression of inducible heme oxygenase with subsequent production of carbon monoxide (CO). We hypothesized that increased production of CO during RDS might be responsible for increased plasma levels of vasodilatory cGMP and, consequently, low blood pressure observed in infants with RDS. Fifty-two infants (no-RDS, n = 21; RDS, n = 31), consecutively admitted to the neonatal intensive care unit (NICU) between January and October 2003 were included. Hemoglobin-bound carbon monoxide (COHb), plasma cGMP, plasma nitric oxide (NOx), and bilirubin were determined at 0-12, 48-72, and at 168 h postnatally, with simultaneous registration of arterial blood pressure. Infants with RDS had higher levels of cGMP and COHb compared with no-RDS infants (RDS vs. no-RDS: cGMP ranging from 76 to 101 vs. 58 to 82 nmol/l; COHb ranging from 1.2 to 1.4 vs. 0.9 to 1.0%). Highest values were reached at 48-72 h [RDS vs. no-RDS mean (SD): cGMP 100 (39) vs. 82 (25) nmol/l (P < 0.001); COHb 1.38 (0.46) vs. 0.91 (0.26)% (P < 0.0001)]. Arterial blood pressure was lower and more blood pressure support was needed in RDS infants at that point of time [RDS vs. no-RDS mean (SD): mean arterial blood pressure 33 (6) vs. 42 (5) mmHg (P < 0.05)]. NOx was not different between groups and did not vary with time. Multiple linear regression analysis showed a significant correlation between cGMP and COHb, suggesting a causal relationship. Mean arterial blood pressure appeared to be primarily correlated to cGMP levels (P < 0.001). We conclude that a CO-mediated increase in cGMP causes systemic vasodilation with a consequent lower blood pressure and increased need for blood pressure support in preterm infants with RDS.

  View details for DOI 10.1152/japplphysiol.00760.2004

  View details for Web of Science ID 000226863100036

  View details for PubMedID 15516362

• Attenuation of heme induction following exposure to tin mesoporphyrin in mice. DeSandre, G. H., Wong, R. J., Morioka, I., Vreman, H. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2005: S125–S125

  View details for Web of Science ID 000226539700280

• Phototherapy: Current methods and future directions SEMINARS IN PERINATOLOGY Vreman, H. J., Wong, R. J., Stevenson, D. K. 2004; 28 (5): 326-333

  Abstract

  Phototherapy is the most common therapeutic intervention used for the treatment of hyperbilirubinemia. Although it has become a mainstay since its introduction in 1958, a better understanding of the photobiology of bilirubin, characteristics of the phototherapy devices, the efficacy and safety considerations of phototherapy applications, and improvements in spectroradiometers and phototherapy devices are necessary for more predictable and improved clinical practices and outcomes. A step forward in instituting consistent, uniform, and effective use of phototherapy is the recent American Academy of Pediatrics clinical guideline on the management of hyperbilirubinemia in the newborn infant 35 or more weeks of gestation, which outlines a clinical strategy for the diagnosis of hyperbilirubinemia and contains direct recommendations for the application of phototherapy. This article reviews the parameters that determine the efficacy of phototherapy, briefly discusses current devices and methods used to deliver phototherapy, and speculates on future directions and studies that are still needed to complement our presently incomplete knowledge of the facets of this common mode of therapy.

  View details for DOI 10.1053/j.semperi.2004.09.003

  View details for PubMedID 15686263

• NICHD Conference on Kernicterus: Research on Prevention of Bilirubin-Induced Brain Injury and Kernicterus: Bench-to-Bedside--Diagnostic Methods and Prevention and Treatment Strategies. Journal of perinatology Stevenson, D. K., Wong, R. J., Vreman, H. J., McDonagh, A. F., Maisels, M. J., Lightner, D. A. 2004; 24 (8): 521-525

  Abstract

  In July 2003, the National Institute of Child Health and Human Development (NICHD) organized a consensus conference, where a group of experts were invited to review and discuss the current state of knowledge regarding neonatal hyperbilirubinemia and identify areas in which where future research should be directed. This paper summarizes the presentations addressing the current methodologies for direct and noninvasive assessments of serum total bilirubin concentrations as well as prevention and treatment strategies for the management of neonatal hyperbilirubinemia.

  View details for PubMedID 15129227

• Transcutaneous bilirubin measurements and serum total bilirubin levels in indigenous African infants PEDIATRICS Slusher, T. M., Angyo, I. A., Bode-Thomas, F., Akor, F., Pam, S. D., Adetunji, A. A., McLaren, D. W., Wong, R. J., Vreman, H. J., Stevenson, D. K. 2004; 113 (6): 1636-1641

  Abstract

  The objective of this study was to determine whether transcutaneous bilirubin (TcB) measurements correlate with serum total bilirubin (STB) levels in indigenous, darkly pigmented African newborns with varying degrees of skin pigmentation, some of which had developed kernicterus.Jaundiced infants who were < or =2 weeks of age and admitted to Baptist Medical Center-Eku (Eku; n = 29) and Jos University Teaching Hospital (Jos; n = 98) in Nigeria were studied. TcB measurements using the BiliChek were made simultaneously with blood sampling for STB measurements by spectrophotometry before phototherapy.Using linear regression analysis, we found that measurements of TcB correlated well with those of STB with r values of.90 and.88 for Eku and Jos, respectively. Mean bias and imprecision of TcB measurements as compared with STB measurements for the total population was 0.5 +/- 7.6 mg/dL using the method of Bland and Altman. At STB > or 12 mg/dL, correlation (r =.84) and bias and imprecision (-1.2 +/- 8.6 mg/dL) of measurements were only slightly poorer. Furthermore, when infants were grouped by degree of skin pigmentation, correlations of TcB and STB measurements remained strong.From these results, we can conclude that TcB measurements are a useful and reliable index for estimating STB levels in pigmented neonates, including those with hyperbilirubinemia and kernicterus. In the absence of reliable STB measurements, the relatively simple and noninvasive TcB measurements can be an important adjunct in directing phototherapy and exchange transfusions, thereby preventing bilirubin-induced morbidity and mortality in low-technology clinical environments.

  View details for Web of Science ID 000221781500012

  View details for PubMedID 15173484

• Induction of heme oxygenase-1 in transgenic mouse fibroblast cells following successive exposures to heme Annual Meeting of the Pediatric-Academic-Societies DeSandre, G. H., Wong, R. J., Vreman, H. J., Stevenson, D. K. NATURE PUBLISHING GROUP. 2004: 473A–473A

  View details for Web of Science ID 000220591102746

• A primer on neonatal jaundice. Advances in pediatrics Stevenson, D. K., Wong, R. J., DeSandre, G. H., Vreman, H. J. 2004; 51: 263-288

  View details for PubMedID 15366777

• Induction of heme oxygenase-1 in mouse fibroblast cells after successive exposures to heme. Western Regional Meeting of the American-Federation-for-Medical-Research DeSandre, G. H., Wong, R. J., Vreman, H. J., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2004: S122–S122

  View details for Web of Science ID 000188254600268

• Characterization of coelenterazine analogs for measurements of Renilla luciferase activity in live cells and living animals. Molecular imaging Zhao, H., Doyle, T. C., Wong, R. J., Cao, Y., Stevenson, D. K., Piwnica-Worms, D., Contag, C. H. 2004; 3 (1): 43-54

  Abstract

  In vivo imaging of bioluminescent reporters relies on expression of light-emitting enzymes, luciferases, and delivery of chemical substrates to expressing cells. Coelenterazine (CLZN) is the substrate for a group of bioluminescent enzymes obtained from marine organisms. At present, there are more than 10 commercially available CLZN analogs. To determine which analog is most suitable for activity measurements in live cells and living animals, we characterized 10 CLZN analogs using Renilla luciferase (Rluc) as the reporter enzyme. For each analog, we monitored enzyme activity, auto-oxidation, and efficiency of cellular uptake. All CLZN analogs tested showed higher auto-oxidation signals in serum than was observed in phosphate buffer or medium, mainly as a result of auto-oxidation by binding to albumin. CLZN-f, -h, and -e analogs showed 4- to 8-fold greater Rluc activity, relative to CLZN-native, in cells expressing the enzyme from a stable integrant. In studies using living mice expressing Rluc in hepatocytes, administration of CLZN-e and -native produced the highest signal. Furthermore, distinct temporal differences in signal for each analog were revealed following intravenous or intraperitoneal delivery. We conclude that the CLZN analogs that are presently available vary with respect to hRluc utilization in culture and in vivo, and that the effective use of CLZN-utilizing enzymes in living animals depends on the selection of an appropriate substrate.

  View details for PubMedID 15142411

• Effects of metalloporphyrins on heme oxygenase-1 transcription: correlative cell culture assays guide in vivo imaging. Molecular imaging Hajdena-Dawson, M., Zhang, W., Contag, P. R., Wong, R. J., Vreman, H. J., Stevenson, D. K., Contag, C. H. 2003; 2 (3): 138-149

  Abstract

  Heme oxygenase (HO) is the rate-limiting step in the heme degradation pathway and is a potential target for the control, or prevention, of pathologic jaundice in neonates. Metalloporphyrins (Mps), a diverse set of synthetic derivatives of heme, can competitively inhibit the HO enzymes. However, certain Mps are phototoxic and some increase transcription of HO-1, the inducible HO isozyme. Therefore, effective development of this class of compounds as therapeutics for treating pathologic jaundice will require rapid and integrated biological screens to identify the most efficacious and safe Mps. To study the safety of these compounds, we assessed their cytotoxic effects and measured luciferase activity by bioluminescent imaging (BLI) as an index of HO-1 transcription, first in live cell cultures and then in living transgenic reporter mice. A total of 12 Mps were first evaluated in the correlative cell culture assay. Based on results from this study, 2 Mps, zinc protoporphyrin (ZnPP) and zinc bis glycol porphyrin (ZnBG), were selected for further studies in the live animal model. In vitro BLI showed ZnPP to be a strong inducer of HO-1 transcription in comparison to ZnBG, which showed minimal induction. Cytotoxicity studies revealed that ZnPP was phototoxic, whereas ZnBG had no effect on cell viability. In vivo BLI showed that both ZnPP and ZnBG had minimal effects on the levels of HO-1 transcription in the animals. Furthermore, serum enzyme assays indicated that neither caused detectable liver toxicity. These findings, and especially those with ZnBG, support the use of selected Mps as therapies for pathologic jaundice. Coupling the high throughput advantage of cell culture with the capability of imaging for whole-body temporal analyses could accelerate and refine the preclinical phases of drug development. Thus, this study serves as a model for understanding the effects of specific compounds in relation to defined targets using an integrated approach.

  View details for PubMedID 14649057

• Heme oxygenase-2 protects against lipid peroxidation-mediated cell loss and impaired motor recovery after traumatic brain injury JOURNAL OF NEUROSCIENCE Chang, E. F., Wong, R. J., Vreman, H. J., Igarashi, T., Galo, E., Sharp, F. R., Stevenson, D. K., Noble-Haeusslein, L. J. 2003; 23 (9): 3689-3696

  Abstract

  After traumatic brain injury (TBI), substantial extracellular heme is released from hemoproteins during hemorrhage and cell injury. Heme oxygenase (HO) isozymes are thought to detoxify the pro-oxidant heme to the potent antioxidant, bilirubin. HO-1, the inducible isozyme, is expressed in glial populations after injury and may play a protective role. However, the role of HO-2, the predominant and constitutively expressed isozyme in the brain, remains unclear after TBI. We used a controlled cortical impact injury model to determine the extent and mechanism of damage between HO-2 knock-out (KO) (-/-) and wild-type (WT) (+/+) mice. The specific cellular and temporal expressions of HO-2 and HO-1 were characterized by immunocytochemistry and Western blots. HO-2 was immunolocalized in neurons both before and after TBI, whereas HO-1 was highly upregulated in glia only after TBI. HO activity determined by gas chromatography using brain sonicates from injured HO-2 KO mice was significantly less than that of HO-2 wild types, despite the induction of HO-1 expression after TBI. Cell loss was significantly greater in KO mice in areas including the cortex, the CA3 region of hippocampus, and the lateral dorsal thalamus. Furthermore, motor recovery after injury, as measured by the rotarod assay and an inclined beam-walking task, was compromised in the KO mice. Finally, brain tissue from injured HO-2 KO mice exhibited decreased ability to reduce oxidative stress, as measured with an Fe(2+)/ascorbic acid-mediated carbon monoxide generation assay for lipid peroxidation susceptibility. These findings demonstrate that HO-2 expression protects neurons against TBI by reducing lipid peroxidation via the catabolism of free heme.

  View details for Web of Science ID 000182700100016

  View details for PubMedID 12736340

• Direct intestinal administration of metalloporphyrins and heme oxygenase expression. Western Regional Meeting of the American-Federation-for-Medical-Research Wong, R. J., Abate, A., Dennery, P. A., Vreman, H. J., Contag, C. H., Stevenson, D. K. LIPPINCOTT WILLIAMS & WILKINS. 2003: S140–S141

  View details for Web of Science ID 000180569600291

• Selection of potential therapeutics based on in vivo spatiotemporal transcription patterns of heme oxygenase-1 JOURNAL OF MOLECULAR MEDICINE-JMM Zhang, W. S., Contag, P. R., Hardy, J., Zhao, H., Vreman, H. J., Hajdena-Dawson, M., Wong, R. J., Stevenson, D. K., Contag, C. H. 2002; 80 (10): 655-664

  Abstract

  Heme oxygenase (HO), a key catabolic enzyme in the conversion of heme to bilirubin, is an ideal target for reducing bilirubin production and preventing pathological jaundice in newborn infants. Metalloporphyrins (Mps) have been well characterized as competitive inhibitors of HO and have been evaluated as potential chemopreventive agents for neonatal jaundice. However, in addition to reducing HO activity, many Mps have been shown to increase HO-1 transcription, which would likely reduce their potential therapeutic utility. The differential effects of Mps on the transcription of HO-1 were therefore evaluated in living transgenic (Tg) reporter mice. Of the compounds evaluated, we observed that zinc bis-glycol porphyrin (ZnBG), a potent inhibitor of HO enzyme activity, did not alter HO-1 transcription patterns in Tg mice. Whole body images of HO-1 transcription patterns did, however; reveal increases in HO-1 transcription in Tg mice after treatment with other Mps, heme and cadmium chloride (CdCl(2)). Intravenous injections of CdCl(2) resulted in expression patterns that differed in tempo and location from those observed in Tg mice treated with intraperitoneal injections. Spatiotemporal analyses of transcriptional regulation in living animals accelerated the assessment of an adverse effect of Mps by revealing different patterns of HO-1 transcription. Among the known inhibitors of HO enzyme activity that were evaluated in this study, ZnBG did not significantly affect HO-1 transcription and therefore may be well suited for the prevention of neonatal jaundice.

  View details for DOI 10.1007/s00109-002-0375-x

  View details for PubMedID 12395150

• The jaundiced newborn. Understanding and managing transitional hyperbilirubinemia. Minerva pediatrica Stevenson, D. K., Wong, R. J., Hintz, S. R., Vreman, H. J. 2002; 54 (5): 373-382

  Abstract

  Neonatal jaundice is one of the most common conditions diagnosed by the pediatrician. This normally benign transitional phenomenon is a dynamic balance between the production and elimination of bilirubin. These processes can be exacerbated by a number of pathophysiologic conditions, which cause either an increase in bilirubin production rates, such as hemolysis, or a decrease in bilirubin elimination rates, such as bilirubin conjugation defects. The most dangerous circumstance for an infant is the combination of increased bilirubin production with impaired elimination. These infants are at considerable risk for developing excessive and potentially dangerous hyperbilirubinemia and subsequent kernicterus. Therefore, the importance of early recognition of the imbalance is paramount. In this review, we will discuss the various risk factors associated with hyperbilirubinemia and describe strategies for the diagnosis and management of transitional hyperbilirubinemia.

  View details for PubMedID 12244276

• Heme oxygenase-1 modulates fetal growth in the rat LABORATORY INVESTIGATION Kreiser, D., Nguyen, X., Wong, R., Seidman, D., Stevenson, D., Quan, S., Abraham, N., Dennery, P. A. 2002; 82 (6): 687-692

  Abstract

  Intrauterine growth restriction is associated with increased perinatal morbidity and mortality as well as with lifelong cardiovascular and metabolic complications. Deficiency of heme oxygenase 1 (HO-1) is associated with growth restriction in mice and in humans, suggesting a role for HO-1 in fetal growth and maintenance of pregnancy. We hypothesized that modulation of HO-1 in the pregnant rat would alter fetal growth. In pregnant dams, placental HO activity was significantly inhibited with zinc deuteroporphyrin IX 2,4 bis glycol, and HO-1 protein was increased by transducing adenoviral human HO-1. Inhibition of HO-1 by zinc deuteroporphyrin IX 2,4 bis glycol resulted in a significant decrease in pup size, whereas transfection with hHO-1 resulted in increased pup size. Furthermore, the expression of IGF binding protein-1 and its receptor paralleled the expression of HO-1 in the placenta and were significantly modulated by modification of HO-1 along with the expression of vascular endothelial growth factor. These observations demonstrate that HO-1 modulates fetal growth by its effects on placental growth factors.

  View details for DOI 10.1097/01.LAB.0000017167.26718.F2

  View details for PubMedID 12065678

• Role of carbon monoxide and nitric oxide in newborn infants with postasphyxial hypoxic-ischemic encephalopathy PEDIATRICS Vreman, H. J., Wong, R. J., Stevenson, D. K., Engel, R. R. 2002; 109 (4): 715-715

  View details for Web of Science ID 000174704000047

  View details for PubMedID 11927721

• Transcutaneous bilirubin measurements correlate well with serum total bilirubin levels in indigenous African infants Slusher, T. M., Angyo, I. A., Bode-Thomas, F., Akor, F., Adetunji, A. A., McLaren, D. W., Wong, R. J., Vreman, H. J. NATURE PUBLISHING GROUP. 2002: 329A–329A

  View details for Web of Science ID 000174714602787

• Alternative metalloporphyrins for the treatment of neonatal jaundice. Journal of perinatology Vreman, H. J., Wong, R. J., Stevenson, D. K. 2001; 21: S108-13

  View details for PubMedID 11803430

• Prediction of hyperbilirubinemia in near-term and term infants PEDIATRICS Stevenson, D. K., Fanaroff, A. A., Maisels, M. J., Young, B. W., Wong, R. J., Vreman, H. J., MacMahon, J. R., Yeung, C. Y., Seidman, D. S., Gale, R., Oh, W., Bhutani, V. K., Johnson, L. H., Kaplan, M., Hammerman, C., Nakamura, H. 2001; 108 (1): 31-39

  Abstract

  The purpose of this study was to determine whether end-tidal carbon monoxide (CO) corrected for ambient CO (ETCOc), as a single measurement or in combination with serum total bilirubin (STB) measurements, can predict the development of hyperbilirubinemia during the first 7 days of life.From 9 multinational clinical sites, 1370 neonates completed this cohort study from February 20, 1998, through February 22, 1999. Measurements of both ETCOc and STB were performed at 30 +/- 6 hours of life; STB also was measured at 96 +/- 12 hours and subsequently following a flow diagram based on a table of hours of age-specific STB. An infant was defined as hyperbilirubinemic if the hours of age-specific STB was greater than or equal to the 95th percentile as defined by the table at any time during the study.A total of 120 (8.8%) of the enrolled infants became hyperbilirubinemic. Mean STB in breastfed infants was 8.92 +/- 4.37 mg/dL at 96 hours versus 7.63 +/- 3.58 mg/dL in those fed formula only. The mean ETCOc at 30 +/- 6 hours for the total population was 1.48 +/- 0.49 ppm, whereas those of nonhyperbilirubinemic and hyperbilirubinemic infants were 1.45 +/- 0.47 ppm and 1.81 +/- 0.59 ppm, respectively. Seventy-six percent (92 of 120) of hyperbilirubinemic infants had ETCOc greater than the population mean. An ETCOc greater than the population mean at 30 +/- 6 hours yielded a 13.0% positive predictive value (PPV) and a 95.8% negative predictive value (NPV) for STB >/=95th percentile. When infants with STB >95th percentile at <36 hours of age were excluded, the STB at 30 +/- 6 hours yielded a 16.7% PPV and a 98.1% NPV for STB >75th percentile. The combination of these 2 measurements at 30 +/- 6 hours (either ETCOc more than the population mean or STB >75th percentile) had a 6.4% PPV with a 99.0% NPV. Conclusions. This prospective cohort study supports previous observations that measuring STB before discharge may provide some assistance in predicting an infant's risk for developing hyperbilirubinemia. The addition of an ETCOc measurement provides insight into the processes that contribute to the condition but does not materially improve the predictive ability of an hours of age-specific STB in this study population. The combination of STB and ETCOc as early as 30 +/- 6 hours may identify infants with increased bilirubin production (eg, hemolysis) or decreased elimination (conjugation defects) as well as infants who require early follow-up after discharge for jaundice or other clinical problems such as late anemia. Depending on the incidence of hyperbilirubinemia within an institution, the criteria for decision making should vary according to its unique population.

  View details for Web of Science ID 000169571400025

  View details for PubMedID 11433051

• Carbon monoxide and bilirubin production in neonates SEMINARS IN PERINATOLOGY Stevenson, D. K., Vreman, H. J., Wong, R. J., Contag, C. H. 2001; 25 (2): 85-93

  Abstract

  Neonatal hyperbilirubinemia is a normal postnatal phenomenon resulting from a transitional imbalance between the production and elimination of bilirubin in the neonate. Bilirubin has been shown to be not only a potent antioxidant, but also toxic at excessive concentrations. As a result, the biology of bilirubin, its production, regulation, and measurements have been the focus of extensive studies. Bilirubin, carbon monoxide, and iron are derived from the degradation of heme, a ubiquitous two-step pathway catalyzed by the enzyme, heme oxygenase. It has been shown that these metabolically active products from the heme catabolic pathway may, in turn, influence many other biologic processes. This report provides a brief overview of these interrelationships in the hope that it may provide insight into the central role this pathway plays in the existence of most organisms.

  View details for PubMedID 11339670

• Exhaled carbon monoxide in asthma JOURNAL OF PEDIATRICS Vreman, H. J., Wong, R. J., Stevenson, D. K. 2000; 137 (6): 889-890

  View details for Web of Science ID 000165876300029

  View details for PubMedID 11113853

• Haem oxygenase activity in human umbilical cord and rat vascular tissues PLACENTA Vreman, H. J., Wong, R. J., Kim, E. C., Nabseth, D. C., Marks, G. S., Stevenson, D. K. 2000; 21 (4): 337-344

  Abstract

  Carbon monoxide (CO) has been shown to affect vascular tone in smooth muscle cells and thus, may regulate regional or systemic blood pressure as well as fetoplacental vascular tone and fetal blood delivery. To assess the potential of vascular tissue to produce CO, we determined haem oxygenase (HO) activity through in vitro quantitation of CO production with gas chromatography and its inhibition by 33-66 microm of chromium mesoporphyrin (CrMP) in homogenate preparations of rat aorta and vena cava and human umbilical cord tissues. We compared these results to HO activity in rat heart and liver. We also discuss normalization of HO activity on a per mg protein as well as per g fresh weight (FW) tissue basis. We found that both rat vascular tissue HO activities (per g FW) were equal, but greater than that of heart (x3) and less than that of liver (x0.2). For human cord tissues, HO activities of artery and vein were equal, but greater than that of Wharton's jelly. Also, HO activity in rat vascular tissues was 3x greater than that of the human cord tissues. HO activity was completely inhibited by CrMP in rat heart (90 per cent) and liver (96 per cent), but incompletely (50-66 per cent) in both rat and human vascular tissues. We established that it is unlikely that other non-haem CO-generating processes account for this unique insensitivity of HO to CrMP inhibition. In fact, high concentrations of other potent metalloporphyrin inhibitors affected vascular tissue HO even less. We found that the degree of in vitro HO inhibition appeared to be related to the concentration of haem in the reaction medium. We conclude that the presence of HO activity in cord tissues supports the possibility that CO plays a role in fetoplacental blood flow regulation.

  View details for Web of Science ID 000087666000006

  View details for PubMedID 10833368

• Carbon monoxide detection and biological investigations. Transactions of the American Clinical and Climatological Association Stevenson, D. K., Vreman, H. J., Wong, R. J., Dennery, P. A., Contag, C. H. 2000; 111: 61-75

  Abstract

  Even though the heme degradation pathway consists of only two reactions, it and its major enzyme (i.e. HO), nonetheless, impact other processes not only through the removal of excess heme, but also through the production of several metabolically active compounds. Thus CO and biliverdin along with reactive iron, Fe2, are the primordial products of this ancient, highly conserved reaction. That every component of the heme catabolic pathway is directly or indirectly related to other reactions involving oxygen or light is, perhaps, no accident of nature. That a fundamentally destructive event can be linked with a multiplicity of synthetic events and various biological effects, depending on the timing and location of the HO activity, is testament to the economy and the ultimate beauty of nature. Furthermore, the interaction of the heme catabolic pathway with that of the NOS system may lead to even more exciting avenues of research. It may be shown that the integrity of the heme catabolic pathway, which is ever present and plays a role in every tissue, is central to the existence of most complex organisms.

  View details for PubMedID 10881332

• Concentration of carbon monoxide (CO) in tissue. Wong, R. J., Vreman, H. J., Kadotani, T., Stevenson, D. K. B C DECKER INC. 2000: 23A–23A

  View details for Web of Science ID 000086346600131

• Validation of the natus CO-Stat (TM) end tidal breath analyzer in children and adults JOURNAL OF CLINICAL MONITORING AND COMPUTING Vreman, H. J., Wong, R. J., Harmatz, P., Fanaroff, A. A., Berman, B., Stevenson, D. K. 1999; 15 (7-8): 421-427

  Abstract

  The performance of a point-of-care, noninvasive end tidal breath carbon monoxide analyzer (CO-Stat End Tidal Breath Analyzer, Natus Medical Inc.) that also reports end tidal carbon dioxide (ETCO2) and respiratory rate (RR), was compared to established, marketed (predicate) devices in children (n = 39) and adults (n = 48) who are normal or at-risk of elevated CO excretion.Concentrations of end tidal breath CO (ETCO), room air CO, ETCO corrected for inhaled CO (ETCOc), ETCO2, and RR were measured with the CO-Stat analyzer and the data compared to those obtained from the same subjects using the Vitalograph BreathCO monitor (Vitalograph, Inc.) for ETCOc and the Pryon CO2 monitor (SC210 and SC300, Pryon Corp) for ETCO2 and RR. Adults and children were studied at three medical centers. The data were analyzed by paired t-tests and linear regression. Bias and imprecision between the CO-Stat analyzer and the predicate devices was calculated by the method of Bland and Altman.Paired t-tests, performed on the three parameters measured with the CO-Stat analyzer and predicate devices showed that only the ETCOc values in the adults and the ETCO2 values in the children were significantly different (lower, p < or = 0.0001, and higher, p < or = 0.0001, respectively). The mean bias and imprecision of the CO-Stat analyzer for adult ETCOc and children ETCO2 measurements were -0.9 +/- 1.2 ppm and 0.4 +/- 0.6%, respectively. Linear regression analysis for the ETCOc results in children and adults had a high degree of correlation (r = 0.91 and 0.98, respectively).We conclude that in a clinical environment the Natus CO-Stat End Tidal Breath Analyzer performs at least as well as predicate devices for the measurements of ETCOc, ETCO2, and RR.

  View details for Web of Science ID 000086962600004

  View details for PubMedID 12578038

• Concentration of carbon monoxide (CO) in tissue Kadotani, T., Vreman, H. J., Wong, R. J., Stevenson, D. K. NATURE PUBLISHING GROUP. 1999: 67A–67A

  View details for Web of Science ID 000079476700389

• In vitro bilirubin degradation by a light emitting diode (LED) device. Vreman, H. J., Stevenson, D. K., Wong, R. J., Reader, S. D., Route, R. K., Fejer, M. M., Gale, R., Seidman, D. S. LIPPINCOTT WILLIAMS & WILKINS. 1998: 157A–157A

  View details for Web of Science ID 000071684700840

• Evaluation of the Ciba Corning 840 blood gas analyzer. Respiratory care Wong, R. J., Mahoney, J. J., van Kessel, A. L. 1995; 40 (6): 638-643

  Abstract

  Because our laboratory had used the Ciba Corning 200 series blood-gas analyzers for a number of years, we were asked to participate in the evaluation of a premarket unit of the Model 840 analyzer (C840). DESCRIPTION OF DEVICE: The C840 is a bench-top instrument that combines a menu-driven user interface with an automated sampler and integrates software for data management and system diagnostics.We compared the performance of the C840 to a laboratory-based Ciba Corning 278 (C278), analyzing a total of 325 blood samples. We also evaluated the software for routine laboratory applications. EVALUATION RESULTS: The bias and imprecision (+/- 2 SD) between the C840 and C278 was calculated for pH (+0.004 +/- 0.014 pH units), PCO2 (+1.8 +/- 3.3 torr), and PO2 (+0.01 +/- 9.0 torr for all PO2 ranges; -0.17 +/- 4.8 torr for PO2 < 150 torr).We conclude that the analytical performance of the C840 is comparable to the C278, and its data storage and interface capabilities should help laboratories meet CLIA-88 requirements.

  View details for PubMedID 10143509

• STATPAL(R) II PH AND BLOOD-GAS ANALYSIS SYSTEM EVALUATED CLINICAL CHEMISTRY Wong, R. J., Mahoney, J. J., Harvey, J. A., VanKessel, A. L. 1994; 40 (1): 124-129

  Abstract

  We evaluated a new portable instrument, the PPG StatPal II pH and Blood Gas Analysis System, designed for "point-of-care" measurements of blood gases and pH. Inaccuracy (% of target value) and imprecision (CV%) were assessed by blood tonometry and comparison with a Corning 178. Within-day results for PCO2 inaccuracy and imprecision ranged from 98.2% to 102.9% and 3.3% to 3.9%, respectively; for PO2, these were 95.5% to 102.3% and 2.3% to 3.0%, respectively. Between-day results for PCO2 inaccuracy and imprecision ranged from 99.2% to 99.3% and from 2.9% to 3.2%, respectively; for PO2, the ranges were 96.2% to 98.2% and 2.6% to 3.0%, respectively. Two PCO2 outliers (in 645 samples = 0.3%) were observed. In general, tonometry recovery, measurement stability, and pH bias results for the StatPal II and Corning 178 were comparable. We conclude that the StatPal II performs within acceptable ranges of inaccuracy and imprecision.

  View details for Web of Science ID A1994NA65900023

  View details for PubMedID 8287519

• REDUCED BOVINE HEMOGLOBIN SOLUTION EVALUATED FOR USE AS A BLOOD-GAS QUALITY-CONTROL MATERIAL CLINICAL CHEMISTRY Mahoney, J. J., Wong, R. J., VanKessel, A. L. 1993; 39 (5): 874-879

  Abstract

  We evaluated a new commercially available partially purified reduced bovine hemoglobin solution (RBHS) and a new tonometer for use in the quality control of blood gas analyzers. RBHS is manufactured in three different formulations, each corresponding to three different bicarbonate-buffering capacities and concentrations of total hemoglobin (ctHb). The P50 for each formulation of RBHS was determined to be as follows: 30.6 mmHg (4.08 kPa) for Level 1, 29.1 mmHg (3.88 kPa) for Level 2, and 28.2 mmHg (3.76 kPa) for Level 3. When RBHS and human blood samples were tonometered at three clinically significant values of PO2 and PCO2, the recovered values for PO2 and PCO2 in RBHS were comparable with those of the tonometered whole blood. Each level of RBHS also produced precise pH (SD < or = 0.006 pH units) and ctHb (SD = 1.0 g/L) values. In addition, when the temperature of a sample chamber was intentionally altered, the changes in RBHS blood gas values closely approximated the changes seen with human blood. RBHS shows more thermal sensitivity than either perfluorocarbon emulsions or aqueous buffers, which are currently being used as quality-control and proficiency testing materials.

  View details for Web of Science ID A1993LB54600023

  View details for PubMedID 8485880

• CHANGES IN OXYGEN MEASUREMENTS WHEN WHOLE-BLOOD IS STORED IN ICED PLASTIC OR GLASS SYRINGES CLINICAL CHEMISTRY Mahoney, J. J., Harvey, J. A., Wong, R. J., VanKessel, A. L. 1991; 37 (7): 1244-1248

  Abstract

  Tonometered whole-blood and plasma specimens were tested in plastic and glass syringes to determine whether clinically significant changes in gas tensions occur during sample storage. When whole blood was tonometered with 60 and 100 mL/L (6% and 10%) oxygen and then stored for 30 min in iced plastic syringes, the pO2 of the samples remained stable (mean change = +0.4 and +0.8 mmHg, respectively). However, for 140 mL/L (14%) oxygen tonometry, the pO2 increased significantly (mean change = +8.4 mmHg; P less than 0.0001). When tonometered plasma was stored in iced plastic syringes, the pO2 increased progressively at all three concentrations, with the smallest change occurring at 140 mL/L (mean change = +12.6 mmHg) and the greatest at 60 mL/L oxygen (mean change = +20.9 mmHg). In contrast, when iced glass syringes were used for storing plasma or whole blood, no clinically significant changes in pO2 were found at any of the tonometered oxygen values for 60 min. When whole blood was stored in plastic syringes at ambient temperature for 30 min, again no clinically significant changes in pO2 were found at these tonometry conditions. Apparently, some blood gas samples stored in iced plastic syringes may yield clinically significant errors in oxygen tension.

  View details for Web of Science ID A1991FX18200018

  View details for PubMedID 1823532

• FETAL HEMOGLOBIN OF TRANSFUSED NEONATES AND SPECTROPHOTOMETRIC MEASUREMENTS OF OXYHEMOGLOBIN AND CARBOXYHEMOGLOBIN JOURNAL OF CLINICAL MONITORING Mahoney, J. J., Wong, R. J., Vreman, H. J., Stevenson, D. K. 1991; 7 (2): 154-160

  Abstract

  The records of 32 neonates in an intensive care unit were examined retrospectively to determine if fetal hemoglobin concentrations could be predicted on the basis of gestational or postnatal age, or on the volume of red blood cell transfusions. In nontransfused neonates, the correlation between measured concentrations of fetal hemoglobin and post-natal age was r = 0.53 with a 17.2 standard error of prediction. In these same neonates, the correlation between measured fetal hemoglobin divided by birth weight and gestational age was r = 0.70, with a 9.6 standard error of prediction. A three-variable regression equation (the latter two variables plus calculated fetal hemoglobin) was found to have a high correlation with data for measured fetal hemoglobin (r = 0.97) and a relatively low 8.4 standard error of prediction. In transfused neonates, however, measured hemoglobin concentrations divided by birth weight correlated poorly with gestational age (r = 0.30 and a 12.4 standard error of prediction). In addition, the transfused neonates had low correlations when fetal hemoglobin concentrations alone were compared with the total volume of red blood cell transfusions (r = 0.35) and with postnatal age (r = 0.18) and the standard errors of prediction were all approximately 17. The correlations found between concentrations of fetal hemoglobin and age in transfused neonates were poorer than those reported in earlier nontransfused infant studies. Previous studies have also shown that neonatal blood containing fetal hemoglobin interferes with the spectrophotometric measurements of carboxyhemoglobin and oxyhemoglobin. Because of the imprecision in the predictions of fetal hemoglobin using age, weight, or the volume of transfusion, we conclude that fetal hemoglobin should be measured if accurate spectrophotometric determinations of carboxyhemoglobin and oxyhemoglobin are desired.

  View details for Web of Science ID A1991FH21600004

  View details for PubMedID 1712833

• INCREASED LUNG WATER AND ASCITES AFTER MASSIVE COCAINE OVERDOSAGE IN MICE AND IMPROVED SURVIVAL RELATED TO BETA-ADRENERGIC BLOCKAGE ANNALS OF INTERNAL MEDICINE Robin, E. D., Wong, R. J., PTASHNE, K. A. 1989; 110 (3): 202-207

  Abstract

  To determine the effect of massive cocaine intoxication on lung water and ascites accumulation and the effect of beta- and alpha-adrenergic blockade on survival in massive cocaine intoxication in the mouse.The effect of massive cocaine intoxication on lung water, ascitic fluid accumulation, and survival following LD 100 doses of intravenous cocaine with and without alpha- and beta-adrenergic blockade was determined.Cocaine hydrochloride (0.15 mg/g body weight) was administered intravenously with no other interventions; with propranolol hydrochloride intravenously (0.5 mg per mouse) before and after cocaine; and with phentolamine intravenously (10.5 micrograms per mouse) before cocaine.Intravenous cocaine hydrochloride resulted in an increase in lung water (saline controls, 4.17 +/- 1.3 [standard deviation] mg water per g mouse; cocaine hydrochloride, 5.94 +/- 0.9 mg water per g mouse; P less than 0.002). Cocaine hydrochloride always resulted in the accumulation of transudative ascitic fluid (saline controls, no measurable ascitic fluid; cocaine administration, 20.2 +/- 12.9 micrograms per mouse; ascitic fluid protein concentration, 23.5 +/- 8.5 g/L). Propranolol hydrochloride administered before or after intravenous cocaine hydrochloride resulted in a striking reduction in mortality (84 of 84 mice without propranolol died [mortality = 100%]; 7 of 39 mice with propranolol died [mortality = 18%]; P less than 0.001).Massive cocaine intoxication is associated with increased lung water and transudative ascites. Fluid accumulation is not prevented by either alpha- or beta-adrenergic blockers. Propranolol, administered either before or after cocaine, sharply reduces mortality. The results should be extrapolated to treatment in humans with caution.

  View details for Web of Science ID A1989T106000006

  View details for PubMedID 2912360

• MITOCHONDRIAL-DNA MOLECULES AND VIRTUAL NUMBER OF MITOCHONDRIA PER CELL IN MAMMALIAN-CELLS JOURNAL OF CELLULAR PHYSIOLOGY Robin, E. D., Wong, R. 1988; 136 (3): 507-513

  Abstract

  A new biochemical method for estimating the virtual number of mitochondria (mt) per cell was developed and used together with a plasmid probe to measure mt DNA/mitochondrion and mt DNA/cell. These methods were used in five cell types from four mammalian species. Mt DNA/mitochondrion was essentially constant in all cell types (mean 2.6 +/- 0.30 SE mitochondrial DNA molecules/mt). Mt DNA molecules/cell encompassed an eight-fold range between various cell types (low 220 +/- 6.2; high 1,720 +/- 162 mt DNA molecules/cell). Virtual mt number/cell ranged from 83 +/- 17 to 677 +/- 80 (SE) mt/cell in various cell types. All five mammalian virtual mitochondria contained the same genomic mass. The number of virtual mitochondria per cell and amount of mt DNA per cell appear to be closely regulated within a given cell type but differ widely from cell type to cell type.

  View details for Web of Science ID A1988Q306400015

  View details for PubMedID 3170646

• ADAPTATIONS OF NEUROTRANSMITTER SYNTHESIS TO CHRONIC HYPOXIA IN CELL-CULTURE BIOCHIMICA ET BIOPHYSICA ACTA Feinsilver, S. H., Wong, R., RAYBIN, D. M. 1987; 928 (1): 56-62

  Abstract

  Tyrosine hydroxylase and tryptophan hydroxylase are widely held to be rate-limiting for the synthesis of the catecholamines and serotonin, respectively. Both enzymes are oxygen-requiring and kinetic properties suggest that oxygen availability may limit synthesis of these neurotransmitters in the brain. Using pheochromocytoma cells as a cell culture model for catecholamine synthesis, and neuroblastoma cells as a model for serotonin synthesis, enzyme activity was measured under control and hypoxic conditions. Both tyrosine hydroxylase and tryptophan hydroxylase activity increased substantially with chronic exposure but not with acute exposure. In the case of tyrosine hydroxylase, increased enzyme content with hypoxia accounts for increased activity. This suggests a mechanism for the maintenance of neurotransmitter synthesis with chronic hypoxia. Measurement of intracellular metabolites revealed no change in dopamine or norepinephrine in hypoxic pheochromocytoma cells, consistent with a simple adaptive mechanism. However, in neuroblastoma cells, hypoxia was associated with an increase in serotonin concentration. The reasons for this are still unclear.

  View details for Web of Science ID A1987G860200008

  View details for PubMedID 2881582

• NEUROGENIC HYPERACUTE ASCITES IN MICE CLINICAL SCIENCE Nelson, D. P., Robin, E. D., Wong, R. J., MORIN, M. E., Bensch, K. G., MURPHY, B. J., Theodore, J. 1986; 71 (3): 327-330

  Abstract

  Four groups of mice were subjected to controlled fatal head trauma and then evaluated for the presence of ascites (neurogenic hyperacute ascites, NHA). The animals died virtually instantaneously and without evidence of maintained pain or suffering. The volume of ascites was determined in one group of animals. Two of the traumatized groups were pretreated, one with the beta-blocker propranolol and the other with the alpha-blocker phentolamine. A fifth, non-traumatized, group which was killed with either inhalation served as a control group. Two more groups of non-traumatized mice were administered either the alpha-adrenergic agonist methoxamine hydrochloride or the beta-agonist isoethrane mesylate before killing by ether inhalation, and then evaluated for ascites. Transudative ascitic fluid was found in 87-100% of untreated traumatized mice and in no control animals. Pretreatment with phentolamine had no effect on the prevalence of ascites. Pretreatment with propranolol produced a significant decrease in the prevalence of ascites compared with trauma alone (P less than 0.001). Isoethrane (beta-agonist) administration caused ascites in 100% of the treated animals. Methoxamine (alpha-agonist) administration did not cause ascites. A previously undescribed consequence of acute brain trauma is described (NHA) which appears to be mediated by beta-sympathetic activity of central origin. NHA is inhibited by beta-blockade and can be simulated with beta-agonist administration.

  View details for Web of Science ID A1986D509900016

  View details for PubMedID 2875822

• HYPOXIC COORDINATE REGULATION OF MITOCHONDRIAL-ENZYMES IN MAMMALIAN-CELLS SCIENCE MURPHY, B. J., Robin, E. D., TAPPER, D. P., Wong, R. J., Clayton, D. A. 1984; 223 (4637): 707-709

  Abstract

  The effect of hypoxic exposure on various mitochondrial enzymes and on cell mitochondrial genomic content was studied in two types of mammalian cells. Hypoxia depressed the activity of six enzymes to the same degree. The kinetics of depression and of recovery during reexposure to normoxia were statistically similar for three marker enzymes. Despite the global and symmetrical decrease in enzyme activities, mitochondrial DNA remained constant. This suggests either symmetrical loss of mitochondrial enzymes from all mitochondria or complete loss of enzymes from a subpopulation of mitochondria with retention of an intact mitochondrial genome.

  View details for Web of Science ID A1984SC10900028

  View details for PubMedID 6320368