A universal system for highly efficient cardiac differentiation of human induced pluripotent stem cells that eliminates interline variability |
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Authors: | Burridge Paul W Thompson Susan Millrod Michal A Weinberg Seth Yuan Xuan Peters Ann Mahairaki Vasiliki Koliatsos Vassilis E Tung Leslie Zambidis Elias T |
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Institution: | Johns Hopkins Institute for Cell Engineering, The Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America. paul.burridge@jhmi.edu |
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Abstract: | BackgroundThe production of cardiomyocytes from human induced pluripotent stem cells
(hiPSC) holds great promise for patient-specific cardiotoxicity drug
testing, disease modeling, and cardiac regeneration. However, existing
protocols for the differentiation of hiPSC to the cardiac lineage are
inefficient and highly variable. We describe a highly efficient system for
differentiation of human embryonic stem cells (hESC) and hiPSC to the
cardiac lineage. This system eliminated the variability in cardiac
differentiation capacity of a variety of human pluripotent stem cells
(hPSC), including hiPSC generated from CD34+ cord blood
using non-viral, non-integrating methods.Methodology/Principal FindingsWe systematically and rigorously optimized >45 experimental variables to
develop a universal cardiac differentiation system that produced contracting
human embryoid bodies (hEB) with an improved efficiency of
94.7±2.4% in an accelerated nine days from four hESC and seven
hiPSC lines tested, including hiPSC derived from neonatal
CD34+ cord blood and adult fibroblasts using
non-integrating episomal plasmids. This cost-effective differentiation
method employed forced aggregation hEB formation in a chemically defined
medium, along with staged exposure to physiological (5%) oxygen, and
optimized concentrations of mesodermal morphogens BMP4 and FGF2, polyvinyl
alcohol, serum, and insulin. The contracting hEB derived using these methods
were composed of high percentages (64–89%) of cardiac troponin
I+ cells that displayed ultrastructural properties of
functional cardiomyocytes and uniform electrophysiological profiles
responsive to cardioactive drugs.Conclusion/SignificanceThis efficient and cost-effective universal system for cardiac
differentiation of hiPSC allows a potentially unlimited production of
functional cardiomyocytes suitable for application to hPSC-based drug
development, cardiac disease modeling, and the future generation of
clinically-safe nonviral human cardiac cells for regenerative medicine. |
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