scholarly journals Identifying the Transcriptome Signatures of Calcium Channel Blockers in Human Induced Pluripotent Stem Cell–Derived Cardiomyocytes

2019 ◽  
Vol 125 (2) ◽  
pp. 212-222 ◽  
Author(s):  
Chi Keung Lam ◽  
Lei Tian ◽  
Nadjet Belbachir ◽  
Alexa Wnorowski ◽  
Rajani Shrestha ◽  
...  
Keyword(s):  
Stem Cell ◽  
Calcium Channel ◽  
Calcium Channel Blockers ◽  
Pluripotent Stem Cell ◽  
Contractile Function ◽  
Induced Pluripotent Stem Cell ◽  
Channel Blockers ◽  
Calcium Kinetics ◽  
Human Cardiomyocytes ◽  
Induced Pluripotent

Rationale: Calcium channel blockers (CCBs) are an important class of drugs in managing cardiovascular diseases. Patients usually rely on these medications for the remainder of their lives after diagnosis. Although the acute pharmacological actions of CCBs in the hearts are well-defined, little is known about the drug-specific effects on human cardiomyocyte transcriptomes and physiological alterations after long-term exposure. Objective: This study aimed to simulate chronic CCB treatment and to examine both the functional and transcriptomic changes in human cardiomyocytes. Methods and Results: We differentiated cardiomyocytes and generated engineered heart tissues from 3 human induced pluripotent stem cell lines and exposed them to 4 different CCBs—nifedipine, amlodipine, diltiazem, and verapamil—at their physiological serum concentrations for 2 weeks. Without inducing cell death and damage to myofilament structure, CCBs elicited line-specific inhibition on calcium kinetics and contractility. While all 4 CCBs exerted similar inhibition on calcium kinetics, verapamil applied the strongest inhibition on cardiomyocyte contractile function. By profiling cardiomyocyte transcriptome after CCB treatment, we identified little overlap in their transcriptome signatures. Verapamil is the only inhibitor that reduced the expression of contraction-related genes, such as MYH (myosin heavy chain) and troponin I, consistent with its depressive effects on contractile function. The reduction of these contraction-related genes may also explain the responsiveness of patients with hypertrophic cardiomyopathy to verapamil in managing left ventricular outflow tract obstruction. Conclusions: This is the first study to identify the transcriptome signatures of different CCBs in human cardiomyocytes. The distinct gene expression patterns suggest that although the 4 inhibitors act on the same target, they may have distinct effects on normal cardiac cell physiology.

scholarly journals Determination of Appropriate Stage of Human-Induced Pluripotent Stem Cell–Derived Cardiomyocytes for Drug Screening and Pharmacological Evaluation In Vitro

2012 ◽  
Vol 17 (9) ◽  
pp. 1192-1203 ◽  
Author(s):  
Tadahiro Shinozawa ◽  
Kenichi Imahashi ◽  
Hiroshi Sawada ◽  
Hatsue Furukawa ◽  
Kenji Takami
Keyword(s):  
Stem Cell ◽  
Sarcoplasmic Reticulum ◽  
Pluripotent Stem Cell ◽  
Microelectrode Array ◽  
Contractile Function ◽  
Induced Pluripotent Stem Cell ◽  
Channel Blockers ◽  
Electrophysiological Properties ◽  
Induced Pluripotent

Human-induced pluripotent stem cell–derived cardiomyocytes (hiPS-CMs) at different stages (approximate days 30, 60, and 90) were used to determine the appropriate stage for functional and morphological assessment of drug effects in vitro. The hiPS-CMs had spontaneous beating activity, and β-adrenergic function was comparable in all stages of differentiation. Microelectrode array analyses using ion channel blockers indicated that the electrophysiological properties of these ion channels were comparable at all differentiation stages. Ultrastructural analysis using electron microscopy showed that myofibrillar structures at days 60 and 90 were similarly distributed and more mature than that at day 30. Analysis of motion vectors in contracting cells showed that the velocity of contraction was the highest at day 90 and was the most mature among the three stages. Gene expression analysis demonstrated that expression of some genes related to myofilament and sarcoplasmic reticulum increased with maturation of morphological and contractile properties. In conclusion, day 30 cardiomyocytes are useful for basic screening such as the assessment of electrophysiological properties, and days 60 and 90 are the appropriate differentiation stage for morphological assays. For the assay of contractile function associated with subcellular components such as sarcoplasmic reticulum, day 90 cardiomyocytes are the most suitable.

scholarly journals Ion Channel Expression and Electrophysiology of Singular Human (Primary and Induced Pluripotent Stem Cell-Derived) Cardiomyocytes

Cells ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 3370
Author(s):  
Christina Schmid ◽  
Najah Abi-Gerges ◽  
Michael Georg Leitner ◽  
Dietmar Zellner ◽  
Georg Rast
Keyword(s):  
Stem Cell ◽  
Single Cell ◽  
Pluripotent Stem Cell ◽  
Quantitative Polymerase Chain Reaction ◽  
Drug Effects ◽  
Induced Pluripotent Stem Cell ◽  
Ionic Currents ◽  
Human Cardiomyocytes ◽  
Ventricular Cells ◽  
Induced Pluripotent

Subtype-specific human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are promising tools, e.g., to assess the potential of drugs to cause chronotropic effects (nodal hiPSC-CMs), atrial fibrillation (atrial hiPSC-CMs), or ventricular arrhythmias (ventricular hiPSC-CMs). We used single-cell patch-clamp reverse transcriptase-quantitative polymerase chain reaction to clarify the composition of the iCell cardiomyocyte population (Fujifilm Cellular Dynamics, Madison, WI, USA) and to compare it with atrial and ventricular Pluricytes (Ncardia, Charleroi, Belgium) and primary human atrial and ventricular cardiomyocytes. The comparison of beating and non-beating iCell cardiomyocytes did not support the presence of true nodal, atrial, and ventricular cells in this hiPSC-CM population. The comparison of atrial and ventricular Pluricytes with primary human cardiomyocytes showed trends, indicating the potential to derive more subtype-specific hiPSC-CM models using appropriate differentiation protocols. Nevertheless, the single-cell phenotypes of the majority of the hiPSC-CMs showed a combination of attributes which may be interpreted as a mixture of traits of adult cardiomyocyte subtypes: (i) nodal: spontaneous action potentials and high HCN4 expression and (ii) non-nodal: prominent INa-driven fast inward current and high expression of SCN5A. This may hamper the interpretation of the drug effects on parameters depending on a combination of ionic currents, such as beat rate. However, the proven expression of specific ion channels supports the evaluation of the drug effects on ionic currents in a more realistic cardiomyocyte environment than in recombinant non-cardiomyocyte systems.

scholarly journals Pharmacological Identification of an L-Type Calcium Channel Impedance Signal in Human Induced Pluripotent Stem-Cell Derived Cardiomyocytes

2017 ◽  
Vol 112 (3) ◽  
pp. 236a
Author(s):  
Carlos A. Obejero-Paz ◽  
Leslie Ellison ◽  
James Kramer ◽  
Arthur M. Brown ◽  
Andrew Bruening-Wright
Keyword(s):  
Stem Cell ◽  
Calcium Channel ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Induced Pluripotent

scholarly journals Deep learning detects cardiotoxicity in a high-content screen with induced pluripotent stem cell-derived cardiomyocytes

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Francis Grafton ◽  
Jaclyn Ho ◽  
Sara Ranjbarvaziri ◽  
Farshad Farshidfar ◽  
Anastasiia Budan ◽  
...  
Keyword(s):  
Stem Cell ◽  
Deep Learning ◽  
Pluripotent Stem Cell ◽  
Kinase Inhibitors ◽  
Early Stage ◽  
Induced Pluripotent Stem Cell ◽  
Channel Blockers ◽  
Drug Induced ◽  
Induced Pluripotent

Drug-induced cardiotoxicity and hepatotoxicity are major causes of drug attrition. To decrease late-stage drug attrition, pharmaceutical and biotechnology industries need to establish biologically relevant models that use phenotypic screening to detect drug-induced toxicity in vitro. In this study, we sought to rapidly detect patterns of cardiotoxicity using high-content image analysis with deep learning and induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). We screened a library of 1280 bioactive compounds and identified those with potential cardiotoxic liabilities in iPSC-CMs using a single-parameter score based on deep learning. Compounds demonstrating cardiotoxicity in iPSC-CMs included DNA intercalators, ion channel blockers, epidermal growth factor receptor, cyclin-dependent kinase, and multi-kinase inhibitors. We also screened a diverse library of molecules with unknown targets and identified chemical frameworks that show cardiotoxic signal in iPSC-CMs. By using this screening approach during target discovery and lead optimization, we can de-risk early-stage drug discovery. We show that the broad applicability of combining deep learning with iPSC technology is an effective way to interrogate cellular phenotypes and identify drugs that may protect against diseased phenotypes and deleterious mutations.

scholarly journals Maturation of human induced pluripotent stem cell-derived cardiomyocytes for modeling hypertrophic cardiomyopathy

2020 ◽  
Author(s):  
Walter E. Knight ◽  
Yingqiong Cao ◽  
Ying-Hsi Lin ◽  
Genevieve C. Sparagna ◽  
Betty Bai ◽  
...  
Keyword(s):  
Cardiovascular Disease ◽  
Stem Cell ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Simple Method ◽  
Human Cardiomyocytes ◽  
Pathological Hypertrophy ◽  
Induced Pluripotent ◽  
Metabolic Behavior

AbstractRationaleHuman induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) are a powerful platform for biomedical research. However, they are immature, which is a barrier to modeling adult-onset cardiovascular disease.ObjectiveWe sought to develop a simple method which could drive cultured hiPSC-CMs towards maturity across a number of phenotypes.Methods and resultsCells were cultured in fatty acid-based media and plated on micropatterned surfaces to promote alignment and elongation. These cells display many characteristics of adult human cardiomyocytes, including elongated cell morphology, enhanced maturity of sarcomeric structures, metabolic behavior, and increased myofibril contractile force. Most notably, hiPSC-CMs cultured under optimal maturity-inducing conditions recapitulate the pathological hypertrophy caused by either a pro-hypertrophic agent or genetic mutations.ConclusionsThe more mature hiPSC-CMs produced by the methods described here will serve as a useful in vitro platform for characterizing cardiovascular disease.

Abstract 20459: Epigenetic Activation of Phosphodiesterase Subtypes Lead to Compromised Beta-Adrenergic Signaling in Induced Pluripotent Stem Cell-Derived Cardiomyocytes From Dilated Cardiomyopathy Patients

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Haodi Wu ◽  
Jaecheol Lee ◽  
Mingxia Gu ◽  
Feng Lan ◽  
Jared Churko ◽  
...  
Keyword(s):  
Stem Cell ◽  
Dilated Cardiomyopathy ◽  
Pluripotent Stem Cell ◽  
Contractile Function ◽  
Induced Pluripotent Stem Cell ◽  
Epigenetic Mechanism ◽  
Patient Specific ◽  
Beta Adrenergic ◽  
Adrenergic Signaling ◽  
Induced Pluripotent

Introduction: Familial dilated cardiomyopathy (DCM) has been modeled by human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). However, the mechanisms of compromised signaling transduction and contractile function in DCM iPSC-CMs are still not well understood. Methods and Results: Beating iPSC-CMs were generated from healthy individuals and DCM patients. RNA-seq and real-time PCR showed strict regulation of the main beta-adrenergic signaling proteins in iPSC-CMs during maturation. Confocal imaging of spontaneous calcium activity and hydrogel-based traction force microscopy (TFM) technology demonstrated beta-adrenergic stimulation induced both inotropic and chronotropic regulation in the contractility of iPSC-CMs. Following extended in vitro maturation of iPSC-CMs, we observed a transition in the beta-adrenergic receptor (beta-AR) subtype dependence from beta-2 AR dominance at early stage (day 30) to beta-1/2 AR co-existence at late stage (day 60). Comparison of the beta-adrenergic responsiveness between iPSC-CMs from DCM patients and their familial controls showed compromised beta-adrenergic signaling in DCM cells. Microarray data and expression profiling indicated up-regulated phosphodiesterases (PDE) 2A, 3A and 5A in DCM iPSC-CMs, which impaired cAMP generation and blunted the beta-adrenergic response. By blocking PDE2A, 3A or 5A, beta-adrenergic signaling reactivity and contractile function in DCM iPSC-CMs were both greatly improved. To further elucidate underlying mechanism of PDE regulation, we conducted chromatin immunoprecipitation (CHIP) assays, which showed significant up-regulation of activation histone marker and down-regulation of repressive histone marker in the PDE promoters during maturation process of DCM iPSC-CMs, which closely recapitulated the epigenetic modulation in the ventricle tissues harvested from DCM patients. Conclusions: Patient-specific DCM iPSC-CMs recapitulated impaired beta-adrenergic responsiveness and contractility in diseased heart. Studies on iPSC-CM model revealed a novel epigenetic mechanism that underlies PDE subtype specific regulation and signaling deficiency in DCM pathogenesis, which may serve as a new therapeutic target in the future.

scholarly journals Dynamic Current Clamp Experiments Define the Functional Roles of IK1 and Ito,f in Human Induced Pluripotent Stem Cell Derived Cardiomyocytes

2017 ◽  
Author(s):  
Scott B. Marrus ◽  
Steven Springer ◽  
Eric Johnson ◽  
Rita J. Martinez ◽  
Edward J. Dranoff ◽  
...  
Keyword(s):  
Stem Cell ◽  
Action Potential ◽  
Action Potentials ◽  
Pluripotent Stem Cell ◽  
Phase 1 ◽  
Induced Pluripotent Stem Cell ◽  
Resting Membrane Potential ◽  
Current Clamp ◽  
Human Cardiomyocytes ◽  
Induced Pluripotent

AbstractThe transient outward potassium current (Ito) plays a key, albeit incompletely defined, role in cardiomyocyte physiology and pathophysiology. In light of the technical challenges of studying adult human cardiomyocytes, this study examines the use of induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) as a system which potentially preserves the native cellular milieu of human cardiomyocytes. ISPC-CMs express a robust Ito with slow recovery kinetics and fail to express the rapidly recovering Ito,f which is implicated in human disease. Overexpression of the accessory subunit KChIP2 (which is not expressed in iPSC-CMs) resulted in restoration of a rapid component of recovery. To define the functional role of Ito, dynamic current clamp was used to introduce computationally modeled currents into iPSC-CMs while recording action potentials. However, iPSC-CMs exhibit action potentials with multiple immature physiological properties, including slow upstroke velocity, heterogeneous action potential waveforms, and the absence of a phase 1 notch, thus potentially limiting the utility of these cells as a model of adult cardiomyocytes. Importantly, the introduction of modeled inwardly rectified current (IK1) ameliorated these immature properties by restoring a hyperpolarized resting membrane potential. In this context of normalized action potential morphologies, dynamic current clamp experiments introducing Ito,f demonstrated that there is significant cell-to-cell heterogeneity and that the functional effect of Ito,f is highly sensitive to the action potential plateau voltage in each cell.

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