Abstract 19780: Mammalian Cardiac Renewal by Preexisting Cardiomyocytes

Abstract

Although recent studies have revealed that adult heart cells are generated after birth in mammals, the frequency and source of these new heart cells is unclear. Here we combined two different pulse-chase approaches--genetic fate-mapping and stable isotope labeling with Multi-isotope Imaging Mass Spectrometry (MIMS)--to determine the origin of cardiomyocytes in adult mice. MIMS allows quantitative imaging of stable isotope reporters in domains smaller than a micron cubed. The nuclear incorporation of a 15N stable isotope-enriched tracer here as 15N-thymidine is detectable with high precision by an increase in the 15N/14N ratio above natural abundance (0.37%) in a pattern resembling chromatin evident in cells having divided during a labeling period. We administered 15N-thymidine for 8 weeks to three age groups of C57BL6 mice starting at day-4 (neonate), at 10-weeks (young adult) and at 22-months (old adult) and extrapolated DNA synthesis yields a yearly rate of 5.5% in the young adult that declines further to 2.6% in advanced aged mice. Fig1a. To evaluate the origin of newly generated cardiomyocytes, we performed 15N-thymidine labeling of double-transgenic MerCreMer/ZEG mice, which we previously developed for genetic lineage mapping. Fig1b. After treatment with 4OH-tamoxifen, we show that DNA synthesis occurs at a low rate in predominantly pre-existing cardiomyocytes [77% expressed GFP, a frequency essentially identical to that of surrounding 15N- cardiomyocytes (15N, 77% vs 15N-, 84%; Fisher’s exact=n.s.)]. Fig1c. Cell cycle activity in cardiomyocytes led to polyploidy and multinucleation, and to new diploid, mononucleated cardiomyocytes [17% of total; predominantly GFP+ (83% vs 82%, p=n.s.)]. Fig1d. Our results support a low rate of cell division of cardiomyocytes as the origin of mammalian adult cardiogenesis and are inconsistent with a high rate of stem cell activity in the normal mammalian myocardium.