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

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 BaCl2 (n = 22). Patch-clamp techniques were used to record IKr and IK1 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. BaCl2, at concentrations that selectively block IK1 (50–100 µM), failed to depolarize the majority of clusters (13/22). Patch-clamp experiments revealed very low or negligible IK1 in 53% (20/38) of the cells studied, but presence of IKr in all (11/11). Consistent with the electrophysiological data, RT-PCR and immunohistochemistry studies showed relatively poor mRNA and protein expression of IK1 in the majority of cells, but robust expression of IKr. 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 IKr due to the absence of IK1. Thus, efforts should be directed at producing more specialized and mature hiPSC-CM for future therapeutic applications.

hERG 1b is critical for human cardiac repolarization

“…Despite their utility in human arrhythmia models, iPSC-CMs differ from adult tissue-derived cardiomyocytes (29)(30)(31)(32)(33). APs recorded from iPSC-CMs display depolarized diastolic potentials, myogenic AP firing, and a reduced maximum rise rate of the AP upstroke (dV/dt MAX ) compared with APs from adult ventricular tissue (Fig.…”

Characterization of human iPS cell-derived cardiomyocyte sheets as a model to detect drug-induced conduction disturbance

“…In addition, the extent of expression of a gene SLC8A1 encoding Na + / Ca 2+ exchanger 1 (NCX1) has been reported to be similar between adult heart tissue and structured hiPSC-CMs (Rao et al, 2013). Meanwhile, inward rectifier K + current (I K1 ) was shown to be hardly expressed in the hiP-SC-CMs (Ma et al, 2011;Doss et al, 2012;Meijer van Putten et al, 2015;Uesugi et al, 2014). Since I K1 plays an important role in the repolarization process in phase 3 (Meijer van Putten et al, 2015), the low current density of I K1 may partly explain the long field potential duration of the cell sheet in this study.…”

IK1-enhanced human-induced pluripotent stem cell-derived cardiomyocytes: an improved cardiomyocyte model to investigate inherited arrhythmia syndromes

“…However, despite advances in differentiation and culturing techniques, currently available iPS-CMs have several features of immature cardiomyocytes, which limit their application for modeling cellular arrhythmia mechanisms and inherited arrhythmic disease. The electrophysiological limitations occur predominately due to action potential (AP) diastolic properties including a relatively depolarized resting membrane potential and spontaneous automaticity due to a small inward rectifier (I K1 ) density and unopposed pacemaker current (I f ) (5,18). In turn, this results in partial inactivation of ion channels such as the cardiac sodium channel to reduce current (I Na ) amplitude.…”