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Functional expression and pharmaceutical efficacy of cardiac-specific ion channels in human embryonic stem cell-derived cardiomyocytes.

ABSTRACT: Cardiomyocytes differentiated from human pluripotent stem cells provide promising tools for screening of cardiotoxic drugs. For evaluation of human pluripotent stem cell-derived cardiomyocytes for cardiotoxicity test, in the present study, human embryonic stem cells (hESCs) were differentiated to cardiomyocytes, followed by metabolic selection to enrich the differentiated cardiomyocytes. The highly purified hESC-derived cardiomyocytes (hESC-CMs) expressed several cardiomyocyte-specific markers including cTnT, MLC2a, and ?-SA, but not pluripotency markers, such as OCT4 and NANOG. Patch clamp technique and RT-PCR revealed the expression of cardiomyocyte-specific Na⁺, Ca²⁺, and K⁺ channels and cardiac action potential in hESC-CMs. To explore the potential use of hESC-CMs as functional cardiomyocytes for drug discovery and cardiotoxicity screening, we examined the effects of bisindolylmaleimide (BIM) (I), which inhibits native cardiac Ca²⁺ channels, on the Ca²⁺ channel activity of hESC-CMs. We observed a similar response for the BIM (I)-induced modulation of Ca²⁺ channels between hESC-CMs and native cardiomyocytes through L-type Ca²⁺ channel current. These results suggest that hESC-CMs can be useful for evaluation of pharmaceutical efficacy and safety of novel drug candidate in cardiac research.

SUBMITTER: Kim HS

PROVIDER: S-EPMC5653792 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:RATIONALE:Heart failure is characterized by electrical remodeling that contributes to arrhythmic risk. The unfolded protein response (UPR) is active in heart failure and can decrease protein levels by increasing mRNA decay, accelerating protein degradation, and inhibiting protein translation. OBJECTIVE:Therefore, we investigated whether the UPR downregulated cardiac ion channels that may contribute to arrhythmogenic electrical remodeling. METHODS:Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were used to study cardiac ion channels. Action potentials (APs) and ion channel currents were measured by patch clamp recording. The mRNA and protein levels of channels and the UPR effectors were determined by quantitative RT-PCR and Western blotting. Tunicamycin (TM, 50?ng/mL and 5??g/mL), GSK2606414 (GSK, 300?nmol/L), and 4?8C (5??mol/L) were utilized to activate the UPR, inhibit protein kinase-like ER kinase (PERK) and inositol-requiring protein-1 (IRE1), respectively. RESULTS:TM-induced activation of the UPR caused significant prolongation of the AP duration (APD) and a reduction of the maximum upstroke velocity (dV/dtmax) of the AP phase 0 in both acute (20-24?h) and chronic treatment (6?days). These changes were explained by reductions in the sodium, L-type calcium, the transient outward and rapidly/slowly activating delayed rectifier potassium currents. Nav1.5, Cav1.2, Kv4.3, and KvLQT1 channels showed concomitant reductions in mRNA and protein levels under activated UPR. Inhibition of PERK or IRE1 shortened the APD and reinstated dV/dtmax. The PERK branch regulated Nav1.5, Kv4.3, hERG, and KvLQT1. The IRE1 branch regulated Nav1.5, hERG, KvLQT1, and Cav1.2. CONCLUSIONS:Activated UPR downregulates all major cardiac ion currents and results in electrical remodeling in hiPSC-CMs. Both PERK and IRE1 branches downregulate Nav1.5, hERG, and KvLQT1. The PERK branch specifically downregulates Kv4.3, while the IRE1 branch downregulates Cav1.2. Therefore, the UPR contributed to electrical remodeling, and targeting the UPR might be anti-arrhythmic.

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Functional expression and pharmaceutical efficacy of cardiac-specific ion channels in human embryonic stem cell-derived cardiomyocytes.

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