Cardiomyocyte Differentiation from hiPSCs

The hiPSC line was adopted in this study after obtaining them from the Stanford Cardiovascular Institute (SCVI) Biobank, Stanford University. It was generated thorough reprograming of peripheral blood mononuclear cells (PBMCs) from an anonymous healthy individual with Sendai virus. The pluripotent cell lines were grown in Matrigel (Corning, CA)-coated 12-well plates in Essential 8™ Medium (Thermo Fisher Scientific, MA) at 37°C in 5% CO2 in compressed air and high humidity. Cardiomyocyte differentiation was conducted using a monolayer differentiation chemically defined method (Burridge et al., 2014 (link)). Briefly, iPSCs were kept in culture until 80%–90% confluence. For the differentiation, iPSCs were treated with 6 µM CHIR99021 in RPMI + B27 (minus insulin) for 2 days, fresh RPMI + B27 (minus insulin) for 1 day, followed by 5 µM IWR-1 treatment for 2 days, and then fresh RPMI + B27 (minus insulin) for another 2 days. Afterward, the cells will be supplied with fresh RPMI + B27 every other day. Beating cardiomyocytes will normally appear after 9–10 days, and the cells can be further treated with glucose free RPMI + B27 for 2–3 rounds.