A universal system for highly efficient cardiac differentiation of human induced pluripotent stem cells that eliminates interline variability

Background

The production of cardiomyocytes from human induced pluripotent stem cells(hiPSC) holds great promise for patient-specific cardiotoxicity drugtesting, disease modeling, and cardiac regeneration. However, existingprotocols for the differentiation of hiPSC to the cardiac lineage areinefficient and highly variable. We describe a highly efficient system fordifferentiation of human embryonic stem cells (hESC) and hiPSC to thecardiac lineage. This system eliminated the variability in cardiacdifferentiation capacity of a variety of human pluripotent stem cells(hPSC), including hiPSC generated from CD34⁺ cord bloodusing non-viral, non-integrating methods.

Methodology/Principal Findings

We systematically and rigorously optimized >45 experimental variables todevelop a universal cardiac differentiation system that produced contractinghuman embryoid bodies (hEB) with an improved efficiency of94.7±2.4% in an accelerated nine days from four hESC and sevenhiPSC lines tested, including hiPSC derived from neonatalCD34⁺ cord blood and adult fibroblasts usingnon-integrating episomal plasmids. This cost-effective differentiationmethod employed forced aggregation hEB formation in a chemically definedmedium, along with staged exposure to physiological (5%) oxygen, andoptimized concentrations of mesodermal morphogens BMP4 and FGF2, polyvinylalcohol, serum, and insulin. The contracting hEB derived using these methodswere composed of high percentages (64–89%) of cardiac troponinI⁺ cells that displayed ultrastructural properties offunctional cardiomyocytes and uniform electrophysiological profilesresponsive to cardioactive drugs.

Conclusion/Significance

This efficient and cost-effective universal system for cardiacdifferentiation of hiPSC allows a potentially unlimited production offunctional cardiomyocytes suitable for application to hPSC-based drugdevelopment, cardiac disease modeling, and the future generation ofclinically-safe nonviral human cardiac cells for regenerative medicine.