scholarly journals SUN-556 Cardiac Phenotype in Familial Partial Lipodystrophy

2020 ◽  
Vol 4 (Supplement_1) ◽  
Author(s):  
Abdelwahab Jalal Eldin ◽  
Baris Akinci ◽  
Andre Monteiro da Rocha ◽  
Rasimcan Meral ◽  
Ilgin Yildirim Simsir ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Stem Cell ◽  
Action Potential Duration ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Conduction System ◽  
Partial Lipodystrophy ◽  
Pathogenic Variants ◽  
Familial Partial Lipodystrophy ◽  
Induced Pluripotent

Abstract Background Pathogenic variants in Lamin A/C (LMNA) gene are the most common monogenic etiology in Familial Partial Lipodystrophy (FPLD) causing FPLD2. LMNA pathogenic variants have been previously associated with cardiomyopathy, familial arrhythmias or conduction system abnormalities independent of lipodystrophy. We aimed to assess cardiac impacts of FPLD, and to explore the extent of overlap between cardiolaminopathies and FPLD. Methods We conducted a retrospective review of an established cohort of 122 patients (age range: 13-77, M/F 21/101) with FPLD from Michigan (n = 83) and Turkey (n = 39) with an accessible cardiac evaluation. Also, functional syncytia of mature human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) from a FLPD2 patient was studied for assessment of autonomous rhythm and action potential duration with optical mapping using a voltage sensitive dye. Results In the whole study cohort, 95 (78%) patients had cardiac alterations (25% ischemic heart disease, 36% arrhythmia, 16% conduction abnormality, 20% prolonged QT interval, 11% cardiomyopathy, and 15% congestive heart failure). The likelihood of having an arrhythmia (OR; 3.95, 95% CI: 1.49-10.49) and conduction disease (OR: 3.324, 95% CI: 1.33-8.31) was significantly higher in patients with LMNA pathogenic variants. Patients with LMNA pathogenic variants were at high risk for atrial fibrillation/flutter (OR: 6.77, 95% CI: 1.27- 39.18). The time to first arrhythmia was significantly shorter in the LMNA group with a higher hazard rate of 3.04 (95% CI: 1.29-7.17, p = 0.032). Non-482 LMNA pathogenic variants were more likely to be associated with cardiac events (vs. 482 LMNA: OR: 4.74, 95% CI: 1.41- 15.98 for arrhythmia; OR: 17.67, 95% CI: 2.44- 127.68 for atrial fibrillation/flutter; OR: 5.71, 95% CI: 1.37- 23.76 for conduction disease. hiPSC-CMs from a FPLD2 patient had higher frequency of autonomous activity, and shorter Fridericia corrected action potential duration at 80% repolarization compared to control cardiomyocytes. Furthermore, FPLD2 functional syncytia of mature hiPSC-CMs presented several rhythm alterations such as early after-depolarizations, spontaneous quiescence and spontaneous tachyarrhythmia; none of those were observed in the control cell lines. Finally, FPLD2 hiPSC-CMs presented significantly slower recovery in chronotropic changes induced by isoproterenol exposure; which indicates disrupted beta-adrenergic response. Conclusion Our results suggest the need for vigilant cardiac monitoring in FPLD, especially in patients with FPLD2 who have an increased risk to develop cardiac arrhythmias and conduction system diseases. In addition, study of human induced pluripotent stem cell-derived cardiomyocytes may prove useful to understand the mechanism of cardiac disease and arrhythmias and to create precision therapy opportunities in the future.

Unlocking Personalized Biomedicine and Drug Discovery with Human Induced Pluripotent Stem Cell–Derived Cardiomyocytes: Fit for Purpose or Forever Elusive?

2020 ◽  
Vol 60 (1) ◽  
pp. 529-551 ◽  
Author(s):  
Tessa de Korte ◽  
Puspita A. Katili ◽  
Nurul A.N. Mohd Yusof ◽  
Berend J. van Meer ◽  
Umber Saleem ◽  
...  
Keyword(s):  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
Genome Engineering ◽  
Induced Pluripotent Stem Cell ◽  
Pathogenic Variants ◽  
Development Costs ◽  
Regulatory Guidelines ◽  
Fit For Purpose ◽  
Safety Assessments ◽  
Induced Pluripotent

In recent decades, drug development costs have increased by approximately a hundredfold, and yet about 1 in 7 licensed drugs are withdrawn from the market, often due to cardiotoxicity. This review considers whether technologies using human induced pluripotent stem cell–derived cardiomyocytes (hiPSC-CMs) could complement existing assays to improve discovery and safety while reducing socioeconomic costs and assisting with regulatory guidelines on cardiac safety assessments. We draw on lessons from our own work to suggest a panel of 12 drugs that will be useful in testing the suitability of hiPSC-CM platforms to evaluate contractility. We review issues, including maturity versus complexity, consistency, quality, and cost, while considering a potential need to incorporate auxiliary approaches to compensate for limitations in hiPSC-CM technology. We give examples on how coupling hiPSC-CM technologies with Cas9/CRISPR genome engineering is starting to be used to personalize diagnosis, stratify risk, provide mechanistic insights, and identify new pathogenic variants for cardiovascular disease.

scholarly journals Development of a Cardiac Sarcomere Functional Genomics Platform to Enable Scalable Interrogation of Human TNNT2 Variants

Circulation ◽  
2020 ◽  
Vol 142 (23) ◽  
pp. 2262-2275
Author(s):  
Anthony M. Pettinato ◽  
Feria A. Ladha ◽  
David J. Mellert ◽  
Nicholas Legere ◽  
Rachel Cohn ◽  
...  
Keyword(s):  
Heart Failure ◽  
Stem Cell ◽  
Hypertrophic Cardiomyopathy ◽  
Dilated Cardiomyopathy ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Pathogenic Variants ◽  
Calcium Affinity ◽  
Uncertain Significance ◽  
Induced Pluripotent

Background: Pathogenic TNNT2 variants are a cause of hypertrophic and dilated cardiomyopathies, which promote heart failure by incompletely understood mechanisms. The precise functional significance for 87% of TNNT2 variants remains undetermined, in part, because of a lack of functional genomics studies. The knowledge of which and how TNNT2 variants cause hypertrophic and dilated cardiomyopathies could improve heart failure risk determination, treatment efficacy, and therapeutic discovery, and provide new insights into cardiomyopathy pathogenesis, as well. Methods: We created a toolkit of human induced pluripotent stem cell models and functional assays using CRISPR/Cas9 to study TNNT2 variant pathogenicity and pathophysiology. Using human induced pluripotent stem cell–derived cardiomyocytes in cardiac microtissue and single-cell assays, we functionally interrogated 51 TNNT2 variants, including 30 pathogenic/likely pathogenic variants and 21 variants of uncertain significance. We used RNA sequencing to determine the transcriptomic consequences of pathogenic TNNT2 variants and adapted CRISPR/Cas9 to engineer a transcriptional reporter assay to assist prediction of TNNT2 variant pathogenicity. We also studied variant-specific pathophysiology using a thin filament–directed calcium reporter to monitor changes in myofilament calcium affinity. Results: Hypertrophic cardiomyopathy–associated TNNT2 variants caused increased cardiac microtissue contraction, whereas dilated cardiomyopathy–associated variants decreased contraction. TNNT2 variant–dependent changes in sarcomere contractile function induced graded regulation of 101 gene transcripts, including MAPK (mitogen-activated protein kinase) signaling targets, HOPX , and NPPB . We distinguished pathogenic TNNT2 variants from wildtype controls using a sarcomere functional reporter engineered by inserting tdTomato into the endogenous NPPB locus. On the basis of a combination of NPPB reporter activity and cardiac microtissue contraction, our study provides experimental support for the reclassification of 2 pathogenic/likely pathogenic variants and 2 variants of uncertain significance. Conclusions: Our study found that hypertrophic cardiomyopathy–associated TNNT2 variants increased cardiac microtissue contraction, whereas dilated cardiomyopathy–associated variants decreased contraction, both of which paralleled changes in myofilament calcium affinity. Transcriptomic changes, including NPPB levels, directly correlated with sarcomere function and can be used to predict TNNT2 variant pathogenicity.

Abstract 16472: Deciphering a Mechanistic Link Between Enhanced Late Sodium Current and Triggered Atrial Fibrillation Using Patient-specific and Gene-corrected Induced Pluripotent Stem Cell-derived Atrial Cardiomyocytes

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Liang HONG ◽  
Olivia T Ly ◽  
Hanna Chen ◽  
Arvind Sridhar ◽  
Meihong Zhang ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Atrial Fibrillation ◽  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
Sodium Current ◽  
Induced Pluripotent Stem Cell ◽  
Patient Specific ◽  
Gain Of Function ◽  
Late Sodium Current ◽  
Induced Pluripotent

Introduction: Gain-of-function mutations in SCN5A, which encodes the cardiac sodium channel, have been linked with familial atrial fibrillation (AF). However, the mechanistic link between the late sodium current (I Na,L ) and triggered arrhythmia remains unclear. Hypothesis: To characterize the electrophysiological (EP) phenotype of gain-of-function AF-linked SCN5A mutations, elucidate the underlying cellular mechanisms using patient-specific and gene-corrected (GC) induced pluripotent stem cell-derived atrial cardiomyocytes (iPSC-aCMs). Methods: We generated iPSC-aCMs from two families carrying SCN5A mutations (E428K and N470K) and control subjects. Whole-cell patch clamp and multi-electrode arrays were recorded to assess the EP phenotypes of the atrial iPSC-CMs. We corrected the E428K iPSC-aCMs using CRISPR/Cas9 gene editing approach (isogenic control). Results: The SCN5A mutation lines displayed abnormal EP properties including increased beating frequency and irregularity with triggered beats characteristic of AF ( Fig. 1 ). E428K iPSC-aCMs displayed spontaneous arrhythmogenic activity with beat-to-beat irregularity ( Fig. 1 A-D ) with the prolonged APD ( Fig. 1 E-H ) associated with enhanced I Na,L ( Fig. 1 I-L ). In contrast, expression of SCN5A -E428K in heterologous expression system failed to show enhanced I Na,L . The gene-corrected E428K iPSC-aCMs normalized the aberrant EP phenotype. Gene expression profiling revealed differential expression of calcium and potassium channel homeostasis and nitric oxide mediated signal transduction which could result in EP remodeling of atrial CMs. Conclusions: Patient-specific and gene-corrected iPSC-aCMs exhibited striking ex-vivo EP phenotype of an AF-causing SCN5A gain-of-function mutation that produced minimal changes in-vitro . We established a mechanistic link between enhanced I Na,L , ion channel remodeling and nitric oxide signaling pathways, and triggered AF.

scholarly journals Human-induced pluripotent stem cell-derived cardiomyocytes have limited IKs for repolarization reserve as revealed by specific KCNQ1/KCNE1 blocker

2019 ◽  
Vol 8 ◽  
pp. 204800401985491 ◽  
Author(s):  
Haoyu Zeng ◽  
Jixin Wang ◽  
Holly Clouse ◽  
Armando Lagrutta ◽  
Frederick Sannajust
Keyword(s):  
Stem Cell ◽  
Action Potential ◽  
Action Potential Duration ◽  
Pluripotent Stem Cell ◽  
Synergistic Effects ◽  
Induced Pluripotent Stem Cell ◽  
Human Cardiomyocytes ◽  
Repolarization Reserve ◽  
Beating Rate ◽  
Induced Pluripotent

Objective We investigated if there is IKs, and if there is repolarization reserve by IKs in human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Design We used a specific KCNQ1/KCNE1 channel blocker, L-000768673, with an IC50 of 9 nM, and four hERG-specific blockers, astemizole, cisapride, dofetilide, and E-4031 to investigate the issue. Results L-000768673 concentration-dependently prolonged feature point duration (FPD)―a surrogate signal of action potential duration―from 1 to 30 nM without pacing or paced at 1.2 Hz, resulting from IKs blockade in hiPSC-CMs. At higher concentrations, the effect of L-000768673 on IKs was mitigated by its effect on ICa-L, resulting in shortened FPD, reduced impedance amplitude, and increased beating rate at 1 µM and above, recapitulating the self-limiting properties of L-000768673 on action potentials. All four hERG-specific blockers prolonged FPD as expected. Co-application of L-000768673 at sub-threshold (0.1 and 0.3 nM) and threshold (1 nM) concentrations failed to synergistically enhance the effects of hERG blockers on FPD prolongation, rather it showed additive effects, inconsistent with the repolarization reserve role of IKs in mature human myocytes that enhanced IKr response, implying a difference between hiPSC-CMs used in this study and mature human cardiomyocytes. Conclusion There was IKs current in hiPSC-CMs, and blockade of IKs current caused prolongation of action potential of hiPSC-CMs. However, we could not demonstrate any synergistic effects on action potential duration prolongation of hiPSC-CMs by blocking hERG current and IKs current simultaneously, implying little or no repolarization reserve by IKs current in hiPSC-CMs used in this study.

Use of 3D culture systems to generate human induced pluripotent stem cell-derived [beta]-cells in vitro

2018 ◽  
Author(s):  
Fantuzzi Federica ◽  
Toivonen Sanna ◽  
Schiavo Andrea Alex ◽  
Pachera Nathalie ◽  
Rajaei Bahareh ◽  
...  
Keyword(s):  
Stem Cell ◽  
Beta Cells ◽  
Pluripotent Stem Cell ◽  
3D Culture ◽  
Induced Pluripotent Stem Cell ◽  
Culture Systems ◽  
Induced Pluripotent
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