Abstract
Human (h) embryonic stem cells (ESC) represent an unlimited source of cardiomyocytes (CMs); however, these differentiated cells are immature. Thus far, gene profiling studies have been performed with non-purified or non-chamber specific CMs. Here we took a combinatorial approach of using systems biology to guide functional discoveries of novel biological properties of purified hESC-derived ventricular (V) CMs. We profiled the transcriptomes of hESCs, hESC-, fetal (hF) and adult (hA) VCMs, and showed that hESC-VCMs displayed a unique transcriptomic signature. Not only did a detailed comparison between hESC-VCMs and hF-VCMs confirm known expression changes in metabolic and contractile genes, it further revealed novel differences in genes associated with reactive oxygen species (ROS) metabolism, migration and cell cycle, as well as potassium and calcium ion transport. Following these guides, we functionally confirmed that hESC-VCMs expressed IKATP with immature properties, and were accordingly vulnerable to hypoxia/reoxygenation-induced apoptosis. For mechanistic insights, our coexpression and promoter analyses uncovered a novel transcriptional hierarchy involving select transcription factors (GATA4, HAND1, NKX2.5, PPARGC1A and TCF8), and genes involved in contraction, calcium homeostasis and metabolism. These data highlight novel expression and functional differences between hESC-VCMs and their fetal counterparts, and offer insights into the underlying cell developmental state. These findings may lead to mechanism-based methods for in vitro driven maturation.
Highlights
The innate regenerative capacity of the adult mammalian heart is insufficient to restore function damaged by myocardial injury or heart failure
Our results show that hESC-VCMs have a unique transcriptomic signature, contractility and metabolic parameters that are most analogous to fetal cells; but, we discovered a range of novel changes in cell cycle, reactive oxygen species (ROS) metabolism and migration that have not been previously reported
HESC-VCMs were identified by the expression of a reporter under the transcriptional control of the MLC2v promoter [19] (Figure 1B) and were isolated by flowactivated cell sorting
Summary
The innate regenerative capacity of the adult mammalian heart is insufficient to restore function damaged by myocardial injury or heart failure. Sarco/endoplasmic reticulum (SR) function remains rudimentary, as the cells exhibit a modest Ca2+ transient with slow kinetics, moderate SR Ca2+ATPase (SERCA) levels, low and disorganized ryanodine receptors and delayed phospholamban (PLN) expression [8,12] without the T-tubule system [9]. To overcome these limitations, an understanding of the molecular and cellular processes responsible for the development of a more physiological adultlike phenotype is required
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