c-kit expression identifies cardiovascular precursors in the neonatal heart

Abstract

Directed differentiation of embryonic stem cells indicates that mesodermal lineages in the mammalian heart (cardiac, endothelial, and smooth muscle cells) develop from a common, multipotent cardiovascular precursor. To isolate and characterize the lineage potential of a resident pool of cardiovascular progenitor cells (CPcs), we developed BAC transgenic mice in which enhanced green fluorescent protein (EGFP) is placed under control of the c-kit locus (c-kitBAC-EGFP mice). Discrete c-kit-EGFP+ cells were observed at different stages of differentiation in embryonic hearts, increasing in number to a maximum at about postnatal day (PN) 2; thereafter, EGFP + cells declined and were rarely observed in the adult heart. EGFP+ cells purified from PN 0-5 hearts were nestin+ and expanded in culture; 67% of cells were fluorescent after 9 days. Purified cells differentiated into endothelial, cardiac, and smooth muscle cells, and differentiation could be directed by specific growth factors. CPc-derived cardiac myocytes displayed rhythmic beating and action potentials characteristic of multiple cardiac cell types, similar to ES cell-derived cardiomyocytes. Single-cell dilution studies confirmed the potential of individual CPcs to form all 3 cardiovascular lineages. In adult hearts, cryoablation resulted in c-kit-EGFP+ expression, peaking 7 days postcryolesion. Expression occurred in endothelial and smooth muscle cells in the revascularizing infarct, and in terminally differentiated cardiomyocytes in the border zone surrounding the infarct. Thus, c-kit expression marks CPc in the neonatal heart that are capable of directed differentiation in vitro; however, c-kit expression in cardiomyocytes in the adult heart after injury does not identify cardiac myogenesis.