Non-cell autonomous cues for enhanced functionality of human embryonic stem cell-derived cardiomyocytes via maturation of sarcolemmal and mitochondrial K ATP channels

Human embryonic stem cells (hESCs) is a potential unlimited ex vivo source of ventricular ( V ) cardiomyocytes (CMs), but hESC- V CMs and their engineered tissues display immature traits. In adult V CMs, sarcolemmal (sarc) and mitochondrial (mito) ATP-sensitive potassium (K ATP ) channels play crucial roles in excitability and cardioprotection. In this study, we aim to investigate the biological roles and use of sarcK ATP and mitoK ATP in hESC- V CM. We showed that SarcI K, ATP in single hESC- V CMs was dormant under baseline conditions, but became markedly activated by cyanide (CN) or the known opener P1075 with a current density that was ~8-fold smaller than adult; These effects were reversible upon washout or the addition of GLI or HMR1098. Interestingly, sarcI K, ATP displayed a ~3-fold increase after treatment with hypoxia (5% O 2 ). MitoI K, ATP was absent in hESC- V CMs. However, the thyroid hormone T3 up-regulated mitoI K, ATP, conferring diazoxide protective effect on T3-treated hESC- V CMs. When assessed using a multi-cellular engineered 3D ventricular cardiac micro-tissue (hvCMT) system, T3 substantially enhanced the developed tension by 3-folds. Diazoxide also attenuated the decrease in contractility induced by simulated ischemia (1% O 2 ). We conclude that hypoxia and T3 enhance the functionality of hESC- V CMs and their engineered tissues by selectively acting on sarc and mitoI K, ATP .