Author + information
- †Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania
- ‡Cardiovascular Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania
- ↵∗Reprint requests and correspondence:
Dr. Kenneth B. Margulies, Department of Medicine, Heart Failure and Transplant Program, Perelman School of Medicine, University of Pennsylvania, Translational Research Center, Room 11-101, 3400 Civic Center Boulevard, Building 421, Philadelphia, Pennsylvania 19104.
Cardiovascular diseases, including myocardial infarction and heart failure, remain the leading causes of morbidity and mortality in the developed world. Because the loss of cardiac myocytes contributes to most types of heart failure, strategies designed to achieve therapeutic cardiac regeneration are being aggressively sought. Since the first report in 2006 (1), the refinement of methods for reprogramming differentiated somatic cells into human-induced pluripotent stem cells (iPSCs) with regenerative capacity comparable to that of embryonic stem cells has further energized basic inquiries into the factors shaping cell fate and has fueled translational efforts toward therapeutic cardiac regeneration.
Given that all of a person’s somatic cells have essentially the same DNA sequence, it has been increasingly appreciated that determination of cell fate and phenotypic features is strongly affected by which parts of the DNA sequence are available to regulate gene expression. This availability for transcription is largely controlled by so-called epigenetic modifications, including DNA methylation and histone modifications. In general, embryonic stem cells and iPSCs have epigenetic modifications restricting lineage-specific gene expression and thus are pluripotent. Conversely, somatic cells have epigenetic marks that restrict expression of some genes involved in maintaining self-renewal or pluripotency, and this contributes to their cell type–specific biology. Thus, reprogramming of somatic cells to a pluripotent state entails reversing the process of cell differentiation through tissue-specific epigenetic modifications and re-establishing an embryonic epigenome, which can include alterations in chromatin chemistry and structure and/or methylation of cytosine residues in DNA.
It has been shown that iPSCs can be generated from a wide variety of somatic cells (2,3), but distinctions among iPSCs generated from alternative cell types are still being defined. For example, studies diverge on whether iPSCs derived from a somatic source retain “epigenetic memory” and exhibit a preference for differentiation into their original cell lineages. One study reported an increased ability to differentiate into insulin-producing beta cells when human pancreatic beta cells were used to derive iPSCs, compared with beta cells derived from embryonic stem cells or alternate iPSC sources (4). In another study, when iPSCs were generated from bone marrow stromal cells and dermal fibroblasts from the same donor, cell type–specific genes were equally silenced when they were reprogrammed into iPSCs. However, gene expression differences in iPSC clones from a single cell type were greater than the differences observed between the 2 cell types (5). Notably, continuous passaging of iPSCs largely attenuates these differences.
In this issue of the Journal, Sanchez-Freire et al. (6) report comparisons of 2 different iPSC sources from the same donor: skin fibroblasts and cardiac-derived progenitor cells (CPCs) expressing the surface marker Sca1. Using uniform protocols for both iPSC induction and subsequent cardiac myocyte differentiation, these investigators compared the yield and functional characteristics of cardiac myocytes originating from Sca1+ CPCs (CPC-iPSC-CMs) and dermal fibroblasts (Fib-iPSC-CMs) from 2 fetal donors and 1 adult donor. The major distinctions observed were higher rates of cardiac myocyte differentiation and early activation of the cardiac transcription factor Nkx 2.5 and other myogenic transcription factors using the CPCs as the somatic cell source, as opposed to greater DNA methylation of the Nkx 2.5 promoter when skin fibroblasts were used as the somatic cell source. The investigators suggest that the reduced cardiac myocyte differentiation efficiency of fibroblast-derived iPSCs results from epigenetic memory, such as the persistent DNA methylation and consequent reduced expression of the Nkx 2.5 gene. These investigators found that CPC-derived iPSCs also differentiate into endothelial cells and smooth muscle cells more efficiently than do the fibroblast-derived iPSCs. Despite differences in efficiency of differentiation into cardiac cell types, the investigators established that cardiac myocytes derived indirectly from fibroblasts and CPCs are functionally equivalent in vitro, based on electrophysiological and calcium (Ca2+) handling properties. They also demonstrated that CPC-iPSC-CMs and Fib-iPSC-CMs have equivalent potential for therapeutic efficacy in vivo, based on similarly improved cardiac function following cell delivery to mouse hearts with experimental myocardial infarction.
Despite the rigor of these studies, the number of cases is small, and the data supporting a functional role of Nkx 2.5 DNA methylation are correlative. Studies relating specific manipulation of Nkx 2.5 methylation status to cardiac cell differentiation efficiency would be more compelling. Another shortcoming is the intrinsic difference in the cells selected from each location: fully differentiated dermal fibroblasts versus multipotent CPCs. From this perspective, a comparison of dermal fibroblasts with cardiac fibroblasts could allow more straightforward interpretation. Nevertheless, these findings support the concept that early-passage iPSCs retain epigenetic memory, which manifests in differential gene expression and altered differentiation capacity with preference to the cell type of origin.
The implications of these studies vary with one’s view of the most promising therapeutic cardiac myogenesis strategies. The functional equivalency of Fib-iPSC-CMs and CPC-iPSC-CMs would favor using an easily accessible somatic cell, such as dermal fibroblasts or blood cells, for iPSC generation and delivery to infarcted hearts. In this application, the lower efficiency of cardiac myocyte formation could be overcome by greater passaging. An alternate vision of therapeutic myogenesis for heart disease focuses on direct reprogramming of somatic cells into functional cardiomyocytes in situ without an iPSC intermediate. Direct reprogramming of cardiac fibroblasts in situ has already been demonstrated in murine hearts by 2 separate laboratories (7,8). From this perspective, the retained epigenetic memory of cardiac cells suggested by Sanchez-Freire et al. (6) could enhance the propensity for resident cardiac cells to transdifferentiate into new cardiac myocytes following appropriate direct reprogramming agents. Of course, the appreciation of epigenetic memory also suggests an additional myogenic strategy: manipulation of adult cardiac myocytes to allow transient re-entry into the cell cycle and proliferation while maintaining a cardiac myocyte phenotype throughout the process (9,10).
Ultimately, further understanding of cell type–specific epigenetic mechanisms in the regulation of reprogramming, maintenance of pluripotency, and induction of differentiation will be crucial for our utilization of iPSC biology and the achievement of therapeutic cardiac regeneration.
↵∗ Editorials published in the Journal of the American College of Cardiology reflect the views of the authors and do not necessarily represent the views of JACC or the American College of Cardiology.
Dr. Margulies has reported unpaid advisory committee membership for Novo Nordisk; advisory committee membership for AstraZeneca; research grant support from the U.S. National Institutes of Health; and research grant support from Juventis Therapeutics, Celladon Corporation, Thoratec Corporation, and Innolign Biomedical, LLC (R43 HL1117543). Ms. Alvarez has reported that she has no relationships relevant to the contents of this paper to disclose.
- American College of Cardiology Foundation