Abstract 18448: Ckit Marks Cardiac Neural Crest Progenitors in the Developing Mouse Heart

Abstract

Introduction: Cardiac stem cells expressing the receptor tyrosine kinase c-Kit are a promising cell-based therapy for heart regeneration. However, there is ongoing controversy regarding the embryogenic origins and identity of these cells. We addressed this issue by using a tamoxifen-inducible cKitCreERT2/+ allele to establish the genetic fate-map of c-Kit in the developing mouse heart. Hypothesis: c-Kit marks an embryonic myocardial progenitor cell population which contributes extensively to mammalian cardiogenesis. Methods: cKitCreERT2/+ mice were crossed with R26RLacZ or IRG Cre-reporter mice. To induce recombination in c-Kit expressing cells, females pregnant with cKitCreERT2/R26RLacZ or cKitCreERT2/IRG fetuses were administered intraperitoneal injections of tamoxifen (100μl of 20mg/ml) from 7.5-8.5 (n=10) or 9.5-12.5 (n=15) days post coitum (dpc). Embryos were analyzed at 18.5dpc. Results: Confocal analysis, as well as x-gal staining, revealed that during 7.5-8.5dpc (the period which first and second heart field myocardial progenitors contribute to cardiogenesis) few to no labeled cells were seen in the heart, indicating that c-Kit does not mark progenitors of the cardiogenic mesoderm. In contrast, there was extensive cardiomyogenic contribution from cells expressing c-Kit during 9.5-12.5dpc. A mean of 171.4±21 c-Kit derivatives/field were detected throughout the myocardium [cells/field: 15.2±5.5 in outflow tract (OFT); 5.8±1.0 in left atrium; 16.1±4.7 in right atrium; 39.4±6.3 in right ventricle; 30.1±3.8 in left ventricle; 72.6±11.0 in interventricular septum], ~30% of which were cardiomyocytes (48.1±5.8 per field). Importantly, c-Kit contributed to all expected cardiac neural crest derivatives, including OFT, cardiac and aortic valves, cardiac conduction system, epicardium, endocardium and myocardium. Consistent with a neural crest origins, contribution to endothelium and smooth muscle of the OFT was documented, although coronary vascular cell differentiation was not observed Conclusions: Collectively, our findings strongly support that adult c-Kit+ CSCs are postnatal derivatives of c-Kit+ cardioblasts of neural crest origin, an original source of cardiomyocytes during cardiogenesis.