Maximum Diastolic Potential of Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes Depends Critically on I _Kr

Abstract

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) hold promise for therapeutic applications. To serve these functions, the hiPSC-CM must recapitulate the electrophysiologic properties of native adult cardiomyocytes. This study examines the electrophysiologic characteristics of hiPSC-CM between 11 and 121 days of maturity. Embryoid bodies (EBs) were generated from hiPS cell line reprogrammed with Oct4, Nanog, Lin28 and Sox2. Sharp microelectrodes were used to record action potentials (AP) from spontaneously beating clusters (BC) micro-dissected from the EBs (n = 103; 37°C) and to examine the response to 5 µM E-4031 (n = 21) or BaCl ₂ (n = 22). Patch-clamp techniques were used to record I _Kr and I _K1 from cells enzymatically dissociated from BC (n = 49; 36°C). Spontaneous cycle length (CL) and AP characteristics varied widely among the 103 preparations. E-4031 (5 µM; n = 21) increased Bazett-corrected AP duration from 291.8±81.2 to 426.4±120.2 msec (p<0.001) and generated early afterdepolarizations in 8/21 preparations. In 13/21 BC, E-4031 rapidly depolarized the clusters leading to inexcitability. BaCl ₂, at concentrations that selectively block I _K1 (50–100 µM), failed to depolarize the majority of clusters (13/22). Patch-clamp experiments revealed very low or negligible I _K1 in 53% (20/38) of the cells studied, but presence of I _Kr in all (11/11). Consistent with the electrophysiological data, RT-PCR and immunohistochemistry studies showed relatively poor mRNA and protein expression of I _K1 in the majority of cells, but robust expression of I _Kr. In contrast to recently reported studies, our data point to major deficiencies of hiPSC-CM, with remarkable diversity of electrophysiologic phenotypes as well as pharmacologic responsiveness among beating clusters and cells up to 121 days post-differentiation (dpd). The vast majority have a maximum diastolic potential that depends critically on I _Kr due to the absence of I _K1. Thus, efforts should be directed at producing more specialized and mature hiPSC-CM for future therapeutic applications.