Sarco/endoplasmic reticulum Ca²⁺-ATPase is a more effective calcium remover than sodium-calcium exchanger in human embryonic stem cell-derived cardiomyocytes

Human pluripotent stem cell (hPSCs)-derived ventricular (V) cardiomyocytes (CMs) display immature Ca²⁺-handing properties with smaller transient amplitudes and slower kinetics due to such differences in crucial Ca²⁺-handling proteins as the poor sarco/endoplasmic reticulum Ca²⁺-ATPase (SERCA) pump but robust Na⁺-Ca²⁺ exchanger (NCX) activities in human embryonic stem cell (ESC)-derived VCMs compared with adult. Despite their fundamental importance in excitation-contraction coupling, the relative contribution of SERCA and NCX to Ca²⁺-handling of hPSC-VCMs remains unexplored. We systematically altered the activities of SERCA and NCX in human embryonic stem cell-derived ventricular cardiomyocytes (hESC-VCMs) and their engineered microtissues, followed by examining the resultant phenotypic consequences. SERCA overexpression in hESC-VCMs shortened the decay of Ca²⁺ transient at low frequencies (0.5 Hz) without affecting the amplitude, SR Ca²⁺ content and Ca²⁺ baseline. Interestingly, short hairpin RNA-based NCX suppression did not prolong the transient decay, indicating a compensatory response for Ca²⁺ removal. Although hESC-VCMs and their derived microtissues exhibited negative frequency-transient/force responses, SERCA overexpression rendered them less negative at high frequencies (>2 Hz) by accelerating Ca²⁺ sequestration. We conclude that for hESC-VCMs and their microtissues, SERCA, rather than NCX, is the main Ca²⁺ remover during diastole; poor SERCA expression is the leading cause for immature negative-frequency/force responses, which can be partially reverted by forced expression. Combinatorial approach to mature calcium handling in hESC-VCMs may help shed further mechanistic insights.NEW & NOTEWORTHY In this study of human pluripotent stem cell-derived cardiomyocytes, we studied the role of sarco/endoplasmic reticulum Ca²⁺-ATPase (SERCA) and Na⁺-Ca²⁺ exchanger (NCX) in Ca²⁺ handling. Our data support the notion that SERCA is more effective in cytosolic calcium removal than the NCX.