HEK293AD (BioCat; AD-100-GVO-CB), A431 (ATCC; CRL-1555), and H9c2 (2, 1) (ATCC; CRL-1446) cells were maintained in Dulbecco’s modified Eagle’s medium (PAN-Biotech) supplemented with 10% (vol/vol) fetal bovine serum (FBS) (Sigma-Aldrich), 2 mM l-glutamine (PAN-Biotech), penicillin (100 U/mL; Gibco), and streptomycin (100 μg/mL; Gibco) at 37 °C and 5% CO2. For passaging, HEK293AD and H9c2 (2, 1) cells were detached using trypsin 0.05%/(ethylenedinitrilo)tetraacetic acid (EDTA) 0.02% in phosphate buffered saline (PBS) (PAN-Biotech) or 0.25% trypsin-EDTA (Gibco) in case of A431 cells.

hiPSCs (line BIHi005-A) were received from and validated by the stem cell core facility of the Max Delbrück Center for Molecular Medicine. While pluripotent cultures were grown at 37 °C with 5% CO2 and 5% O2, differentiated cultures were maintained at 5% CO2 and atmospheric (21%) O2. Monolayers of hiPSC were differentiated into CM-hiPSC by modulating Wnt signaling following a small molecule-based cardiac differentiation strategy (56) and enriched for cardiac myocytes by metabolic lactate selection as previously described (57). Further method details for cell culture and materials used can be found in SI Appendix, Supplementary Materials and Methods.

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