Comparable calcium handling of human iPSC-derived cardiomyocytes generated by multiple laboratories

Abstract

Cardiomyocytes (CMs) derived from human induced pluripotent stem cells (hiPSCs) are being increasingly used to model human heart diseases. hiPSC-CMs generated by earlier aggregation-based methods (i.e., embryoid body) often lack functional sarcoplasmic reticulum (SR) Ca stores characteristic of mature mammalian CMs. Newer monolayer-based cardiac differentiation methods (i.e., Matrigel sandwich or small molecule-based differentiation) produce hiPSC-CMs of high purity and yield, but their Ca handling has not been comprehensively investigated. Here, we studied Ca handling and cytosolic Ca buffering properties of hiPSC-CMs generated independently from multiple hiPSC lines at Stanford University, Vanderbilt University and University of Wisconsin. hiPSC-CMs were cryopreserved at each university. Frozen aliquots were shipped, recovered from cryopreservation, plated at low density and compared 3-5 days after plating with acutely-isolated adult rabbit and mouse ventricular CMs. Although hiPSC-CM cell volume was significantly smaller, cell capacitance to cell volume ratio and cytoplasmic Ca buffering were not different from rabbit-CMs. hiPSC-CMs from all three laboratories exhibited robust L-type Ca currents, twitch Ca transients and caffeine-releasable SR Ca stores comparable to adult CMs. Ca transport by sarcoendoplasmic reticulum Ca ATPase (SERCA) and Na/Ca exchanger (NCX) was similar in all hiPSC-CM lines, but slower compared to rabbit-CMs. However, the relative contribution of SERCA and NCX to Ca transport of hiPSC-CMs was comparable to rabbit-CMs. Ca handling maturity of hiPSC-CMs increased from 15 to 21 days post-induction. We conclude that hiPSC-CMs generated independently from multiple iPSC lines using monolayer-based methods can be reproducibly recovered from cryopreservation and exhibit comparable and functional SR Ca handling.

Integrating cardiomyocytes from human pluripotent stem cells in safety pharmacology: has the time come?

“…This particularly impacts I Ks and I K1 , whose contribution is essential for a correct prediction of drug effects. Only few attempts have been made to promote phenotypic uniformity in hiPSC‐CMs (Hwang et al , 2015a; Zhu et al , 2016), and we believe that the reproducibility of outcomes would benefit from standardized conditions to provide better input for computational models. On the other hand, as highlighted by Rodriguez et al , the cross‐institutional limitations of accessing in vitro propriety data profoundly slows the development of comprehensive computational models (Rodríguez et al , 2015).…”

“…Even though Ca 2 + ‐handling features appear conserved in independent hiPSC‐CM cultures from different research groups (Hwang et al , 2015a, 2015b; Kane and Terracciano, 2015), heterogeneity has been widely observed in other electrophysiological properties such as the APD (Figure 3), upstroke velocity and current densities (Hoekstra et al , 2012). Inconsistent results on the mechanism of hiPSC‐CM automaticity have also been noted: Kim et al reported a Ca 2 + clock mechanism, consistent with ivabradine having no effect on beating frequency (Kim et al , 2015).…”

Comparative Histological Study on the Effect of Stem Cells, and Gene Modified Stem Cells in Experimentally-Induced Diabetes Type 1 Cardiomyopathy

“…Calcium transport by SERCA was found to be similar in stem cells-cardiac myocyte lines [5] . Mesenchymal stem cells (MSCs) are widely used for both allogeneic and autologous transplantations.…”

Induced pluripotent stem cells: at the heart of cardiovascular precision medicine

“…162, 163 The gene expression profiles, especially those of contractile proteins, simulate fetal cardiomyocytes. 164 Furthermore, the hiPSC-CMs have poorly developed SR and altered calcium handling at early stages of differentiation, 165 nonexistent t-tubules, 166 automaticity, 167 and preference for glucose metabolism over fatty acid metabolism, 168 which are all consistent with immature phenotypes. Although various maturation techniques (e.g., electromechanical stimulation, 153, 169 overexpression of cardiac genes 101 or miR-499, 170 exposure to fatty acid supplementation, 90 and incorporation of cardiomyocytes into 3D tissue constructs 105 ) have all been tried with varying degrees of success, it is unclear whether the most mature hiPSC-CMs developed in an artificial cell culture environment can achieve the same level of maturity as adult cardiomyocytes in vivo .…”