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 α-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.

Kinin B2-Receptor in human iPSC differentiation into cardiomyocytes

“…This rate is comparable to that observed in other studies, which report 21-130 beats per minute (118,123). Variations on the average AP firing rate as a consequence of more prolonged time in culture have been reported, although the magnitude and direction of these observations are not necessarily consistent among studies (123 (110,113,117). In addition to this, a persistent expression of the HCN4 channel and of the I f has largely been reported in hPSC-CM, and could possibly explain the high automaticity observed in these cells (37).…”

“…Nevertheless, as the AP shape and kinetic parameters are consistent with those observed in the literature (42,117,118,121,122), it seems reasonable to assume that it is properly regulated by the ion channels that most commonly coordinate the cardiac AP.…”

“…Our results reveal only a tendency to increase dV/dT max (Fig.9b) in older hiPSC-CM, but an analysis of each ion current that could confirm or refute this observed tendency has not been performed. The magnitude of dV/dT max found in the present study is smaller than that reported by a series of studies which observe upstroke velocities ranging from 11-15V/s on hiPSC-CM (42,117,118,121), but is in line with that reported by Burridge (2015) (122).…”

“…Peinkofer and co-workers (112) The morphology of APs is the result of a net balance of all ion currents that enter and leave the cell (116). Because of this, several studies have characterized the expression and activity of ion channels in in vitro differentiated cardiomyocytes and reported all inward and outward currents depicted in Fig.1a, except from the I k1 current, which is often absent or present on very low densities on PSC-CM (65,113,(117)(118)(119)(120). Although the study of ion currents is important to assess the functionality of hiPSC-CM and validate their use as platform for in vitro drug testing, such characterization was beyond the scope of this project.…”

“…The average RMP observed in this study are consistently more hyperpolarized than -77mV, with many observations averaging around -85mV (Fig.9d). This is an interesting achievement, considering that although the native cardiac RMP is -85mV (41), most ventricular-like hiPSC-CM in the literature display way more depolarized RMP, ranging from 68mV to -50mV (115,117,121,122,125). In spite of their more hyperpolarized RMP, the AP amplitudes of the hiPSC-CM analysed in this study (Fig.9c) still fall short of that seen in native APs (>100mV) (2), being coherent with what was verified by other authors (24,103).…”