Karim Sallam, MD

All Publications

• CCL2-mediated endothelial injury drives cardiac dysfunction in long COVID. Nature cardiovascular research Thomas, D., Noishiki, C., Gaddam, S., Wu, D., Manhas, A., Liu, Y., Tripathi, D., Kathale, N., Adkar, S. S., Garhyan, J., Liu, C., Xu, B., Ross, E. G., Dalman, R. L., Wang, K. C., Oro, A. E., Sallam, K., Lee, J. T., Wu, J. C., Sayed, N. 2024; 3 (10): 1249-1265

  Abstract

  Evidence linking the endothelium to cardiac injury in long coronavirus disease (COVID) is well documented, but the underlying mechanisms remain unknown. Here we show that cytokines released by endothelial cells (ECs) contribute to long-COVID-associated cardiac dysfunction. Using thrombotic vascular tissues from patients with long COVID and induced pluripotent stem cell-derived ECs (iPSC-ECs), we modeled endotheliitis and observed similar dysfunction and cytokine upregulation, notably CCL2. Cardiac organoids comprising iPSC-ECs and iPSC-derived cardiomyocytes showed cardiac dysfunction after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) exposure, driven by CCL2. Profiling of chromatin accessibility and gene expression at a single-cell resolution linked CCL2 to 'phenotype switching' and cardiac dysfunction, validated by high-throughput proteomics. Disease modeling of cardiac organoids and exposure of human ACE2 transgenic mice to SARS-CoV-2 spike proteins revealed that CCL2-induced oxidative stress promoted post-translational modification of cardiac proteins, leading to cardiac dysfunction. These findings suggest that EC-released cytokines contribute to cardiac dysfunction in long COVID, highlighting the importance of early vascular health monitoring in patients with long COVID.

  View details for DOI 10.1038/s44161-024-00543-8

  View details for PubMedID 39402206

  View details for PubMedCentralID 7899720

• Validating the Association Between Composite Metrics of Guideline-Directed Medical Therapy (GDMT) and Clinical Outcomes for Patients with Heart Failure with Reduced Ejection Fraction (HFrEF). Journal of cardiac failure Steverson, A., Calma, J., Hsiao, S., Sallam, K., Varshney, A. S., Golbus, J. R., Heidenreich, P. A., Sandhu, A. T. 2024

  View details for DOI 10.1016/j.cardfail.2024.08.054

  View details for PubMedID 39357668

• One-year real-world experience with mavacamten and its physiologic effects on obstructive hypertrophic cardiomyopathy FRONTIERS IN CARDIOVASCULAR MEDICINE Kim, D., Chu, E. L., Keamy-Minor, E. E., Paranjpe, I., Tang, W. L., O'Sullivan, J. W., Desai, Y. B., Liu, M. B., Munsey, E., Hecker, K., Cuenco, I., Kao, B., Bacolor, E., Bonnett, C., Linder, A., Lacar, K., Robles, N., Lamendola, C., Smith, A., Knowles, J. W., Perez, M. V., Kawana, M., Sallam, K. I., Weldy, C. S., Wheeler, M. T., Parikh, V. N., Salisbury, H., Ashley, E. A., Stanford Ctr Inherited Cardiovasc Dis 2024; 11

  View details for DOI 10.3389/fcvm.2024.1429230

  View details for Web of Science ID 001312496500001

• One-year real-world experience with mavacamten and its physiologic effects on obstructive hypertrophic cardiomyopathy. Frontiers in cardiovascular medicine Kim, D. S., Chu, E. L., Keamy-Minor, E. E., Paranjpe, I. D., Tang, W. L., O'Sullivan, J. W., Desai, Y. B., Liu, M. B., Munsey, E., Hecker, K., Cuenco, I., Kao, B., Bacolor, E., Bonnett, C., Linder, A., Lacar, K., Robles, N., Lamendola, C., Smith, A., Knowles, J. W., Perez, M. V., Kawana, M., Sallam, K. I., Weldy, C. S., Wheeler, M. T., Parikh, V. N., Salisbury, H., Ashley, E. A. 2024; 11: 1429230

  Abstract

  Mavacamten is a first-in-class cardiac myosin ATPase inhibitor, approved by the United States Food and Drug Administration for the treatment of hypertrophic cardiomyopathy with obstructive physiology (oHCM). Here, we present the real-world use of mavacamten in 50 patients with oHCM at a tertiary care referral center. In both our highlighted case and in our aggregate data, we report significant improvement in wall thickness, mitral regurgitation, left ventricular outflow tract obstruction and New York Heart Association symptom class. Moreover, in our center's experience, neither arrhythmia burden, nor contractility have worsened in the vast majority of patients: we note a clinically insignificant mean decrease in left ventricular ejection fraction (LVEF), with only two patients requiring temporary mavacamten discontinuance for LVEF < 50%. Adverse events were rare, unrelated to mavacamten itself, and seen solely in patients with disease too advanced to have been represented in clinical trials. Moreover, our multidisciplinary pathway enabled us to provide a large number of patients with a novel closely-monitored therapeutic within just a few months of commercial availability. These data lead us to conclude that mavacamten, as a first-in-class cardiac myosin inhibitor, is safe and efficacious in real-world settings.

  View details for DOI 10.3389/fcvm.2024.1429230

  View details for PubMedID 39314763

  View details for PubMedCentralID PMC11417615

• Generation of two iPSC lines from dilated cardiomyopathy patients with pathogenic variants in the SCN5A gene. Stem cell research Dexheimer, R., Manhas, A., Wu, D., Tripathi, D., Yu Chan, S., Li, J., Yu, R., Sayed, N., Wu, J. C., Sallam, K. 2024; 80: 103498

  Abstract

  Dilated cardiomyopathy (DCM) is a disorder of cardiac ventricular dilation and contractile dysfunction that often progresses to heart failure. Multiple genes have been associated with DCM, including SCN5A which has been linked to 2 % of all DCM cases. Peripheral mononuclear blood cells from DCM patients with SCN5A variants (c.2440C>T and c.665G>A) were utilized to generate two human induced pluripotent stem cell (iPSC) lines. Both lines exhibited typical iPSC morphology, expressed pluripotency markers, normal karyotypes, and trilineage differentiation capabilities. These lines offer valuable resources for investigating the mechanism of SCN5A-associated DCM, facilitating studies of ion channel protein involvement in the disease.

  View details for DOI 10.1016/j.scr.2024.103498

  View details for PubMedID 39067410

• Generation of two iPSC lines from vascular Ehlers-Danlos Syndrome (vEDS) patients carrying a missense mutation in COL3A1 gene. Stem cell research Manhas, A., Tripathi, D., Noishiki, C., Wu, D., Liu, L., Sallam, K., Lee, J. T., Fukaya, E., Sayed, N. 2024; 79: 103485

  Abstract

  Vascular Ehlers-Danlos Syndrome (vEDS) is an inherited connective tissue disorder caused by COL3A1 gene, mutations that encodes type III collagen, a crucial component of blood vessels. vEDS can be life-threatening as these patients can have severe internal bleeding due to arterial rupture. Here, we generated induced pluripotent stem cell (iPSC) lines from two vEDS patients carrying a missense mutation in the COL3A1 (c.226A > G, p.Asn76Asp) gene. These lines exhibited typical iPSC characteristics including morphology, expression of pluripotency markers, and could differentiate to all three germ layer. These iPSC lines can serve as valuable tools for elucidating the pathophysiology underlying vEDS.

  View details for DOI 10.1016/j.scr.2024.103485

  View details for PubMedID 38944978

• Artificial intelligence predictive analytics in heart failure: results of the pilot phase of a pragmatic randomized clinical trial. Journal of the American Medical Informatics Association : JAMIA Sideris, K., Weir, C. R., Schmalfuss, C., Hanson, H., Pipke, M., Tseng, P. H., Lewis, N., Sallam, K., Bozkurt, B., Hanff, T., Schofield, R., Larimer, K., Kyriakopoulos, C. P., Taleb, I., Brinker, L., Curry, T., Knecht, C., Butler, J. M., Stehlik, J. 2024

  Abstract

  We conducted an implementation planning process during the pilot phase of a pragmatic trial, which tests an intervention guided by artificial intelligence (AI) analytics sourced from noninvasive monitoring data in heart failure patients (LINK-HF2).A mixed-method analysis was conducted at 2 pilot sites. Interviews were conducted with 12 of 27 enrolled patients and with 13 participating clinicians. iPARIHS constructs were used for interview construction to identify workflow, communication patterns, and clinician's beliefs. Interviews were transcribed and analyzed using inductive coding protocols to identify key themes. Behavioral response data from the AI-generated notifications were collected.Clinicians responded to notifications within 24 hours in 95% of instances, with 26.7% resulting in clinical action. Four implementation themes emerged: (1) High anticipatory expectations for reliable patient communications, reduced patient burden, and less proactive provider monitoring. (2) The AI notifications required a differential and tailored balance of trust and action advice related to role. (3) Clinic experience with other home-based programs influenced utilization. (4) Responding to notifications involved significant effort, including electronic health record (EHR) review, patient contact, and consultation with other clinicians.Clinician's use of AI data is a function of beliefs regarding the trustworthiness and usefulness of the data, the degree of autonomy in professional roles, and the cognitive effort involved.The implementation planning analysis guided development of strategies that addressed communication technology, patient education, and EHR integration to reduce clinician and patient burden in the subsequent main randomized phase of the trial. Our results provide important insights into the unique implications of implementing AI analytics into clinical workflow.

  View details for DOI 10.1093/jamia/ocae017

  View details for PubMedID 38341800

• Almanac - Retrieval-Augmented Language Models for Clinical Medicine. NEJM AI Zakka, C., Shad, R., Chaurasia, A., Dalal, A. R., Kim, J. L., Moor, M., Fong, R., Phillips, C., Alexander, K., Ashley, E., Boyd, J., Boyd, K., Hirsch, K., Langlotz, C., Lee, R., Melia, J., Nelson, J., Sallam, K., Tullis, S., Vogelsong, M. A., Cunningham, J. P., Hiesinger, W. 2024; 1 (2)

  Abstract

  Large language models (LLMs) have recently shown impressive zero-shot capabilities, whereby they can use auxiliary data, without the availability of task-specific training examples, to complete a variety of natural language tasks, such as summarization, dialogue generation, and question answering. However, despite many promising applications of LLMs in clinical medicine, adoption of these models has been limited by their tendency to generate incorrect and sometimes even harmful statements.We tasked a panel of eight board-certified clinicians and two health care practitioners with evaluating Almanac, an LLM framework augmented with retrieval capabilities from curated medical resources for medical guideline and treatment recommendations. The panel compared responses from Almanac and standard LLMs (ChatGPT-4, Bing, and Bard) versus a novel data set of 314 clinical questions spanning nine medical specialties.Almanac showed a significant improvement in performance compared with the standard LLMs across axes of factuality, completeness, user preference, and adversarial safety.Our results show the potential for LLMs with access to domain-specific corpora to be effective in clinical decision-making. The findings also underscore the importance of carefully testing LLMs before deployment to mitigate their shortcomings. (Funded by the National Institutes of Health, National Heart, Lung, and Blood Institute.).

  View details for DOI 10.1056/aioa2300068

  View details for PubMedID 38343631

  View details for PubMedCentralID PMC10857783

• Uptake of sodium-glucose cotransporter-2 inhibitors in hospitalized patients with heart failure: insights from the veterans affairs healthcare system. Journal of cardiac failure Varshney, A. S., Calma, J., Kalwani, N. M., Hsiao, S., Sallam, K., Cao, F., Din, N., Schirmer, J., Bhatt, A. S., Ambrosy, A. P., Heidenreich, P., Sandhu, A. T. 2024

  Abstract

  The use of sodium-glucose cotransporter-2 inhibitor (SGLT2i) in Veteran Affairs (VA) patients hospitalized with heart failure (HF) has not been previously reported.VA electronic health record data were used to identify patients hospitalized for HF (primary or secondary diagnosis) from 01/2019-11/2022. Patients with SGLT2i allergy, advanced/end-stage chronic kidney disease (CKD), or advanced HF therapies were excluded. We identified factors associated with discharge SGLT2i prescription among hospitalizations in 2022. We also compared SGLT2i and angiotensin receptor-neprilysin inhibitor (ARNI) prescription rates. Hospital-level variation in SGLT2i prescription was assessed via the median odds ratio.A total of 69,680 patients were hospitalized for HF; 10.3% were prescribed SGLT2i at discharge (4.4% newly prescribed, 5.9% continued pre-admission therapy). SGLT2i prescription increased over time and was higher in patients with HFrEF and primary HF. Among 15,762 patients hospitalized in 2022, SGLT2i prescription was more likely in patients with diabetes (adjusted odds ratio [aOR] 2.27; 95% confidence interval [CI]: 2.09-2.47) and ischemic heart disease (aOR 1.14; 95% CI: 1.03-1.26). Patients with increased age (aOR 0.77 per 10 years; 95% CI: 0.73-0.80) and lower systolic blood pressure (aOR 0.94 per 10mmHg; 95% CI: 0.92-0.96) were less likely to be prescribed SGLT2i, and SGLT2i prescription was not more likely in patients with CKD (aOR 1.07; 95% CI 0.98-1.16). The adjusted median odds ratio suggested a 1.8-fold variation in the likelihood that similar patients at 2 random VA sites were prescribed SGLT2i (range 0%-21.0%). In patients with EF ≤40%, 30.9% were prescribed SGLT2i while 26.9% were prescribed ARNI (p<0.01).One-tenth of VA patients hospitalized for HF were prescribed SGLT2i at discharge. Opportunities exist to reduce variation in SGLT2i prescription across hospitals and promote use in patients with CKD and older age.

  View details for DOI 10.1016/j.cardfail.2023.12.018

  View details for PubMedID 38281540

• Generation of induced pluripotent stem cell line from a patient suffering from arterial calcification due to deficiency of CD73 (ACDC). Stem cell research Tripathi, D., Manhas, A., Noishiki, C., Wu, D., Adkar, S., Sallam, K., Fukaya, E., Leeper, N. J., Sayed, N. 2023; 75: 103285

  Abstract

  Arterial calcification due to deficiency of CD73 (ACDC) is an adult onset, rare genetic vascular disorder signified by calcium deposition in lower extremity arteries and joints of hands and feet. Mutations in NT5E gene has been shown to be responsible for the inactivation of enzyme CD73 causing calcium buildup. Here, we report a iPSC line generated from a patient showing signs of ACDC and carrying a missense mutation in NT5E (c.1126AG,p.T376A) gene. This iPSC line shows normal morphology, pluripotency, karyotype, and capability to differentiate into three germ layers, making it useful for disease modeling and investigating pathological mechanisms of ACDC.

  View details for DOI 10.1016/j.scr.2023.103285

  View details for PubMedID 38199067

• Generation of two induced pluripotent stem cell lines from hereditary amyloidosis patients with polyneuropathy carrying heterozygous transthyretin (TTR) mutation. Stem cell research Melesio, J., Bonilauri, B., Li, A., Pang, P. D., Liao, R., Witteles, R. M., Wu, J. C., Sallam, K. 2023; 74: 103265

  Abstract

  Hereditary transthyretin amyloidosis with polyneuropathy (ATTR-PN) results from specific TTR gene mutations. In this study, we generated two induced pluripotent stem cell (iPSC) lines derived from ATTR-PN patients with heterozygous TTR gene mutations (Ala97Ser and Phe64Leu). These iPSC lines exhibited normal morphology, karyotype, high pluripotency marker expression, and differentiation into cells representing all germ layers. The generation of these iPSC lines serve as a valuable tool for investigating the mechanisms of ATTR-PN across various cell types and facilitating patient-specific in vitro amyloidosis modeling.

  View details for DOI 10.1016/j.scr.2023.103265

  View details for PubMedID 38100909

• Generation of two induced pluripotent stem cell lines from patients with cardiac amyloidosis carrying heterozygous transthyretin (TTR) mutation STEM CELL RESEARCH Bonilauri, B., Shin, H., Htet, M., Yan, C. D., Witteles, R. M., Sallam, K., Wu, J. C. 2023; 72

  View details for Web of Science ID 001090482900002

• Continuous Wearable Monitoring Analytics To Improve Outcomes In Heart Failure: Vanguard Phase Results And Study Design Of The Randomized Phase Of Link-HF2 Multicenter Study Sideris, K., Weir, C., Schmalfuss, C., Bozkurt, B., Lewis, N., Sallam, K., Hanff, T., Schofield, R., Pipke, M., Larimer, K., Kyriakopoulos, C. P., Davis, C., Beauchamp, B., Hanson, H., Stehlik, J. CHURCHILL LIVINGSTONE INC MEDICAL PUBLISHERS. 2023: 597

  View details for Web of Science ID 001009258900127

• Generation of two induced pluripotent stem cell lines from dilated cardiomyopathy patients carrying heterozygous FLNC mutations. Stem cell research Kojic, A., Kim, H., Guevara, J. V., Ravada, S., Sallam, K., Wu, J. C. 2022; 64: 102928

  Abstract

  Dilated cardiomyopathy (DCM) is a heterogeneous cardiac disorder characterized by left ventricular dilatation and dysfunction. Mutations in dozens of cardiac genes have been connected to the development of DCM including the filamin C gene (FLNC). We generated two induced pluripotent stem cell (iPSCs) lines from DCM patients carrying single missense heterozygote FLNC mutations (c.6689G > A and c.3745G > A). Both lines expressed high levels of pluripotency markers, differentiated into derivatives of the three germ layers and possessed normal karyotypes. The derived iPSC lines can serve as powerful tools to model DCM in vitro and as a platform for therapeutic development.

  View details for DOI 10.1016/j.scr.2022.102928

  View details for PubMedID 36194907

• Generation of two induced pluripotent stem cell lines carrying the phospholamban R14del mutation for modeling ARVD/C. Stem cell research Vera, C. D., Manhas, A., Shenoy, S. P., Wheeler, M. T., Sallam, K., Wu, J. C. 2022; 63: 102834

  Abstract

  The phospholamban (PLN) R14del mutation is associated with arrhythmogenic right ventricular dysplasia (ARVD/C). ARVD/C is a cardiac disease characterized by arrhythmias and structural abnormalities in the right ventricle. Because PLN is a regulator of calcium release, this mutation can have deleterious effects on tissue integrity and contraction. This mutation is a trinucleotide (AGA) deletion that leads to an arginine deletion at position 14 of the PLN structure. Here we show two lines carrying this mutation with typical iPSC morphology, pluripotency, karyotype, ability to differentiate into the three germ layers in vitro, and readily availability for studying pathological mechanisms or ARVD/C.

  View details for DOI 10.1016/j.scr.2022.102834

  View details for PubMedID 35700631

• Modeling Effects of Immunosuppressive Drugs on Human Hearts Using Induced Pluripotent Stem Cell-Derived Cardiac Organoids and Single-Cell RNA Sequencing. Circulation Sallam, K., Thomas, D., Gaddam, S., Lopez, N., Beck, A., Beach, L., Rogers, A. J., Zhang, H., Chen, I. Y., Ameen, M., Hiesinger, W., Teuteberg, J. J., Rhee, J. W., Wang, K. C., Sayed, N., Wu, J. C. 2022; 145 (17): 1367-1369

  View details for DOI 10.1161/CIRCULATIONAHA.121.054317

  View details for PubMedID 35467958

• Heterozygous LMNA mutation-carrying iPSC lines from three cardiac laminopathy patients. Stem cell research Cho, S., Lee, C., Lai, C., Zhuge, Y., Haddad, F., Fowler, M., Sallam, K., Wu, J. C. 1800; 59: 102657

  Abstract

  LMNA-related dilated cardiomyopathy (LMNA-DCM) is caused by pathogenic variants in the LMNA gene and is characterized by left ventricular chamber enlargement, reduced systolic function, and arrhythmia. Here, we generated three human induced pluripotent stem cell (iPSC) lines from peripheral blood mononuclear cells (PBMCs) of three DCM patients carrying the same single heterozygous mutation, c.398 G>A, in LMNA. All lines exhibited normal iPSC morphology, expressed high levels of pluripotency markers, showed normal karyotypes, and could differentiate into the three germ layers. These patient-specific iPSC lines can serve as invaluable tools to model in vitro pathological mechanisms of LMNA-DCM.

  View details for DOI 10.1016/j.scr.2022.102657

  View details for PubMedID 34999423

• Cardiovascular effects of immunosuppression agents. Frontiers in cardiovascular medicine Elezaby, A., Dexheimer, R., Sallam, K. 2022; 9: 981838

  Abstract

  Immunosuppressive medications are widely used to treat patients with neoplasms, autoimmune conditions and solid organ transplants. Key drug classes, namely calcineurin inhibitors, mammalian target of rapamycin (mTOR) inhibitors, and purine synthesis inhibitors, have direct effects on the structure and function of the heart and vascular system. In the heart, immunosuppressive agents modulate cardiac hypertrophy, mitochondrial function, and arrhythmia risk, while in vasculature, they influence vessel remodeling, circulating lipids, and blood pressure. The aim of this review is to present the preclinical and clinical literature examining the cardiovascular effects of immunosuppressive agents, with a specific focus on cyclosporine, tacrolimus, sirolimus, everolimus, mycophenolate, and azathioprine.

  View details for DOI 10.3389/fcvm.2022.981838

  View details for PubMedID 36211586

• Generation of two induced pluripotent stem cell lines from Brugada syndrome affected patients carrying SCN5A mutations. Stem cell research Belbachir, N., Lai, C., Rhee, J., Zhuge, Y., Perez, M. V., Sallam, K., Wu, J. C. 2021; 57: 102605

  Abstract

  SCN5A gene loss-of-function mutations are commonly associated with Brugada syndrome, which represents a risk of lethal arrhythmias and sudden cardiac death. The present report describes the generation of two human induced pluripotent stem cell (iPSC) lines reprogrammed from two Brugada syndrome affected patients carrying SCN5A mutations, c.53506 G>A and c.2102 C>T, respectively. Pluripotency markers, karyotype stability, and differentiation capability into derivatives of the three germ layers were assessed and described in the present report. These lines can be used as a reliable cell model for Brugada syndrome investigations and characterization of leading cellular mechanisms.

  View details for DOI 10.1016/j.scr.2021.102605

  View details for PubMedID 34856468

• Coronary Artery Vasospasm Requiring Cardiac Autotransplantation Yet Controlled With Tobacco. JACC. Case reports Tran, M. V., Marceau, E., Liu, Y., Sallam, K., Medina, P., Liu, C., Sayed, N., Muller, M. D., Liang, D. H., Chen, I. Y. 2021; 3 (9): 1177-1181

  Abstract

  Coronary artery vasospasm is typically managed through avoidance of triggers and with symptomatic treatments with calcium channel blockers and long-acting nitrates. Here, we report a rare case of medically refractory coronary artery vasospasm associated with genetic predispositions that initially required cardiac autotransplantation followed paradoxically by nicotine for long-term symptomatic control. (Level of Difficulty: Intermediate.).

  View details for DOI 10.1016/j.jaccas.2021.03.018

  View details for PubMedID 34401754

• Generation of three induced pluripotent stem cell lines from hypertrophic cardiomyopathy patients carrying MYH7 mutations. Stem cell research Cao, X., Jahng, J. W., Lee, C., Zha, Y., Wheeler, M. T., Sallam, K., Wu, J. C. 2021; 55: 102455

  Abstract

  MYH7 heterozygous mutations are common genetic causes of hypertrophic cardiomyopathy (HCM). HCM is characterized by hypertrophy of the left ventricle and diastolic dysfunction. We generated three human induced pluripotent stem cell (iPSC) lines from three HCM patients each carrying a single heterozygous mutation in MYH7, c.2167C>T, c.4066G>A, and c.5135G>A, respectively. All lines expressed high levels of pluripotent markers, had normal karyotype, and possessed capability of differentiation into derivatives of the three germ layers, which can serve as valuable tools for modeling HCM in vitro and investigating the pathological mechanisms related to MYH7 mutations.

  View details for DOI 10.1016/j.scr.2021.102455

  View details for PubMedID 34352619

• A protocol for transdifferentiation of human cardiac fibroblasts into endothelial cells via activation of innate immunity. STAR protocols Liu, C., Medina, P., Thomas, D., Chen, I. Y., Sallam, K., Sayed, D., Sayed, N. 2021; 2 (2): 100556

  Abstract

  Endothelial cells (ECs) have emerged as key pathogenic players in cardiac disease due to their proximity with cardiomyocytes. Induced pluripotent stem cells (iPSCs) have been employed to generate ECs. However, it may be more clinically relevant to transdifferentiate fibroblasts into ECs directly without introducing pluripotent or virally driven transcription factors. Here, we present a protocol that describes the direct conversion of human cardiac fibroblasts into ECs by leveraging the innate immune system. Our protocol produces bona fide human ECs with 95%-98% purity by first passage. For complete details on the use and execution of this protocol, please refer to Liu etal. (2020) and Sayed etal. (2015).

  View details for DOI 10.1016/j.xpro.2021.100556

  View details for PubMedID 34151292

• The Unfolded Protein Response as a Compensatory Mechanism and Potential Therapeutic Target in PLN R14del Cardiomyopathy. Circulation Feyen, D. A., Perea-Gil, I., Maas, R. G., Harakalova, M., Gavidia, A. A., Arthur Ataam, J., Wu, T., Vink, A., Pei, J., Vadgama, N., Suurmeijer, A. J., Te Rijdt, W. P., Vu, M., Amatya, P. L., Prado, M., Zhang, Y., Dunkenberger, L., Sluijter, J. P., Sallam, K., Asselbergs, F. W., Mercola, M., Karakikes, I. 2021

  Abstract

  Background: Phospholamban (PLN) is a critical regulator of calcium cycling and contractility in the heart. The loss of arginine at position 14 in PLN (R14del) is associated with dilated cardiomyopathy (DCM) with a high prevalence of ventricular arrhythmias. How the R14 deletion causes DCM is poorly understood and there are no disease-specific therapies. Methods: We used single-cell RNA sequencing to uncover PLN R14del disease-mechanisms in human induced pluripotent stem cells (hiPSC-CMs). We utilized both 2D and 3D functional contractility assays to evaluate the impact of modulating disease relevant pathways in PLN R14del hiPSC-CMs. Results: Modeling of the PLN R14del cardiomyopathy with isogenic pairs of hiPSC-CMs recapitulated the contractile deficit associated with the disease in vitro. Single-cell RNA sequencing revealed the induction of the unfolded protein response pathway (UPR) in PLN R14del compared to isogenic control hiPSC-CMs. The activation of UPR was also evident in the hearts from PLN R14del patients. Silencing of each of the three main UPR signaling branches (IRE1, ATF6, or PERK) by siRNA exacerbated the contractile dysfunction of PLN R14del hiPSC-CMs. We explored the therapeutic potential of activating the UPR with a small molecule activator, BiP protein Inducer X (BiX). PLN R14del hiPSC-CMs treated with BiX showed a dose-dependent amelioration of the contractility deficit of in both 2D cultures and 3D engineered heart tissues without affecting calcium homeostasis. Conclusions: Together, these findings suggest that the UPR exerts a protective effect in the setting of PLN R14del cardiomyopathy and that modulation of the UPR might be exploited therapeutically.

  View details for DOI 10.1161/CIRCULATIONAHA.120.049844

  View details for PubMedID 33928785

• Generation of three induced pluripotent stem cell lines, SCVIi003-A, SCVIi004-A, SCVIi005-A, from patients with ARVD/C caused by heterozygous mutations in the PKP2 gene. Stem cell research Jahng, J. W., Black, K. E., Liu, L., Bae, H. R., Perez, M., Ashley, E. A., Sallam, K., Wu, J. C. 2021; 53: 102284

  Abstract

  Arrhythmogenic right ventricular dysplasia/cardiomyopathy (ARVD/C) is an inherited heart disease which can cause life-threatening ventricular arrhythmias and cardiac dysfunction. The autosomal dominant form of ARVD/C is caused by mutations in the cardiac desmosome, such as those in the plakoglobin plakophilin-2 (PKP2) gene. Here, we generated three human induced pluripotent stem cell (iPSC) lines from the peripheral blood mononuclear cells (PBMCs) of three ARVD/C patients carrying pathogenic variants in their PKP2 genes (c.2065_2070delinsG; c.235C>T; c.1725_1728dup). All lines show the typical morphology of pluripotent stem cells, demonstrate high expression of pluripotent markers, display normal karyotype, and differentiate into all three germ layers in vitro. These lines are valuable resources for studying the pathological mechanisms of ARVD/C caused by PKP2 mutation.

  View details for DOI 10.1016/j.scr.2021.102284

  View details for PubMedID 33743362

• Sirolimus Adverse Event Profile in a Non-Clinical Trial Cohort of Heart Transplantation Patients. Annals of transplantation Sallam, K., Bhumireddy, G. P., Evuri, V. D., Abella, J. P., Haddad, F., Valentine, H. A., Nguyen, P. K., Pham, M. X. 2021; 26: e923536

  Abstract

  BACKGROUND Sirolimus has been used increasingly in heart transplantation for its ability to reduce acute rejection, prevent the progression of cardiac allograft vasculopathy (CAV), and preserve renal function. We sought to assess the adverse reactions associated with the use of sirolimus compared to mycophenolate mofetil (MMF). MATERIAL AND METHODS We retrospectively reviewed the charts of 221 adult heart transplant patients who received either sirolimus or MMF as part of their immunosuppression from June 1, 2001 to April 1, 2005. Patients were assigned to 2 groups based upon immunosuppression use. The prevalence and types of complications were recorded in each group. RESULTS Sirolimus was received by 109 patients and 112 patients received MMF during the study period. Seventy-seven patients (71%) in the sirolimus group experienced adverse reactions compared to 45 patients (40%) in the MMF group (P<0.01). Compared to MMF, the use of sirolimus was associated with a higher prevalence of elevated triglyceride levels, lower-extremity edema, and oral ulcerations. Sirolimus was discontinued due to adverse reactions in 22% of patients, whereas no patients in the MMF group experienced adverse effects requiring drug discontinuation. CONCLUSIONS Compared to MMF, sirolimus use is associated with a higher prevalence of adverse reactions requiring drug discontinuation, but most patients were able to stay on therapy despite adverse effects.

  View details for DOI 10.12659/AOT.923536

  View details for PubMedID 33462174

• Clinical trial in a dish using iPSCs shows lovastatin improves endothelial dysfunction and cellular cross-talk in LMNA cardiomyopathy. Science translational medicine Sayed, N., Liu, C., Ameen, M., Himmati, F., Zhang, J. Z., Khanamiri, S., Moonen, J., Wnorowski, A., Cheng, L., Rhee, J., Gaddam, S., Wang, K. C., Sallam, K., Boyd, J. H., Woo, Y. J., Rabinovitch, M., Wu, J. C. 2020; 12 (554)

  Abstract

  Mutations in LMNA, the gene that encodes lamin A and C, causes LMNA-related dilated cardiomyopathy (DCM) or cardiolaminopathy. LMNA is expressed in endothelial cells (ECs); however, little is known about the EC-specific phenotype of LMNA-related DCM. Here, we studied a family affected by DCM due to a frameshift variant in LMNA Human induced pluripotent stem cell (iPSC)-derived ECs were generated from patients with LMNA-related DCM and phenotypically characterized. Patients with LMNA-related DCM exhibited clinical endothelial dysfunction, and their iPSC-ECs showed decreased functionality as seen by impaired angiogenesis and nitric oxide (NO) production. Moreover, genome-edited isogenic iPSC lines recapitulated the EC disease phenotype in which LMNA-corrected iPSC-ECs showed restoration of EC function. Simultaneous profiling of chromatin accessibility and gene expression dynamics by combining assay for transposase-accessible chromatin using sequencing (ATAC-seq) and RNA sequencing (RNA-seq) as well as loss-of-function studies identified Kruppel-like factor 2 (KLF2) as a potential transcription factor responsible for the EC dysfunction. Gain-of-function studies showed that treatment of LMNA iPSC-ECs with KLF2 agonists, including lovastatin, rescued the EC dysfunction. Patients with LMNA-related DCM treated with lovastatin showed improvements in clinical endothelial dysfunction as indicated by increased reactive hyperemia index. Furthermore, iPSC-derived cardiomyocytes (iPSC-CMs) from patients exhibiting the DCM phenotype showed improvement in CM function when cocultured with iPSC-ECs and lovastatin. These results suggest that impaired cross-talk between ECs and CMs can contribute to the pathogenesis of LMNA-related DCM, and statin may be an effective therapy for vascular dysfunction in patients with cardiolaminopathy.

  View details for DOI 10.1126/scitranslmed.aax9276

  View details for PubMedID 32727917

• Modeling Secondary Iron Overload Cardiomyopathy with Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes. Cell reports Rhee, J. W., Yi, H. n., Thomas, D. n., Lam, C. K., Belbachir, N. n., Tian, L. n., Qin, X. n., Malisa, J. n., Lau, E. n., Paik, D. T., Kim, Y. n., Choi, B. S., Sayed, N. n., Sallam, K. n., Liao, R. n., Wu, J. C. 2020; 32 (2): 107886

  Abstract

  Excessive iron accumulation in the heart causes iron overload cardiomyopathy (IOC), which initially presents as diastolic dysfunction and arrhythmia but progresses to systolic dysfunction and end-stage heart failure when left untreated. However, the mechanisms of iron-related cardiac injury and how iron accumulates in human cardiomyocytes are not well understood. Herein, using human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), we model IOC and screen for drugs to rescue the iron overload phenotypes. Human iPSC-CMs under excess iron exposure recapitulate early-stage IOC, including oxidative stress, arrhythmia, and contractile dysfunction. We find that iron-induced changes in calcium kinetics play a critical role in dysregulation of CM functions. We identify that ebselen, a selective divalent metal transporter 1 (DMT1) inhibitor and antioxidant, could prevent the observed iron overload phenotypes, supporting the role of DMT1 in iron uptake into the human myocardium. These results suggest that ebselen may be a potential preventive and therapeutic agent for treating patients with secondary iron overload.

  View details for DOI 10.1016/j.celrep.2020.107886

  View details for PubMedID 32668256

• HIF1α Regulates Early Metabolic Changes due to Activation of Innate Immunity in Nuclear Reprogramming. Stem cell reports Liu, C. n., Ruan, H. n., Himmati, F. n., Zhao, M. T., Chen, C. C., Makar, M. n., Chen, I. Y., Sallam, K. n., Mocarski, E. S., Sayed, D. n., Sayed, N. n. 2020; 14 (2): 192–200

  Abstract

  Innate immune signaling has recently been shown to play an important role in nuclear reprogramming, by altering the epigenetic landscape and thereby facilitating transcription. However, the mechanisms that link innate immune activation and metabolic regulation in pluripotent stem cells remain poorly defined, particularly with regard to key molecular components. In this study, we show that hypoxia-inducible factor 1α (HIF1α), a central regulator of adaptation to limiting oxygen tension, is an unexpected but crucial regulator of innate immune-mediated nuclear reprogramming. HIF1α is dramatically upregulated as a consequence of Toll-like receptor 3 (TLR3) signaling and is necessary for efficient induction of pluripotency and transdifferentiation. Bioenergetics studies reveal that HIF1α regulates the reconfiguration of innate immune-mediated reprogramming through its well-established role in throwing a glycolytic switch. We believe that results from these studies can help us better understand the influence of immune signaling in tissue regeneration and lead to new therapeutic strategies.

  View details for DOI 10.1016/j.stemcr.2020.01.006

  View details for PubMedID 32048999

• Targeted and Selective Treatment of Pluripotent Stem Cell-derived Teratomas Using External Beam Radiation in a Small-animal Model. Journal of visualized experiments : JoVE Sallam, K., Rhee, J., Chour, T., D'addabbo, J., Lee, A. S., Graves, E., Nguyen, P. K. 2019

  Abstract

  The growing number of victims of "stem cell tourism," the unregulated transplantation of stem cells worldwide, has raised concerns about the safety of stem cell transplantation. Although the transplantation of differentiated rather than undifferentiated cells is common practice, teratomas can still arise from the presence of residual undifferentiated stem cells at the time of transplant or from spontaneous mutations in differentiated cells. Because stem cell therapies are often delivered into anatomically sensitive sites, even small tumors can be clinically devastating, resulting in blindness, paralysis, cognitive abnormalities, and cardiovascular dysfunction. Surgical access to these sites may also be limited, leaving patients with few therapeutic options. Controlling stem cell misbehavior is, therefore, critical for the clinical translation of stem cell therapy. External beam radiation offers an effective means of delivering targeted therapy to decrease the teratoma burden while minimizing injury to surrounding organs. Additionally, this method avoids genetic manipulation or viral transduction of stem cells-which are associated with additional clinical safety and efficacy concerns. Here, we describe a protocol to create pluripotent stem cell-derived teratomas in mice and to apply external beam radiation therapy to selectively ablate these tumors in vivo.

  View details for PubMedID 30829317

• Targeted and Selective Treatment of Pluripotent Stem Cell-derived Teratomas Using External Beam Radiation in a Small-animal Model JOVE-JOURNAL OF VISUALIZED EXPERIMENTS Sallam, K., Rhee, J., Chour, T., D'addabbo, J., Lee, A. S., Graves, E., Nguyen, P. K. 2019

  View details for DOI 10.3791/58115

  View details for Web of Science ID 000462905000010

• Electronic Cigarettes: Where There Is Smoke There Is Disease. Journal of the American College of Cardiology Wu, J. C., Rhee, J. W., Sallam, K. n. 2019; 74 (25): 3121–23

  View details for DOI 10.1016/j.jacc.2019.10.029

  View details for PubMedID 31856968

• Modelling diastolic dysfunction in induced pluripotent stem cell-derived cardiomyocytes from hypertrophic cardiomyopathy patients. European heart journal Wu, H. n., Yang, H. n., Rhee, J. W., Zhang, J. Z., Lam, C. K., Sallam, K. n., Chang, A. C., Ma, N. n., Lee, J. n., Zhang, H. n., Blau, H. M., Bers, D. M., Wu, J. C. 2019

  Abstract

  Diastolic dysfunction (DD) is common among hypertrophic cardiomyopathy (HCM) patients, causing major morbidity and mortality. However, its cellular mechanisms are not fully understood, and presently there is no effective treatment. Patient-specific induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) hold great potential for investigating the mechanisms underlying DD in HCM and as a platform for drug discovery.In the present study, beating iPSC-CMs were generated from healthy controls and HCM patients with DD. Micropatterned iPSC-CMs from HCM patients showed impaired diastolic function, as evidenced by prolonged relaxation time, decreased relaxation rate, and shortened diastolic sarcomere length. Ratiometric Ca2+ imaging indicated elevated diastolic [Ca2+]i and abnormal Ca2+ handling in HCM iPSC-CMs, which were exacerbated by β-adrenergic challenge. Combining Ca2+ imaging and traction force microscopy, we observed enhanced myofilament Ca2+ sensitivity (measured as dF/Δ[Ca2+]i) in HCM iPSC-CMs. These results were confirmed with genome-edited isogenic iPSC lines that carry HCM mutations, indicating that cytosolic diastolic Ca2+ overload, slowed [Ca2+]i recycling, and increased myofilament Ca2+ sensitivity, collectively impairing the relaxation of HCM iPSC-CMs. Treatment with partial blockade of Ca2+ or late Na+ current reset diastolic Ca2+ homeostasis, restored diastolic function, and improved long-term survival, suggesting that disturbed Ca2+ signalling is an important cellular pathological mechanism of DD. Further investigation showed increased expression of L-type Ca2+channel (LTCC) and transient receptor potential cation channels (TRPC) in HCM iPSC-CMs compared with control iPSC-CMs, which likely contributed to diastolic [Ca2+]i overload.In summary, this study recapitulated DD in HCM at the single-cell level, and revealed novel cellular mechanisms and potential therapeutic targets of DD using iPSC-CMs.

  View details for DOI 10.1093/eurheartj/ehz326

  View details for PubMedID 31219556

• Activation of PDGF pathway links LMNA mutation to dilated cardiomyopathy. Nature Lee, J. n., Termglinchan, V. n., Diecke, S. n., Itzhaki, I. n., Lam, C. K., Garg, P. n., Lau, E. n., Greenhaw, M. n., Seeger, T. n., Wu, H. n., Zhang, J. Z., Chen, X. n., Gil, I. P., Ameen, M. n., Sallam, K. n., Rhee, J. W., Churko, J. M., Chaudhary, R. n., Chour, T. n., Wang, P. J., Snyder, M. P., Chang, H. Y., Karakikes, I. n., Wu, J. C. 2019

  Abstract

  Lamin A/C (LMNA) is one of the most frequently mutated genes associated with dilated cardiomyopathy (DCM). DCM related to mutations in LMNA is a common inherited cardiomyopathy that is associated with systolic dysfunction and cardiac arrhythmias. Here we modelled the LMNA-related DCM in vitro using patient-specific induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). Electrophysiological studies showed that the mutant iPSC-CMs displayed aberrant calcium homeostasis that led to arrhythmias at the single-cell level. Mechanistically, we show that the platelet-derived growth factor (PDGF) signalling pathway is activated in mutant iPSC-CMs compared to isogenic control iPSC-CMs. Conversely, pharmacological and molecular inhibition of the PDGF signalling pathway ameliorated the arrhythmic phenotypes of mutant iPSC-CMs in vitro. Taken together, our findings suggest that the activation of the PDGF pathway contributes to the pathogenesis of LMNA-related DCM and point to PDGF receptor-β (PDGFRB) as a potential therapeutic target.

  View details for DOI 10.1038/s41586-019-1406-x

  View details for PubMedID 31316208

• The Incremental Value of Right Ventricular Size and Strain in the Risk Assessment of Right Heart Failure Post - Left Ventricular Assist Device Implantation. Journal of cardiac failure Aymami, M., Amsallem, M., Adams, J., Sallam, K., Moneghetti, K., Wheeler, M., Hiesinger, W., Teuteberg, J., Weisshaar, D., Verhoye, J., Woo, Y. J., Ha, R., Haddad, F., Banerjee, D. 2018; 24 (12): 823–32

  Abstract

  BACKGROUND: Right heart failure (RHF) after left ventricular assist device (LVAD) implantation is associated with high morbidity and mortality. Existing risk scores include semiquantitative evaluation of right ventricular (RV) dysfunction. This study aimed to determine whether quantitative evaluation of both RV size and function improve risk stratification for RHF after LVAD implantation beyond validated scores.METHODS AND RESULTS: From 2009 to 2015, 158 patients who underwent implantation of continuous-flow devices who had complete echocardiographic and hemodynamic data were included. Quantitative RV parameters included RV end-diastolic (RVEDAI) and end-systolic area index, RV free-wall longitudinal strain (RVLS), fractional area change, tricuspid annular plane systolic excursion, and right atrial area and pressure. Independent correlates of early RHF (<30 days) were determined with the use of logistic regression analysis. Mean age was 56 ± 13 years, with 79% male; 49% had INTERMACS profiles ≤2. RHF occurred in 60 patients (38%), with 20 (13%) requiring right ventricular assist device. On multivariate analysis, INTERMACS profiles (adjusted odds ratio 2.38 [95% confidence interval [CI] 1.47-3.85]), RVEDAI (1.61 [1.08-2.32]), and RVLS (2.72 [1.65-4.51]) were independent correlates of RHF (all P < .05). Both RVLS and RVEDAI were incremental to validated risk scores (including the EUROMACS score) for early RHF after LVAD (all P < .01).CONCLUSIONS: RV end-diastolic and strain are complementary prognostic markers of RHF after LVAD implantation.

  View details for PubMedID 30539717

• Genome Editing of Induced PluripotentStem Cells to Decipher CardiacChannelopathy Variant. Journal of the American College of Cardiology Garg, P., Oikonomopoulos, A., Chen, H., Li, Y., Lam, C. K., Sallam, K., Perez, M., Lux, R. L., Sanguinetti, M. C., Wu, J. C. 2018; 72 (1): 62–75

  Abstract

  BACKGROUND: The long QT syndrome (LQTS) is an arrhythmogenic disorder of QT interval prolongation that predisposes patients to life-threatening ventricular arrhythmias such as Torsades de pointes and sudden cardiac death. Clinical genetic testing has emerged as the standard of care to identify genetic variants in patients suspected of having LQTS. However, these results are often confounded by the discovery of variants of uncertain significance (VUS), for which there is insufficient evidence of pathogenicity.OBJECTIVES: The purpose of this study was to demonstrate that genome editing of patient-specific induced pluripotent stem cells (iPSCs) can be a valuable approach to delineate the pathogenicity of VUS in cardiac channelopathy.METHODS: Peripheral blood mononuclear cells were isolated from a carrier with a novel missense variant (T983I) in the KCNH2 (LQT2) gene and an unrelated healthy control subject. iPSCs were generated using an integration-free Sendai virus and differentiated to iPSC-derived cardiomyocytes (CMs).RESULTS: Whole-cell patch clamp recordings revealed significant prolongation of the action potential duration (APD) and reduced rapidly activating delayed rectifier K+ current (IKr) density in VUS iPSC-CMs compared with healthy control iPSC-CMs. ICA-105574, a potent IKr activator, enhanced IKr magnitude and restored normal action potential duration in VUS iPSC-CMs. Notably, VUS iPSC-CMs exhibited greater propensity to proarrhythmia than healthy control cells in response to high-risk torsadogenic drugs (dofetilide, ibutilide, and azimilide), suggesting a compromised repolarization reserve. Finally, the selective correction of the causal variant in iPSC-CMs using CRISPR/Cas9 gene editing (isogenic control) normalized the aberrant cellular phenotype, whereas the introduction of the homozygous variant in healthy control cells recapitulated hallmark features of the LQTS disorder.CONCLUSIONS: The results suggest that the KCNH2T983I VUS may be classified as potentially pathogenic.

  View details for PubMedID 29957233

• RIGHT HEART MALADAPTIVE PHENOTYPES AND PREDICTION OF RIGHT HEART FAILURE FOLLOWING CONTINUOUS-FLOW LEFT VENTRICULAR ASSIST DEVICE IMPLANTATION Aymami, M., Haddad, F., Amsallem, M., Wheeler, M., Moneghetti, K., Adams, J., Verhoye, J., Sallam, K., Woo, Y., Ha, R., Banerjee, D. ELSEVIER SCIENCE INC. 2018: 652

  View details for DOI 10.1016/S0735-1097(18)31193-8

  View details for Web of Science ID 000429659701502

• Transcriptomic and epigenomic differences in human induced pluripotent stem cells generated from six reprogramming methods NATURE BIOMEDICAL ENGINEERING Churko, J. M., Lee, J., Ameen, M., Gu, M., Venkatasubramanian, M., Diecke, S., Sallam, K., Im, H., Wang, G., Gold, J. D., Salomonis, N., Snyder, M. P., Wu, J. C. 2017; 1 (10): 826–37

  View details for DOI 10.1038/s41551-017-0141-6

  View details for Web of Science ID 000418860600003

• Transcriptomic and epigenomic differences in human induced pluripotent stem cells generated from six reprogramming methods. Nature biomedical engineering Churko, J. M., Lee, J., Ameen, M., Gu, M., Venkatasubramanian, M., Diecke, S., Sallam, K., Im, H., Wang, G., Gold, J. D., Salomonis, N., Snyder, M. P., Wu, J. C. 2017; 1 (10): 826-837

  Abstract

  Many reprogramming methods can generate human induced pluripotent stem cells (hiPSCs) that closely resemble human embryonic stem cells (hESCs). This has led to assessments of how similar hiPSCs are to hESCs, by evaluating differences in gene expression, epigenetic marks and differentiation potential. However, all previous studies were performed using hiPSCs acquired from different laboratories, passage numbers, culturing conditions, genetic backgrounds and reprogramming methods, all of which may contribute to the reported differences. Here, by using high-throughput sequencing under standardized cell culturing conditions and passage number, we compare the epigenetic signatures (H3K4me3, H3K27me3 and HDAC2 ChIP-seq profiles) and transcriptome differences (by RNA-seq) of hiPSCs generated from the same primary fibroblast population by using six different reprogramming methods. We found that the reprogramming method impacts the resulting transcriptome and that all hiPSC lines could terminally differentiate, regardless of the reprogramming method. Moreover, by comparing the differences between the hiPSC and hESC lines, we observed a significant proportion of differentially expressed genes that could be attributed to polycomb repressive complex targets.

  View details for DOI 10.1038/s41551-017-0141-6

  View details for PubMedID 30263871

  View details for PubMedCentralID PMC6155993

• Simultaneous ramp right heart catheterization and echocardiography in a ReliantHeart left ventricular assist device. World journal of cardiology Banerjee, D., Dutt, D., Duclos, S., Sallam, K., Wheeler, M., Ha, R. 2017; 9 (1): 55-59

  Abstract

  Many clinicians caring for patients with continuous flow left ventricular assist devices (CF-LVAD) use ramp right heart catheterization (RHC) studies to optimize pump speed and also to troubleshoot CF-LVAD malfunction. An investigational device, the ReliantHeart Heart Assist 5 (Houston, TX), provides the added benefit of an ultrasonic flow probe on the outflow graft that directly measures flow through the CF-LVAD. We performed a simultaneous ramp RHC and echocardiogram on a patient who received the above CF-LVAD to optimize pump parameters and investigate elevated flow through the CF-LVAD as measured by the flow probe. We found that the patient's hemodynamics were optimized at their baseline pump speed, and that the measured cardiac output via the Fick principle was lower than that measured by the flow probe. Right heart catheterization may be useful to investigate discrepancies between flow measured by a CF-LVAD and a patient's clinical presentation, particularly in investigational devices where little clinical experience exists. More data is needed to elucidate the correlation between the flow measured by an ultrasonic probe and cardiac output as measured by RHC.

  View details for DOI 10.4330/wjc.v9.i1.55

  View details for PubMedID 28163837

  View details for PubMedCentralID PMC5253195

• Patient-Specific Induced Pluripotent Stem Cell-Based Disease Model for Pathogenesis Studies and Clinical Pharmacotherapy. Circulation. Arrhythmia and electrophysiology Li, Y. n., Sallam, K. n., Schwartz, P. J., Wu, J. C. 2017; 10 (6)

  View details for PubMedID 28630175

• Patient-Specific and Genome-Edited Induced Pluripotent Stem Cell-Derived Cardiomyocytes Elucidate Single-Cell Phenotype of Brugada Syndrome. Journal of the American College of Cardiology Liang, P., Sallam, K., Wu, H., Li, Y., Itzhaki, I., Garg, P., Zhang, Y., Vermglinchan, V., Lan, F., Gu, M., Gong, T., Zhuge, Y., He, C., Ebert, A. D., Sanchez-Freire, V., Churko, J., Hu, S., Sharma, A., Lam, C. K., Scheinman, M. M., Bers, D. M., Wu, J. C. 2016; 68 (19): 2086-2096

  Abstract

  Brugada syndrome (BrS), a disorder associated with characteristic electrocardiogram precordial ST-segment elevation, predisposes afflicted patients to ventricular fibrillation and sudden cardiac death. Despite marked achievements in outlining the organ level pathophysiology of the disorder, the understanding of human cellular phenotype has lagged due to a lack of adequate human cellular models of the disorder.The objective of this study was to examine single cell mechanism of Brugada syndrome using induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs).This study recruited 2 patients with type 1 BrS carrying 2 different sodium voltage-gated channel alpha subunit 5 variants as well as 2 healthy control subjects. We generated iPSCs from their skin fibroblasts by using integration-free Sendai virus. We used directed differentiation to create purified populations of iPSC-CMs.BrS iPSC-CMs showed reductions in inward sodium current density and reduced maximal upstroke velocity of action potential compared with healthy control iPSC-CMs. Furthermore, BrS iPSC-CMs demonstrated increased burden of triggered activity, abnormal calcium (Ca(2+)) transients, and beating interval variation. Correction of the causative variant by genome editing was performed, and resultant iPSC-CMs showed resolution of triggered activity and abnormal Ca(2+) transients. Gene expression profiling of iPSC-CMs showed clustering of BrS compared with control subjects. Furthermore, BrS iPSC-CM gene expression correlated with gene expression from BrS human cardiac tissue gene expression.Patient-specific iPSC-CMs were able to recapitulate single-cell phenotype features of BrS, including blunted inward sodium current, increased triggered activity, and abnormal Ca(2+) handling. This novel human cellular model creates future opportunities to further elucidate the cellular disease mechanism and identify novel therapeutic targets.

  View details for DOI 10.1016/j.jacc.2016.07.779

  View details for PubMedID 27810048

• A Rapid, High-Quality, Cost-Effective, Comprehensive and Expandable Targeted Next-Generation Sequencing Assay for Inherited Heart Diseases. Circulation research Wilson, K. D., Shen, P., Fung, E., Karakikes, I., Zhang, A., Inanloorahatloo, K., Odegaard, J., Sallam, K., Davis, R. W., Lui, G. K., Ashley, E. A., Scharfe, C., Wu, J. C. 2015; 117 (7): 603-611

  Abstract

  Thousands of mutations across >50 genes have been implicated in inherited cardiomyopathies. However, options for sequencing this rapidly evolving gene set are limited because many sequencing services and off-the-shelf kits suffer from slow turnaround, inefficient capture of genomic DNA, and high cost. Furthermore, customization of these assays to cover emerging targets that suit individual needs is often expensive and time consuming.We sought to develop a custom high throughput, clinical-grade next-generation sequencing assay for detecting cardiac disease gene mutations with improved accuracy, flexibility, turnaround, and cost.We used double-stranded probes (complementary long padlock probes), an inexpensive and customizable capture technology, to efficiently capture and amplify the entire coding region and flanking intronic and regulatory sequences of 88 genes and 40 microRNAs associated with inherited cardiomyopathies, congenital heart disease, and cardiac development. Multiplexing 11 samples per sequencing run resulted in a mean base pair coverage of 420, of which 97% had >20× coverage and >99% were concordant with known heterozygous single nucleotide polymorphisms. The assay correctly detected germline variants in 24 individuals and revealed several polymorphic regions in miR-499. Total run time was 3 days at an approximate cost of $100 per sample.Accurate, high-throughput detection of mutations across numerous cardiac genes is achievable with complementary long padlock probe technology. Moreover, this format allows facile insertion of additional probes as more cardiomyopathy and congenital heart disease genes are discovered, giving researchers a powerful new tool for DNA mutation detection and discovery.

  View details for DOI 10.1161/CIRCRESAHA.115.306723

  View details for PubMedID 26265630

• Epigenetic Regulation of Phosphodiesterases 2A and 3A Underlies Compromised ß-Adrenergic Signaling in an iPSC Model of Dilated Cardiomyopathy. Cell stem cell Wu, H., Lee, J., Vincent, L. G., Wang, Q., Gu, M., Lan, F., Churko, J. M., Sallam, K. I., Matsa, E., Sharma, A., Gold, J. D., Engler, A. J., Xiang, Y. K., Bers, D. M., Wu, J. C. 2015; 17 (1): 89-100

  Abstract

  β-adrenergic signaling pathways mediate key aspects of cardiac function. Its dysregulation is associated with a range of cardiac diseases, including dilated cardiomyopathy (DCM). Previously, we established an iPSC model of familial DCM from patients with a mutation in TNNT2, a sarcomeric protein. Here, we found that the β-adrenergic agonist isoproterenol induced mature β-adrenergic signaling in iPSC-derived cardiomyocytes (iPSC-CMs) but that this pathway was blunted in DCM iPSC-CMs. Although expression levels of several β-adrenergic signaling components were unaltered between control and DCM iPSC-CMs, we found that phosphodiesterases (PDEs) 2A and PDE3A were upregulated in DCM iPSC-CMs and that PDE2A was also upregulated in DCM patient tissue. We further discovered increased nuclear localization of mutant TNNT2 and epigenetic modifications of PDE genes in both DCM iPSC-CMs and patient tissue. Notably, pharmacologic inhibition of PDE2A and PDE3A restored cAMP levels and ameliorated the impaired β-adrenergic signaling of DCM iPSC-CMs, suggesting therapeutic potential.

  View details for DOI 10.1016/j.stem.2015.04.020

  View details for PubMedID 26095046

• Finding the rhythm of sudden cardiac death: new opportunities using induced pluripotent stem cell-derived cardiomyocytes. Circulation research Sallam, K., Li, Y., Sager, P. T., Houser, S. R., Wu, J. C. 2015; 116 (12): 1989-2004

  Abstract

  Sudden cardiac death is a common cause of death in patients with structural heart disease, genetic mutations, or acquired disorders affecting cardiac ion channels. A wide range of platforms exist to model and study disorders associated with sudden cardiac death. Human clinical studies are cumbersome and are thwarted by the extent of investigation that can be performed on human subjects. Animal models are limited by their degree of homology to human cardiac electrophysiology, including ion channel expression. Most commonly used cellular models are cellular transfection models, which are able to mimic the expression of a single-ion channel offering incomplete insight into changes of the action potential profile. Induced pluripotent stem cell-derived cardiomyocytes resemble, but are not identical, adult human cardiomyocytes and provide a new platform for studying arrhythmic disorders leading to sudden cardiac death. A variety of platforms exist to phenotype cellular models, including conventional and automated patch clamp, multielectrode array, and computational modeling. Induced pluripotent stem cell-derived cardiomyocytes have been used to study long QT syndrome, catecholaminergic polymorphic ventricular tachycardia, hypertrophic cardiomyopathy, and other hereditary cardiac disorders. Although induced pluripotent stem cell-derived cardiomyocytes are distinct from adult cardiomyocytes, they provide a robust platform to advance the science and clinical care of sudden cardiac death.

  View details for DOI 10.1161/CIRCRESAHA.116.304494

  View details for PubMedID 26044252

• Establishing disease causality for a novel gene variant in familial dilated cardiomyopathy using a functional in-vitro assay of regulated thin filaments and human cardiac myosin. BMC medical genetics Pan, S., Sommese, R. F., Sallam, K. I., Nag, S., Sutton, S., Miller, S. M., Spudich, J. A., Ruppel, K. M., Ashley, E. A. 2015; 16 (1): 97-?

  Abstract

  As next generation sequencing for the genetic diagnosis of cardiovascular disorders becomes more widely used, establishing causality for putative disease causing variants becomes increasingly relevant. Diseases of the cardiac sarcomere provide a particular challenge in this regard because of the complexity of assaying the effect of genetic variants in human cardiac contractile proteins.In this study we identified a novel variant R205Q in the cardiac troponin T gene (TNNT2). Carriers of the variant allele exhibited increased chamber volumes associated with decreased left ventricular ejection fraction. To clarify the causal role of this variant, we generated recombinant variant human protein and examined its calcium kinetics as well as the maximally activated ADP release of human β-cardiac myosin with regulated thin filaments containing the mutant troponin T. We found that the R205Q mutation significantly decreased the calcium sensitivity of the thin filament by altering the effective calcium dissociation kinetics.The development of moderate throughput post-genomic assays is an essential step in the realization of the potential of next generation sequencing. Although technically challenging, biochemical and functional assays of human cardiac contractile proteins of the thin filament can be achieved and provide an orthogonal source of information to inform the question of causality for individual variants.

  View details for DOI 10.1186/s12881-015-0243-5

  View details for PubMedID 26498512

  View details for PubMedCentralID PMC4620603

• Human induced pluripotent stem cell-derived cardiomyocytes as an in vitro model for coxsackievirus B3-induced myocarditis and antiviral drug screening platform. Circulation research Sharma, A., Marceau, C., Hamaguchi, R., Burridge, P. W., Rajarajan, K., Churko, J. M., Wu, H., Sallam, K. I., Matsa, E., Sturzu, A. C., Che, Y., Ebert, A., Diecke, S., Liang, P., Red-Horse, K., Carette, J. E., Wu, S. M., Wu, J. C. 2014; 115 (6): 556-566

  Abstract

  Rationale: Viral myocarditis is a life-threatening illness that may lead to heart failure or cardiac arrhythmias. A major causative agent for viral myocarditis is the B3 strain of coxsackievirus, a positive-sense RNA enterovirus. However, human cardiac tissues are difficult to procure in sufficient enough quantities for studying the mechanisms of cardiac-specific viral infection. Objective: This study examined whether human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) could be used to model the pathogenic processes of coxsackievirus-induced viral myocarditis and to screen antiviral therapeutics for efficacy. Methods and Results: Human iPSC-CMs were infected with a luciferase-expressing coxsackievirus B3 strain (CVB3-Luc). Brightfield microscopy, immunofluorescence, and calcium imaging were utilized to characterize virally-infected hiPSC-CMs for alterations in cellular morphology and calcium handling. Viral proliferation in hiPSC-CMs was quantified using bioluminescence imaging. Antiviral compounds including interferon beta 1 (IFNβ1), ribavirin, pyrrolidine dithiocarbamate, and fluoxetine were tested for their capacity to abrogate CVB3-Luc proliferation in hiPSC-CMs in vitro. The ability of these compounds to reduce CVB3-Luc proliferation in hiPSC-CMs was consistent with reported drug effects in previous studies. Mechanistic analyses via gene expression profiling of hiPSC-CMs infected with CVB3-Luc revealed an activation of viral RNA and protein clearance pathways after IFNβ1 treatment. Conclusions: This study demonstrates that hiPSC-CMs express the coxsackievirus and adenovirus receptor, are susceptible to coxsackievirus infection, and can be used to predict antiviral drug efficacy. Our results suggest that the hiPSC-CM/CVB3-Luc assay is a sensitive platform that can screen novel antiviral therapeutics for their effectiveness in a high-throughput fashion.

  View details for DOI 10.1161/CIRCRESAHA.115.303810

  View details for PubMedID 25015077

• Modeling inherited cardiac disorders. Circulation journal Sallam, K., Kodo, K., Wu, J. C. 2014; 78 (4): 784-794

  Abstract

  Advances in the understanding and treatment of cardiac disorders have been thwarted by the inability to study beating human cardiac cells in vitro. Induced pluripotent stem cells (iPSCs) bypass this hurdle by enabling the creation of patient-specific iPSC-derived cardiomyocytes (iPSC-CMs). These cells provide a unique platform to study cardiac diseases in vitro, especially hereditary cardiac conditions. To date, iPSC-CMs have been used to successfully model arrhythmic disorders, showing excellent recapitulation of cardiac channel function and electrophysiologic features of long QT syndrome types 1, 2, 3, and 8, and catecholaminergic polymorphic ventricular tachycardia (CPVT). Similarly, iPSC-CM models of dilated cardiomyopathy (DCM) and hypertrophic cardiomyopathy (HCM) have shown robust correlation of predicted morphologic, contractile, and electrical phenotypes. In addition, iPSC-CMs have shown some features of the respective phenotypes for arrhythmogenic right ventricular dysplasia/cardiomyopathy (ARVD/C), LEOPARD syndrome, Pompe's disease, and Friedriech's ataxia. In this review, we examine the progress of utilizing iPSC-CMs as a model for cardiac conditions and analyze the potential for the platform in furthering the biology and treatment of cardiac disorders.  

  View details for PubMedID 24632794

• Cardiac stem cell biology: glimpse of the past, present, and future. Circulation research Matsa, E., Sallam, K., Wu, J. C. 2014; 114 (1): 21-27

  Abstract

  Cardiac regeneration strategies and de novo generation of cardiomyocytes have long been significant areas of research interest in cardiovascular medicine. In this review, we outline a variety of common cell sources and methods used to regenerate cardiomyocytes and highlight the important role that key Circulation Research articles have played in this flourishing field.

  View details for DOI 10.1161/CIRCRESAHA.113.302895

  View details for PubMedID 24385505

• Concomitant ECG findings and J wave patterns. Journal of electrocardiology Sallam, K., Froelicher, V. 2013; 46 (5): 399-403

  View details for DOI 10.1016/j.jelectrocard.2013.06.022

  View details for PubMedID 23981305

• Drug screening using a library of human induced pluripotent stem cell-derived cardiomyocytes reveals disease-specific patterns of cardiotoxicity. Circulation Liang, P., Lan, F., Lee, A. S., Gong, T., Sanchez-Freire, V., Wang, Y., Diecke, S., Sallam, K., Knowles, J. W., Wang, P. J., Nguyen, P. K., Bers, D. M., Robbins, R. C., Wu, J. C. 2013; 127 (16): 1677-1691

  Abstract

  Cardiotoxicity is a leading cause for drug attrition during pharmaceutical development and has resulted in numerous preventable patient deaths. Incidents of adverse cardiac drug reactions are more common in patients with preexisting heart disease than the general population. Here we generated a library of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) from patients with various hereditary cardiac disorders to model differences in cardiac drug toxicity susceptibility for patients of different genetic backgrounds.Action potential duration and drug-induced arrhythmia were measured at the single cell level in hiPSC-CMs derived from healthy subjects and patients with hereditary long QT syndrome, familial hypertrophic cardiomyopathy, and familial dilated cardiomyopathy. Disease phenotypes were verified in long QT syndrome, hypertrophic cardiomyopathy, and dilated cardiomyopathy hiPSC-CMs by immunostaining and single cell patch clamp. Human embryonic stem cell-derived cardiomyocytes (hESC-CMs) and the human ether-a-go-go-related gene expressing human embryonic kidney cells were used as controls. Single cell PCR confirmed expression of all cardiac ion channels in patient-specific hiPSC-CMs as well as hESC-CMs, but not in human embryonic kidney cells. Disease-specific hiPSC-CMs demonstrated increased susceptibility to known cardiotoxic drugs as measured by action potential duration and quantification of drug-induced arrhythmias such as early afterdepolarizations and delayed afterdepolarizations.We have recapitulated drug-induced cardiotoxicity profiles for healthy subjects, long QT syndrome, hypertrophic cardiomyopathy, and dilated cardiomyopathy patients at the single cell level for the first time. Our data indicate that healthy and diseased individuals exhibit different susceptibilities to cardiotoxic drugs and that use of disease-specific hiPSC-CMs may predict adverse drug responses more accurately than the standard human ether-a-go-go-related gene test or healthy control hiPSC-CM/hESC-CM screening assays.

  View details for DOI 10.1161/CIRCULATIONAHA.113.001883

  View details for PubMedID 23519760

  View details for PubMedCentralID PMC3870148

• Natural History of Early Repolarization in the Inferior Leads ANNALS OF NONINVASIVE ELECTROCARDIOLOGY Stein, R., Sallam, K., Adhikarla, C., Boga, M., Wood, A. D., Froelicher, V. F. 2012; 17 (4): 331-339

  Abstract

  Though early repolarization (ER) in the inferior leads has been associated with increased cardiovascular risk, its natural history is uncertain. We aimed to study the serial electrocardiographic behavior of inferior ER and understand factors associated with that behavior.We selected electrocardiograms (ECGs) from patients with the greatest amplitude of ER in AVF from ECGs of 29,281 ambulatory patients recorded between 1987 and 1999 at the Palo Alto Veterans Affairs Hospital. Starting from the highest amplitude, we reviewed the ECGs and medical records from the first 85%. From this convenience sample, 36 were excluded for abnormal patterns similar to ER. The remaining 257 patients were searched for another ECG at least 5 months later, of whom, 136 satisfied this criteria. These ECGs were paired for comparison and coded by four interpreters.The average time between the first and second ECGs was 10 years. Of the 136 subjects, 47% retained ER while 53% no longer fulfilled the amplitude criteria. While no significant differences were found in initial heart rate (HR) or time interval between ECGs, those who lost the ER pattern had a greater difference in HR between the ECGs. There was no significant difference in the incidence of cardiovascular events or deaths.In conclusion, the ECG pattern of ER was lost over 10 years in over half of the cohort. The loss of ER was partially explained by changes in HR, but not higher incidence of cardiovascular events or death, suggesting the entity is a benign finding.

  View details for DOI 10.1111/j.1542-474X.2012.00537.x

  View details for Web of Science ID 000310248100005

  View details for PubMedID 23094879

• Prognostic Implications of Q Waves and T-Wave Inversion Associated With Early Repolarization MAYO CLINIC PROCEEDINGS Uberoi, A., Sallam, K., Perez, M., Jain, N. A., Ashley, E., Froelicher, V. 2012; 87 (7): 614-619

  Abstract

  To evaluate the prevalence of early polarization (ER) in a stable population and to evaluate the prognostic significance of the association or absence of Q waves or T-wave inversion (TWI).In this retrospective study performed at the university-affiliated Palo Alto Veterans Affairs Health Care Center from March 1, 1987, through December 31, 1999, we evaluated outpatient electrocardiograms. Vital status and cause of death were determined in all patients, with a mean ± SD follow-up of 7.6±3.8 years.Of the 29,281 patients, 87% were men and 13% were African American. Inferior or lateral ER was present in 664 patients (2.3%): in inferior leads in 185 (0.6%), in lateral leads in 479 (1.6%) , and in both inferior and lateral leads in 163 (0.6%). Only when Q waves or TWI accompanied ER was there an increased risk of cardiovascular death (Cox proportional hazards regression model, 5.0; 95% confidence interval, 3.4-7.2; P<.001).Common patterns of ER without concomitant Q waves or TWI are not associated with increased risk of cardiovascular death; however, when either occurs with ER, there is a hazard ratio of 5.0. These findings confirm that ER is a benign entity; however, the presence of Q waves or TWI with ER is predictive of increased cardiovascular death.

  View details for DOI 10.1016/j.mayocp.2012.04.009

  View details for PubMedID 22766081

• COMPARISON OF EARLY REPOLARIZATION IN INFERIOR AND LATERAL LEADS 61st Annual Scientific Session and Expo of the American-College-of-Cardiology (ACC)/Conference on ACC-i2 with TCT Downing, N. L., Abhimanyu, Uberoi, M., Sallam, K., Sadik, J., Adhikarla, C., Froelicher, V. ELSEVIER SCIENCE INC. 2012: E1940–E1940

  View details for Web of Science ID 000302326702151

• Embryonic stem cell biology: insights from molecular imaging. Methods in molecular biology (Clifton, N.J.) Sallam, K., Wu, J. C. 2010; 660: 185-199

  Abstract

  Embryonic stem (ES) cells have therapeutic potential in disorders of cellular loss such as myocardial infarction, type I diabetes and neurodegenerative disorders. ES cell biology in living subjects was largely poorly understood until incorporation of molecular imaging into the field. Reporter gene imaging works by integrating a reporter gene into ES cells and using a reporter probe to induce a signal detectable by normal imaging modalities. Reporter gene imaging allows for longitudinal tracking of ES cells within the same host for a prolonged period of time. This has advantages over postmortem immunohistochemistry and traditional imaging modalities. The advantages include expression of reporter gene is limited to viable cells, expression is conserved between generations of dividing cells, and expression can be linked to a specific population of cells. These advantages were especially useful in studying a dynamic cell population such as ES cells and proved useful in elucidating the biology of ES cells. Reporter gene imaging identified poor integration of differentiated ES cells transplanted into host tissue as well as delayed donor cell death as reasons for poor long-term survival in vivo. This imaging technology also confirmed that ES cells indeed have immunogenic properties that factor into cell survival and differentiation. Finally, reporter gene imaging improved our understanding of the neoplastic risk of undifferentiated ES cells in forming teratomas. Despite such advances, much remains to be understood about ES cell biology to translate this technology to the bedside, and reporter gene imaging will certainly play a key role in formulating this understanding.

  View details for DOI 10.1007/978-1-60761-705-1_12

  View details for PubMedID 20680820