Abstract
BackgroundIt has long been thought that mammalian cardiomyocytes are terminally-differentiated and unable to proliferate. However, myocytes in more primitive animals such as zebrafish are able to dedifferentiate and proliferate to regenerate amputated cardiac muscle.Methodology/Principal FindingsHere we test the hypothesis that mature mammalian cardiomyocytes retain substantial cellular plasticity, including the ability to dedifferentiate, proliferate, and acquire progenitor cell phenotypes. Two complementary methods were used: 1) cardiomyocyte purification from rat hearts, and 2) genetic fate mapping in cardiac explants from bi-transgenic mice. Cardiomyocytes isolated from rodent hearts were purified by multiple centrifugation and Percoll gradient separation steps, and the purity verified by immunostaining and RT-PCR. Within days in culture, purified cardiomyocytes lost their characteristic electrophysiological properties and striations, flattened and began to divide, as confirmed by proliferation markers and BrdU incorporation. Many dedifferentiated cardiomyocytes went on to express the stem cell antigen c-kit, and the early cardiac transcription factors GATA4 and Nkx2.5. Underlying these changes, inhibitory cell cycle molecules were suppressed in myocyte-derived cells (MDCs), while microRNAs known to orchestrate proliferation and pluripotency increased dramatically. Some, but not all, MDCs self-organized into spheres and re-differentiated into myocytes and endothelial cells in vitro. Cell fate tracking of cardiomyocytes from 4-OH-Tamoxifen-treated double-transgenic MerCreMer/ZEG mouse hearts revealed that green fluorescent protein (GFP) continues to be expressed in dedifferentiated cardiomyocytes, two-thirds of which were also c-kit+.Conclusions/SignificanceContradicting the prevailing view that they are terminally-differentiated, postnatal mammalian cardiomyocytes are instead capable of substantial plasticity. Dedifferentiation of myocytes facilitates proliferation and confers a degree of stemness, including the expression of c-kit and the capacity for multipotency.
Highlights
Generations of physicians have been taught that the heart is a static organ, incapable of self-renewal
As an alternative to flow cytometry, which is not well-suited for cells as large as adult cardiomyocytes [25], we employed high-density tile scanning confocal microscopy: myocyte preparations of more than 100,000 cells were cyto-spun onto 22 mm culture glasses
No cells expressing c-kit, CD31 (PECAM, an endothelial cell marker), CD34 or CD90 were observed in purified cardiomyocyte samples containing as many as 500,000 counted cells (Figure S1) [2]
Summary
Generations of physicians have been taught that the heart is a static organ, incapable of self-renewal. That dogma has been undermined by the recognition that the adult mammalian heart contains its own reservoir of progenitor (or stem) cells [1,2,3,4,5]. Cardiomyocytes in the human heart are renewed throughout life [6]. In contrast to the prevailing view for mammals, hearts from amphibians and zebrafish exhibit surprising potential to regenerate cardiac muscle by partial dedifferentiation [7,8], and possibly by stem cell-mediated regeneration as well [9]. Epidermal cells, pancreatic beta cells and adipose stromal cells dedifferentiate and exhibit stem cell phenotypes [11,12,13,14,15]. Myocytes in more primitive animals such as zebrafish are able to dedifferentiate and proliferate to regenerate amputated cardiac muscle
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