Cardiac Bmi1(+) cells contribute to myocardial renewal in the murine adult heart

Introduction: The mammalian adult heart maintains a continuous, low cardiomyocyte turnover rate throughout life. Although many cardiac stem cell populations have been studied, the natural source for homeostatic repair has not yet been defined. The Polycomb protein BMI1 is the most representative marker of mouse adult stem cell systems. We have evaluated the relevance and role of cardiac Bmi1(+) cells in cardiac physiological homeostasis. Methods: Bmi1(CreER/+); Rosa26(YFP/+) (Bmi1-YFP) mice were used for lineage tracing strategy. After tamoxifen (TM) induction, yellow fluorescent protein (YFP) is expressed under the control of Rosa26 regulatory sequences in Bmi1(+) cells. These cells and their progeny were tracked by FACS, immunofluorescence and RT-qPCR techniques from 5 days to 1 year. Results: FACS analysis of non-cardiomyocyte compartment from TM-induced Bmi1-YFP mice showed a Bmi1 (+)-expressing cardiac progenitor cell (Bmi1-CPC: B-CPC) population, SCA-1 antigen-positive (95.9 +/- 0.4 \%) that expresses some stemness-associated genes. B-CPC were also able to differentiate in vitro to the three main cardiac lineages. Pulse-chase analysis showed that B-CPC remained quite stable for extended periods (up to 1 year), which suggests that this Bmi1(+) population contains cardiac progenitors with substantial self-maintenance potential. Specific immunostaining of Bmi1-YFP hearts serial sections 5 days post-TM induction indicated broad distribution of B-CPC, which were detected in variably sized clusters, although no YFP+ cardiomyocytes (CM) were detected at this time. Between 2 to 12 months after TM induction, YFP+ CM were clearly identified (3 +/- 0.6 \% to 6.7 +/- 1.3 \%) by immunohistochemistry of serial sections and by flow cytometry of total freshly isolated CM. B-CPC also contributed to endothelial and smooth muscle (SM) lineages in vivo. Conclusions: High Bmi1 expression identifies a non-cardiomyocyte resident cardiac population (B-CPC) that contributes to the main lineages of the heart in vitro and in vivo.We wish to thank M. Torres, J.M. Perez-Pomares and B.G. Galvez for critical discussions of the manuscript, A. M. Santos for assistance with confocal microscopy and dynamic imaging, R.M. Carmona for help with the animal colony management, F.S. Cabo for bioinformatics and statistical support, J.M Ligos for the sorting strategy, and K. McCreath and C. Mark for editorial support. This study was supported by grants to A.B. from the Ministry of Science and Innovation (SAF2012-34327; PLE2009-0147 and PSE-010000-2009-3), the Research Program of the Comunidad Autonoma de Madrid (S2010/BMD-2420), the Instituto de Salud Carlos III (RETICS-RD12/0019/0018 and RETICS-RD12/0019/0023) and the European Commission (Proposal 242038). The CNB-CSIC and CNIC are supported by the Spanish Ministry of Economy and Competitiveness.S