Clinical Focus

  • Primary Immunodeficiency
  • Pediatrics

Administrative Appointments

  • Acting Associate Professor, Stanford University, Department of Pediatrics (1997 - 1997)
  • Associate Professor, Stanford University, Department of Pediatrics (1997 - 2005)
  • Director of the Jeffrey Modell Primary Immunodeficiency Center at Stanford, Supported by the Jeffrey Modell Foundation and the Lucile Packard Fund of Children's Health (2002 - Present)
  • Professor, Stanford University, Department of Pediatrics (2005 - Present)
  • Chief, Stanford Department of Pediatrics, Division of Immunology and Allergy (2008 - Present)

Honors & Awards

  • Henry J. Kaiser Award for Excellence in Preclinical Teaching (Immunology), Kaiser Foundation (2001)
  • Henry J. Kaiser Award for Excellence in Preclinical Teaching (Immunology), Kaiser Foundation (2005)

Professional Education

  • Fellowship: Seattle Children's Hospital Pediatric Infectious Diseases Fellowship (1987) WA
  • Residency: UCSF Pediatric Residency (1984) CA
  • Internship: Seattle Childrens Hospital Pediatric Residency (1982) WA
  • Medical Education: University of California at San Francisco School of Medicine (1981) CA
  • Board Certification: American Board of Pediatrics, Pediatrics (1986)
  • M.D., Univ.Calif at San Francisco, Medicine (1981)
  • B.S., Yale University, Biology (1976)

Current Research and Scholarly Interests

A long-standing interest is to understand the cellular and molecular basis for this vulnerability of the human neonate to infection with intracellular pathogens that require T helper 1 (Th1) cells [CD4 T cell producing interferon-gamma (IFN-gamma)] for effective immune control. We have previously shown that CD4 T cells of the newborn have a unique limitation in the ability to produce certain effector molecules, such as CD40-ligand (CD154) and IFN-gamma compared to these cells in adults due to both reduced gene transcriptional and impaired signals that lead to gene transcription. Recently, we have shown that these limitations apply to physiological T-cell activation, e.g., using allogeneic dendritic cells. Defining the molecular mechanisms for decreased IFN-gamma production by neonatal CD4 T cells is a current focus.
We have also found that recent thymic emigrants, which predominate in the newborn and young infant, are less able to differentiate into T helper 1 cells, which produce IFN-gamma. These studies required the development of a novel marker for recent thymic emigrants (RTEs) of the CD4 T-cell lineage in humans. Using a combination of approaches, we have identified protein tyrosine kinase 7 (PTK7) as such a marker. In progress are to studies to define the role of PTK7, an orphan member (no known ligand) of the receptor tyrosine kinase family, in T-cell development and immunity, and to determine how this marker can be used to follow the output of recent thymic emigrants in health and disease. We are also interesed in determining the molecular mechanisms for the reduced RTE function and to what extent these mechanisms are shared by neonatal CD4 T cells and CD4+CD8-CD3+ thymocytes, the immediate precursors of antigenically naive CD4 T cells.

We have also found that limitations in T-cell immunogenicity to viruses and viral vaccines extend beyond the neonatal period to childhood. These studies highlight a need to develop more potent vaccines to overcome developmental and other factors, such as genetic inheritance, in mounting adaptive immunity. With this as an ultimate goal, we have previously examined the ability of a novel adjuvant, cationic liposome DNA complexes (CLDC)(Juvaris Biotherapeutics), to induce durable CD4 and CD8 T-cell immunity and humoral immunity to influenza A. The molecular and cellular components of the innate immune system that are required for immunogenicity are of particular interest. We are currently embarking on studies of vaccine immunity using novel influenza A virus antigens produced by our collaborators at Sutrovax. We are also beginning studies to determine if universal anti-influenza A viral protection can be achieved using catalytically inactive Cas proteins combined with appropriate guide RNAs. This work is being carried out in collaboration with Stanley Qi's laboratory at Stanford.

As part of an on-going collaboration with Dr. Neal Boerkoel, University of British Columbia, we are defining the mechanism of T-cell lymphopenia in genetic deficiency of SMARCAL1, a protein that plays a novel role in relieving stalled DNA replication forks. Patients with SMARCAL1 deficiency (Schimke immuno-osseous dysplasia) suffer from not only T-cell immunodeficiency but also progressive renal dysfunction due to collapsing variant focal glomerulosclerosis, short stature due to growth place abnormalities, and vascular disease, with an increased risk of TIAs and strokes. How a block in DNA repair selective influences these disease programs is unclear and is a major current focus of research. This research is being supported in part by funds from the Kruzn' for a Kure Foundation, a philanthropic foundation that was started by parents of two children with SIOD.

2023-24 Courses

Stanford Advisees

Graduate and Fellowship Programs

All Publications

  • Single-cell Transcriptomics Reveal Different Maturation Stages and Sublineages Commitment of Human Thymic Invariant Natural Killer T cells. Journal of leukocyte biology Maas-Bauer, K., Köhler, N., Stell, A. V., Zwick, M., Acharya, S., Rensing-Ehl, A., König, C., Kroll, J., Baker, J., Koßmann, S., Pradier, A., Wang, S., Docquier, M., Lewis, D. B., Negrin, R. S., Simonetta, F. 2023


    Invariant natural killer T (iNKT) cells are a rare, heterogeneous T-cell subset with cytotoxic and immunomodulatory properties. During thymic development, murine iNKT cells go through different maturation stages differentiating into distinct sublineages, namely iNKT1, iNKT2, and iNKT17 cells. Recent reports indicate that iNKT2 cells display immature properties and give rise to other subsets, whereas iNKT1 cells seem to be terminally differentiated. Whether human iNKT cells follow a similar differentiation model is still unknown. To define the maturation stages and assess the sublineage commitment of human iNKT cells during thymic development, in this study we performed scRNAseq analysis on human Vα24+ Vβ11 + iNKT cells isolated from thymocytes. We show that these iNKT cells displayed heterogeneity and our unsupervised analysis identified five clusters representing different maturation stages, from an immature profile with high expression of genes important for iNKT cell development and proliferation to a mature, fully differentiated profile with high levels of cytotoxic effector molecules. Evaluation of expression of sublineage-defining gene sets revealed mainly cells with an iNKT2 signature in the most immature cluster, whereas the more differentiated ones displayed an iNKT1 signature. Combined analysis with a publicly available scRNAseq dataset of human iNKT cells from peripheral blood suggested that the two main subsets exist both in thymus and in the periphery, while a third more immature one was restricted to the thymus. Our data point to the existence of different maturation stages of human thymic iNKT cells and provide evidence for sublineage commitment of iNKT cells in the human thymus.

    View details for DOI 10.1093/jleuko/qiad113

    View details for PubMedID 37742056

  • Sequential Stem Cell-Kidney Transplantation in Schimke Immuno-osseous Dysplasia. The New England journal of medicine Bertaina, A., Grimm, P. C., Weinberg, K., Parkman, R., Kristovich, K. M., Barbarito, G., Lippner, E., Dhamdhere, G., Ramachandran, V., Spatz, J. M., Fathallah-Shaykh, S., Atkinson, T. P., Al-Uzri, A., Aubert, G., van der Elst, K., Green, S. G., Agarwal, R., Slepicka, P. F., Shah, A. J., Roncarolo, M. G., Gallo, A., Concepcion, W., Lewis, D. B. 2022; 386 (24): 2295-2302


    Lifelong immunosuppression is required for allograft survival after kidney transplantation but may not ultimately prevent allograft loss resulting from chronic rejection. We developed an approach that attempts to abrogate immune rejection and the need for post-transplantation immunosuppression in three patients with Schimke immuno-osseous dysplasia who had both T-cell immunodeficiency and renal failure. Each patient received sequential transplants of alphabeta T-cell-depleted and CD19 B-cell-depleted haploidentical hematopoietic stem cells and a kidney from the same donor. Full donor hematopoietic chimerism and functional ex vivo T-cell tolerance was achieved, and the patients continued to have normal renal function without immunosuppression at 22 to 34 months after kidney transplantation. (Funded by the Kruzn for a Kure Foundation.).

    View details for DOI 10.1056/NEJMoa2117028

    View details for PubMedID 35704481

  • Broad-spectrum CRISPR-mediated inhibition of SARS-CoV-2 variants and endemic coronaviruses in vitro. Nature communications Zeng, L., Liu, Y., Nguyenla, X. H., Abbott, T. R., Han, M., Zhu, Y., Chemparathy, A., Lin, X., Chen, X., Wang, H., Rane, D. A., Spatz, J. M., Jain, S., Rustagi, A., Pinsky, B., Zepeda, A. E., Kadina, A. P., Walker, J. A., Holden, K., Temperton, N., Cochran, J. R., Barron, A. E., Connolly, M. D., Blish, C. A., Lewis, D. B., Stanley, S. A., La Russa, M. F., Qi, L. S. 2022; 13 (1): 2766


    A major challenge in coronavirus vaccination and treatment is to counteract rapid viral evolution and mutations. Here we demonstrate that CRISPR-Cas13d offers a broad-spectrum antiviral (BSA) to inhibit many SARS-CoV-2 variants and diverse human coronavirus strains with >99% reduction of the viral titer. We show that Cas13d-mediated coronavirus inhibition is dependent on the crRNA cellular spatial colocalization with Cas13d and target viral RNA. Cas13d can significantly enhance the therapeutic effects of diverse small molecule drugs against coronaviruses for prophylaxis or treatment purposes, and the best combination reduced viral titer by over four orders of magnitude. Using lipid nanoparticle-mediated RNA delivery, we demonstrate that the Cas13d system can effectively treat infection from multiple variants of coronavirus, including Omicron SARS-CoV-2, in human primary airway epithelium air-liquid interface (ALI) cultures. Our study establishes CRISPR-Cas13 as a BSA which is highly complementary to existing vaccination and antiviral treatment strategies.

    View details for DOI 10.1038/s41467-022-30546-7

    View details for PubMedID 35589813

  • Immunogenicity of a Third COVID-19 mRNA Vaccine Dose in PID Patients with Functional B-cell Defects. The journal of allergy and clinical immunology. In practice Gernez, Y., Murugesan, K., Cortales, C. R., Banaei, N., Hoyte, L., Pinsky, B. A., Lewis, D. B., Pham, M. 2022

    View details for DOI 10.1016/j.jaip.2022.02.030

    View details for PubMedID 35259538

  • Increased double-negative alpha beta plus T cells reveal adult-onset autoimmune lymphoproliferative syndrome in a patient with IgG4-related disease HAEMATOLOGICA Brar, N., Spinner, M. A., Baker, M. C., Advani, R. H., Natkunam, Y., Lewis, D. B., Silva, O. 2022; 107 (1): 347-350
  • Immunogenicity and Tolerability of COVID-19 mRNA Vaccines in PID patients with functional B-cell defects. The Journal of allergy and clinical immunology Pham, M. N., Murugesan, K., Banaei, N., Pinsky, B. A., Tang, M., Hoyte, E., Lewis, D. B., Gernez, Y. 1800


    BACKGROUND: Data on the safety and efficacy of COVID-19 vaccination in people with a range of primary immune deficiencies are lacking since these patients were excluded from COVID-19 vaccine trials. This information may help in clinical management of this vulnerable patient group.OBJECTIVE: To assess humoral and T-cell immune responses after two doses of SARS-CoV-2 mRNA vaccines in patients with PIDs and functional B-cell defects.METHODS: A double-center retrospective review of patients with PID who completed COVID-19 mRNA vaccination and who had humoral responses assessed through SARS-CoV-2 spike protein receptor binding domain (RBD) IgG antibody levels with reflex assessment of the antibody to block RBD binding to ACE2 (hereafter referred to as ACE2 receptor blocking activity, as a surrogate test for neutralization) and T-cell response evaluated by an interferon-gamma release assay (IGRA). Immunization reactogenicity was also reviewed.RESULTS: A total of 33 patients with humoral defect were evaluated. 69.6% received BNT162b2 vaccine (Pfizer-BioNTech) and 30.3% received mRNA-1273 (Moderna). The mRNA vaccines were generally well tolerated without severe reactions. The IGRA was positive in 77.4% of our patients (24 of 31). About half of our subjects (16 of 33) had detectable RBD-specific IgG responses but only two of these 16 subjects had an ACE2 receptor blocking activity level of >50%.CONCLUSION: Vaccination of this cohort of PID patients with COVID-19 mRNA vaccines was safe and cellular immunity was stimulated in a majority. However, antibody responses to the spike protein RBD were less consistent, and, when detected, was not effective at ACE2 blocking.CLINICAL IMPLICATION: mRNA vaccination may be less effective at preventing acquisition of SARS-CoV-2 in our cohort of PID patients with functional B-cell defects. The Induction of SARS-CoV-2 spike protein-specific T-cell immunity by vaccination might help reduce the severity of disease in these patients.

    View details for DOI 10.1016/j.jaci.2021.11.022

    View details for PubMedID 34952033

  • Single-Cell Analysis of the Neonatal Immune System Across the Gestational Age Continuum. Frontiers in immunology Peterson, L. S., Hedou, J., Ganio, E. A., Stelzer, I. A., Feyaerts, D., Harbert, E., Adusumelli, Y., Ando, K., Tsai, E. S., Tsai, A. S., Han, X., Ringle, M., Houghteling, P., Reiss, J. D., Lewis, D. B., Winn, V. D., Angst, M. S., Aghaeepour, N., Stevenson, D. K., Gaudilliere, B. 2021; 12: 714090


    Although most causes of death and morbidity in premature infants are related to immune maladaptation, the premature immune system remains poorly understood. We provide a comprehensive single-cell depiction of the neonatal immune system at birth across the spectrum of viable gestational age (GA), ranging from 25 weeks to term. A mass cytometry immunoassay interrogated all major immune cell subsets, including signaling activity and responsiveness to stimulation. An elastic net model described the relationship between GA and immunome (R=0.85, p=8.75e-14), and unsupervised clustering highlighted previously unrecognized GA-dependent immune dynamics, including decreasing basal MAP-kinase/NFκB signaling in antigen presenting cells; increasing responsiveness of cytotoxic lymphocytes to interferon-α; and decreasing frequency of regulatory and invariant T cells, including NKT-like cells and CD8+CD161+ T cells. Knowledge gained from the analysis of the neonatal immune landscape across GA provides a mechanistic framework to understand the unique susceptibility of preterm infants to both hyper-inflammatory diseases and infections.

    View details for DOI 10.3389/fimmu.2021.714090

    View details for PubMedID 34497610

    View details for PubMedCentralID PMC8420969

  • Increased double-negative αβ+ T-cells reveal adult-onset autoimmune lymphoproliferative syndrome in a patient with IgG4-related disease. Haematologica Brar, N., Spinner, M. A., Baker, M. C., Advani, R. H., Natkunam, Y., Lewis, D. B., Silva, O. 2021


    Not available.

    View details for DOI 10.3324/haematol.2021.278772

    View details for PubMedID 34474549

  • Disseminated Pneumocystis jirovecii Infection with Osteomyelitis in a Patient with CTLA-4 Haploinsufficiency. Journal of clinical immunology Siddiqi, A. E., Liu, A. Y., Charville, G. W., Kunder, C. A., Uzel, G. n., Sadighi Akha, A. A., Oak, J. n., Martin, B. n., Sacha, J. n., Lewis, D. B., Gernez, Y. n. 2020

    View details for DOI 10.1007/s10875-020-00748-z

    View details for PubMedID 31955317

  • Development of CRISPR as an Antiviral Strategy to Combat SARS-CoV-2 and Influenza. Cell Abbott, T. R., Dhamdhere, G. n., Liu, Y. n., Lin, X. n., Goudy, L. n., Zeng, L. n., Chemparathy, A. n., Chmura, S. n., Heaton, N. S., Debs, R. n., Pande, T. n., Endy, D. n., La Russa, M. F., Lewis, D. B., Qi, L. S. 2020


    The coronavirus disease 2019 (COVID-19) pandemic, caused by the SARS-CoV-2 virus, has highlighted the need for antiviral approaches that can target emerging viruses with no effective vaccines or pharmaceuticals. Here, we demonstrate a CRISPR-Cas13-based strategy, PAC-MAN (prophylactic antiviral CRISPR in human cells), for viral inhibition that can effectively degrade RNA from SARS-CoV-2 sequences and live influenza A virus (IAV) in human lung epithelial cells. We designed and screened CRISPR RNAs (crRNAs) targeting conserved viral regions and identified functional crRNAs targeting SARS-CoV-2. This approach effectively reduced H1N1 IAV load in respiratory epithelial cells. Our bioinformatic analysis showed that a group of only six crRNAs can target more than 90% of all coronaviruses. With the development of a safe and effective system for respiratory tract delivery, PAC-MAN has the potential to become an important pan-coronavirus inhibition strategy.

    View details for DOI 10.1016/j.cell.2020.04.020

    View details for PubMedID 32353252

  • COVID-19 and cardiovascular disease: from basic mechanisms to clinical perspectives. Nature reviews. Cardiology Nishiga, M. n., Wang, D. W., Han, Y. n., Lewis, D. B., Wu, J. C. 2020


    Coronavirus disease 2019 (COVID-19), caused by a strain of coronavirus known as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a global pandemic that has affected the lives of billions of individuals. Extensive studies have revealed that SARS-CoV-2 shares many biological features with SARS-CoV, the zoonotic virus that caused the 2002 outbreak of severe acute respiratory syndrome, including the system of cell entry, which is triggered by binding of the viral spike protein to angiotensin-converting enzyme 2. Clinical studies have also reported an association between COVID-19 and cardiovascular disease. Pre-existing cardiovascular disease seems to be linked with worse outcomes and increased risk of death in patients with COVID-19, whereas COVID-19 itself can also induce myocardial injury, arrhythmia, acute coronary syndrome and venous thromboembolism. Potential drug-disease interactions affecting patients with COVID-19 and comorbid cardiovascular diseases are also becoming a serious concern. In this Review, we summarize the current understanding of COVID-19 from basic mechanisms to clinical perspectives, focusing on the interaction between COVID-19 and the cardiovascular system. By combining our knowledge of the biological features of the virus with clinical findings, we can improve our understanding of the potential mechanisms underlying COVID-19, paving the way towards the development of preventative and therapeutic solutions.

    View details for DOI 10.1038/s41569-020-0413-9

    View details for PubMedID 32690910

  • Severe autoinflammation in four patients with C-terminal variants in CDC42 successfully treated with IL-1beta inhibition. The Journal of allergy and clinical immunology Gernez, Y., de Jesus, A. A., Alsaleem, H., Macaubas, C., Roy, A., Lovell, D., Jagadeesh, K. A., Alehashemi, S., Erdman, L., Grimley, M., Talarico, S., Bacchetta, R., Lewis, D. B., Canna, S. W., Laxer, R. M., Mellins, E. D., Goldbach-Mansky, R., Weinacht, K. G. 2019


    Four patients with novel variants in the C-terminal domain of CDC42, severe autoinflammation, constitutive elevation of serum interleukin 18, and predisposition to macrophage activation syndrome respond to treatment with interleukin-1beta signaling inhibition.

    View details for DOI 10.1016/j.jaci.2019.06.017

    View details for PubMedID 31271789

  • 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


    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: information: Supplementary data are available at Bioinformatics online.

    View details for PubMedID 30561547

  • 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
  • Durable multitransgene expression in vivo using systemic, nonviral DNA delivery. Science advances Handumrongkul, C. n., Ye, A. L., Chmura, S. A., Soroceanu, L. n., Mack, M. n., Ice, R. J., Thistle, R. n., Myers, M. n., Ursu, S. J., Liu, Y. n., Kashani-Sabet, M. n., Heath, T. D., Liggitt, D. n., Lewis, D. B., Debs, R. n. 2019; 5 (11): eaax0217


    Recombinant adeno-associated virus (AAV) vectors are transforming therapies for rare human monogenic deficiency diseases. However, adaptive immune responses to AAV and its limited DNA insert capacity, restrict their therapeutic potential. HEDGES (high-level extended duration gene expression system), a nonviral DNA- and liposome-based gene delivery platform, overcomes these limitations in immunocompetent mice. Specifically, one systemic HEDGES injection durably produces therapeutic levels of transgene-encoded human proteins, including FDA-approved cytokines and monoclonal antibodies, without detectable integration into genomic DNA. HEDGES also controls protein production duration from <3 weeks to >1.5 years, does not induce anti-vector immune responses, is reexpressed for prolonged periods following reinjection, and produces only transient minimal toxicity. HEDGES can produce extended therapeutic levels of multiple transgene-encoded therapeutic human proteins from DNA inserts >1.5-fold larger than AAV-based therapeutics, thus creating combinatorial interventions to effectively treat common polygenic diseases driven by multigenic abnormalities.

    View details for DOI 10.1126/sciadv.aax0217

    View details for PubMedID 31807699

    View details for PubMedCentralID PMC6881169

  • Paraneoplastic and Therapy-Related Immune Complications in Thymic Malignancies. Current treatment options in oncology Lippner, E. A., Lewis, D. B., Robinson, W. H., Katsumoto, T. R. 2019; 20 (7): 62


    The thymus is a key organ involved in establishing central immune tolerance. Thymic epithelial tumors (TETs) include thymomas and thymic carcinomas. Thymomas, which are histologically distinct from thymic carcinomas, lead to dysregulated thymopoiesis via decreased thymic epithelial expression of AIRE and MHC Class II, as well as via alterations in thymic architecture, thereby resulting in autoimmune complications that manifest as paraneoplastic disorders (PNDs). Although progress has been made in elucidating the mechanisms underlying thymoma-associated PNDs, there remains a great need to further define the underlying mechanisms and to identify additional immune biomarkers, such as novel antibodies (in "seronegative" cases) to facilitate diagnosis and monitoring of patients. In addition, a better understanding of the pathogenesis of PNDs could lead to improved treatment strategies for both thymomas and their immune complications. In advanced, refractory cases of TETs (both thymoma and thymic carcinoma), additional therapeutic approaches are needed. Immune checkpoint inhibitors have revolutionized the treatment of several malignancies and hold promise in the treatment of TETs; however, the risks for immune-related adverse events (especially for inducing PNDs as well as in the setting of pre-existing PNDs) underscore the need to optimize patient selection and improve clinical management before there can be widespread acceptance of checkpoint inhibitor therapy in patients with TETs.

    View details for DOI 10.1007/s11864-019-0661-2

    View details for PubMedID 31227926

  • Allergic Diseases and Immune-Mediated Food Disorders in Pediatric Acute-Onset Neuropsychiatric Syndrome. Pediatric allergy, immunology, and pulmonology Rosa, J. S., Hernandez, J. D., Sherr, J. A., Smith, B. M., Brown, K. D., Farhadian, B., Mahony, T., McGhee, S. A., Lewis, D. B., Thienemann, M., Frankovich, J. D. 2018; 31 (3): 158-165


    Background: The prevalence and impact of allergic and immune-mediated food disorders in pediatric acute-onset neuropsychiatric syndrome (PANS) are mostly unknown. Objective: We sought to explore the prevalence of atopic dermatitis (AD), asthma, allergic rhinitis (AR), IgE-mediated food allergies (FAs), and other immune-mediated food disorders requiring food avoidance in patients with PANS. In addition, to further understand the extent of food restriction in this population, we investigated the empiric use of dietary measures to improve PANS symptoms. Methods: Pediatric patients in a PANS Clinic and Research Program were given surveys regarding their caregiver burdens, allergic and food-related medical history, and whether food elimination resulted in perception of improvement of PANS symptoms. A review of health records was conducted to confirm that all responses in the survey were concordant with documentation of each patient's medical chart. Results: Sixty-nine (ages 4-20 years) of 80 subjects who fulfilled PANS criteria completed the surveys. Thirteen (18.8%) had AD, 11 (15.9%) asthma, 33 (47.8%) AR, 11 (15.9%) FA, 1 (1.4%) eosinophilic gastrointestinal disorders, 1 (1.4%) food protein-induced enterocolitis syndrome, 3 (4.3%) milk protein-induced proctocolitis syndrome, and 3 (4.3%) celiac disease. Thirty subjects (43.5%) avoided foods due to PANS; elimination of gluten and dairy was most common and was associated with perceived improvement of PANS symptoms (by parents). This perceived improvement was not confirmed with objective data. Conclusions: The prevalence of allergic and immune-mediated food disorders in PANS is similar to the general population as reported in the literature, with the exception of AR that appears to be more prevalent in our PANS cohort. More research will be required to establish whether diet or allergies influence PANS symptoms.

    View details for DOI 10.1089/ped.2018.0888

    View details for PubMedID 30283713

    View details for PubMedCentralID PMC6154445

  • Allergic Diseases and Immune-Mediated Food Disorders in Pediatric Acute-Onset Neuropsychiatric Syndrome PEDIATRIC ALLERGY IMMUNOLOGY AND PULMONOLOGY Rosa, J. S., Hernandez, J. D., Sherr, J. A., Smith, B. M., Brown, K. D., Farhadian, B., Mahony, T., McGhee, S. A., Lewis, D. B., Thienemann, M., Frankovich, J. D. 2018; 31 (3): 158–65
  • The ratio of circulating regulatory cluster of differentiation 4 T cells to endothelial progenitor cells predicts clinically significant acute rejection after heart transplantation JOURNAL OF HEART AND LUNG TRANSPLANTATION Choi, D., Chmura, S. A., Ramachandran, V., Dionis-Petersen, K. Y., Kobayashi, Y., Nishi, T., Luikart, H., Dimbil, S., Kobashigawa, J., Khush, K., Lewis, D. B., Fearon, W. F. 2018; 37 (4): 496–502
  • Novel nonsense gain-of-function NFKB2 mutations associated with a combined immunodeficiency phenotype BLOOD Kuehn, H., Niemela, J. E., Sreedhara, K., Stoddard, J. L., Grossman, J., Wysocki, C. A., de la Morena, M., Garofalo, M., Inlora, J., Snyder, M. P., Lewis, D. B., Stratakis, C. A., Fleisher, T. A., Rosenzweig, S. D. 2017; 130 (13): 1553–64


    NF-κB signaling through its NFKB1-dependent canonical and NFKB2-dependent noncanonical pathways plays distinctive roles in a diverse range of immune processes. Recently, mutations in these 2 genes have been associated with common variable immunodeficiency (CVID). While studying patients with genetically uncharacterized primary immunodeficiencies, we detected 2 novel nonsense gain-of-function (GOF) NFKB2 mutations (E418X and R635X) in 3 patients from 2 families, and a novel missense change (S866R) in another patient. Their immunophenotype was assessed by flow cytometry and protein expression; activation of canonical and noncanonical pathways was examined in peripheral blood mononuclear cells and transfected HEK293T cells through immunoblotting, immunohistochemistry, luciferase activity, real-time polymerase chain reaction, and multiplex assays. The S866R change disrupted a C-terminal NF-κΒ2 critical site affecting protein phosphorylation and nuclear translocation, resulting in CVID with adrenocorticotropic hormone deficiency, growth hormone deficiency, and mild ectodermal dysplasia as previously described. In contrast, the nonsense mutations E418X and R635X observed in 3 patients led to constitutive nuclear localization and activation of both canonical and noncanonical NF-κΒ pathways, resulting in a combined immunodeficiency (CID) without endocrine or ectodermal manifestations. These changes were also found in 2 asymptomatic relatives. Thus, these novel NFKB2 GOF mutations produce a nonfully penetrant CID phenotype through a different pathophysiologic mechanism than previously described for mutations in NFKB2.

    View details for PubMedID 28778864

    View details for PubMedCentralID PMC5620416

  • An immune clock of human pregnancy SCIENCE IMMUNOLOGY Aghaeepour, N., Ganio, E. A., Mcilwain, D., Tsai, A. S., Tingle, M., Van Gassen, S., Gaudilliere, D. K., Baca, Q., McNeil, L., Okada, R., Ghaemi, M. S., Furman, D., Wong, R. J., Winn, V. D., Druzin, M. L., El-Sayed, Y. Y., Quaintance, C., Gibbs, R., Darmstadt, G. L., Shaw, G. M., Stevenson, D. K., Tibshirani, R., Nolan, G. P., Lewis, D. B., Angst, M. S., Gaudilliere, B. 2017; 2 (15)
  • A Case Report of Hypoglycemia and Hypogammaglobulinemia: DAVID syndrome in a patient with a novel NFKB2 mutation. journal of clinical endocrinology and metabolism Lal, R. A., Bachrach, L. K., Hoffman, A. R., Inlora, J., Rego, S., Snyder, M. P., Lewis, D. B. 2017


    DAVID syndrome (Deficient Anterior pituitary with Variable Immune Deficiency) is a rare disorder in which children present with symptomatic ACTH deficiency preceded by hypogammaglobulinemia from B-cell dysfunction with recurrent infections, termed common variable immunodeficiency (CVID). Subsequent whole exome sequencing studies have revealed germline heterozygous C-terminal mutations of NFKB2 as either a cause of DAVID syndrome or of CVID without clinical hypopituitarism. However, to the best of our knowledge there have been no cases in which the endocrinopathy has presented in the absence of a prior clinical history of CVID.A previously healthy 7 year-old boy with no history of clinical immunodeficiency, presented with profound hypoglycemia and seizures. He was found to have secondary adrenal insufficiency and was started on glucocorticoid replacement. An evaluation for autoimmune disease, including for anti-pituitary antibodies, was negative. Evaluation unexpectedly revealed hypogammaglobulinemia (decreased IgG, IgM, and IgA). He had moderately reduced serotype-specific IgG responses following pneumococcal polysaccharide vaccine. Subsequently, he was found to have growth hormone (GH) deficiency. Six years after initial presentation, whole exome sequencing revealed a novel de novo heterozygous NFKB2 missense mutation c.2596A>C (p.Ser866Arg) in the C-terminal region predicted to abrogate the processing of the p100 NFKB2 protein to its active p52 form.Isolated early-onset ACTH deficiency is rare and C-terminal region NFKB2 mutations should be considered as an etiology even in the absence of a clinical history of CVID. Early immunologic evaluation is indicated in the diagnosis and management of isolated ACTH deficiency.

    View details for DOI 10.1210/jc.2017-00341

    View details for PubMedID 28472507

  • Angiotensin-Converting Enzyme Inhibition Early After Heart Transplantation. Journal of the American College of Cardiology Fearon, W. F., Okada, K. n., Kobashigawa, J. A., Kobayashi, Y. n., Luikart, H. n., Sana, S. n., Daun, T. n., Chmura, S. A., Sinha, S. n., Cohen, G. n., Honda, Y. n., Pham, M. n., Lewis, D. B., Bernstein, D. n., Yeung, A. C., Valantine, H. A., Khush, K. n. 2017; 69 (23): 2832–41


    Cardiac allograft vasculopathy (CAV) remains a leading cause of mortality after heart transplantation (HT). Angiotensin-converting enzyme inhibitors (ACEIs) may retard the development of CAV but have not been well studied after HT.This study tested the safety and efficacy of the ACEI ramipril on the development of CAV early after HT.In this prospective, multicenter, randomized, double-blind, placebo-controlled trial, 96 HT recipients were randomized to undergo ramipril or placebo therapy. They underwent coronary angiography, endothelial function testing; measurements of fractional flow reserve (FFR) and coronary flow reserve (CFR) and the index of microcirculatory resistance (IMR); and intravascular ultrasonography (IVUS) of the left anterior descending coronary artery, within 8 weeks of HT. At 1 year, the invasive assessment was repeated. Circulating endothelial progenitor cells (EPCs) were quantified at baseline and 1 year.Plaque volumes at 1 year were similar between the ramipril and placebo groups (162.1 ± 70.5 mm(3) vs. 177.3 ± 94.3 mm(3), respectively; p = 0.73). Patients receiving ramipril had improvement in microvascular function as shown by a significant decrease in IMR (21.4 ± 14.7 to 14.4 ± 6.3; p = 0.001) and increase in CFR (3.8 ± 1.7 to 4.8 ± 1.5; p = 0.017), from baseline to 1 year. This did not occur with IMR (17.4 ± 8.4 to 21.5 ± 20.0; p = 0.72) or CFR (4.1 ± 1.8 to 4.1 ± 2.2; p = 0.60) in the placebo-treated patients. EPCs decreased significantly at 1 year in the placebo group but not in the ramipril group.Ramipril does not slow development of epicardial plaque volume but does stabilize levels of endothelial progenitor cells and improve microvascular function, which have been associated with improved long-term survival after HT. (Angiotensin Converting Enzyme [ACE] Inhibition and Cardiac Allograft Vasculopathy; NCT01078363).

    View details for PubMedID 28595700

  • 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)


    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

  • 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


    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

  • A cationic liposome-DNA complexes adjuvant (JVRS-100) enhances the immunogenicity and cross-protective efficacy of pre-pandemic influenza A (H5N1) vaccine in ferrets VIROLOGY Liu, F., Sun, X., Fairman, J., Lewis, D. B., Katz, J. M., Levine, M., Tumpey, T. M., Lu, X. 2016; 492: 197-203


    Influenza A (H5N1) viruses continue to pose a public health threat. As inactivated H5N1 vaccines are poorly immunogenic, adjuvants are needed to improve the immunogenicity of H5N1 vaccine in humans. Here, we investigated the immunogenicity and cross-protective efficacy in ferrets of a clade 2.2-derived vaccine with addition of JVRS-100, an adjuvant consisting of cationic liposome-DNA complexes (CLDC). After the first vaccination, significantly higher levels of hemagglutination-inhibition (HAI) and neutralizing antibody titers were detected in ferrets immunized with adjuvanted vaccine compared to unadjuvanted vaccine. Following a second dose of adjuvanted vaccine, HAI antibody titers of ≥ 40 were detected against viruses from multiple H5N1 clades. HAI antibodies against newly isolated H5N2 and H5N8 viruses were also augmented by JVRS-100. Ferrets were challenged with a heterologous H5N1 virus. All ferrets that received two doses of adjuvanted vaccine exhibited mild illness, significantly reduced nasal wash virus titers and protection from lethal challenge. In contrast, ferrets that received unadjuvanted vaccine showed greater weight loss, high viral titers and 3 of 6 animals succumbed to the lethal challenge. Our results indicate that the addition of JVRS-100 to H5N1 vaccine enhanced immunogenicity and cross-protection against lethal H5N1 virus disease in ferrets. JVRS-100 warrants further investigation as a potential adjuvant for influenza vaccines.

    View details for DOI 10.1016/j.virol.2016.02.024

    View details for Web of Science ID 000374209900022

    View details for PubMedID 26967975

  • Longitudinal Kinetics of Cytomegalovirus-Specific T-Cell Immunity and Viral Replication in Infants With Congenital Cytomegalovirus Infection. Journal of the Pediatric Infectious Diseases Society Chen, S. F., Holmes, T. H., Slifer, T., Ramachandran, V., Mackey, S., Hebson, C., Arvin, A. M., Lewis, D. B., Dekker, C. L. 2016; 5 (1): 14-20


    Congenital cytomegalovirus (CMV) is reported to affect up to 1% of all live births in the United States. T-cell immunity may be important for controlling CMV replication in congenital CMV-infected infants. We describe the natural history of CMV-specific T-cell evolution and CMV replication in infants with congenital CMV infection.Cytomegalovirus viral load, CMV urine culture, and CMV-specific CD4 and CD8 T-cell responses were assessed in a prospective longitudinal cohort of 51 infants with congenital CMV infection who were observed from birth to 3 years of age.We found a kinetic pattern of decreasing urinary CMV replication and increasing CMV-specific CD4 and CD8 T-cell responses during the first 3 years of life. We also found higher CMV-specific CD8 T-cell responses were associated with subsequent reduction of urine CMV viral load.For infants with congenital CMV infection, our data suggest an age-related maturation of both CMV-specific CD4 and CD8 T-cell immunity that is associated with an age-related decline in urinary CMV replication.

    View details for DOI 10.1093/jpids/piu089

    View details for PubMedID 26908487

  • Treatment of CGD-associated Colitis with the IL-23 Blocker Ustekinumab. Journal of clinical immunology Butte, M. J., Park, K. T., Lewis, D. B. 2016; 36 (7): 619–20

    View details for PubMedID 27465505

    View details for PubMedCentralID PMC5018915

  • Lack of IL7Ra expression in T cells is a hallmark of T-cell immunodeficiency in Schimke immuno-osseous dysplasia (SIOD). Clinical immunology Sanyal, M., Morimoto, M., Baradaran-Heravi, A., Choi, K., Kambham, N., Jensen, K., Dutt, S., Dionis-Petersen, K. Y., Liu, L. X., Felix, K., Mayfield, C., Dekel, B., Bokenkamp, A., Fryssira, H., Guillen-Navarro, E., Lama, G., Brugnara, M., Lücke, T., Olney, A. H., Hunley, T. E., Polat, A. I., Yis, U., Bogdanovic, R., Mitrovic, K., Berry, S., Najera, L., Najafian, B., Gentile, M., Nur Semerci, C., Tsimaratos, M., Lewis, D. B., Boerkoel, C. F. 2015; 161 (2): 355-365


    Schimke immuno-osseous dysplasia (SIOD) is an autosomal recessive, fatal childhood disorder associated with skeletal dysplasia, renal dysfunction, and T-cell immunodeficiency. This disease is linked to biallelic loss-of-function mutations of the SMARCAL1 gene. Although recurrent infection, due to T-cell deficiency, is a leading cause of morbidity and mortality, the etiology of the T-cell immunodeficiency is unclear. Here, we demonstrate that the T cells of SIOD patients have undetectable levels of protein and mRNA for the IL-7 receptor alpha chain (IL7Rα) and are unresponsive to stimulation with IL-7, indicating a loss of functional receptor. No pathogenic mutations were detected in the exons of IL7R in these patients; however, CpG sites in the IL7R promoter were hypermethylated in SIOD T cells. We propose therefore that the lack of IL7Rα expression, associated with hypermethylation of the IL7R promoter, in T cells and possibly their earlier progenitors, restricts T-cell development in SIOD patients.

    View details for DOI 10.1016/j.clim.2015.10.005

    View details for PubMedID 26499378

  • Single-cell systems-level analysis of human Toll-like receptor activation defines a chemokine signature in patients with systemic lupus erythematosus JOURNAL OF ALLERGY AND CLINICAL IMMUNOLOGY O'Gorman, W. E., Hsieh, E. W., Savig, E. S., Gherardini, P. F., Hernandez, J. D., Hansmann, L., Balboni, I. M., Utz, P. J., Bendall, S. C., Fantl, W. J., Lewis, D. B., Nolan, G. P., Davis, M. M. 2015; 136 (5): 1326-1336


    Activation of Toll-like receptors (TLRs) induces inflammatory responses involved in immunity to pathogens and autoimmune pathogenesis, such as in patients with systemic lupus erythematosus (SLE). Although TLRs are differentially expressed across the immune system, a comprehensive analysis of how multiple immune cell subsets respond in a system-wide manner has not been described.We sought to characterize TLR activation across multiple immune cell subsets and subjects, with the goal of establishing a reference framework against which to compare pathologic processes.Peripheral whole-blood samples were stimulated with TLR ligands and analyzed by means of mass cytometry simultaneously for surface marker expression, activation states of intracellular signaling proteins, and cytokine production. We developed a novel data visualization tool to provide an integrated view of TLR signaling networks with single-cell resolution. We studied 17 healthy volunteer donors and 8 patients with newly diagnosed and untreated SLE.Our data revealed the diversity of TLR-induced responses within cell types, with TLR ligand specificity. Subsets of natural killer cells and T cells selectively induced nuclear factor κ light chain enhancer of activated B cells in response to TLR2 ligands. CD14(hi) monocytes exhibited the most polyfunctional cytokine expression patterns, with more than 80 distinct cytokine combinations. Monocytic TLR-induced cytokine patterns were shared among a group of healthy donors, with minimal intraindividual and interindividual variability. Furthermore, autoimmune disease altered baseline cytokine production; newly diagnosed untreated SLE patients shared a distinct monocytic chemokine signature, despite clinical heterogeneity.Mass cytometry defined a systems-level reference framework for human TLR activation, which can be applied to study perturbations in patients with inflammatory diseases, such as SLE.

    View details for DOI 10.1016/j.jaci.2015.04.008

    View details for Web of Science ID 000364787200023

    View details for PubMedID 26037552

  • Heightened risk of preterm birth and growth restriction after a first-born son ANNALS OF EPIDEMIOLOGY Bruckner, T. A., Mayo, J. A., Gould, J. B., Stevenson, D. K., Lewis, D. B., Shaw, G. M., Carmichael, S. L. 2015; 25 (10): 743-747


    In Scandinavia, delivery of a first-born son elevates the risk of preterm delivery and intrauterine growth restriction of the next-born infant. External validity of these results remains unclear. We test this hypothesis for preterm delivery and growth restriction using the linked California birth cohort file. We examined the hypothesis separately by race and/or ethnicity.We retrieved data on 2,852,976 births to 1,426,488 mothers with at least two live births. Our within-mother tests applied Cox proportional hazards (preterm delivery, defined as less than 37 weeks gestation) and linear regression models (birth weight for gestational age percentiles).For non-Hispanic whites, Hispanics, Asians, and American Indian and/or Alaska Natives, analyses indicate heightened risk of preterm delivery and growth restriction after a first-born male. The race-specific hazard ratios for preterm delivery range from 1.07 to 1.18. Regression coefficients for birth weight for gestational age percentile range from -0.73 to -1.49. The 95% confidence intervals for all these estimates do not contain the null. By contrast, we could not reject the null for non-Hispanic black mothers.Whereas California findings generally support those from Scandinavia, the null results among non-Hispanic black mothers suggest that we do not detect adverse outcomes after a first-born male in all racial and/or ethnic groups.

    View details for DOI 10.1016/j.annepidem.2015.07.002

    View details for PubMedID 26265442

  • 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


    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

  • 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-?


    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

  • Bayesian immunological model development from the literature: Example investigation of recent thymic emigrants. Journal of immunological methods Holmes, T. H., Lewis, D. B. 2014; 414: 32-50


    Bayesian estimation techniques offer a systematic and quantitative approach for synthesizing data drawn from the literature to model immunological systems. As detailed here, the practitioner begins with a theoretical model and then sequentially draws information from source data sets and/or published findings to inform estimation of model parameters. Options are available to weigh these various sources of information differentially per objective measures of their corresponding scientific strengths. This approach is illustrated in depth through a carefully worked example for a model of decline in T-cell receptor excision circle content of peripheral T cells during development and aging. Estimates from this model indicate that 21years of age is plausible for the developmental timing of mean age of onset of decline in T-cell receptor excision circle content of peripheral T cells.

    View details for DOI 10.1016/j.jim.2014.08.001

    View details for PubMedID 25179832

  • Somatic reversion in dedicator of cytokinesis 8 immunodeficiency modulates disease phenotype. journal of allergy and clinical immunology Jing, H., Zhang, Q., Zhang, Y., Hill, B. J., Dove, C. G., Gelfand, E. W., Atkinson, T. P., Uzel, G., Matthews, H. F., Mustillo, P. J., Lewis, D. B., Kavadas, F. D., Hanson, I. C., Kumar, A. R., Geha, R. S., Douek, D. C., Holland, S. M., Freeman, A. F., Su, H. C. 2014; 133 (6): 1667-1675


    Autosomal recessive loss-of-function mutations in dedicator of cytokinesis 8 (DOCK8) cause a combined immunodeficiency characterized by atopy, recurrent infections, and cancer susceptibility. A genotype-phenotype explanation for the variable disease expression is lacking.We investigated whether reversions contributed to the variable disease expression.Patients followed at the National Institutes of Health's Clinical Center were studied. We performed detailed genetic analyses and intracellular flow cytometry to detect DOCK8 protein expression within lymphocyte subsets.We identified 17 of 34 DOCK8-deficient patients who had germline mutations with variable degrees of reversion caused by somatic repair. Somatic repair of the DOCK8 mutations resulted from second-site mutation, original-site mutation, gene conversion, and intragenic crossover. Higher degrees of reversion were associated with recombination-mediated repair. DOCK8 expression was restored primarily within antigen-experienced T cells or natural killer cells but less so in naive T or B cells. Several patients exhibited multiple different repair events. Patients who had reversions were older and had less severe allergic disease, although infection susceptibility persisted. No patients were cured without hematopoietic cell transplantation.In patients with DOCK8 deficiency, only certain combinations of germline mutations supported secondary somatic repair. Those patients had an ameliorated disease course with longer survival but still had fatal complications or required hematopoietic cell transplantation. These observations support the concept that some DOCK8-immunodeficient patients have mutable mosaic genomes that can modulate disease phenotype over time.

    View details for DOI 10.1016/j.jaci.2014.03.025

    View details for PubMedID 24797421

    View details for PubMedCentralID PMC4132167

  • ICOS Regulates the Generation and Function of Human CD4(+) Treg in a CTLA-4 Dependent Manner PLOS ONE Zheng, J., Chan, P., Liu, Y., Qin, G., Xiang, Z., Lam, K., Lewis, D. B., Lau, Y., Tu, W. 2013; 8 (12)


    Inducible co-stimulator (ICOS) is a member of CD28/Cytotoxic T-lymphocyte Antigen-4 (CTLA-4) family and broadly expressed in activated CD4(+) T cells and induced regulatory CD4(+) T cells (CD4(+) iTreg). ICOS-related signal pathway could be activated by the interaction between ICOS and its ligand (ICOSL). In our previous work, we established a cost-effective system to generate a novel human allo-antigen specific CD4(hi) Treg by co-culturing their naïve precursors with allogeneic CD40-activated B cells in vitro. Here we investigate the role of ICOS in the generation and function of CD4(hi) Treg by interrupting ICOS-ICOSL interaction with ICOS-Ig. It is found that blockade of ICOS-ICOSL interaction impairs the induction and expansion of CD4(hi) Treg induced by allogeneic CD40-activated B cells. More importantly, CD4(hi) Treg induced with the addition of ICOS-Ig exhibits decreased suppressive capacity on alloantigen-specific responses. Dysfunction of CD4(hi) Treg induced with ICOS-Ig is accompanied with its decreased exocytosis and surface CTLA-4 expression. Through inhibiting endocytosis with E64 and pepstatin A, surface CTLA-4 expression and suppressive functions of induced CD4(hi) Treg could be partly reversed. Conclusively, our results demonstrate the beneficial role of ICOS-ICOSL signal pathway in the generation and function of CD4(hi) Treg and uncover a novel relationship between ICOS and CTLA-4.

    View details for DOI 10.1371/journal.pone.0082203

    View details for Web of Science ID 000327944500124

    View details for PubMedID 24312642

    View details for PubMedCentralID PMC3846688

  • Defective sphingosine 1-phosphate receptor 1 (S1P1) phosphorylation exacerbates TH17-mediated autoimmune neuroinflammation. Nature immunology Garris, C. S., Wu, L., Acharya, S., Arac, A., Blaho, V. A., Huang, Y., Moon, B. S., Axtell, R. C., Ho, P. P., Steinberg, G. K., Lewis, D. B., Sobel, R. A., Han, D. K., Steinman, L., Snyder, M. P., Hla, T., Han, M. H. 2013; 14 (11): 1166-1172

    View details for DOI 10.1038/ni.2730

    View details for PubMedID 24076635

  • Defective sphingosine 1-phosphate receptor 1 (S1P1) phosphorylation exacerbates TH17-mediated autoimmune neuroinflammation. Nature immunology Garris, C. S., Wu, L., Acharya, S., Arac, A., Blaho, V. A., Huang, Y., Moon, B. S., Axtell, R. C., Ho, P. P., Steinberg, G. K., Lewis, D. B., Sobel, R. A., Han, D. K., Steinman, L., Snyder, M. P., Hla, T., Han, M. H. 2013; 14 (11): 1166-1172


    Sphingosine 1-phosphate (S1P) signaling regulates lymphocyte egress from lymphoid organs into systemic circulation. The sphingosine phosphate receptor 1 (S1P1) agonist FTY-720 (Gilenya) arrests immune trafficking and prevents multiple sclerosis (MS) relapses. However, alternative mechanisms of S1P-S1P1 signaling have been reported. Phosphoproteomic analysis of MS brain lesions revealed S1P1 phosphorylation on S351, a residue crucial for receptor internalization. Mutant mice harboring an S1pr1 gene encoding phosphorylation-deficient receptors (S1P1(S5A)) developed severe experimental autoimmune encephalomyelitis (EAE) due to autoimmunity mediated by interleukin 17 (IL-17)-producing helper T cells (TH17 cells) in the peripheral immune and nervous system. S1P1 directly activated the Jak-STAT3 signal-transduction pathway via IL-6. Impaired S1P1 phosphorylation enhances TH17 polarization and exacerbates autoimmune neuroinflammation. These mechanisms may be pathogenic in MS.

    View details for DOI 10.1038/ni.2730

    View details for PubMedID 24076635

  • Newborn screening for severe combined immunodeficiency and T-cell lymphopenia in California: Results of the first 2 years JOURNAL OF ALLERGY AND CLINICAL IMMUNOLOGY Kwan, A., Church, J. A., Cowan, M. J., Agarwal, R., Kapoor, N., Kohn, D. B., Lewis, D. B., McGhee, S. A., Moore, T. B., Stiehm, E. R., Porteus, M., Aznar, C. P., Currier, R., Lorey, F., Puck, J. M. 2013; 132 (1): 140-U245


    Assay of T-cell receptor excision circles (TRECs) in dried blood spots obtained at birth permits population-based newborn screening (NBS) for severe combined immunodeficiency (SCID).We sought to report the first 2 years of TREC NBS in California.Since August 2010, California has conducted SCID NBS. A high-throughput TREC quantitative PCR assay with DNA isolated from routine dried blood spots was developed. Samples with initial low TREC numbers had repeat DNA isolation with quantitative PCR for TRECs and a genomic control, and immunophenotyping was performed within the screening program for infants with incomplete or abnormal results. Outcomes were tracked.Of 993,724 infants screened, 50 (1/19,900 [0.005%]) had significant T-cell lymphopenia. Fifteen (1/66,250) required hematopoietic cell or thymus transplantation or gene therapy; these infants had typical SCID (n = 11), leaky SCID or Omenn syndrome (n = 3), or complete DiGeorge syndrome (n = 1). Survival to date in this group is 93%. Other T-cell lymphopenic infants had variant SCID or combined immunodeficiency (n = 6), genetic syndromes associated with T-cell impairment (n = 12), secondary T-cell lymphopenia (n = 9), or preterm birth (n = 8). All T-cell lymphopenic infants avoided live vaccines and received appropriate interventions to prevent infections. TREC test specificity was excellent: only 0.08% of infants required a second test, and 0.016% required lymphocyte phenotyping by using flow cytometry.TREC NBS in California has achieved early diagnosis of SCID and other conditions with T-cell lymphopenia, facilitating management and optimizing outcomes. Furthermore, NBS has revealed the incidence, causes, and follow-up of T-cell lymphopenia in a large diverse population.

    View details for DOI 10.1016/j.jaci.2013.04.024

    View details for Web of Science ID 000321052300019

    View details for PubMedID 23810098

  • Retinoic Acid Hypersensitivity Promotes Peripheral Tolerance in Recent Thymic Emigrants JOURNAL OF IMMUNOLOGY Bhaumik, S., Giffon, T., Bolinger, D., Kirkman, R., Lewis, D. B., Weaver, C. T., Randolph, D. A. 2013; 190 (6): 2603-2613


    Whereas thymic education eliminates most self-reactive T cells, additional mechanisms to promote tolerance in the periphery are critical to prevent excessive immune responses against benign environmental Ags and some self-Ags. In this study we show that murine CD4(+) recent thymic emigrants (RTEs) are programmed to facilitate tolerance in the periphery. Both in vitro and in vivo, naive RTEs more readily upregulate Foxp3 than do mature naive cells after stimulation under tolerogenic conditions. In RTEs, a relatively high sensitivity to retinoic acid contributes to decreased IFN-γ production, permitting the expression of Foxp3. Conversely, mature naive CD4 cells have a lower sensitivity to retinoic acid, resulting in increased IFN-γ production and subsequent IFN-γ-mediated silencing of Foxp3 expression. Enhanced retinoic acid signaling and Foxp3 induction in RTEs upon Ag encounter in the periphery may serve as form of secondary education that complements thymic education and helps avoid inappropriate immune responses. This mechanism for tolerance may be particularly important in settings where RTEs comprise a large fraction of the peripheral T cell pool, such as in newborns or after umbilical cord blood transplant.

    View details for DOI 10.4049/jimmunol.1200852

    View details for Web of Science ID 000315657200018

    View details for PubMedID 23401586

    View details for PubMedCentralID PMC3594456

  • Human Neonatal Naive CD4(+) T Cells Have Enhanced Activation-Dependent Signaling Regulated by the MicroRNA miR-181a JOURNAL OF IMMUNOLOGY Palin, A. C., Ramachandran, V., Acharya, S., Lewis, D. B. 2013; 190 (6): 2682-2691


    Compared with older children and adults, human neonates have reduced and delayed CD4(+) T cell immunity to certain pathogens, but the mechanisms for these developmental differences in immune function remain poorly understood. We investigated the hypothesis that impaired human neonatal CD4(+) T cell immunity was due to reduced signaling by naive CD4(+) T cells following engagement of the αβ-TCR/CD3 complex and CD28. Surprisingly, calcium flux following engagement of CD3 was significantly higher in neonatal naive CD4(+) T cells from umbilical cord blood (CB) compared with naive CD4(+) T cells from adult peripheral blood. Enhanced calcium flux was also observed in adult CD4(+) recent thymic emigrants. Neonatal naive CD4(+) T cells also had higher activation-induced Erk phosphorylation. The microRNA miR-181a, which enhances activation-induced calcium flux in murine thymocytes, was expressed at significantly higher levels in CB naive CD4(+) T cells compared with adult cells. Overexpression of miR-181a in adult naive CD4(+) T cells increased activation-induced calcium flux, implying that the increased miR-181a levels of CB naive CD4(+) T cells contributed to their enhanced signaling. In contrast, AP-1-dependent transcription, which is downstream of Erk and required for full T cell activation, was decreased in CB naive CD4(+) T cells compared with adult cells. Thus, CB naive CD4(+) T cells have enhanced activation-dependent calcium flux, indicative of the retention of a thymocyte-like phenotype. Enhanced calcium signaling and Erk phosphorylation are decoupled from downstream AP-1-dependent transcription, which is reduced and likely contributes to limitations of human fetal and neonatal CD4(+) T cell immunity.

    View details for DOI 10.4049/jimmunol.1202534

    View details for Web of Science ID 000315657200026

    View details for PubMedID 23408835

  • Human CD8(+) Regulatory T Cells Inhibit GVHD and Preserve General Immunity in Humanized Mice SCIENCE TRANSLATIONAL MEDICINE Zheng, J., Liu, Y., Liu, Y., Liu, M., Xiang, Z., Lam, K., Lewis, D. B., Lau, Y., Tu, W. 2013; 5 (168)


    Graft-versus-host disease (GVHD) is a lethal complication of allogeneic bone marrow transplantation (BMT). Immunosuppressive agents are currently used to control GVHD but may cause general immune suppression and limit the effectiveness of BMT. Adoptive transfer of regulatory T cells (T(regs)) can prevent GVHD in rodents, suggesting a therapeutic potential of T(regs) for GVHD in humans. However, the clinical application of T(reg)-based therapy is hampered by the low frequency of human T(regs) and the lack of a reliable model to test their therapeutic effects in vivo. Recently, we successfully generated human alloantigen-specific CD8(hi) T(regs) in a large scale from antigenically naïve precursors ex vivo using allogeneic CD40-activated B cells as stimulators. We report a human allogeneic GVHD model established in humanized mice to mimic GVHD after BMT in humans. We demonstrate that ex vivo-induced CD8(hi) T(regs) controlled GVHD in an allospecific manner by reducing alloreactive T cell proliferation as well as decreasing inflammatory cytokine and chemokine secretion within target organs through a CTLA-4-dependent mechanism in humanized mice. These CD8(hi) T(regs) induced long-term tolerance effectively without compromising general immunity and graft-versus-tumor activity. Our results support testing of human CD8(hi) T(regs) in GVHD in clinical trials.

    View details for DOI 10.1126/scitranslmed.3004943

    View details for Web of Science ID 000313739200006

    View details for PubMedID 23325802

  • Dental Abnormalities in Schimke Immuno-osseous Dysplasia JOURNAL OF DENTAL RESEARCH Morimoto, M., Kerouredan, O., Gendronneau, M., Shuen, C., Baradaran-Heravi, A., Asakura, Y., Basiratnia, M., Bogdanovic, R., Bonneau, D., Buck, A., Charrow, J., Cochat, P., DeHaai, K. A., Fenkci, M. S., Frange, P., Fruend, S., Fryssira, H., Keller, K., Kirmani, S., Kobelka, C., Kohler, K., Lewis, D. B., Massella, L., MCLEOD, D. R., Milford, D. V., Nobili, F., Olney, A. H., Semerci, C. N., Stajic, N., Stein, A., Taque, S., Zonana, J., Luecke, T., Hendson, G., Bonnaure-Mallet, M., Boerkoel, C. F. 2012; 91 (7): S29-S37


    Described for the first time in 1971, Schimke immuno-osseous dysplasia (SIOD) is an autosomal-recessive multisystem disorder that is caused by bi-allelic mutations of SMARCAL1, which encodes a DNA annealing helicase. To define better the dental anomalies of SIOD, we reviewed the records from SIOD patients with identified bi-allelic SMARCAL1 mutations, and we found that 66.0% had microdontia, hypodontia, or malformed deciduous and permanent molars. Immunohistochemical analyses showed expression of SMARCAL1 in all developing teeth, raising the possibility that the malformations are cell-autonomous consequences of SMARCAL1 deficiency. We also found that stimulation of cultured skin fibroblasts from SIOD patients with the tooth morphogens WNT3A, BMP4, and TGFβ1 identified altered transcriptional responses, raising the hypothesis that the dental malformations arise in part from altered responses to developmental morphogens. To the best of our knowledge, this is the first systematic study of the dental anomalies associated with SIOD.

    View details for DOI 10.1177/0022034512450299

    View details for Web of Science ID 000307189700006

    View details for PubMedCentralID PMC3383106

  • Dental abnormalities in Schimke immuno-osseous dysplasia. Journal of dental research Morimoto, M., Kérourédan, O., Gendronneau, M., Shuen, C., Baradaran-Heravi, A., Asakura, Y., Basiratnia, M., Bogdanovic, R., Bonneau, D., Buck, A., Charrow, J., Cochat, P., DeHaai, K. A., Fenkçi, M. S., Frange, P., Fründ, S., Fryssira, H., Keller, K., Kirmani, S., Kobelka, C., Kohler, K., Lewis, D. B., Massella, L., MCLEOD, D. R., Milford, D. V., Nobili, F., Olney, A. H., Semerci, C. N., Stajic, N., Stein, A., Taque, S., Zonana, J., Lücke, T., Hendson, G., Bonnaure-Mallet, M., Boerkoel, C. F. 2012; 91 (7): 29S-37S


    Described for the first time in 1971, Schimke immuno-osseous dysplasia (SIOD) is an autosomal-recessive multisystem disorder that is caused by bi-allelic mutations of SMARCAL1, which encodes a DNA annealing helicase. To define better the dental anomalies of SIOD, we reviewed the records from SIOD patients with identified bi-allelic SMARCAL1 mutations, and we found that 66.0% had microdontia, hypodontia, or malformed deciduous and permanent molars. Immunohistochemical analyses showed expression of SMARCAL1 in all developing teeth, raising the possibility that the malformations are cell-autonomous consequences of SMARCAL1 deficiency. We also found that stimulation of cultured skin fibroblasts from SIOD patients with the tooth morphogens WNT3A, BMP4, and TGFβ1 identified altered transcriptional responses, raising the hypothesis that the dental malformations arise in part from altered responses to developmental morphogens. To the best of our knowledge, this is the first systematic study of the dental anomalies associated with SIOD.

    View details for PubMedID 22699664

  • Genotype, phenotype, and outcomes of nine patients with T-B plus NK plus SCID PEDIATRIC TRANSPLANTATION Yu, G. P., Nadeau, K. C., Berk, D. R., de Saint Basile, G., Lambert, N., Knapnougel, P., Roberts, J., Kavanau, K., Dunn, E., Stiehm, E. R., Lewis, D. B., Umetsu, D. T., Puck, J. M., Cowan, M. J. 2011; 15 (7): 733-741


    There are few reports of clinical presentation, genotype, and HCT outcomes for patients with T-B+NK+ SCID. Between 1981 and 2007, eight of 84 patients with SCID who received and/or were followed after HCT at UCSF had the T-B+NK+ phenotype. One additional patient with T-B+NK+ SCID was identified as the sibling of a patient treated at UCSF. Chart reviews were performed. Molecular analyses of IL7R, IL2RG, JAK3, and the genes encoding the CD3 T-cell receptor components δ (CD3D), ε (CD3E), and ζ (CD3Z) were carried out. IL7R mutations were documented in four patients and CD3D mutations in two others. Three patients had no defects found. Only two of nine patients had an HLA-matched related HCT donor. Both survived, and neither developed GVHD. Five of seven recipients of haploidentical grafts survived. Although the majority of reported cases of T-B+NK+ SCID are caused by defects in IL7R, CD3 complex defects were also found in this series and should be considered when evaluating patients with T-B+NK+ SCID. Additional genes, mutations in which account for T-B+NK+ SCID, remain to be found. Better approaches to early diagnosis and HCT treatment are needed for patients lacking an HLA-matched related donor.

    View details for DOI 10.1111/j.1399-3046.2011.01563.x

    View details for Web of Science ID 000296049000020

    View details for PubMedID 21883749

    View details for PubMedCentralID PMC3196791

  • Protein tyrosine kinase 7: a novel surface marker for human recent thymic emigrants with potential clinical utility JOURNAL OF PERINATOLOGY Lewis, D. B., Haines, C., Ross, D. 2011; 31: S72-S81


    Recent thymic emigrants (RTEs) are antigenically naive T cells that have recently completed intrathymic maturation and have emigrated from the thymus to the periphery. RTEs are clinically and immunologically important as they are essential for maintaining peripheral T cells in sufficient numbers in order to recognize, by their αβT-cell receptors (TCRs), a diverse array of foreign peptide antigens. However, RTE frequency and function has been poorly understood because of a lack of surface markers to distinguish them from older non-RTE naive T cells. This review summarizes the biology of the intrathymic generation and function of RTEs, including the recent identification of protein tyrosine kinase 7 (PTK7) as a novel marker for human RTEs of the CD4 (helper) T-cell lineage. PTK7+ RTEs in adults have a reduced capacity for activation-induced proliferation and cytokine production (interleukin-2 and interferon-γ) than older PTK7- naive CD4 T cells. Importantly, this immaturity in CD4 RTE effector function may contribute to the reduced adaptive immune responses observed in situations in which CD4 RTEs predominate, including the fetus, neonate and young infant, and following immune reconstitution, such as post-hematopoietic stem cell transplant. The ability to identify viable CD4+ RTEs based on PTK7 surface staining may be particularly useful in the infant for better defining the impact of nutritional and environmental factors on thymic output, peripheral T-cell function and adaptive immune responses to vaccination and infection.

    View details for DOI 10.1038/jp.2010.187

    View details for Web of Science ID 000289236900012

    View details for PubMedID 21448210

  • Generation of human Th1-like regulatory CD4(+) T cells by an intrinsic IFN-gamma- and T-bet-dependent pathway EUROPEAN JOURNAL OF IMMUNOLOGY Zheng, J., Liu, Y., Qin, G., Lam, K., Guan, J., Xiang, Z., Lewis, D. B., Lau, Y., Tu, W. 2011; 41 (1): 128-139


    Murine Foxp3(+) Treg have recently been shown to express T-bet, a transcription factor characteristic of Th1 effector cells. A human Treg phenotype equivalent has not been reported. Here, we show that naïve human CD4(+) T cells incubated with low numbers of CD40-activated allogeneic B cells preferentially differentiate into alloantigen-specific CD4(hi) CD25(hi) Treg. These differentiated cells potently suppress effector T-cell responses and express T-bet, IFN-γ, and CXCR3, the features of Th1 effector cells. In contrast, co-culture of naïve CD4(+) T cells with high numbers of allogeneic B cells results in CD4(+) CD25(+) T cells that promote, rather than inhibit, effector T-cell responses, demonstrating the plasticity of CD4(+) T-cell differentiation in response to alloantigen-presenting B cells. The optimal accumulation of CD4(hi) CD25(hi) Treg induced using higher T cell:B cell co-culture ratios was dependent on the expression of T-bet and endogenously produced IFN-γ. Induction of Treg-mediated suppression function in the Treg population was not. As CXCR3 confers the preferential trafficking of T cells to tissue sites of IFN-γ, these human Th1-like Treg might be useful for modulating pathological Th1 effector responses, such as that occurring during graft-versus-host disease or graft rejection.

    View details for DOI 10.1002/eji.201040724

    View details for Web of Science ID 000285933000014

    View details for PubMedID 21182084

  • Toxicopathology of the Developing Immune System: Investigative and Development Strategies TOXICOLOGIC PATHOLOGY Weinstock, D., Lewis, D. B., Parker, G. A., Beyer, J., Collinge, M., Brown, T. P., Dybdal, N. 2010; 38 (7): 1111-1117


    Developmental immunotoxicity (DIT) has gained attention with the recognition that environmental chemicals can potentially affect the developing immune system and the incidence of childhood allergic diseases. Preclinical safety assessment of pharmaceuticals for men and women of childbearing potential as well as for pediatric and juvenile indications may require DIT assessments. Draft documents from environmental and chemical regulatory agencies propose strategies that use the rat as a test species and incorporate histopathology and functional testing as endpoints. While there are no guidelines for DIT assessment of pharmaceuticals, current discussions suggest that combining immunotoxicity and developmental and reproductive toxicology studies may serve this purpose. Knowledge of the principles and applications of DIT will facilitate participation in strategy development and effective conduct of relevant studies.

    View details for DOI 10.1177/0192623310382436

    View details for Web of Science ID 000293378600011

    View details for PubMedID 20962107

  • Abstract Submission Acharya, S., Petersen, K. D., Golubkov, V., Kwong, M., Adams, C. M., Jackson, P. K., Lewis, D. B. AMER ASSOC CANCER RESEARCH. 2015
  • Acute Subclinical Maternal Inflammation in Late Pregnancy Ozen, M., Zhao, H., Winn, V. D., Lewis, D. B., Kalish, F., Palmer, T. D., Wong, R. J., Stevenson, D. K. SAGE PUBLICATIONS INC. 2015: 89A–90A
  • Protein tyrosine kinase 7 promotes PI3K/AKT signaling and survival in primary and neoplastic CD4+T-lineage cells. Acharya, S., Dionis-Petersen, K., Jackson, P., Admas, C., Lewis, D. AMER ASSOC IMMUNOLOGISTS. 2014
  • Defective sphingosine-1-phosphate receptor 1 (S1PR(1)) internalization exacerbates autoimmune neuroinflammation by enhancing Stat3-mediated Th17 polarization Garris, C. S., Acharya, S., Axtell, R., Lewis, D., Sobel, R. A., Steinman, L., Hla, T., Han, M. H. SAGE PUBLICATIONS LTD. 2012: 510
  • Reduced elastogenesis: a clue to the arteriosclerosis and emphysematous changes in Schimke immuno-osseous dysplasia? ORPHANET JOURNAL OF RARE DISEASES Morimoto, M., Yu, Z., Stenzel, P., Clewing, J. M., Najafian, B., Mayfield, C., Hendson, G., Weinkauf, J. G., Gormley, A. K., Parham, D. M., Ponniah, U., Andre, J., Asakura, Y., Basiratnia, M., Bogdanovic, R., Bokenkamp, A., Bonneau, D., Buck, A., Charrow, J., Cochat, P., Cordeiro, I., Deschenes, G., Fenkci, M. S., Frange, P., Fruend, S., Fryssira, H., Guillen-Navarro, E., Keller, K., Kirmani, S., Kobelka, C., Lamfers, P., Levtchenko, E., Lewis, D. B., Massella, L., McLeod, D. R., Milford, D. V., Nobili, F., Saraiva, J. M., Semerci, C. N., Shoemaker, L., Stajic, N., Stein, A., Taha, D., Wand, D., Zonana, J., Luecke, T., Boerkoel, C. F. 2012; 7


    Arteriosclerosis and emphysema develop in individuals with Schimke immuno-osseous dysplasia (SIOD), a multisystem disorder caused by biallelic mutations in SMARCAL1 (SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily a-like 1). However, the mechanism by which the vascular and pulmonary disease arises in SIOD remains unknown.We reviewed the records of 65 patients with SMARCAL1 mutations. Molecular and immunohistochemical analyses were conducted on autopsy tissue from 4 SIOD patients.Thirty-two of 63 patients had signs of arteriosclerosis and 3 of 51 had signs of emphysema. The arteriosclerosis was characterized by intimal and medial hyperplasia, smooth muscle cell hyperplasia and fragmented and disorganized elastin fibers, and the pulmonary disease was characterized by panlobular enlargement of air spaces. Consistent with a cell autonomous disorder, SMARCAL1 was expressed in arterial and lung tissue, and both the aorta and lung of SIOD patients had reduced expression of elastin and alterations in the expression of regulators of elastin gene expression.This first comprehensive study of the vascular and pulmonary complications of SIOD shows that these commonly cause morbidity and mortality and might arise from impaired elastogenesis. Additionally, the effect of SMARCAL1 deficiency on elastin expression provides a model for understanding other features of SIOD.

    View details for DOI 10.1186/1750-1172-7-70

    View details for Web of Science ID 000312257000001

    View details for PubMedID 22998683

    View details for PubMedCentralID PMC3568709

  • Role of protein tyrosine kinase seven in human recent thymic emigrant cell survival Acharya, S., Ramachandran, V., Dionis, K., Lewis, D. AMER ASSOC IMMUNOLOGISTS. 2012
  • Enhanced in vivo immunity and protection provided by cationic lipid-DNA complex adjuvanted influenza vaccination is independent of Toll-like receptor 9 Botham, C., Hong, D., Giffon, T., Draghi, M., Fairman, J., Lewis, D. AMER ASSOC IMMUNOLOGISTS. 2011
  • Neonatal naive CD4+T cells have increased calcium flux and higher expression of miR-181a relative to adult naive CD4+T cells Paulin, A., Venkatasubrahmanyam, S., Butte, A., Lewis, D. AMER ASSOC IMMUNOLOGISTS. 2011


    Prepandemic intravenous immunoglobulin (IVIG) and sera from Kawasaki disease patients treated with this IVIG were analyzed for 2009 H1N1-specific microneutralization and hemagglutination inhibition antibodies. All 6 different IVIG preparations tested had significant levels of cross-reactive-specific antibody at a concentration of 2.0 g/dL of immunoglobulin. Sera from 18 of 19 Kawasaki disease patients had significant increases of cross-reactive-specific antibody after 2.0 g/kg of prepandemic IVIG. These results suggest a role for adjunctive IVIG therapy for severe and/or drug-resistant 2009 H1N1 virus and other highly antigenically drifted influenza strains, particularly in the immunocompromised.

    View details for DOI 10.1097/INF.0b013e3181f127be

    View details for PubMedID 20724956

  • Cationic Lipid/DNA Complex-Adjuvanted Influenza A Virus Vaccination Induces Robust Cross-Protective Immunity JOURNAL OF VIROLOGY Hong, D. K., Chang, S., Botham, C. M., Giffon, T. D., Fairman, J., Lewis, D. B. 2010; 84 (24): 12691-12702


    Influenza A virus is a negative-strand segmented RNA virus in which antigenically distinct viral subtypes are defined by the hemagglutinin (HA) and neuraminidase (NA) major viral surface proteins. An ideal inactivated vaccine for influenza A virus would induce not only highly robust strain-specific humoral and T-cell immune responses but also cross-protective immunity in which an immune response to antigens from a particular viral subtype (e.g., H3N2) would protect against other viral subtypes (e.g., H1N1). Cross-protective immunity would help limit outbreaks from newly emerging antigenically novel strains. Here, we show in mice that the addition of cationic lipid/noncoding DNA complexes (CLDC) as adjuvant to whole inactivated influenza A virus vaccine induces significantly more robust adaptive immune responses both in quantity and quality than aluminum hydroxide (alum), which is currently the most widely used adjuvant in clinical human vaccination. CLDC-adjuvanted vaccine induced higher total influenza virus-specific IgG, particularly for the IgG2a/c subclass. Higher levels of multicytokine-producing influenza virus-specific CD4 and CD8 T cells were induced by CLDC-adjuvanted vaccine than with alum-adjuvanted vaccine. Importantly, CLDC-adjuvanted vaccine provided significant cross-protection from either a sublethal or lethal influenza A viral challenge with a different subtype than that used for vaccination. This superior cross-protection afforded by the CLDC adjuvant required CD8 T-cell recognition of viral peptides presented by classical major histocompatibility complex class I proteins. Together, these results suggest that CLDC has particular promise for vaccine strategies in which T cells play an important role and may offer new opportunities for more effective control of human influenza epidemics and pandemics by inactivated influenza virus vaccine.

    View details for DOI 10.1128/JVI.00769-10

    View details for PubMedID 20943978

  • Lack of IL-7 Receptor Alpha Chain (CD127) Expression In T Cells Is a Hallmark of T-Cell Immunodeficiency In Schimke Immuno-Osseous Dysplasia (SIOD) Sanyal, M., Dionis, K. Y., Baradaran-Heravi, A., Dekel, B., Boekenkamp, A., Boerkoel, C. F., Lewis, D. B. AMER SOC HEMATOLOGY. 2010: 1141
  • Revisiting Human IL-12R beta 1 Deficiency A Survey of 141 Patients From 30 Countries MEDICINE de Beaucoudrey, L., Samarina, A., Bustamante, J., Cobat, A., Boisson-Dupuis, S., Feinberg, J., Al-Muhsen, S., Janniere, L., Rose, Y., de Suremain, M., Kong, X., Filipe-Santos, O., Chapgier, A., Picard, C., Fischer, A., Dogu, F., Ikinciogullari, A., Tanir, G., Al-Hajjar, S., Al-Jumaah, S., Frayha, H. H., Alsum, Z., Al-Ajaji, S., Alangari, A., Al-Ghonaium, A., Adimi, P., Mansouri, D., Ben-Mustapha, I., Yancoski, J., Garty, B., Rodriguez-Gallego, C., Caragol, I., Kutukculer, N., Kumararatne, D. S., Patel, S., Doffinger, R., Exley, A., Jeppsson, O., Reichenbach, J., Nadal, D., Boyko, Y., Pietrucha, B., Anderson, S., Levin, M., Schandene, L., Schepers, K., Efira, A., Mascart, F., Matsuoka, M., Sakai, T., Siegrist, C., Frecerova, K., Blueetters-Sawatzki, R., Bernhoeft, J., Freihorst, J., Baumann, U., Richter, D., Haerynck, F., De Baets, F., Novelli, V., Lammas, D., Vermylen, C., Tuerlinckx, D., Nieuwhof, C., Pac, M., Haas, W. H., Mueller-Fleckenstein, I., Fleckenstein, B., Levy, J., Raj, R., Cohen, A. C., Lewis, D. B., Holland, S. M., Yang, K. D., Wang, X., Wang, X., Jiang, L., Yang, X., Zhu, C., Xie, Y., Lee, P. P., Chan, K. W., Chen, T., Castro, G., Natera, I., Codoceo, A., King, A., Bezrodnik, L., Di Giovani, D., Isabel Gaillard, M., de Moraes-Vasconcelos, D., Grumach, A. S., da Silva Duarte, A. J., Aldana, R., Javier Espinosa-Rosales, F., Bejaoui, M., Bousfiha, A. A., El Baghdadi, J., Ozbek, N., Aksu, G., Keser, M., Somer, A., Hatipoglu, N., Aydogmus, C., Asilsoy, S., Camcioglu, Y., Gulle, S., Ozgur, T. T., Ozen, M., Oleastro, M., Bernasconi, A., Mamishi, S., Parvaneh, N., Rosenzweig, S., Barbouche, R., Pedraza, S., Lau, Y. L., Ehlayel, M. S., Fieschi, C., Abel, L., Sanal, O., Casanova, J. 2010; 89 (6): 381-402


    Interleukin-12 receptor β1 (IL-12Rβ1) deficiency is the most common form of Mendelian susceptibility to mycobacterial disease (MSMD). We undertook an international survey of 141 patients from 102 kindreds in 30 countries. Among 102 probands, the first infection occurred at a mean age of 2.4 years. In 78 patients, this infection was caused by Bacille Calmette-Guérin (BCG; n = 65), environmental mycobacteria (EM; also known as atypical or nontuberculous mycobacteria) (n = 9) or Mycobacterium tuberculosis (n = 4). Twenty-two of the remaining 24 probands initially presented with nontyphoidal, extraintestinal salmonellosis. Twenty of the 29 genetically affected sibs displayed clinical signs (69%); however 8 remained asymptomatic (27%). Nine nongenotyped sibs with symptoms died. Recurrent BCG infection was diagnosed in 15 cases, recurrent EM in 3 cases, recurrent salmonellosis in 22 patients. Ninety of the 132 symptomatic patients had infections with a single microorganism. Multiple infections were diagnosed in 40 cases, with combined mycobacteriosis and salmonellosis in 36 individuals. BCG disease strongly protected against subsequent EM disease (p = 0.00008). Various other infectious diseases occurred, albeit each rarely, yet candidiasis was reported in 33 of the patients (23%). Ninety-nine patients (70%) survived, with a mean age at last follow-up visit of 12.7 years ± 9.8 years (range, 0.5-46.4 yr). IL-12Rβ1 deficiency is characterized by childhood-onset mycobacteriosis and salmonellosis, rare recurrences of mycobacterial disease, and more frequent recurrence of salmonellosis. The condition has higher clinical penetrance, broader susceptibility to infections, and less favorable outcome than previously thought.

    View details for DOI 10.1097/MD.0b013e3181fdd832

    View details for Web of Science ID 000283840300002

    View details for PubMedID 21057261

    View details for PubMedCentralID PMC3129625

  • Alternate Mechanisms of Initial Pattern Recognition Drive Differential Immune Responses to Related Poxviruses CELL HOST & MICROBE O'Gorman, W. E., Sampath, P., Simonds, E. F., Sikorski, R., O'Malley, M., Krutzik, P. O., Chen, H., Panchanathan, V., Chaudhri, G., Karupiah, G., Lewis, D. B., Thorne, S. H., Nolan, G. P. 2010; 8 (2): 174-185


    Vaccinia immunization was pivotal to successful smallpox eradication. However, the early immune responses that distinguish poxvirus immunization from pathogenic infection remain unknown. To address this, we developed a strategy to map the activation of key signaling networks in vivo and applied this approach to define and compare the earliest signaling events elicited by immunizing (vaccinia) and lethal (ectromelia) poxvirus infections in mice. Vaccinia induced rapid TLR2-dependent responses, leading to IL-6 production, which then initiated STAT3 signaling in dendritic and T cells. In contrast, ectromelia did not induce TLR2 activation, and profound mouse strain-dependent responses were observed. In resistant C57BL/6 mice, the STAT1 and STAT3 pathways were rapidly activated, whereas in susceptible BALB/c mice, IL-6-dependent STAT3 activation did not occur. These data link early immune signaling events to infection outcome and suggest that activation of different pattern-recognition receptors early after infection may be important in determining vaccine efficacy.

    View details for DOI 10.1016/j.chom.2010.07.008

    View details for Web of Science ID 000281169600007

    View details for PubMedID 20709294

    View details for PubMedCentralID PMC2940993

  • Adult-onset presentations of genetic immunodeficiencies: genes can throw slow curves CURRENT OPINION IN INFECTIOUS DISEASES Nelson, K. S., Lewis, D. B. 2010; 23 (4): 359-364


    The molecular and genetic mechanisms behind adult presentations of primary immunodeficiency diseases are examined, with particular emphasis on cases where this was heralded by severe, recurrent, or opportunistic infection.A detailed analysis over the last two decades of the relationship between genotype and clinical phenotype for a number of genetic immunodeficiencies has revealed multiple mechanisms that can account for the delayed presentation of genetic disorders that typically present in childhood, including hypomorphic gene mutations and X-linked gene mutations with age-related skewing in random X-chromosome inactivation. Adult-onset presentations of chronic granulomatous disease, X-linked agammaglobulinemia, IL-12/Th1/IFN-gamma and IL-23/Th17/IL-17 pathway defects, and X-linked lymphoproliferative disorder are used to illustrate these mechanisms. Finally, certain genetic types of common variable immunodeficiency are used to illustrate that inherited null mutations can take decades to manifest immunologically.Both genetic mechanisms and environmental factors can account for adult-onset infectious and noninfectious complications as manifestations of disorders that are typically present in childhood. This emphasizes the potential complexity in the relationship between genotype and phenotype with natural human mutations.

    View details for DOI 10.1097/QCO.0b013e32833bc1b0

    View details for Web of Science ID 000279397600010

    View details for PubMedID 20581672

  • Surveying the Influenza-specific Cytotoxic T Cell Response in Humans and Mice Using Mass-cytometry (CyTOF) and Combinatorial Tetramer Staining Newell, E., Bendall, S., Hong, D., Lewis, D., Nolan, G., Davis, M. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2010: S104
  • A new genetic subgroup of chronic granulomatous disease with autosomal recessive mutations in p40(phox) and selective defects in neutrophil NADPH oxidase activity BLOOD Matute, J. D., Arias, A. A., Wright, N. A., Wrobel, I., Waterhouse, C. C., Li, X. J., Marchal, C. C., Stull, N. D., Lewis, D. B., Steele, M., Kellner, J. D., Yu, W., Meroueh, S. O., Nauseef, W. M., Dinauer, M. C. 2009; 114 (15): 3309-3315


    Chronic granulomatous disease (CGD), an immunodeficiency with recurrent pyogenic infections and granulomatous inflammation, results from loss of phagocyte superoxide production by recessive mutations in any 1 of 4 genes encoding subunits of the phagocyte NADPH oxidase. These include gp91(phox) and p22(phox), which form the membrane-integrated flavocytochrome b, and cytosolic subunits p47(phox) and p67(phox). A fifth subunit, p40(phox), plays an important role in phagocytosis-induced superoxide production via a phox homology (PX) domain that binds to phosphatidylinositol 3-phosphate (PtdIns(3)P). We report the first case of autosomal recessive mutations in NCF4, the gene encoding p40(phox), in a boy who presented with granulomatous colitis. His neutrophils showed a substantial defect in intracellular superoxide production during phagocytosis, whereas extracellular release of superoxide elicited by phorbol ester or formyl-methionyl-leucyl-phenylalanine (fMLF) was unaffected. Genetic analysis of NCF4 showed compound heterozygosity for a frameshift mutation with premature stop codon and a missense mutation predicting a R105Q substitution in the PX domain. Parents and a sibling were healthy heterozygous carriers. p40(phox)R105Q lacked binding to PtdIns(3)P and failed to reconstitute phagocytosis-induced oxidase activity in p40(phox)-deficient granulocytes, with premature loss of p40(phox)R105Q from phagosomes. Thus, p40(phox) binding to PtdIns(3)P is essential for phagocytosis-induced oxidant production in human neutrophils and its absence can be associated with disease.

    View details for DOI 10.1182/blood-2009-07-231498

    View details for Web of Science ID 000270595700024

    View details for PubMedID 19692703

    View details for PubMedCentralID PMC2759653

  • Efficient Induction and Expansion of Human Alloantigen-Specific CD8 Regulatory T Cells from Naive Precursors by CD40-Activated B Cells JOURNAL OF IMMUNOLOGY Zheng, J., Liu, Y., Qin, G., Chan, P., Mao, H., Lam, K., Lewis, D. B., Lau, Y., Tu, W. 2009; 183 (6): 3742-3750


    Although recent studies have focused on CD4(+) regulatory T cells (Treg), CD8(+) Treg have also been reported to play important roles in the induction and maintenance of immune tolerance. Adoptive transfer of CD8(+) Treg in rodents or induction of CD8(+) Treg in humans can prevent or treat allograft rejection and autoimmune diseases. However, no approaches have been reported for the generation of human Ag-specific CD8(+) Treg at a practical scale for clinical use. Here, we found that two novel CD8(+) T cell subsets with different levels of CD8 surface expression, CD8(high) and CD8(low), could be induced from naive CD8(+) precursors in vitro by allogeneic CD40-activated B cells, whereas only CD8(high) T cells were alloantigen-specific Treg with relatively poor alloantigen-specific cytotoxicity. Importantly, alloantigen-specific CD8(high) Treg could be induced and expanded from naive CD8(+)CD25(-) T cells at a large scale after 3 wk of culture without exogenous cytokines. These induced alloantigen-specific Treg were CD45RO(+) and CCR7(-) memory cells, and they expressed Foxp3, CD25, CD27, CD28, and CD62L. The induction and expansion of CD8(high) Treg by CD40-activated B cells were dependent on endogenously expressed IFN-gamma, IL-2, IL-4, and CTLA-4. This approach may facilitate the clinical application of CD8(+) Treg-based immunotherapy in transplantation and autoimmune diseases.

    View details for DOI 10.4049/jimmunol.0901329

    View details for Web of Science ID 000270179700025

    View details for PubMedID 19684082

  • Cationic lipid/DNA complexes (JVRS-100) combined with influenza vaccine (Fluzone (R)) increases antibody response, cellular immunity, and antigenically drifted protection VACCINE Lay, M., Callejo, B., Chang, S., Hong, D. K., Lewis, D. B., Carroll, T. D., Matzinger, S., Fritts, L., Miller, C. J., Warner, J. F., Liang, L., Fairman, J. 2009; 27 (29): 3811-3820


    Safe and effective adjuvants for influenza vaccines that could increase both the levels of neutralizing antibody, including against drifted viral subtypes, and T-cell immunity would be a major advance in vaccine design. The JVRS-100 adjuvant, consisting of DOTIM/cholesterol cationic liposome-DNA complexes, is particularly promising for vaccines that require induction of high levels of antibody and T-cell immunity, including CD8(+) cytotoxic T lymphocytes (CTL). Inclusion of protein antigens with JVRS-100 results in the induction of enhanced humoral and cell-mediated (i.e., CD4(+) and CD8(+) T cells) immune responses. The JVRS-100 adjuvant combined with a split trivalent influenza vaccine (Fluzone-sanofi pasteur) elicited increased antibody and T-cell responses in mice and non-human primates compared to vaccination with Fluzone alone. Mice vaccinated with JVRS-100-Fluzone and challenged with antigenically drifted strains of H1N1 (PR/8/34) and influenza B (B/Lee/40) viruses had higher grade protection, as measured by attenuation of weight loss and increased survival, compared to recipients of unadjuvanted vaccine. The results indicate that the JVRS-100 adjuvant substantially increases immunogenicity and protection from drifted-strain challenge using an existing influenza vaccine.

    View details for DOI 10.1016/j.vaccine.2009.04.054

    View details for Web of Science ID 000267581300003

    View details for PubMedID 19406188

    View details for PubMedCentralID PMC2690618

  • Human CD4(+) T cell recent thymic emigrants are identified by protein tyrosine kinase 7 and have reduced immune function JOURNAL OF EXPERIMENTAL MEDICINE Haines, C. J., Giffon, T. D., Lu, L., Lu, X., Tessier-Lavigne, M., Ross, D. T., Lewis, D. B. 2009; 206 (2): 275-285


    CD4(+) recent thymic emigrants (RTEs) comprise a clinically and immunologically important T cell population that indicates thymic output and that is essential for maintaining a diverse alphabeta-T cell receptor (TCR) repertoire of the naive CD4(+) T cell compartment. However, their frequency and function are poorly understood because no known surface markers distinguish them from older non-RTE naive CD4(+) T cells. We demonstrate that protein tyrosine kinase 7 (PTK7) is a novel marker for human CD4(+) RTEs. Consistent with their recent thymic origin, human PTK7(+) RTEs contained higher levels of signal joint TCR gene excision circles and were more responsive to interleukin (IL)-7 compared with PTK7(-) naive CD4(+) T cells, and rapidly decreased after complete thymectomy. Importantly, CD4(+) RTEs proliferated less and produced less IL-2 and interferon-gamma than PTK7(-) naive CD4(+) T cells after alphabeta-TCR/CD3 and CD28 engagement. This immaturity in CD4(+) RTE effector function may contribute to the reduced CD4(+) T cell immunity observed in contexts in which CD4(+) RTEs predominate, such as in the fetus and neonate or after immune reconstitution. The ability to identify viable CD4(+) RTEs by PTK7 staining should be useful for monitoring thymic output in both healthy individuals and in patients with genetic or acquired CD4(+) T cell immunodeficiencies.

    View details for DOI 10.1084/jem.20080996

    View details for Web of Science ID 000266008800004

    View details for PubMedID 19171767

    View details for PubMedCentralID PMC2646563

  • CMV-Specific T-Cell Responses in Lung Transplant Patients Using a Monocyte-Derived Dendritic Cell (MDDC) Assay. Chen, S. F., Liu, L. X., Dhillon, G., de Leon, V., Weill, D., Lewis, D. B. WILEY-BLACKWELL PUBLISHING, INC. 2009: 551
  • Efficient generation of human alloantigen-specific CD4(+) regulatory T cells from naive precursors by CD40-activated B cells BLOOD Tu, W., Lau, Y., Zheng, J., Liu, Y., Chan, P., Mao, H., Dionis, K., Schneider, P., Lewis, D. B. 2008; 112 (6): 2554-2562


    CD4(+)CD25(+)Foxp3(+) regulatory T cells (Treg) play an important role in the induction and maintenance of immune tolerance. Although adoptive transfer of bulk populations of Treg can prevent or treat T cell-mediated inflammatory diseases and transplant allograft rejection in animal models, optimal Treg immunotherapy in humans would ideally use antigen-specific rather than polyclonal Treg for greater specificity of regulation and avoidance of general suppression. However, no robust approaches have been reported for the generation of human antigen-specific Treg at a practical scale for clinical use. Here, we report a simple and cost-effective novel method to rapidly induce and expand large numbers of functional human alloantigen-specific Treg from antigenically naive precursors in vitro using allogeneic nontransformed B cells as stimulators. By this approach naive CD4(+)CD25(-) T cells could be expanded 8-fold into alloantigen-specific Treg after 3 weeks of culture without any exogenous cytokines. The induced alloantigen-specific Treg were CD45RO(+)CCR7(-) memory cells, and had a CD4(high), CD25(+), Foxp3(+), and CD62L (L-selectin)(+) phenotype. Although these CD4(high)CD25(+)Foxp3(+) alloantigen-specific Treg had no cytotoxic capacity, their suppressive function was cell-cell contact dependent and partially relied on cytotoxic T lymphocyte antigen-4 expression. This approach may accelerate the clinical application of Treg-based immunotherapy in transplantation and autoimmune diseases.

    View details for DOI 10.1182/blood-2008-04-152041

    View details for Web of Science ID 000259088000052

    View details for PubMedID 18599794

    View details for PubMedCentralID PMC2532818

  • Focus on FOCIS: The continuing diagnostic challenge of autosomal recessive chronic granulomatous disease CLINICAL IMMUNOLOGY Yu, G., Hong, D. K., Dionis, K. Y., Rae, J., Heyworth, P. G., Curnutte, J. T., Lewis, D. B. 2008; 128 (2): 117-126


    Chronic granulomatous disease (CGD) is a primary immunodeficiency of defective neutrophil oxidative burst activity due to mutations in the genes CYBA, NCF-1, NCF-2, and CYBB, which respectively encode the p22-phox, p47-phox, p67-phox, and gp91-phox subunits. CGD usually presents in early childhood with recurrent or severe infection with catalase-positive bacteria and fungi. We present an unusual case of CGD in which Burkholderia cepacia lymphadenitis developed in a previously healthy 10-year-old girl. Flow cytometric analysis of dihydrorhodamine (DHR)-labeled neutrophils performed by a CLIA-approved outside reference laboratory was reported as normal. However, we found that this patient's neutrophil oxidative burst activity in DHR assays was substantially reduced but not absent. A selective decrease in intracellular staining for p67-phox suggested the diagnosis of autosomal recessive CGD due to NCF-2 gene mutations, and a novel homozygous and hypomorphic NCF-2 gene mutation was found. The potential mechanisms for this delayed and mild presentation of CGD are discussed.

    View details for DOI 10.1016/j.clim.2008.05.008

    View details for PubMedID 18625437

  • Maternal T-cell engraftment associated with severe hemophagocytosis of the bone marrow in untreated X-linked severe combined immunodeficiency JOURNAL OF PEDIATRIC HEMATOLOGY ONCOLOGY Dvorak, C. C., Sandford, A., Fong, A., Cowan, M. J., George, T. L., Lewis, D. B. 2008; 30 (5): 396-400


    Maternal engraftment of T cells in severe combined immunodeficiency can lead to graft-versus-host disease of the skin and liver. We report the case of an infant with X-linked severe combined immunodeficiency, confirmed by DNA sequencing of the common gamma chain gene locus, in which this disorder's characteristic peripheral lymphocyte phenotype [T(-)B(+)NK(-)] was obscured by the postnatal onset of hemophagocytic syndrome that included severe B-cell lymphopenia, neutropenia, and anemia. Hemophagocytosis was most likely owing to maternal graft-versus-host disease, as perforin-expressing CD8 T cells, presumably of maternal origin, were prominent in the bone marrow and there was no concurrent severe infection.

    View details for PubMedID 18458578

  • Immunotherapy of Acute Leukemia with Cationic Lipid DNA Complexes (JVRS-100) Chang, S., Herse, Z., Claxton, D., Giffon, T., Lewis, D., Fairman, J. FEDERATION AMER SOC EXP BIOL. 2008
  • Inverse relationship between innate and acquired immune responsiveness in mice initially dosed with cationic lipid DNA complexes (JVRS-100) Lay, M., Callejo, B., Giffon, T., Lewis, D., Chang, S., Fairman, J. FEDERATION AMER SOC EXP BIOL. 2008
  • Asymmetric dimethylarginine and cardiac allograft vasculopathy progression: Modulation by sirolimus TRANSPLANTATION Potena, L., Fearon, W. F., Sydow, K., Holweg, C., Luikart, H., Chin, C., Weisshaar, D., Mocarski, E. S., Lewis, D. B., Valantine, H. A., Cooke, J. P. 2008; 85 (6): 827-833


    Cardiac allograft vasculopathy (CAV) is a major cause of death after heart transplantation (HT). The reduced bioavailability of endothelium-derived nitric oxide may play a role in endothelial vasodilator dysfunction and thus in the structural changes characterizing CAV. A potential contributor to endothelial pathobiology is asymmetric dimethylarginine (ADMA), an endogenous nitric oxide synthase inhibitor. It was hypothesized that ADMA concentrations may influence CAV progression during the first postoperative year.Thirty-two consecutive HT recipients underwent intravascular ultrasound evaluation at month 1 and year 1 after HT. Immunosuppression included mycophenolate mofetil (MMF, n=16) and sirolimus (n=16). Change in intimal volume greater than the median and vascular remodeling were major outcome measures.Plasma ADMA levels were associated with subsequent development of intimal hyperplasia (risk ratio [95% confidence interval] =2.72 [1.06-6.94]; P=0.038), and plasma ADMA levels greater than 0.70 micromol/L most accurately identified patients who would have developed intimal hyperplasia. However, ADMA levels did not correlate with negative coronary remodeling. Treatment with sirolimus, as compared with MMF, was associated with significantly lower ADMA levels (0.65+/-0.12 vs. 0.77+/-0.10 micromol/L; P<0.01) and less intimal hyperplasia (risk ratio [95% confidence interval] = 0.08 [0.01-0.56]; P=0.01).Elevated plasma ADMA is associated with coronary intimal hyperplasia, supporting the importance of nitric oxide synthase inhibition in CAV pathogenesis. Treatment with sirolimus (rather than MMF) is associated with lower ADMA levels and reduced risk of accelerated CAV.

    View details for DOI 10.1097/TP.0b013e318166a3a4

    View details for PubMedID 18360263

  • A novel mutation associated with autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy ANNALS OF ALLERGY ASTHMA & IMMUNOLOGY Bhui, R. D., Lewis, D. B., Nadeau, K. C. 2008; 100 (2): 169-169

    View details for Web of Science ID 000253136500012

    View details for PubMedID 18320920

  • Influenza A virus elevates active cathepsin B in primary murine DC INTERNATIONAL IMMUNOLOGY Burster, T., Giffon, T., Dahl, M. E., Bjorck, P., Bogy, M., Weber, E., Mahmood, K., Lewis, D. B., Mellins, E. D. 2007; 19 (5): 645-655


    Dendritic cells (DCs) act as a first-line recognition system for invading pathogens, such as influenza A. The interaction of DC with influenza A virus results in DC activation via endosomal Toll-like receptors and also leads to presentation of viral peptides on MHC class II molecules. Prior work demonstrated that influenza A virus (A/HKx31; H3N2) infection of BALB/c mice activates lung DCs for antigen presentation, and that the enhanced function of these cells persists long after viral clearance and resolution of the virus-induced inflammatory response. Whether influenza A virus has acute or longer-lasting effects on the endo/lysosomal antigen-processing machinery of DCs has not been studied. Here, we show that antigen presentation from intact protein antigen, but not peptide presentation, results in increased T cell stimulation by influenza-exposed lung DCs, suggesting increased antigen processing/loading in these DCs. We find that cathepsin (Cat) B levels and activity are substantially up-regulated in murine lung DCs, harvested 30 days after A/HKx31 infection. CatB levels and activity are also increased in murine splenic and bone marrow-derived DCs, following short-term in vitro exposure to UV-inactivated influenza A virus. Modest effects on CatX are also seen during in vivo and in vitro exposure to influenza A virus. Using a cell permeable Cat inhibitor, we show Cats in influenza-exposed DCs to be functional and required for generation of a T cell epitope from intact ovalbumin. Our findings indicate that influenza A virus affects the MHC class II antigen-processing pathway, an essential pathway for CD4(+) T cell activation.

    View details for DOI 10.1093/intimm/dxm030

    View details for Web of Science ID 000246964500007

    View details for PubMedID 17446210

  • [Coronary allograft vasculopathy: pathophysiological interaction between the immune system, infections and metabolic syndrome]. Giornale italiano di cardiologia (2006) Potena, L., Ferrara, R., Mocarski, E. S., Lewis, D. B., Cooke, J. P., Grigioni, F., Coccolo, F., Magnani, G., Fallani, F., Magelli, C., Valantine, H. A., Branzi, A. 2007; 8 (2): 73-82


    Cardiac allograft vasculopathy is still the main cause of long-term graft loss after heart transplantation. Indeed, recent advances in immunosuppression management led to a significant improvement in short-term survival, while long-term death rate did not change significantly in the last 20 years. In this paper, we will review the latest advances in the understanding of this peculiar form of atherosclerosis, focusing on the mechanisms that can be potentially targeted by specific therapeutic interventions.

    View details for PubMedID 17402351

  • Humoral and cellular immune responses in children given annual immunization with trivalent inactivated influenza vaccine PEDIATRIC INFECTIOUS DISEASE JOURNAL Zeman, A. M., Holmes, T. H., Stamatis, S., Tu, W., He, X., Bouvier, N., Kemble, G., Greenberg, H. B., Lewis, D. B., Arvin, A. M., Dekker, C. L. 2007; 26 (2): 107-115


    There have been no prior reports of the frequency of circulating influenza-specific, interferon gamma-producing memory CD4+ and CD8+ T-cells in healthy children who have received multiple influenza immunizations.We evaluated 21 previously immunized children, ages 3 to 9 years, before and 1 month after administration of trivalent inactivated influenza vaccine. Frequencies of influenza-specific CD4+ and CD8+ T-cells stimulated with trivalent inactivated influenza vaccine or A/Panama (H3N2) virus were determined by flow cytometry, and antibody responses to vaccine strains and a drifted H3N2 strain were measured by hemagglutination inhibition assay and neutralizing antibody assays.Mean change in CD4+ and in CD8+ T-cell frequencies after immunization was 0.01% (P > 0.39) with postimmunization CD4+ frequencies higher than CD8+ frequencies. Children with more previous vaccinations had a higher baseline frequency of CD4+ T-cells (P = 0.0002) but a smaller increase or even a decline from baseline after immunization (P = 0.003). An association between age and change in frequency was not detected. Baseline geometric mean titers (GMTs) and seroprotection rates were significantly higher in older children against A/Panama (neutralizing baseline GMT, P = 0.0488) and A/New Caledonia (hemagglutination inhibition baseline GMT and seroprotection, P < 0.0297). Baseline GMTs against B/Hong Kong were not associated with age or quantity of prior vaccinations.These findings suggest that children may plateau in CD4+ T-cell responses to influenza antigens with repeated exposures and that the number of exposures may play a large role in building a memory CD4+ T-cell response to influenza A, perhaps independently from age.

    View details for DOI 10.1097/01.inf.0000253251.03785.9b

    View details for Web of Science ID 000243985700002

    View details for PubMedID 17259871

  • A novel missense mutation in CYBB gene in chronic granulomatous disease 63rd Annual Meeting of the American-Academy-of-Allergy-Asthma-and-Immunology DeMore, J. P., Dong, Y., Lewis, D. B., Gern, J. E., Seroogy, C. M. MOSBY-ELSEVIER. 2007: S257–S257
  • Severe hypogammaglobulinemia and absent B cells in an adult patient: A case report Iyengar, S. R., Blackbum, B. G., Lewis, D. B. MOSBY-ELSEVIER. 2007: S256
  • Cellular immune responses in children and adults receiving inactivated or live attenuated influenza vaccines JOURNAL OF VIROLOGY He, X., Holmes, T. H., Zhang, C., Mahmood, K., Kemble, G. W., Lewis, D. B., Dekker, C. L., Greenberg, H. B., Arvin, A. M. 2006; 80 (23): 11756-11766


    The patterns of cellular immune responses induced by live attenuated influenza vaccine (LAIV) versus those of the trivalent inactivated influenza vaccine (TIV) have not been studied extensively, especially in children. The goals of this study were to evaluate the effects of TIV and LAIV immunization on cellular immunity to live influenza A virus in children and adults and to explore factors associated with variations in responses to influenza vaccines among individuals. A gamma interferon (IFN-gamma) flow cytometry assay was used to measure IFN-gamma-producing (IFN-gamma+) NK and T cells in peripheral blood mononuclear cell cultures stimulated with a live influenza A virus strain before and after LAIV or TIV immunization of children and adults. The mean percentages of influenza A virus-specific IFN-gamma+ CD4 and CD8 T cells increased significantly after LAIV, but not TIV, immunization in children aged 5 to 9 years. No increases in the mean levels of influenza A virus-reactive IFN-gamma+ T cells and NK cells were observed in adults given LAIV or TIV. TIV induced a significant increase in influenza A virus-reactive T cells in 6-month- to 4-year-old children; LAIV was not evaluated in this age group. The postvaccination changes (n-fold) in the percentages of influenza A virus-reactive IFN-gamma+ T and NK cells in adults were highly variable and correlated inversely with the prevaccination percentages, in particular with that of the CD56(dim) NK cell subset. In conclusion, our findings identify age, type of vaccine, and prevaccination levels of immune reactivity to influenza A virus as factors significantly associated with the magnitude of cellular immune responses to influenza vaccines.

    View details for DOI 10.1128/JVI.01460-06

    View details for Web of Science ID 000242222200032

    View details for PubMedID 16971435

    View details for PubMedCentralID PMC1642596

  • T-cell immunity to subclinical cytomegalovirus infection reduces cardiac allograft disease CIRCULATION Tu, W., Potena, L., Stepick-Biek, P., Liu, L., Dionis, K. Y., Luikart, H., Fearon, W. F., Holmes, T. H., Chin, C., Cooke, J. P., Valantine, H. A., Mocarski, E. S., Lewis, D. B. 2006; 114 (15): 1608-1615


    Asymptomatic cytomegalovirus (CMV) replication is frequent after cardiac transplantation in recipients with pretransplantation CMV infection. How subclinical viral replication influences cardiac allograft disease remains poorly understood, as does the importance of T-cell immunity in controlling such replication.Thirty-nine cardiac recipients who were pretransplantation CMV antibody positive were longitudinally studied for circulating CMV-specific CD4 and CD8 T-cell responses, CMV viral load in blood neutrophils, and allograft rejection during the first posttransplantation year. Nineteen of these recipients were also analyzed for changes of coronary artery intimal, lumen, and whole-vessel area. All recipients received early prophylactic therapy with ganciclovir. No recipients developed overt CMV disease. Those with detectable levels of CMV-specific CD4 T cells in the first month after transplantation were significantly protected from high mean and peak posttransplantation viral load (P<0.05), acute rejection (P<0.005), and loss of allograft coronary artery lumen (P<0.05) and of whole-vessel area (P<0.05) compared with those who lacked this immune response. The losses of lumen and vessel area were both significantly correlated with the time after transplantation at which a CD4 T-cell response was first detected (P<0.05) and with the cumulative graft rejection score (P<0.05).The early control of subclinical CMV replication after transplantation by T-cell immunity may limit cardiac allograft rejection and vascular disease. Interventions to increase T-cell immunity might be clinically useful in limiting these adverse viral effects.

    View details for DOI 10.1161/CIRCULATIONAHA.105.607549

    View details for PubMedID 17015794

  • Cutting edge: Decreased accumulation and regulatory function of CD4(+)CD25(high) T cells in human STAT 5b deficiency JOURNAL OF IMMUNOLOGY Cohen, A. C., Nadeau, K. C., Tu, W., Hwa, V., Dionis, K., Bezrodnik, L., Teper, A., Gaillard, M., Heinrich, J., Krensky, A. M., Rosenfeld, R. G., Lewis, D. B. 2006; 177 (5): 2770-2774


    We show that STAT5b is important for the in vivo accumulation of CD4+ CD25(high) T cells with regulatory cell function. A patient homozygous for a missense A630P STAT5b mutation displayed immune dysregulation and decreased numbers of CD4+ CD25(high) T cells. STAT5b(A630P/A630P) CD4+ CD25(high) T cells had low expression of forkhead box P3 and an impaired ability to suppress the proliferation of or to kill CD4+ CD25- T cells. Expression of CD25, a component of the high-affinity IL-2R, was also reduced in response to IL-2 or after in vitro propagation. The impact of the STAT5b mutation was selective in that IL-2-mediated up-regulation of the common gamma-chain cytokine receptor and perforin, and activation-induced expressions of CD154 and IFN-gamma were normal. These results indicate that STAT5b propagates an important IL-2-mediated signal for the in vivo accumulation of functional regulatory T cells.

    View details for Web of Science ID 000240002800007

    View details for PubMedID 16920911

  • Acute rejection and cardiac allograft vascular disease is reduced by suppression of subclinical cytomegalovirus infection TRANSPLANTATION Potena, L., Holweg, C. T., Chin, C., Luikart, H., Weisshaar, D., Narasimhan, B., Fearon, W. F., Lewis, D. B., Cooke, J. P., Mocarski, E. S., Valantine, H. A. 2006; 82 (3): 398-405


    Anticytomegalovirus (CMV) prophylaxis prevents the acute disease but its impact on subclinical infection and allograft outcome is unknown. We sought to determine whether CMV prophylaxis administered for three months after heart transplant would improve patient outcomes.This prospective cohort study of 66 heart transplant recipients compared aggressive CMV prophylaxis (n = 21, CMV hyperimmune globulin [CMVIG] plus four weeks of intravenous ganciclovir followed by two months of valganciclovir); with standard prophylaxis (n = 45, intravenous ganciclovir for four weeks). Prophylaxis was based on pretransplant donor (D) and recipient (R) CMV serology: R-/D+ received aggressive prophylaxis; R+ received standard prophylaxis. Outcome measures were: CMV infection assessed by DNA-polymerase chain reaction on peripheral blood polymorphonuclear leukocytes, acute rejection, and cardiac allograft vascular disease (CAV) assessed by intravascular ultrasound. All patients completed one year of follow-up. RESULTS.: CMV infection was subclinical in all but four patients (two in each group). Aggressively treated patients had a lower incidence of CMV infection (73 +/- 10% vs. 94 +/- 4%; P = 0.038), and an independent reduced relative risk for acute rejection graded > or =3A (relative risk [95% CI] = 0.55 [0.26-0.96]; P = 0.03), as compared with the standard prophylaxis group. Aggressively prophylaxed patients also showed a slower progression of CAV, in terms of coronary artery lumen loss (lumen volume change=-21 +/- 13% vs. -10+/-14%; P = 0.05); and vessel shrinkage (vessel volume change = -15 +/- 11% vs. -3 +/- 18%; P = 0.03).Prolonged (val)ganciclovir plus CMVIG reduces viral levels, acute rejection, and allograft vascular disease, suggesting a role for chronic subclinical infection in the pathophysiology of the most common diseases affecting heart transplant recipients.

    View details for DOI 10.1097/

    View details for PubMedID 16906040

  • Newborn immunology: relevance to the clinician. Current problems in pediatric and adolescent health care Lewis, D. B., Gern, J. E., Hill, H. R., Friedlander, S. L., La Pine, T. R., Lemanske, R. F., Stiehm, E. R. 2006; 36 (5): 189-204

    View details for PubMedID 16631097

  • Impaired allogeneic activation and T-helper differentiation of human cord blood naive CD4 T cells BIOLOGY OF BLOOD AND MARROW TRANSPLANTATION Chen, L., Cohen, A. C., Lewis, D. B. 2006; 12 (2): 160-171


    CD4 T cells, particularly those of the T-helper 1 (Th1) subset, are important effectors in alloimmune diseases, such as graft-versus-host disease, and in controlling infections with intracellular pathogens. Thus, it is plausible that impaired neonatal CD4 T-cell immunity might contribute to the low incidence of acute graft-versus-host disease after allogeneic transplantation of hematopoietic stem cells using cord blood (CB) compared with adult sources of hematopoietic stem cells. In support of this hypothesis, we found that CB naive CD4 T cells had reduced activation and impaired early Th1 differentiation compared with adult peripheral blood naive CD4 T cells after stimulation by allogeneic dendritic cells derived from adult monocytes. Early Th1 polarization was dependent on interleukin-12 and CD154, and CB CD4 T cell/dendritic cell co-cultures had impaired expression of both proteins. CB naive CD4 T cells had low basal levels of signal transduction and activation of transcription 4 messenger RNA and protein, and, after alloantigen stimulation, reduced interleukin-12-induced signal transduction and activation of transcription 4 tyrosine phosphorylation, compared with adult peripheral blood naive T cells. Lastly, FoxP3 protein expression, a marker for regulatory CD25(high) CD4 T cells, was lower for naive CD4 T cells of CB compared with those of adult peripheral blood, which argued against increased T-regulatory activity as a mechanism for the decreased Th1 differentiation of CB CD4 T cells. Together, these intrinsic limitations in T-cell activation and Th1 differentiation may impair the ability of T cells in CB and the neonate to respond to allogeneic or infectious challenges.

    View details for DOI 10.1016/j.bbmt.2005.10.027

    View details for Web of Science ID 000235284900005

    View details for PubMedID 16443514

  • Prevention of subclinical CMV infection reduces cardiac allograft disease progression by positively affecting coronary remodeling Potena, L., Fearon, W., Holweg, C., Luikart, H., Chin, C., Cooke, J., Lewis, D., Mocarski, E., Valantine, H. ELSEVIER SCIENCE INC. 2006: S138–S139
  • Chemotactic responses of IL-4-, IL-10-, and IFN-gamma-producing CD4(+) T cells depend on tissue origin and microbial stimulus JOURNAL OF IMMUNOLOGY Debes, G. F., Dahl, M. E., Mahiny, A. J., Bonhagen, K., Campbell, D. J., Siegmund, K., Erb, K. J., Lewis, D. B., Kamradt, T., Hamann, A. 2006; 176 (1): 557-566


    Th1- and Th2-polarized immune responses are crucial in the defense against pathogens but can also promote autoimmunity and allergy. The chemokine receptors CXCR3 and CCR4 have been implicated in differential trafficking of IFN-gamma- and IL-4-producing T cells, respectively, but also in tissue and inflammation-specific homing independent of cytokine responses. Here, we tested whether CD4+ T cells isolated from murine tissues under homeostatic or inflammatory conditions exhibit restricted patterns of chemotactic responses that correlate with their production of IFN-gamma, IL-4, or IL-10. In uninfected mice, IL-4-producing T cells preferentially migrated to the CCR4 ligand, CCL17, whereas IFN-gamma-expressing T cells as well as populations of IL-4+ or IL-10+ T cells migrated to the CXCR3 ligand, CXCL9. All cytokine-producing T cell subsets strongly migrated to the CXCR4 ligand, CXCL12. We assessed chemotaxis of T cells isolated from mice infected with influenza A virus or the nematode Nippostrongylus brasiliensis, which induce a strong Th1 or Th2 response in the lung, respectively. Unexpectedly, the chemotactic responses of IL-4+ T cells and T cells expressing the immunosuppressive cytokine IL-10 were influenced not only by the strongly Th1- or Th2-polarized environments but also by their anatomical localization, i.e., lung or spleen. In contrast, IFN-gamma+ T cells exhibited robust chemotaxis toward CXCL9 and had the most consistent migration pattern in both infection models. The results support a model in which the trafficking responses of many effector and regulatory T cells are regulated as a function of the infectious and tissue environments.

    View details for Web of Science ID 000234262600064

    View details for PubMedID 16365450

  • Decreased generation and function of CD4+CD25(hi) T regulatory cells in human STAT5b deficiency. Cohen, A., Nadeau, K., Tu, W., Hwa, V., Dionis, K., Bezrodnik, L., Teper, A., Gaillard, M., Heinrich, J., Krensky, A., Rosenfeld, R., Lewis, D. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2006: S135
  • Avian flu to human influenza ANNUAL REVIEW OF MEDICINE Lewis, D. B. 2006; 57: 139-154


    Influenza A viral infection causes substantial annual morbidity and mortality worldwide, particularly for infants, the elderly, and the immunocompromised. The virus mainly replicates in the respiratory tract and is spread by respiratory secretions. A growing concern is the recent identification of H5N1 strains of avian influenza A in Asia that were previously thought to infect only wild birds and poultry, but have now infected humans, cats, pigs, and other mammals, often with fatal results, in an ongoing outbreak. A human pandemic with H5N1 virus could potentially be catastrophic because most human populations have negligible antibody-mediated immunity to the H5 surface protein and this viral subtype is highly virulent. Whether an H5N1 influenza pandemic will occur is likely to hinge on whether the viral strains involved in the current outbreak acquire additional mutations that facilitate efficient human-to-human transfer of infection. Although there is no historical precedent for an H5N1 avian strain causing widespread human-to-human transmission, some type of influenza A pandemic is very likely in the near future. The possibility of an H5N1 influenza pandemic has highlighted the many current limitations of treatment with antiviral agents and of vaccine production and immunogenicity. Future vaccine strategies that may include more robust induction of T-cell responses, such as cytotoxic T lymphocytes, may provide better protection than is offered by current vaccines, which rely solely or mainly on antibody neutralization of infection.

    View details for DOI 10.1146/

    View details for Web of Science ID 000235981900009

    View details for PubMedID 16409141

  • Transient deficiencies of T-cell-mediated immunity in the neonate Course on Infection and Immunity in Children 2005 Randolph, D. A., Lewis, D. B. SPRINGER-VERLAG BERLIN. 2006: 55–69

    View details for Web of Science ID 000238206400006

    View details for PubMedID 16802619

  • Decreased generation and function of CD4+CD25(hi) T regulatory cells in human STAT5b deficiency. Cleary, A., Nadeau, K., Tu, W. W., Hwa, Dionis, K. Y., Bezrodnik, L., Teper, A., Gaillard, M., Heinrich, J. J., Krensky, A. M., Rosenfeld, R. G., Lewis, D. B. AMER SOC HEMATOLOGY. 2005: 227A
  • Thymic stromal lymphopoietin as a key initiator of allergic airway inflammation in mice NATURE IMMUNOLOGY Zhou, B. H., Comeau, M. R., De Smedt, T., Liggitt, H. D., Dahl, M. E., Lewis, D. B., Gyarmati, D., Aye, T., Campbell, D. J., Ziegler, S. F. 2005; 6 (10): 1047-1053


    The cytokine thymic stromal lymphopoietin (TSLP) has been linked to human allergic inflammatory diseases. We show here that TSLP expression was increased in the lungs of mice with antigen-induced asthma, whereas TSLP receptor-deficient mice had considerably attenuated disease. Lung-specific expression of a Tslp transgene induced airway inflammation and hyperreactivity characterized by T helper type 2 cytokines and increased immunoglobulin E. The lungs of Tslp-transgenic mice showed massive infiltration of leukocytes, goblet cell hyperplasia and subepithelial fibrosis. TSLP was capable of activating bone marrow-derived dendritic cells to upregulate costimulatory molecules and produce the T helper type 2 cell-attracting chemokine CCL17. These findings suggest that TSLP is an important factor necessary and sufficient for the initiation of allergic airway inflammation.

    View details for DOI 10.1038/ni1247

    View details for Web of Science ID 000232027200024

    View details for PubMedID 16142237

  • Subclinical cytomegalovirus infection modulates systemic inflammation and graft coronary remodeling one year after heart transplantation. Potena, L., Biadi, O., Holweg, C., Fearon, W., Cooke, J., Lewis, D., Mocarski, E., Valantine, H. BLACKWELL PUBLISHING. 2005: 382
  • Cytomegalovirus-specific CD4 T-cell immunity is associated with protection from human cardiac allograft rejection and negative coronary-artery remodeling. Tu, W., Potena, L., Stepick-Biek, P., Liu, L., Dionis, K. Y., Bashyam, L., Fearon, W., Valantine, T. A., Mocarski, E. S., Lewis, D. B. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2005: S277
  • ICOS engagement of CD4 T-cells following pulmonary influenza infection: Evidence for negative regulation of T-cell effector function. Dahl, M. E., Coyle, A., Lewis, D. B. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2005: S35
  • Immunodeficiency associated with a STAT-5b mutation and growth hormone insensitivity. Cleary, A. M., Hwa, Tu, W. W., Dionis, K., Bezrodnik, L., Teper, A., Gaillard, M., Heinrich, J. J., Rosenfeld, R. G., Lewis, D. B. AMER SOC HEMATOLOGY. 2004: 374A
  • The role of IL-12 and IL-23 in the generation and maintenance of memory CD4+T cells Cleary, A. M., Lewis, D. B. AMER SOC HEMATOLOGY. 2004: 728A
  • Systemic inflammation links impaired glucose metabolism to cardiac allograft vasculopathy development 77th Scientific Meeting of the American-Heart-Association Potena, L., Biadi, O., Fearon, W. B., Holweg, C. T., Luikart, H. I., Mocarski, E. S., Lewis, D. B., Valantine, H. A. LIPPINCOTT WILLIAMS & WILKINS. 2004: 753–53
  • Antiviral CD8 T cells in the control of primary human cytomegalovirus infection in early childhood 40th Annual Meeting of the Infectious-Diseases-Society-of-America Chen, S. F., Tu, W. W., Sharp, M. A., Tongson, E. C., He, X. S., Greenberg, H. B., Holmes, T. H., Wang, Z. T., Kemble, G., Manganello, A. M., Adler, S. P., Dekker, C. L., Levvis, D. B., Arvin, A. M. UNIV CHICAGO PRESS. 2004: 1619–27


    Human cytomegalovirus (CMV) establishes persistent infection, with control of replication thought to be mediated by CMV-specific CD8 T cells. Primary CMV infection commonly affects young children and causes prolonged viral shedding in saliva and urine. We investigated whether this virus-host interaction pattern reflects a developmental deficiency of antiviral CD8 T cell-mediated immunity during childhood. CMV-specific CD8 T cell responses in asymptomatic children with active infection were not different from adults with recent or long-term infection in frequency and functional analyses. High urine CMV concentrations were detected, despite these CMV-specific CD8 T cell responses. We conclude that delayed resolution of primary CMV infection in young children is not caused by a deficient CMV-specific CD8 T cell response. Because these healthy children continue to have local CMV replication, we suggest that CD8 T cells may function primarily to prevent symptomatic, disseminated disease.

    View details for PubMedID 15116298

  • Cytomegalovirus infectious burden is proportional to cardiac allograft vasculopathy in heart transplant recipients 53rd Annual Scientific Session of the American-College-of-Cardiology Potena, L., Magelli, C., Ortolani, P., Fearon, W. F., Grigioni, F., Magnani, G., Coccolo, F., Yeung, A. C., Luikart, H. I., Hunt, S. A., Mocarski, E. S., Cooke, J. P., Lewis, D. B., Branzi, A., Valantine, H. A. ELSEVIER SCIENCE INC. 2004: 185A–185A
  • Viral-induced T helper type 1 responses enhance allergic disease by effects on lung dendritic cells NATURE IMMUNOLOGY Dahl, M. E., Dabbagh, K., Liggitt, D., Kim, S., Lewis, D. B. 2004; 5 (3): 337-343


    It is widely accepted that T helper type 1 (T(H)1) cytokines such as interferon-gamma (IFN-gamma) antagonize allergic diseases mediated by T(H)2 cytokines. The 'hygiene hypothesis' has also proposed that decreased childhood exposure to pathogen-derived T(H)1 cytokines may underlie the recent increased prevalence of asthma, a T(H)2-mediated disease. We show here that influenza A viral infection, which induces large amounts of intrapulmonary IFN-gamma production, unexpectedly enhanced later allergen-specific asthma and promoted dual allergen-specific T(H)1 and T(H)2 responses. Pulmonary dendritic cells obtained from the lung after viral clearance and resolution of acute inflammation conferred enhanced allergic disease and concurrent T(H)1 and T(H)2 immune responses, and these effects were dependent on IFN-gamma secreted during the acute viral infection. Thus, respiratory viral infection and the acute T(H)1 response can positively regulate T(H)2-dependent allergic pulmonary disease in vivo, at least in part, by altering pulmonary dendritic cell function.

    View details for DOI 10.1038/ni1041

    View details for Web of Science ID 000189344600017

    View details for PubMedID 14973436

  • Persistent and selective deficiency of CD4(+) T cell immunity to cytomegalovirus in immunocompetent young children JOURNAL OF IMMUNOLOGY Tu, W. W., Chen, S., Sharp, M., Dekker, C., Manganello, A. M., Tongson, E. C., Maecker, H. T., Holmes, T. H., Wang, Z. T., Kemble, G., Adler, S., Arvin, A., Lewis, D. B. 2004; 172 (5): 3260-3267


    Healthy young children who acquire CMV have prolonged viral shedding into the urine and saliva, but whether this is attributable to limitations in viral-specific immune responses has not been explored. In this study, we found that otherwise immunocompetent young children after recent primary CMV infection accumulated markedly fewer CMV-specific CD4(+) T cells that produced IFN-gamma than did adults. These differences in CD4(+) T cell function persisted for more than 1 year after viral acquisition, and did not apply to CMV-specific IFN-gamma production by CD8(+) T cells. The IFN-gamma-producing CD4(+) T cells of children or adults that were reactive with CMV Ags were mainly the CCR7(low) cell subset of memory (CD45R0(high)CD45RA(low)) cells. The decreased IFN-gamma response to CMV in children was selective, because their CCR7(low) memory CD4(+) T cells and those of adults produced similar levels of this cytokine after stimulation with staphylococcal enterotoxin B superantigen. CD4(+) T cells from children also had reduced CMV-specific IL-2 and CD154 (CD40 ligand) expression, suggesting an early blockade in the differentiation of viral-specific CD4(+) T cells. Following CMV acquisition, children, but not adults, persistently shed virus in urine, and this was observable for at least 29 mo postinfection. Thus, CD4(+) T cell-mediated immunity to CMV in humans is generated in an age-dependent manner, and may have a substantial role in controlling renal viral replication and urinary shedding.

    View details for PubMedID 14978134

  • Asymptomatic cytomegalovirus activation leads to acute rejection in heart transplant recipients despite anti-viral prophylaxis. 5th American Transplant Congress Potena, L., Holweg, C., Luikart, H. I., Mocarski, E. S., Cooke, J. P., Lewis, D. B., Valantine, H. A. WILEY-BLACKWELL. 2004: 453–453
  • Decreased CD154 expression by neonatal CD4(+) T cells is due to limitations in both proximal and distal events of T cell activation INTERNATIONAL IMMUNOLOGY JULLIEN, P., Cron, R. Q., Dabbagh, K., Cleary, A., Chen, L., Tran, P., Stepick-Biek, P., Lewis, D. B. 2003; 15 (12): 1461-1472


    Neonatal CD4(+) T cells express less CD154 protein and mRNA than adult CD4(+) T cells after activation by calcium ionophore and phorbol ester, but the mechanism for this reduced expression and its relevance to the primary immune response remain unclear. We compared expression of CD154 protein and mRNA and CD154 gene promoter activity by purified naive (CD45RA(high)CD45RO(low)) neonatal and adult CD4(+) T cells after activation by calcium ionophore (ionomycin) and phorbol myristate acetate (PMA) treatment or by engagement of alphabeta TCR-CD3 complex. Substantial and consistent reductions in expression by neonatal cells were found in all cases and were paralleled by decreased CD154-dependent activation of a B cell line. CD69 expression by neonatal CD4(+) T cells after alphabeta TCR-CD3 engagement was also reduced compared to adult cells, which suggested that limitations in activation-induced signaling by neonatal CD4(+) T cells occurred at a point upstream of where the signaling pathways leading to CD154 and CD69 expression diverge. Decreased CD154 expression by neonatal cells after alphabeta TCR-CD3 engagement was paralleled by a lower free intracellular calcium concentration, a key event for CD154 gene transcription. Reduced CD154 promoter activity by neonatal cells persisted when proximal signaling events were bypassed using ionomycin and PMA, suggesting an additional and more distal mechanism for decreased transcription. In contrast, CD154 mRNA stability was similar in neonatal and adult cells after either ionomycin and PMA stimulation or engagement of the alphabeta TCR-CD3 complex. We conclude that decreased CD154 production by neonatal CD4(+) T cells is due to limitations in both proximal and distal activation events, which together ultimately limit CD154 gene transcription.

    View details for DOI 10.1093/intimm/dxg145

    View details for Web of Science ID 000187225000007

    View details for PubMedID 14645155

  • Identification of a partial autosomal dominant form of IL-12R beta 1 chain deficiency. Cleary, A. M., Hwang, N., Lewis, D. B. AMER SOC HEMATOLOGY. 2003: 280A
  • A block in Th1 pathway differentiation presenting as recurrent viral infections. Cleary, A. M., Hwang, N., Marincola, J., Zloof, J., Lewis, D. B. AMER SOC HEMATOLOGY. 2003: 48B
  • Toll-like receptors and T-helper-1/T-helper-2 responses CURRENT OPINION IN INFECTIOUS DISEASES Dabbagh, K., Lewis, D. B. 2003; 16 (3): 199-204


    Toll-like receptors (TLRs) are a family of pattern recognition receptors that are activated by specific components of microbes and certain host molecules. They constitute the first line of defense against many pathogens and play a crucial role in the function of the innate immune system. Recently, TLRs were observed to influence the development of adaptive immune responses, presumably by activating antigen-presenting cells. This has important implications for our understanding of how the host tailors its immune response as a function of specific pathogen recognition. The present review discusses the recent studies that demonstrate the role of TLRs in the regulation of adaptive T-helper-1 (Th1) and Th2 responses, and the mechanisms by which the effects are carried out.Most studies have focused on the role of TLRs and components of their signaling pathways in the control of Th1-type immune responses, and on the implications for their use as antimicrobial agents, such as adjuvants in vaccines, or to treat or prevent the Th2-type dominated immune responses seen in allergies. TLR-deficient mice have been described and used to come to these conclusions. Although controversial, there is also evidence that TLRs may be important for Th2-type responses, possibly by augmenting the overall maturity of dendritic cells.A greater understanding of the processes by which TLRs regulate adaptive immunity may yield not only improved ways to treat infectious diseases but also new approaches to the treatment and prevention of allergic and certain autoimmune disorders.

    View details for DOI 10.1097/01.qco.0000073767.11390.47

    View details for Web of Science ID 000183434100003

    View details for PubMedID 12821808

  • Deficient cytomegalovirus-specific Th1 responses in young children with urine virus shedding 90th Annual Meeting of the American-Association-for-Immunologists Tu, W. W., Chen, S. R., Sharp, M., Dekker, C., Manganello, A. M., Maecker, H., Holmes, T. H., Kemble, G., Adler, S., Arvin, A., Lewis, D. B. FEDERATION AMER SOC EXP BIOL. 2003: C301–C301
  • Different patterns of migration and expansion of blood and marrow CD4 T cells in lymphoid and non-lymphoid tissues result in a different capacity to induce graft-vs-host disease Zeng, D. F., Liu, Y. P., Dutt, S., Li, Z. Q., Higgins, J., Lewis, D. B., Strober, S. FEDERATION AMER SOC EXP BIOL. 2003: C59
  • Impaired accumulation and function of memory CD4 T cells in human IL-12 receptor beta 1 deficiency JOURNAL OF IMMUNOLOGY Cleary, A. M., Tu, W. W., Enright, A., Giffon, T., Dewaal-Malefyt, R., Gutierrez, K., Lewis, D. B. 2003; 170 (1): 597-603


    Defects in IL-12 production or IL-12 responsiveness result in a vulnerability to infection with non-viral intracellular organisms, but the immunological mechanisms responsible for this susceptibility remain poorly understood. We present an immunological analysis of a patient with disseminated Salmonella enteritidis and a homozygous splice acceptor mutation in the IL-12Rbeta1-chain gene. This mutation resulted in the absence of IL-12Rbeta1 protein on PBMC and an inability of T cells to specifically bind IL-12 or produce IFN-gamma in response to either IL-12 or IL-23. The accumulation of memory (CD45R0(high)) CD4 T cells that were CCR7(high) (putative central memory cells) was normal or increased for age. Central memory CD4 T cells of the patient and age-matched controls were similar in having a low to undetectable capacity to produce IFN-gamma after polyclonal stimulation. In contrast, the patient had a substantial decrease in the number of CCR7(neg/dull) CD45R0(high) memory CD4 T cells (putative effector memory cells), and these differed from control cells in having a minimal ability to produce IFN-gamma after polyclonal stimulation. Importantly, tetanus toxoid-specific IFN-gamma production by PBMC from the patient was also significantly reduced compared with that in age-matched controls, indicating that signaling via the IL-12Rbeta1-chain is generally necessary for the in vivo accumulation of human memory CD4 T cells with Th1 function. These results are also consistent with a model in which the IL-12Rbeta1 subunit is necessary for the conversion of central memory CD4 T cells into effector memory cells.

    View details for Web of Science ID 000180106600075

    View details for PubMedID 12496448

  • IL-12 receptor beta 1 deficiency: Insights into memory CD4 T cell generation/maintenance and receptor expression. Cleary, A. M., Tu, W. W., de Waal Malefyt, R., Lewis, D. B. AMER SOC HEMATOLOGY. 2002: 188A–189A
  • Allergy immunotherapy and inhibition of Th2 immune responses: a sufficient strategy? CURRENT OPINION IN IMMUNOLOGY Lewis, D. B. 2002; 14 (5): 644-651


    Th2 immune responses mediated by the secretion of IL-4, IL-5 and IL-13 are key in the pathogenesis of atopic disorders, including allergen-induced asthma, rhinoconjunctivitis and anaphylaxis. Although such responses are downregulated to some degree by conventional specific immunotherapy, this approach is only partially effective and has a substantial risk of adverse effects. Many strategies for immunotherapeutic prophylaxis and for treatment of atopic diseases have been devised on the basis of mouse allergy and autoimmune models, including the downregulation of Th2 responses by the induction of regulatory T cell activity, Th2 to Th1 immune deviation, Th1 crossregulation of Th2 immune responses, anergy and immunosuppressive cytokines. The blockade of events that are not allergen-specific, such as T cell costimulation and downstream events dependent on IgE, cytokines and chemokines, has also been pursued. With the exception of monoclonal antibody therapy for the blockade of IgE effector function, the application of most of these strategies to humans is at an early stage. Whether the inhibition of Th2 responses without concurrent downregulation of Th1 responses will be sufficient for allergic immunotherapy, particularly for atopic dermatitis and asthma, is an important but unresolved issue.

    View details for Web of Science ID 000177716100017

    View details for PubMedID 12183167

  • Dynamics of CD4 and CD8 T cell responses to cytomegalovirus in healthy human donors JOURNAL OF INFECTIOUS DISEASES Dunn, H. S., Haney, D. J., Ghanekar, S. A., Stepick-Biek, P., Lewis, D. B., Maecker, H. T. 2002; 186 (1): 15-22


    To study the dynamics of cytomegalovirus (CMV) immunity in healthy immunocompetent hosts, interferon-gamma-producing CD4 and CD8 T cell responses in the presence or absence of CMV antigens were measured from 15 CMV-seropositive donors and 13 CMV-seronegative donors. Cytokine responses in the absence of antigen were significantly higher in CMV-seropositive donors. Also, a disproportionate number of CD69(+) cells isolated ex vivo from CMV-seropositive donors were specific for CMV, suggesting recent reactivation in vivo. To examine changes in cellular responses over time, 10 seropositive donors were tested over a 6-month period. About half of the donors showed significant variability over time, but staphylococcal enterotoxin B responses remained relatively constant. These findings suggest that CMV can present a considerable and recurrent burden to the human immune system. By understanding the normal dynamics of CMV responses over time, it may be possible to better identify aberrant responses to CMV in immunocompromised hosts.

    View details for Web of Science ID 000176307800003

    View details for PubMedID 12089657

  • Simple febrile seizures - Do polymorphisms of the interleukin 1 gene cluster simplify our understanding? ARCHIVES OF PEDIATRICS & ADOLESCENT MEDICINE Lewis, D. B. 2002; 156 (6): 529-530

    View details for Web of Science ID 000176055900001

    View details for PubMedID 12038881

  • Toll-like receptor 4 is required for optimal development of Th2 immune responses: Role of dendritic cells JOURNAL OF IMMUNOLOGY Dabbagh, K., Dahl, M. E., Stepick-Biek, P., Lewis, D. B. 2002; 168 (9): 4524-4530


    LPS potently induces dendritic cell maturation and the production of proinflammatory cytokines, such as IL-12, by activation of Toll-like receptor 4 (TLR4). Since IL-12 is important for the generation and maintenance of Th1 responses and may also inhibit Th2 cell generation from naive CD4 T cell precursors, it has been inferred that TLR4 signaling would have similar effects via the induction of IL-12 secretion. Surprisingly, we found that TLR4-defective mice subjected to sensitization and pulmonary challenge with a protein allergen had reductions in airway inflammation with eosinophils, allergen-specific IgE levels, and Th2 cytokine production, compared with wild-type mice. These reduced responses were attributable, at least in part, to decreased dendritic cell function: Dendritic cells from TLR4-defective mice expressed lower levels of CD86, a costimulatory molecule important for Th2 responses. They also induced less Th2 cytokine production by antigenically naive CD4 T cells in vitro and mediated diminished CD4 T cell Ag-specific pulmonary inflammation in vivo. These results indicate that TLR4 is required for optimal Th2 responses to Ags from nonpathogenic sources and suggest a role for TLR4 ligands, such as LPS derived from commensal bacteria or endogenously derived ligands, in maturation of the innate immune system before pathogen exposure.

    View details for Web of Science ID 000175263000036

    View details for PubMedID 11970998

  • Impaired accumulation and function of effector memory CD4T cells in human IL-12 receptor beta 1 deficiency Cleary, A. M., Tu, W. W., Enright, A., Giffon, T., Dewaal-Malefyt, R., Gutierrez, K., Lewis, D. B. INT PEDIATRIC RESEARCH FOUNDATION, INC. 2002: 11A
  • A T cell-specific enhancer of the human CD40 ligand gene JOURNAL OF BIOLOGICAL CHEMISTRY Schubert, L. A., Cron, R. Q., Cleary, A. M., Brunner, M., Song, A., Lu, L. S., JULLIEN, P., KRENSKY, A. M., Lewis, D. B. 2002; 277 (9): 7386-7395


    We observed that the human CD40 ligand (CD40L) gene 5'-flanking region conferred weak promoter activity in activated CD4 T cells, suggesting that additional regions are required for optimal CD40L gene transcription. We therefore examined a 3'-flanking segment of the CD40L gene, which contained a putative NF-kappaB/Rel cis-element, for its ability to enhance CD40L promoter function. This segment augmented CD40L promoter activity in an orientation-independent manner in CD4 T-lineage cells but not in human B cell or monocyte cell lines. Mapping of CD4 T-lineage cell nuclei identified a DNase I-hypersensitive site in the flanking region near the NF-kappaB/Rel sequence, suggesting a transcriptional regulatory role. This was further supported by truncation analysis and site-directed mutagenesis, which indicated that the CD40L 3'-flanking NF-kappaB/Rel cis-element was critical for enhancer function. Electrophoretic mobility shift assays showed that the cis-element preferentially bound the p50 form of the NF-kappaB1 gene contained in human T cell nuclear protein extracts. This binding also appeared to occur in vivo in CD4 T cells based on chromatin immunoprecipitation assays using NF-kappaB p50-specific antiserum. Together, these results suggest that the CD40L gene 3'-flanking region acts as a T cell-specific classical transcriptional enhancer by a NF-kappaB p50-dependent mechanism.

    View details for DOI 10.1074/jbc.M110350200

    View details for Web of Science ID 000174104300083

    View details for PubMedID 11751888

  • Group A streptococcal meningitis: Report of a case and review of literature since 1976 PEDIATRIC EMERGENCY CARE Shetty, A. K., Frankel, L. R., Maldonado, Y., Falco, D. A., Lewis, D. B. 2001; 17 (6): 430-434


    Group A streptococcal (GAS) invasive disease has become increasingly common in recent years. However, acute bacterial meningitis caused by this pathogen is unusual. We report a case of GAS meningitis in a previously healthy 21/2-year-old child associated with a dramatically rapid course and fatal outcome. A literature review of previously reported cases is presented. This case serves as a reminder that GAS can cause severe meningitis in otherwise healthy hosts.

    View details for Web of Science ID 000173001500007

    View details for PubMedID 11753187

  • Octamer proteins inhibit IL-4 gene transcription in normal human CD4 T cells GENES AND IMMUNITY Cron, R. Q., Zhou, B., Brunvand, M. W., Lewis, D. B. 2001; 2 (8): 464-468


    The balance of Th1 (eg, interleukin-2 (IL-2)) and Th2 (eg, IL-4) cytokines produced by CD4 T cells markedly influences the outcome of the adaptive immune response. Although octamer transcription factor proteins increase IL-2 transcription in T cells, their role in IL-4 gene transcription remains controversial. We have previously shown and now confirm that the proximal octamer binding site of the human IL-4 promoter, which separates the two most proximal NFAT binding sites, is bound prior to, but not after, activation in vivo. Since these two NFAT sites are essential for optimal IL-4 promoter activity, this suggested that prior engagement by octamer proteins might prevent adjacent NFAT binding and inhibit IL-4 gene transcription. In support of this hypothesis, here we show that NFAT proteins are unable to bind to a combined octamer/NFAT site unless the octamer proteins are competed away. Moreover, activity of an IL-4 reporter gene mutated in the proximal octamer binding site is increased compared to the wild-type promoter in human peripheral blood CD4 T cells. In addition, over-expression of either Oct-1 or Oct-2 decreased wild-type IL-4 promoter activity, while increasing IL-2 promoter activity. No decrease in promoter activity was seen when Oct-1 or Oct-2 was over-expressed with the octamer-mutant IL-4 reporter gene. Thus, octamer proteins are candidates to promote a Th1 rather than Th2 pattern of cytokine gene expression by activated CD4 T cells.

    View details for Web of Science ID 000172443000008

    View details for PubMedID 11781715

  • Respiratory infection with influenza A virus interferes with the induction of tolerance to aeroallergens JOURNAL OF IMMUNOLOGY Tsitoura, D. C., Kim, S., Dabbagh, K., Berry, G., Lewis, D. B., Umetsu, D. T. 2000; 165 (6): 3484-3491


    Viral respiratory infections have been implicated in influencing allergen sensitization and the development of asthma, but their exact role remains controversial. Because respiratory exposure to Ag normally engenders T cell tolerance and prevents the development of airway hyperreactivity (AHR) and inflammation, we examined the effects of influenza A virus infection on tolerance induced by exposure to intranasal (i.n.) OVA and the subsequent development of AHR. We found that concurrent infection with influenza A abrogated tolerance induced by exposure to i.n. OVA, and instead led to the development of AHR accompanied by the production of OVA-specific IgE, IL-4, IL-5, IL-13, and IFN-gamma. When both IL-4 and IL-5 were neutralized in this system, AHR was still induced, suggesting that influenza-induced cytokines such as IL-13, or mechanisms unrelated to cytokines, might be responsible for the development of AHR. The length of time between influenza A infection and i.n. exposure to OVA was crucial, because mice exposed to i.n. OVA 15-30 days after viral inoculation developed neither AHR nor OVA-specific tolerance. These mice instead acquired Th1-biased OVA-specific immune responses associated with vigorous OVA-induced T cell proliferation, and reduced production of OVA-specific IgE. The protective effect of influenza A on AHR was dependent on IFN-gamma, because protection was abrogated with a neutralizing anti-IFN-gamma mAb. These results suggest that viral respiratory infection interferes with the development of respiratory allergen-induced tolerance, and that the time interval between viral infection and allergen exposure is critical in determining whether viral infection will enhance, or protect against, the development of respiratory allergen sensitization and AHR.

    View details for Web of Science ID 000165938100073

    View details for PubMedID 10975869

  • Local blockade of allergic airway hyperreactivity and inflammation by the poxvirus-derived pan-CC-chemokine inhibitor vCCI JOURNAL OF IMMUNOLOGY Dabbagh, K., Xiao, Y., Smith, C., Stepick-Biek, P., Kim, S. G., Lamm, W. J., Liggitt, D. H., Lewis, D. B. 2000; 165 (6): 3418-3422


    Allergen-induced asthma is characterized by chronic pulmonary inflammation, reversible bronchoconstriction, and airway hyperreactivity to provocative stimuli. Multiple CC-chemokines, which are produced by pulmonary tissue in response to local allergen challenge of asthmatic patients or experimentally sensitized rodents, chemoattract leukocytes from the circulation into the lung parenchyma and airway, and may also modify nonchemotactic function. To determine the therapeutic potential of local intrapulmonary CC-chemokine blockade to modify asthma, a recombinant poxvirus-derived viral CC-chemokine inhibitor protein (vCCI), which binds with high affinity to rodent and human CC-chemokines in vitro and neutralizes their biological activity, was administered by the intranasal route. Administration of vCCI to the respiratory tract resulted in dramatically improved pulmonary physiological function and decreased inflammation of the airway and the lung parenchyma. In contrast, vCCI had no significant effect on the circulating levels of total or allergen-specific IgE, allergen-specific cytokine production by peripheral lymph node T cells, or peritoneal inflammation after local allergen challenge, indicating that vCCI did not alter systemic Ag-specific immunity or chemoattraction at extrapulmonary sites. Together, these findings emphasize the importance of intrapulmonary CC-chemokines in the pathogenesis of asthma, and the therapeutic potential of generic and local CC-chemokine blockade for this and other chronic diseases in which CC-chemokines are locally produced.

    View details for Web of Science ID 000165938100065

    View details for PubMedID 10975861

  • An immunosuppressive and anti-inflammatory HLA class I-derived peptide binds vascular cell adhesion molecule-1 TRANSPLANTATION Ling, X. F., Tamaki, T., Xiao, Y., Kamangar, S., CLAYBERGER, C., Lewis, D. B., KRENSKY, A. M. 2000; 70 (4): 662-667


    A synthetic peptide corresponding to residues 75-84 of HLA-B2702 modulates immune responses in rodents and humans both in vitro and in vivo.We used a yeast two-hybrid screening, an in vitro biochemical method, and an in vivo animal model.Two cellular receptors for this novel immunomodulatory peptide were identified using a yeast two-hybrid screen: immunoglobulin binding protein (BiP), a member of the heat shock protein 70 family, and vascular cell adhesion molecule (VCAM)-1. Identification of BiP as a ligand for this peptide confirms earlier biochemical findings, while the interaction with VCAM-1 suggests an alternative mechanism of action. Binding to the B2702 peptide but not to closely related variants was confirmed by ligand Western blot analysis and correlated with immunomodulatory activity of each peptide. In mice, an ovalbumin-induced allergic pulmonary response was blocked by in vivo administration of either the B2702 peptide or anti-VLA-4 antibody.We propose that the immunomodulatory effect of the B2702 peptide is caused, in part, by binding to VCAM-1, which then prevents the normal interaction of VCAM-1 with VLA-4.

    View details for Web of Science ID 000088986100021

    View details for PubMedID 10972226

  • Generation of conditional NFAT1 transgenic mice. Cron, R. Q., Lee, P., Lewis, D. B. FEDERATION AMER SOC EXP BIOL. 2000: A1082
  • NFAT1 enhances HIV-1 gene expression in primary human CD4 T cells CLINICAL IMMUNOLOGY Cron, R. Q., Bartz, S. R., Clausell, A., Bort, S. J., Klebanoff, S. J., Lewis, D. B. 2000; 94 (3): 179-191


    Cyclosporin A (CsA) is a potent inhibitor of the NFAT family of transcription factors that enhance T cell activation. The observation that human immunodeficiency virus type 1 (HIV-1)-positive transplant recipients have a reduced HIV-1 viral burden during treatment with CsA suggested that NFAT may play a direct role in enhancing transcription of the HIV-1 viral genome. Two sets of NFAT binding sites were identified in the HIV-1 long terminal repeat (LTR) promoter by in vitro footprinting with full-length recombinant NFAT protein, and gel shift analysis of nuclear protein from polyclonally activated primary CD4 T cells revealed specific binding of NFAT1 to the NFkappaB binding sites of the HIV-1 LTR. Activation of primary CD4 T cells transiently transfected with a HIV-1 LTR luciferase reporter plasmid, lacking the NFAT binding sites in the upstream putative negative regulatory element but maintaining the NFkappaB/NFAT sites, demonstrated increased HIV-1 gene expression when cotransfected with a NFAT1 expression vector. Moreover, CsA, FK506, and a dominant-negative NFAT1 protein independently inhibited HIV-1 LTR promoter activity in CD4 T cells stimulated with phorbol ester and calcium ionophore. In primary human CD4 T cells, CsA also inhibited promoter activity directed by multimers of binding sites for NFAT, while having no effect on NFkappaB multimer-driven promoter activity. Increasing NFAT1 levels in CD4 T cells transiently transfected with a HIV-1 provirus also increased p24 protein expression. Thus, NFAT may be a target for prevention of HIV-1 LTR-directed gene expression in human CD4 T cells.

    View details for Web of Science ID 000085927300004

    View details for PubMedID 10692237

  • Distinct types of diffuse large B-cell lymphoma identified by gene expression profiling NATURE Alizadeh, A. A., Eisen, M. B., Davis, R. E., Ma, C., Lossos, I. S., Rosenwald, A., Boldrick, J. G., Sabet, H., Tran, T., Yu, X., Powell, J. I., Yang, L. M., Marti, G. E., Moore, T., Hudson, J., Lu, L. S., Lewis, D. B., Tibshirani, R., Sherlock, G., Chan, W. C., Greiner, T. C., Weisenburger, D. D., Armitage, J. O., Warnke, R., Levy, R., Wilson, W., Grever, M. R., Byrd, J. C., Botstein, D., Brown, P. O., Staudt, L. M. 2000; 403 (6769): 503-511


    Diffuse large B-cell lymphoma (DLBCL), the most common subtype of non-Hodgkin's lymphoma, is clinically heterogeneous: 40% of patients respond well to current therapy and have prolonged survival, whereas the remainder succumb to the disease. We proposed that this variability in natural history reflects unrecognized molecular heterogeneity in the tumours. Using DNA microarrays, we have conducted a systematic characterization of gene expression in B-cell malignancies. Here we show that there is diversity in gene expression among the tumours of DLBCL patients, apparently reflecting the variation in tumour proliferation rate, host response and differentiation state of the tumour. We identified two molecularly distinct forms of DLBCL which had gene expression patterns indicative of different stages of B-cell differentiation. One type expressed genes characteristic of germinal centre B cells ('germinal centre B-like DLBCL'); the second type expressed genes normally induced during in vitro activation of peripheral blood B cells ('activated B-like DLBCL'). Patients with germinal centre B-like DLBCL had a significantly better overall survival than those with activated B-like DLBCL. The molecular classification of tumours on the basis of gene expression can thus identify previously undetected and clinically significant subtypes of cancer.

    View details for Web of Science ID 000085227300039

    View details for PubMedID 10676951

  • Bone marrow NK1.1(-) and NK1.1(+) T cells reciprocally regulate acute graft versus host disease JOURNAL OF EXPERIMENTAL MEDICINE Zeng, D. F., Lewis, D., Dejbakhsh-Jones, S., Lan, F. S., Garcia-Ojeda, M., Sibley, R., Strober, S. 1999; 189 (7): 1073-1081


    Sorted CD4(+) and CD8(+) T cells from the peripheral blood or bone marrow of donor C57BL/6 (H-2(b)) mice were tested for their capacity to induce graft-versus-host disease (GVHD) by injecting the cells, along with stringently T cell-depleted donor marrow cells, into lethally irradiated BALB/c (H-2(d)) host mice. The peripheral blood T cells were at least 30 times more potent than the marrow T cells in inducing lethal GVHD. As NK1.1(+) T cells represented <1% of all T cells in the blood and approximately 30% of T cells in the marrow, the capacity of sorted marrow NK1.1(-) CD4(+) and CD8(+) T cells to induce GVHD was tested. The latter cells had markedly increased potency, and adding back marrow NK1.1(+) T cells suppressed GVHD. The marrow NK1.1(+) T cells secreted high levels of both interferon gamma (IFN-gamma) and interleukin 4 (IL-4), and the NK1.1(-) T cells secreted high levels of IFN-gamma with little IL-4. Marrow NK1.1(+) T cells obtained from IL-4(-/-) rather than wild-type C57BL/6 donors not only failed to prevent GVHD but actually increased its severity. Together, these results demonstrate that GVHD is reciprocally regulated by the NK1.1(-) and NK1.1(+) T cell subsets via their differential production of cytokines.

    View details for Web of Science ID 000079648700008

    View details for PubMedID 10190898

    View details for PubMedCentralID PMC2193016

  • Lack of NFAT1 correlates with decreased cytokine production by neonatal T cells. Cron, R. Q., Bort, S. J., Lewis, D. B. FEDERATION AMER SOC EXP BIOL. 1999: A617
  • T cell priming enhances IL-4 gene expression by increasing nuclear factor of activated T cells JOURNAL OF IMMUNOLOGY Cron, R. Q., Bort, S. J., Wang, Y. X., Brunvand, M. W., Lewis, D. B. 1999; 162 (2): 860-870


    The repetitive activation of T cells (priming) enhances the expression of many cytokines, such as IL-4, but not others, such as IL-2. Molecular mechanisms underlying selective expression of cytokines by T cells remain poorly understood. Here we show that priming of CD4 T cells selectively enhances IL-4 expression relative to IL-2 expression by a transcriptional mechanism involving nuclear factor of activated T cells (NFAT) proteins. As detected by in vivo footprinting, priming markedly increases the activation-dependent engagement of the P0 and P1 NFAT-binding elements of the IL-4 promoter. Moreover, each proximal P element is essential for optimal IL-4 promoter activity. Activated primed CD4 T cells contain more NFAT1 and support greater NFAT-directed transcription than unprimed CD4 T cells, while activator protein 1 binding and activator protein 1-mediated transcription by both cell types is similar. Increased expression of wild-type NFAT1 substantially increases IL-4 promoter activity in unprimed CD4 T cells, suggesting NFAT1 may be limiting for IL-4 gene expression in this cell type. Furthermore, a truncated form of NFAT1 acts as a dominant-negative, reducing IL-4 promoter activity in primed CD4 T cells and confirming the importance of endogenous NFAT to increased IL-4 gene expression by effector T cells. NFAT1 appears to be the major NFAT family member responsible for the initial increased expression of IL-4 by primed CD4 T cells.

    View details for Web of Science ID 000077951400031

    View details for PubMedID 9916709

  • Identification of an enhancer element located downstream of the CD40-ligand gene. Cron, R. Q., Schubert, L. A., Lewis, D. B. LIPPINCOTT WILLIAMS & WILKINS. 1998: S235
  • Diminished expression of CD40 ligand may contribute to the defective humoral immunity in patients with MHC class II deficiency EUROPEAN JOURNAL OF IMMUNOLOGY Nonoyama, S., Etzioni, A., Toru, H., Ruggerie, D. P., Lewis, D., Pollack, S., Aruffo, A., Yata, J. I., Ochs, H. D. 1998; 28 (2): 589-598


    Major histocompatibility complex (MHC) class II deficiency (bare lymphocyte syndrome, BLS) is a rare primary immunodeficiency classified as a subgroup of severe combined immunodeficiency. We studied T and B lymphocyte function by examining the CD40 ligand/CD40 system in three BLS patients from two unrelated families. CD40 ligand expression by maximally activated BLS T cells was diminished. This abnormality may represent immunological naïveté rather than a general T cell defect, since expression of activation marker CD69 and proliferative responses to PHA or anti-CD3 were normal, and BLS T cells primed and restimulated in vitro expressed normal amounts of CD40 ligand. BLS B cells proliferated and produced IgE if stimulated with anti-CD40 or soluble CD40 ligand and IL-4. Activation of BLS B cells with soluble CD40 ligand and IL-4 induced normal expression of activation markers, although MHC class II expression remained absent. Depressed antibody titers, lack of amplification and failure to undergo isotype switching in response to immunization with bacteriophage phi x 174 demonstrated defective T cell help. We conclude that BLS B cells are functionally normal if appropriately stimulated, and that the defective humoral immunity observed may be related to diminished expression of CD40 ligand on BLS T cells.

    View details for Web of Science ID 000072307700023

    View details for PubMedID 9521069

  • Consistent transient transfection of DNA into non-transformed human and murine T-lymphocytes JOURNAL OF IMMUNOLOGICAL METHODS Cron, R. Q., Schubert, L. A., Lewis, D. B., HUGHES, C. C. 1997; 205 (2): 145-150


    The ability to analyze transcriptional regulation in non-transformed T-cells has been hampered by the inability to reproducibly transiently transfect these cells with DNA constructs. We have previously demonstrated that normal human whole mononuclear and CD4 T-cells can be consistently transiently transfected with plasmid DNA. Human cells were most receptive to plasmid DNA uptake between 19.5 and 20 h after prestimulation with a submitogenic dose of the polyclonal T-cell activator, PHA. Here we report an alteration and optimization of this protocol for non-transformed murine splenic T-cells, using concanavalin A instead of PHA as the preactivation stimulus. When coupled with the high sensitivity of luciferase reporter gene constructs, this protocol facilitates the analysis of a variety of T-cell-specific promoters in non-transformed T-cells. In addition, we directly demonstrate that murine T-cells are specifically transiently transfected among a population of whole mononuclear cells by using an expression vector for green fluorescent protein.

    View details for Web of Science ID A1997XR21700005

    View details for PubMedID 9294595

  • Influence of the route of allergen administration and genetic background on the murine allergic pulmonary response AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE Zhang, Y., Lamm, W. J., Albert, R. K., Chi, E. Y., Henderson, W. R., Lewis, D. B. 1997; 155 (2): 661-669


    We used various ovalbumin sensitization and challenge protocols to determine the importance of the route of allergen administration and the genetic background in modulating the physiologic, inflammatory, and immunologic features characteristic of allergen-induced asthma. In BALB/c mice, induction of maximal airway hyperresponsiveness and airspace eosinophilia required administration of ovalbumin by both the intraperitoneal and the intranasal routes (combination protocol), whereas intraperitoneal immunization alone resulted in maximal ovalbumin-specific IgE plasma levels. Thus, a systemic immune response to allergen, in addition to, or independent of IgE production, as well as local allergen challenge were necessary for maximal induction of pulmonary disease. BALB/c mice treated with ovalbumin by the combination protocol had increased Th2-type cytokine mRNA levels in bronchial lymph node tissue compared with control mice. In contrast, C57BL/6 mice treated with ovalbumin by the combination protocol had significantly decreased responses compared with BALB/c mice for all parameters of allergic pulmonary disease examined, with the exception of airspace eosinophilia. Genetic background has a striking and selective effect on the phenotype of murine allergic pulmonary disease. Further analysis of this murine model should be useful in helping define the critical pathogenetic events in allergen-induced asthma.

    View details for Web of Science ID A1997WH87100041

    View details for PubMedID 9032210

  • The importance of leukotrienes in airway inflammation in a mouse model of asthma JOURNAL OF EXPERIMENTAL MEDICINE Henderson, W. R., Lewis, D. B., Albert, R. K., Zhang, Y., Lamm, W. J., Chiang, G. K., Jones, F., Eriksen, P., Tien, Y. T., Jonas, M., Chi, E. Y. 1996; 184 (4): 1483-1494


    Inhalation of antigen in immunized mice induces an infiltration of eosinophils into the airways and increased bronchial hyperreactivity as are observed in human asthma. We employed a model of late-phase allergic pulmonary inflammation in mice to address the role of leukotrienes (LT) in mediating airway eosinophilia and hyperreactivity to methacholine. Allergen intranasal challenge in OVA-sensitized mice induced LTB4 and LTC4 release into the airspace, widespread mucus occlusion of the airways, leukocytic infiltration of the airway tissue and broncho-alveolar lavage fluid that was predominantly eosinophils, and bronchial hyperreactivity to methacholine. Specific inhibitors of 5-lipoxygenase and 5-lipoxygenase-activating protein (FLAP) blocked airway mucus release and infiltration by eosinophils indicating a key role for leukotrienes in these features of allergic pulmonary inflammation. The role of leukotrienes or eosinophils in mediating airway hyperresponsiveness to aeroallergen could not be established, however, in this murine model.

    View details for Web of Science ID A1996VP23000030

    View details for PubMedID 8879219

  • beta 2-microglobulin-dependent T cells are dispensable for allergen-induced T helper 2 responses JOURNAL OF EXPERIMENTAL MEDICINE Zhang, Y., Rogers, K. H., Lewis, D. B. 1996; 184 (4): 1507-1512


    CD4+ and CD8+ alpha/beta+ T cells of the T helper cell (Th)2 phenotype produce the cytokines IL-4, IL-5, and IL-13 that promote IgE production and eosinophilic inflammation. IL-4 may play an important role in mediating the differentiation of antigenically naive alpha/beta+ T cells into Th2 cells. Murine NK1.1+ (CD4+ or CD4-CD8-) alpha/beta+ T cells comprise a beta 2-microglobulin (beta 2m)-dependent cell population that rapidly produces IL-4 after cell activation in vitro and in vivo and has been proposed as a source of IL-4 for Th2 cell differentiation. alpha/beta+ CD8+ T cells, most of which require beta 2m for their development, have also been proposed as positive regulators of allergen-induced Th2 responses. We tested whether beta 2m-dependent T cells were essential for Th2 cell-mediated allergic reactions by treating wild-type, beta 2m-deficient (beta 2m -/-), and IL-4-deficient (IL-4 -/-) mice of the C57BL/6 genetic background with ovalbumin (OVA), using a protocol that induces robust allergic pulmonary disease in wild-type mice. OVA-treated beta 2m -/- mice had circulating levels of total and OVA-specific IgE, pulmonary eosinophilia, and expression of IL-4, IL-5, and IL-13 mRNA in bronchial lymph node tissue similar to that of OVA-treated wild-type mice. In contrast, these responses in OVA-treated IL-4 -/- mice were all either undetectable or markedly reduced compared with wild-type mice, confirming that IL-4 was required in this allergic model. These results indicate that the NK1.1+ alpha/beta+ T cell population, as well as other beta 2m-dependent populations, such as most peripheral alpha/beta+ CD8+ T cells, are dispensable for the Th2 pulmonary response to protein allergens.

    View details for Web of Science ID A1996VP23000032

    View details for PubMedID 8879221

  • Coexpression of the interleukin-13 and interleukin-4 genes correlates with their physical linkage in the cytokine gene cluster on human chromosome 5q23-31 BLOOD Dolganov, G., Bort, S., LOVETT, M., Burr, J., Schubert, L., Short, D., MCGURN, M., Gibson, C., Lewis, D. B. 1996; 87 (8): 3316-3326


    Interleukin-13 (IL-13) and IL-4 are cytokines produced by T cells that are encoded by the q23-31 region of human chromosome 5. To investigate the regulation of IL-13 gene expression by T cells, we isolated and sequenced the human IL-13 gene, analyzed its 5'-flanking region for potential transcriptional activation elements, and examined its expression in nontransformed T-lineage cell populations. The human IL-13 gene was located 12.5-kb upstream of the IL-4 gene and 2-kb downstream of a CpG island. The IL-13 gene 5' flank region included a segment with sequence homology to P elements of the IL-4 promoter involved in transcriptional activation in T cells. Mutation of the IL-13 P element site significantly reduced IL-13 promoter activity in response to T-cell activation. Oligonucleotides containing the IL-13 or IL-4 P element sites specifically bound the transcriptional activator protein, nuclear factor-activated T cells, preformed (NF-ATp), when incubated with nuclear protein extracts from activated T cells. Similar to IL-4, IL-13 mRNA expression was highest in T-cell populations enriched for cells that had previously been primed in vivo or in vitro, indicating that priming increases the expression of the IL-13 and IL-4 genes in a coordinate manner. Because the primed T cells contain higher levels of nuclear NF-ATp, capable of binding to P elements of the IL-4 and IL-13 promoters, than do freshly-isolated T cells, the NF-AT-binding P elements are attractive candidates to mediate the coordinate expression of these two cytokine genes.

    View details for Web of Science ID A1996UF38100030

    View details for PubMedID 8605348



    gp39, a cytokine expressed on the surface of activated T cells, is essential for T cell-dependent antibody responses in vivo. We cloned and sequenced 1.2 kilobases of the 5' flank region of the human gp39 gene promoter and determined its transcription start site. When used in reporter gene assays, this DNA segment conferred promoter activity in response to T cell activation. gp39 promoter function in transfectants was inhibited by cyclosporin A, as is expression of the endogenous gp39 gene in T-lineage cells. At least 0.5 kilobase of the 5' flank region was required for promoter activity. Two putative binding sites for the NF-AT family of transcriptional activator proteins were identified at -259 to -265 and -62 to -69 with respect to the transcription start site. Both sites contributed significantly and independently to promoter activity in response to T cell activation. Additionally, when incubated in vitro with nuclear protein purified from activated human CD4 T cells, both of these sites preferentially bound the NF-AT family member, NF-ATp. These results suggest that NF-ATp, via binding to at least two cis-elements, is essential for the induction of gp39 gene expression in response to T cell activation.

    View details for Web of Science ID A1995TK38000002

    View details for PubMedID 8530342



    To determine the potential role of methylation in the regulation of interferon-gamma (IFN-gamma) gene transcription by T cells, primary T-lineage cell populations were analyzed for the extent of methylation of three CpG sites within or near transcriptional activator elements in the 5' flank and first intron of the human IFN-gamma gene. A striking correlation was observed between the capacity of the IFN-gamma gene to be expressed and the degree of hypomethylation. The IFN-gamma gene was virtually completely methylated at all sites in thymocytes, neonatal T cells, and adult CD45RAhiCD45R0lo (antigenically naive) CD4 T cells, cell types that all have a low or undetectable capacity to express the IFN-gamma gene. In contrast, there was substantial hypomethylation in T-lineage cell types with relatively high capacities to express the IFN-gamma gene, including adult CD8 T cells and adult CD45RAloCD45R0hi (memory/effector) CD4 T cells. These results suggest that hypomethylation of the IFN-gamma genetic locus may be an important determinant of IFN-gamma gene expression in vivo by T-lineage cells.

    View details for Web of Science ID A1995QY50600017

    View details for PubMedID 7875204

  • DIMINISHED EXPRESSION OF CD40 LIGAND BY ACTIVATED NEONATAL T-CELLS JOURNAL OF CLINICAL INVESTIGATION Nonoyama, S., Penix, L. A., Edwards, C. P., Lewis, D. B., Ito, S., Aruffo, A., Wilson, C. B., Ochs, H. D. 1995; 95 (1): 66-75


    CD40 and CD40 ligand (gp39) mediate contact-dependent T-B cell interaction. We determined the expression of CD40 ligand by activated neonatal T cells and the response of neonatal B cells when activated through CD40. Although expression of CD40 ligand peaked simultaneously in both activated adult and neonatal cells, neonatal T cells expressed significantly less CD40 ligand surface protein and mRNA than adult T cells. Activated thymocytes also expressed far less CD40 ligand than adult T cells. Consistent with these results, activated neonatal T cells exhibited less helper function than activated adult T cells. Neonatal T cells primed and restimulated in vitro expressed CD40 ligand in amounts comparable with adult T cells and provided B cell help more effectively. This suggests that the poor expression of CD40 ligand reflects antigenic naiveté rather than an intrinsic defect of neonatal T cells. Neonatal B cells cultured with soluble CD40 ligand (sgp39) and IL-10 produced IgM in amounts comparable with adult cells, but much less IgG and IgA. Nevertheless, neonatal B cells were capable of proliferation and class switching, since sgp39 and IL-4 induced proliferation and IgE production comparable to adult B cells and production of modest amounts of IgG. Together, these results indicate that diminished CD40 ligand expression, along with decreased production of lymphokines, may be responsible, at least in part, for the transient immunodeficiency observed in human neonates.

    View details for Web of Science ID A1995QA88200012

    View details for PubMedID 7814647

  • DIFFERENTIATION OF THE T-HELPER PHENOTYPES BY ANALYSIS OF THE METHYLATION STATE OF THE IFN-GAMMA GENE JOURNAL OF IMMUNOLOGY Young, H. A., Ghosh, P., Ye, J. P., Lederer, J., Lichtman, A., GERARD, J. R., Penix, L., Wilson, C. B., Melvin, A. J., MCGURN, M. E., Lewis, D. B., Taub, D. D. 1994; 153 (8): 3603-3610


    Th1 and Th2 CD4+ T cell clones have been defined by their ability to produce different lymphokines. However, the processes by which CD4+ T cells differentially regulate lymphokine gene expression have not been well defined. In this report, we demonstrate that the methylation status of a CpG dinucleotide contained within a TATA proximal regulatory element of the IFN-gamma promoter correlates with the transcription of the gene. In murine Th1 clones and two human CD4+ Th0 clones, this site is either completely or partially hypomethylated, whereas in murine Th2 clones this site is > 98% methylated. Treatment of murine Th2 clones with 5-azacytidine, an agent that inhibits methylation of the DNA, converts these cells to IFN-gamma producers. Additional targets for methylation outside the transcriptional control regions of the IFN-gamma genetic locus were found to be hypomethylated in Th2 cells but not in Th1 cells. Electrophoretic mobility shift assays (EMSA) revealed at least five distinct protein-DNA complexes that are formed with an oligonucleotide containing the IFN-gamma promoter TATA proximal regulatory element, and in vitro methylation of this site results in a loss of these three complexes. Furthermore, a comparison of nuclear extracts prepared from Th1 and Th2 clones revealed that the EMSA patterns were qualitatively similar but differed quantitatively. In addition, transient transfection of a murine IFN-gamma promoter-chloramphenicol acetyl transferase (CAT) gene construct into both Th1 and Th2 clones produced CAT activity that was not inducible by anti-CD3, indicating that hypomethylation per se of the promoter alone is not sufficient for inducible gene expression.

    View details for Web of Science ID A1994PM58100026

    View details for PubMedID 7523497

  • OSTEOPOROSIS INDUCED IN MICE BY OVERPRODUCTION OF INTERLEUKIN-4 PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Lewis, D. B., Liggitt, H. D., Effmann, E. L., Motley, S. T., Teitelbaum, S. L., Jepsen, K. J., Goldstein, S. A., Bonadio, J., Carpenter, J., Perlmutter, R. M. 1993; 90 (24): 11618-11622


    Osteoporosis is a common disease in which loss of bone mass results in skeletal fragility. The development of therapies for this disorder has been hampered by the lack of a convenient animal model. Here we describe a disorder in bone homeostasis in transgenic mice that inappropriately express the cytokine interleukin 4 (IL-4) under the direction of the lymphocyte-specific proximal promoter for the lck gene. Bone disease in lck-IL-4 mice appeared to result from markedly decreased bone formation by osteoblasts, features strikingly similar to those observed in cases of severe low-turnover human involutional osteoporosis. By 2 months of age, female and male lck-IL-4 mice invariably developed severe osteoporosis of both cortical and trabecular bone. Osteoporosis was observed in two independently derived founder animals, indicating that this phenotype was directly mediated by the IL-4 transgene.

    View details for Web of Science ID A1993MM51500038

    View details for PubMedID 8265598



    T cell-derived lymphokines mediate or modulate various aspects of the immune response and immunodeficiency states related to abnormalities in lymphokine production or regulation are now being recognized. One example of this is seen in the fetus and neonate, in whom a physiologic immunodeficiency appears to reflect in part deficient production of certain lymphokines, including interferon-gamma, IL-4, and IL-5. The deficiency in production of these lymphokines appears to reflect to a large extent the paucity of memory T cells during these periods of life. Diminished lymphokine production has also recently been implicated as the cause for three cases of primary severe combined immunodeficiency. In disorders associated with excess IgE production, including allergy, hyper IgE syndrome, and Omenn's syndrome, excess IL-4 production relative to the production of interferon-gamma may play a contributory role. Regulation of the production of these and other T cell-derived lymphokines appears to be affected predominantly by control of lymphokine gene transcription, the basis for which is just now becoming understood at a molecular level. The elucidation of these regulatory mechanisms offers the promise for understanding the basis for disordered lymphokine production in immunodeficiency states.

    View details for Web of Science ID A1993LF99900004

    View details for PubMedID 8500278



    T cell precursors are first detected in the thymus at eight weeks of gestation. By 15 to 20 weeks of gestation, T-cell precursors expressing alpha beta and gamma delta T-cell receptors are present in the thymus in numbers relatively similar to those found in postnatal life. However, recent data suggest that T-cell receptor diversity is more limited during fetal and neonatal life than in adults. Additionally, the functional capacity of T cells in the fetus and neonate is immature, in that neonatal T cells express a limited repertoire of lymphokines in response to activation. Specifically, the production of the lymphokines, interferon-gamma and interleukin-4, which participate in the maturation of cytotoxic cells, activation of macrophages, and the maturation and modulation of B cell function and isotype expression, is reduced more than tenfold compared to cells from adults. This appears to result primarily from the lack of memory T cells in the fetus and neonate, reflecting their antigenic naivete. The difference in lymphokine expression is due to diminished transcription of these genes in neonatal T cells in response to activation. Preliminary data indicate that differences in essential promoter elements regulating transcription of these lymphokine genes plays a role in their differential expression in T cells.

    View details for Web of Science ID A1992KH72800005

    View details for PubMedID 1285862

  • HAWKINSINURIA IN 2 FAMILIES AMERICAN JOURNAL OF MEDICAL GENETICS Borden, M., Holm, J., Leslie, J., Sweetman, L., Nyhan, W. L., Fleisher, L., Nadler, H., Lewis, D., Scott, C. R. 1992; 44 (1): 52-56


    Hawkinsinuria, a disorder of tyrosine metabolism has been documented in two families in the United States, in one of which there was clear evidence of autosomal dominant inheritance. Metabolic acidosis and failure to thrive appear to be confined to infancy. Tyrosyl metabolites and 5-oxoproline are also found only in infancy, while 4-hydroxycyclohexylacetic acid was present only with time. The disease may be detected by organic acid analysis or by staining an electropherogram for sulfur containing compounds.

    View details for Web of Science ID A1992JJ63400012

    View details for PubMedID 1519651



    The clinical and microbiologic features of primary cutaneous infections by Yersinia enterocolitica are described in three children. Vesiculobullous lesions developed in two patients, and an intense granulation response followed incision and drainage. In the third child, cellulitis and abscess formation developed at the site of minor skin trauma. Y. enterocolitica isolates from the patients were extensively serologically, biochemically, and molecularly analyzed and compared with virulent Y. enterocolitica strains. The ability of the isolates to adhere to and invade eukaryotic cells was determined using in vitro assays; virulence was assessed by inoculation of suckling mice. The resulting data suggest that primary cutaneous infections by Y. enterocolitica involve strains that are as virulent as pathogenic gastrointestinal isolates.

    View details for Web of Science ID A1992HJ64200023

    View details for PubMedID 1552204



    Impaired production and delivery of neutrophils to the site of infection have been implicated in the increased susceptibility of the neonate to infection. Because granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF) play critical roles in the production of neutrophils from marrow precursors, we assessed the ability of leukocytes from neonates and adults to produce GM-CSF, G-CSF, and, for comparison, macrophage colony-stimulating factor (M-CSF) after stimulation with concanavalin A +/- phorbol myristate acetate [blood mononuclear cells (MC) and T lymphocytes] or lipopolysaccharide (monocytes). MC and monocytes from adult and neonatal subjects produced mRNA for GM-CSF, G-CSF, and M-CSF, whereas T cells produced only GM-CSF mRNA. Neonatal MC and T cells accumulated only approximately 30% as much GM-CSF mRNA as did adult MC and T cells. In contrast, the accumulation of GM-CSF mRNA by neonatal and adult monocytes was similar. Neonatal MC also accumulated similar amounts of G-CSF mRNA and somewhat more M-CSF mRNA than did adult MC; results with monocytes were similar to those with MC. Results of colony-stimulating activity bioassays on supernatants from neonatal and adult MC stimulated with concanavalin A paralleled the mRNA results.(ABSTRACT TRUNCATED AT 250 WORDS)

    View details for Web of Science ID A1992HF34600003

    View details for PubMedID 1373232

  • DEVELOPMENTAL REGULATION OF LCK GENE-EXPRESSION IN LYMPHOCYTES-T JOURNAL OF EXPERIMENTAL MEDICINE Wildin, R. S., Garvin, A. M., Pawar, S., Lewis, D. B., Abraham, K. M., Forbush, K. A., Ziegler, S. F., Allen, J. M., Perlmutter, R. M. 1991; 173 (2): 383-393


    In the mouse and human, mRNA transcripts encoding the lymphocyte-specific protein tyrosine kinase p56lck are derived from two separate promoters resulting in heterogeneity in the 5' untranslated region sequence. The proximal promoter lies just 5' to the coding region for the gene and is active only in thymocytes. In contrast, the distal promoter lies 34 kilobases (kb) 5' in the human, and is active both in thymocytes and mature peripheral T cells. As previously reported, transgenic mice bearing functional proximal promoter sequence juxtaposed with the SV40 large T antigen gene invariably develop lymphoid tumors confined to the thymus. In the current work, transgenic mice bearing a 2.6-kb fragment of the human distal promoter fused to the SV40 large T antigen gene express large T antigen in thymocytes and in peripheral lymphoid cells, and develop tumors of both the thymus and the peripheral lymphoid organs. The ability of the human distal promoter to function appropriately in transgenic mice is consistent with the strong similarity observed between the mouse and human distal promoter sequences. With the exception of a single short interval that serves as a target for binding of nuclear factors, significant sequence similarity is not seen when the distal and proximal promoter sequences are compared. Hence, developmentally regulated, lineage-specific transcription of the lck gene is mediated by distinct promoter sequences that appear to be capable of functioning independently.

    View details for Web of Science ID A1991EV17500013

    View details for PubMedID 1988541

  • INTERLEUKIN-4 EXPRESSED INSITU SELECTIVELY ALTERS THYMOCYTE DEVELOPMENT JOURNAL OF EXPERIMENTAL MEDICINE Lewis, D. B., Yu, C. C., Forbush, K. A., Carpenter, J., Sato, T. A., Grossman, A., Liggitt, D. H., Perlmutter, R. M. 1991; 173 (1): 89-100


    Using a transgenic mouse model we show that increased intrathymic expression of interleukin 4 (IL-4) significantly perturbs the development of thymocytes. Transgenic double-positive (CD4+CD8+) thymocytes, which are present in dramatically reduced numbers, exhibit increased T cell receptor (TCR) expression and increased mobilization of calcium mediated by these receptors. In contrast, transgenic single-positive (CD4+CD8- and CD4-CD8+) thymocytes and peripheral T cells exhibit decreased TCR-mediated calcium mobilization. The development of CD4-CD8+ thymocytes is significantly perturbed by IL-4 expressed in vivo; only peripheral CD4+ T cells are found in significant numbers in transgenic mice, while CD4-CD8+ thymocytes are present in increased numbers, apparently because of their failure to emigrate to the periphery. In contrast to these selective effects on T cell development, no significant differences in the numbers of B cells or mast cells, or in the plasma levels of IgE and IgG1 are observed between transgenic and control mice. These observations suggest that IL-4 in vivo exerts its major effects locally rather than systemically, even when its expression is constitutively increased.

    View details for Web of Science ID A1991EP65900010

    View details for PubMedID 1824637

  • T cell development in the fetus and neonate. Advances in experimental medicine and biology Wilson, C. B., Lewis, D. B., English, B. K. 1991; 310: 17-27

    View details for PubMedID 1808993



    The mechanisms by which T lymphocytes acquire the capacity to produce interleukin 4 (IL-4) and other lymphokines during intrathymic and extrathymic development are poorly understood. To gain insight into this process, we determined the capacity of human neonatal and adult T lineage cell populations to produce IL-4 after polyclonal activation. IL-2 and interferon-gamma (IFN-gamma) production were studied in parallel, since their production by neonatal T cells is known to be similar or diminished, respectively, compared to adult T cells. Production of IL-4 by neonatal CD4+ T cells and IFN-gamma by neonatal CD4+ and CD8+ T cells was markedly lower compared with analogous adult cell populations, whereas IL-2 production was similar. Transcription of IL-4, as determined by nuclear run-on assays, and IL-4 mRNA-containing cells, as determined by in situ hybridization, were undetectable in neonatal T cells, whereas both were detectable in adult T cells. IFN-gamma transcription and IFN-gamma mRNA-containing cells were reduced in neonatal T cells compared with adult T cells. Reduced lymphokine production by neonatal T cells correlated with their lack of a CD45R- (putative memory T cell) population; cells with this surface phenotype comprised 30-40% of the adult CD4+ T cells and were highly enriched for IL-4 and IFN-gamma, but not IL-2 production. IL-4, IFN-gamma, and IL-2 mRNA expression by neonatal CD4+CD8- thymocytes was similar to that found in circulating neonatal CD4+ T cells. Taken together, these findings suggest that the extrathymic generation of memory T cells during postnatal life may result in an increased capacity for IL-4 and IFN-gamma gene expression. In addition, IFN-gamma and IL-2 mRNA were significantly more abundant than IL-4 mRNA in activated neonatal CD4+CD8- thymocytes and CD4+ T cells, as well as adult CD4+ CD45R- T cells. Therefore, the capacity of T lineage cells to express the IL-4 gene may be more restricted compared to other lymphokine genes beginning in intrathymic development. This restricted capacity appears to persist during postnatal extrathymic maturation of T cells.

    View details for Web of Science ID A1991EQ97600026

    View details for PubMedID 1824631


    View details for Web of Science ID A1990DY44800007

    View details for PubMedID 2122411

  • REGULATION OF IMMUNOGLOBULIN (IG)E SYNTHESIS IN THE HYPER-IGE SYNDROME JOURNAL OF CLINICAL INVESTIGATION Vercelli, D., Jabara, H. H., CUNNINGHAMRUNDLES, C., Abrams, J. S., Lewis, D. B., Meyer, J., Schneider, L. C., Leung, D. Y., Geha, R. S. 1990; 85 (5): 1666-1671


    The hyper-IgE (HIE) syndrome is characterized by high IgE serum levels, chronic dermatitis, and recurrent infections. The mechanisms responsible for hyperproduction of IgE in HIE patients are presently unknown. We investigated whether spontaneous in vitro IgE synthesis by PBMC from seven HIE patients was sensitive to signals (cell adhesion, T/B cell cognate interaction and lymphokines: IL-4, IL-6, and IFN-gamma) known to regulate IgE induction in normals. Our results show that, unlike IL-4 dependent IgE synthesis induced in normals, spontaneous IgE production by PBMC from HIE patients was not blocked by monoclonal antibodies to CD2, CD4, CD3, and MHC class II antigens. Furthermore, antibodies to IL-4 and IL-6 did not significantly suppress IgE production. IFN-gamma had no significant effects on spontaneous in vitro IgE synthesis. To test whether an imbalance in lymphokine production might underlie hyperproduction of IgE in HIE patients, mitogen-induced secretion of IL-4 and IFN-gamma by PBMC was assessed. No significant difference was detected between HIE patients and normal controls. Thus, ongoing IgE synthesis in the HIE syndrome is largely independent of cell-cell interactions and endogenous lymphokines, and is due to a terminally differentiated B cell population, no longer sensitive to regulatory signals.

    View details for Web of Science ID A1990DB95800041

    View details for PubMedID 2110192



    The human neonate is unduly susceptible to infection with viruses and other pathogens, such as Toxoplasma and Listeria, that survive and replicate within cells. Cellular immunity is the major mechanism of host defense against these intracellular pathogens. Selective immaturity in certain functions of T lymphocytes appears to be a major factor in the neonate's susceptibility to these infections. Particularly striking is the deficiency in production of interferon-gamma. We review the data regarding the deficiency in the production of interferon-gamma by cells of healthy and infected neonates, discuss what is known regarding the cellular and molecular mechanisms for this deficiency, and review the unique role played by interferon-gamma in host defense against intracellular pathogens. The response of the neonate's effector cells to the immunoenhancing effects of interferon-gamma appears to be variable; diminished enhancement by interferon-gamma of cytotoxic cell function and the production of tumor necrosis factor by macrophages may further compound the effects of diminished production of interferon-gamma.

    View details for Web of Science ID A1990DG33700005

    View details for PubMedID 2114034

  • LYMPHOCYTE-ACTIVATION PROVOKES MODIFICATION OF A LYMPHOCYTE-SPECIFIC PROTEIN TYROSINE KINASE (P56LCK) JOURNAL OF IMMUNOLOGY Marth, J. D., Lewis, D. B., Cooke, M. P., Mellins, E. D., GEARN, M. E., Samelson, L. E., Wilson, C. B., Miller, A. D., Perlmutter, R. M. 1989; 142 (7): 2430-2437


    The protein tyrosine kinase p56lck is implicated in the control of lymphocyte growth by virtue of its overexpression in some lymphoid malignancies and its transforming activity in heterologous systems. Previous studies have demonstrated that levels of lck mRNA and of p56lck decline rapidly after T cell activation. The disappearance of p56lck results primarily from post-translational conversion of p56lck to more slowly migrating forms with apparent sizes of approximately 60 kDa. This modification can be provoked by treatment of lymphocytes with PMA, and has been associated with increased serine phosphorylation of the p56lck molecule. Here we demonstrate that conversion of p56lck to p60lck is a feature of the physiologic activation of T lymphocytes by antigen-presenting cells. In addition, we show that the PMA-induced modification of p56lck proceeds via a mechanism distinct from conventional protein kinase C activation. The rapid conversion of p56lck to p60lck after antigenic stimulation is consistent with the view that this membrane-associated protein tyrosine kinase regulates some aspects of the lymphocyte activation sequence.

    View details for Web of Science ID A1989T802000037

    View details for PubMedID 2784463



    Interleukin 4 (IL-4) is secreted by activated T cells and pleiotropically modulates both B- and T-lymphocyte function. In murine helper (CD4+) T-cell clones IL-4 production appears to be regulated independently of interferon gamma and interleukin 2. To determine whether production of these lymphokines is also differentially regulated in uncloned human T cells, we studied lymphokine production by normal human peripheral T cells and T-cell subsets after in vitro polyclonal activation. After maximal induction of lymphokine expression, IL-4 mRNA was detectable in less than 5% of CD4+ and 1-2% of unfractionated T cells, whereas approximately 33% and 60% of CD4+ cells expressed detectable mRNA for interferon gamma and interleukin 2, respectively. This finding correlated with dramatically lower production of IL-4 mRNA and protein than of interferon gamma and interleukin 2 by peripheral blood and tonsillar T cells. The helper-inducer (CD4+ CD45R-) T-cell subset, which significantly enhances in vitro immunoglobulin production, accounted for the preponderance of IL-4 mRNA accumulation and protein production by CD4+ T cells; nevertheless, cells with detectable IL-4 mRNA constituted less than 10% of the CD4+ CD45R- subset. Limitation of IL-4 production to a comparatively small population of normal human T cells could selectively regulate the effects of this lymphokine in T-cell-mediated immune responses; such selective regulation may be a fundamental mechanism for restricting the potentially pleiotropic effects of certain lymphokines to appropriate responder cells.

    View details for Web of Science ID A1988R518700080

    View details for PubMedID 3144002

  • STRUCTURE AND EXPRESSION OF ICK - TRANSCRIPTS IN HUMAN LYMPHOID-CELLS JOURNAL OF CELLULAR BIOCHEMISTRY Perlmutter, R. M., Marth, J. D., Lewis, D. B., Peet, R., Ziegler, S. F., Wilson, C. B. 1988; 38 (2): 117-126


    The murine lck gene encodes a membrane-associated protein tyrosine kinase that has been implicated in lymphocyte oncogenesis. Here we report the structure of normal human lck transcripts and the pattern of expression of these transcripts in developing thymus and in peripheral T cell subsets. The human lck gene encodes a 509 amino acid polypeptide that is closely related to the murine lck-encoded protein throughout its length. Analysis of the deduced amino acid sequence of human p56lck demonstrates that an amino-terminal domain, widely divergent among the seven known src family members, has been conserved between murine and human p56lck, and thus probably includes sequences crucial to the lymphocyte-specific function of this molecule. Human lck transcripts were detected in CD4+ and CD8+ T cells, in partially purified B cells, and in Epstein-Barr virus-immortalized B cell lines, but not in monocytes, granulocytes, or in nonhematopoetic cell types. Human lck transcripts are readily detectable in fetal thymocytes at 70 days of gestation, but not at 57 days of gestation, indicating that lck expression appears coordinately with the appearance of lymphoid cells in the developing thymus. Thus lck gene expression is a marker for cells of the lymphocyte lineage in man. We conclude that the lck gene probably participates in a signal transduction pathway uniquely present in lymphoid cells.

    View details for Web of Science ID A1988Q654800005

    View details for PubMedID 3265417



    The lymphocyte-specific protein tyrosine kinase pp56lck, encoded by a member of the src gene family, is implicated in the control of T-cell growth and differentiation. Purified resting human T lymphocytes contain appreciable levels of lck mRNA and of pp56lck. Upon activation of these T cells, levels of lck mRNA and of pp56lck promptly decline. These reductions in lck mRNA and protein expression are closely correlated with the induction of lymphokine production. Both require identical stimuli and follow a similar time course of response. Down-regulation of lck expression, however, is not correlated with proliferation. Our results provide an example of regulation of a src-like protein tyrosine kinase in a normal fully differentiated cell population and suggest that modulation of lck RNA and protein expression is an important feature of the lymphocyte activation sequence leading to lymphokine production.

    View details for Web of Science ID A1987J692700030

    View details for PubMedID 3119327



    Protein-tyrosine kinases are implicated in the control of cell growth by virtue of their frequent appearance as products of retroviral oncogenes and as components of growth factor receptors. Here we report the characterization of a novel human protein-tyrosine kinase gene (hck) that is primarily expressed in hematopoietic cells, particularly granulocytes. The hck gene encodes a 505-residue polypeptide that is closely related to pp56lck, a lymphocyte-specific protein-tyrosine kinase. The exon breakpoints of the hck gene, partially defined by using murine genomic clones, demonstrate that hck is a member of the src gene family and has been subjected to strong selection pressure during mammalian evolution. High-level expression of hck transcripts in granulocytes is especially provocative since these cells are terminally differentiated and typically survive in vivo for only a few hours. Thus the hck gene, like other members of the src gene family, appears to function primarily in cells with little growth potential.

    View details for Web of Science ID A1987H488700030

    View details for PubMedID 3453117



    IFN-gamma mRNA levels in human neonatal blood mononuclear cells or highly purified T cells were markedly lower than those of adult cells after incubation with Con A and PMA. In contrast, IL-2, IL-2-R, and T3 delta chain mRNA levels were kinetically and quantitatively similar in neonatal and adult T cells. The peak amount of IFN-gamma and IL-2 mRNA correlated well with IFN-gamma and IL-2 detected in supernatants of both neonatal and adult T cells. These results suggest that reduced IFN-gamma mRNA levels in neonatal T cells is due to an intrinsic deficiency at the pretranslational level and indicate that the magnitude of IL-2 and IFN-gamma gene expression can be independently modulated pretranslationally.

    View details for Web of Science ID A1986A688300019

    View details for PubMedID 3081678



    Human neonatal lymphocytes produced little macrophage activation factor in response to mitogens. This correlated with decreased production of interferon-gamma (IFN gamma): adult lymphokines contained 894.2 +/- 177.1 U/ml, whereas neonatal cord and peripheral lymphokines contained 66.9 +/- 17.0 and 116.7 +/- 29.6 U/ml by bioassay. Results by radioimmunoassay (RIA) for IFN gamma were similar. In contrast, the interleukin 2 content of cord lymphokines was greater (P less than 0.01) and that of neonatal peripheral blood lymphokines similar to that of adults. Interleukin 1 production and interleukin 2 receptor expression and affinity were similar for adult and neonatal cells. Interleukins 1 and 2 in amounts comparable to those in adult lymphokines did not increase production of macrophage activation factor or IFN gamma by neonatal cells. Neonatal cells did not contain intracellular IFN or degrade exogenous IFN. Excess suppressor activity was not found in neonatal cultures. Addition of IFN alpha, 10,000-50,000 U/ml of interleukin 2 or phorbol myristate acetate (PMA) to cord mononuclear cells or of adult monocytes or PMA to cord T cells increased IFN gamma production compared to cells stimulated with concanavalin A (ConA) alone. Nevertheless, under optimal conditions (T cells + PMA + Con A), adult cells produced much more IFN gamma (1,360 +/- 261 U/ml by RIA) than cord cells (122 +/- 37 U/ml). Staphylococcal enterotoxin A (SEA) stimulated cord cell IFN gamma production at low cell densities; nevertheless, adult cells produced more IFN in response to SEA 1,341 +/- 350 U/ml) than cord cells (350 +/- 33 U/ml). Decreased production of IFN gamma by neonatal cells appears to be due both to differences in their intrinsic capacity to produce IFN gamma and to differences in regulatory mechanisms.

    View details for Web of Science ID A1986A542900025

    View details for PubMedID 3081575