Abstract
INTRODUCTION: Human induced pluripotent stem cells (hiPSCs) harboring cardiac myosin heavy chain 6 promoter can differentiate into functional cardiomyocytes called "iCell cardiomyocytes" under blasticidin treatment condition. While iCell cardiomyocytes are expected to be used for predicting cardiotoxicity of drugs, their responses to antiarrhythmic agents remain to be elucidated. We first examined electrophysiological properties of iCell cardiomyocytes and mRNA levels of ion channels and Ca handling proteins, and then evaluated effects of class I antiarrhythmic agents on their Na(+) and Ca(2+) currents. METHODS: iCell cardiomyocytes were cultured for 8-14 days (38-44 days after inducing their differentiation), according to the manufacturer's protocol. We determined their action potentials (APs) and sarcolemmal ionic currents using whole-cell patch clamp techniques, and also mRNA levels of ion channels and Ca handling proteins by RT-PCR. Effects of three class I antiarrhythmic agents, pirmenol, pilsicainide and mexiletine, on Na(+) channel current (I(Na)) and L-type Ca(2+) channel current (I(CaL)) were evaluated by the whole-cell patch clamp. RESULTS: iCell cardiomyocytes revealed sinoatrial node-type (18%), atrial-type (18%) and ventricular-type (64%) spontaneous APs. The maximum peak amplitudes of I(Na), I(CaL), and rapidly-activating delayed-rectifier K(+) channel current were -62.7 ± 13.7, -8.1 ± 0.7, and 3.0 ± 1.0 pA/pF, respectively. The hyperpolarization-activated cation channel and inward-rectifier K(+) channel currents were observed, whereas the T-type Ca(2+) channel or slowly-activating delayed-rectifier K(+) channel current was not detectable. mRNAs of Nav1.5, Cav1.2, Kir2.1, HCN4, KvLQT1, hERG and SERCA2 were detected, while that of HCN1, minK or MiRP was not. The class Ia antiarrhythmic agent pirmenol and class Ic agent pilsicainide blocked I(Na) in a concentration-dependent manner with IC(50) of 0.87 ± 0.37 and 0.88 ± 0.16 μM, respectively; the class Ib agent mexiletine revealed weak I(Na) block with a higher IC(50) of 30.0 ± 3.0 μM. Pirmenol, pilsicainide and mexiletine blocked I(CaL) with IC(50) of 2.00 ± 0.39, 7.7 ± 2.5 and 5.0 ± 0.1 μM, respectively. CONCLUSIONS: In iCell cardiomyocytes, I(Na) was blocked by the class Ia and Ic antiarrhythmic agents and I(CaL) was blocked by all the class I agents within the ranges of clinical concentrations, suggesting their cardiotoxicity.