Abstract
Myocardial infarction leads to myocardial necrosis, and cardiomyocytes are non-renewable. Fatty acid-containing cardiomyocyte maturation medium promotes maturation of stem cell-derived cardiomyocytes. To study the effect palmitic acid on maturation of cardiomyocytes derived from human embryonic stem cells (hESCs) to optimize differentiation for potential treatment of myocardial infarction by hESCs. hESCs were differentiated into cardiomyocytes using standard chemically defined medium 3 (CDM3). Up to day 20 of differentiation, 200Mm palmitic acid were added, and then the culture was continued for another 8 days to mimic the environment in which human cardiomyocytes mainly use fatty acids as the main energy source. Light microscopy, transmission electron microscopy, immunofluorescence, reverse transcription-polymerase chain reaction, and cellular ATP assays, were carried out to analyze the expression of relevant cardiomyocyte-related genes, cell morphology, metabolism levels, and other indicators cardiomyocyte maturity. Cardiomyocytes derived from hESCs under exogenous palmitic acid had an elongated pike shape and a more regular arrangement. Sarcomere stripes were clear, and the cells color was clearly visible. The cell perimeter and elongation rate were also increased. Myogenic fibers were abundant, myofibrillar z-lines were regularly, the numbers of mitochondria and mitochondrial cristae were higher, more myofilaments were observed, and the structure of round-like discs was occasionally seen. Expression of mature cardiomyocyte-associated genes TNNT2, MYL2 and MYH6, and cardiomyocyte-associated genes KCNJ4, RYR2,and PPARα, was upregulated (p <0.05). Expression of MYH7, MYL7, KCND2, KCND3, GJA1 and TNNI1 genes was unaffected (p >0.05). Expression of mature cardiomyocyte-associated sarcomere protein MYL2 was significantly increased (p <0.05), MYH7 protein expression was unaffected (p >0.05). hESC-derived cardiomyocytes exposed to exogenous palmitic acid produced more ATP per unit time (p <0.05). Exogenous palmitic acid induced more mature hESC-CMs in terms of the cellular architecture, expression of cardiomyocyte maturation genes adnprotein, and metabolism.
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