Abstract
Background: We have previously established that induced pluripotent stem (iPS) cells spontaneously differentiate into beating cardiomyocytes. The present study was designed to identify cardiac progenitor cell (PCs) population in iPS cells and determine their cellular and molecular characteristics and myocardial regenerative potential. Methods and Results: PCs were sorted from day-5 embryoid bodies (EBs) on flow cytometer. Surface marker analysis showed that PCs were cKit low Sca1 high Flk high besides positivity for Nkx2.5 and GATA4. When exposed to 1% hypoxia for 24 hours, PCs showed significant increase in gene expression of proangiogenic and cardiomyogenic factors like VEGF, Flt-1, Angiopoietin-1, vWF, and Nkx2.5 as compared to the cells cultured under normoxia. VEGF secretion in the conditioned medium from PCs cultured under hypoxia was 290 pg/ml ( p <0.01 vs cells under normoxia, 126pg/ml) and showed enhanced in vitro angiogenic potential as determined by tube formation assay (39.8±1.9, p <0.01 vs control) and cell invasion assay (220.5±1.8; p <0.01 vs control) using rat aortic endothelial cells. For in vivo determination of cardiogenic potential of PCs, 20µl DMEM without cells (group1, n=6) or containing 5.0x10 5 male cKit low Sca1 high Flk high PCs labeled with PKH26 and cultured under normoxia (group2, n=8) or 1% hypoxia for 24 hours (group3, n=8) were injected intramyocardially in a female mouse model of acute coronary artery ligation. Real time PCR on day7 for Sry gene (n=4/group) showed higher survival in group-3 ( p <0.05 vs group-2). Echocardiography at 4 weeks after cell transplantation, significant improvement in global heart function was observed in group-3 (LVEF =55.9%, LVFS =21%; p <0.05 vs group-2). Fluorescence immunostaining of the histological sections for myogenic and endothelial cell specific markers showed cardiac differentiation and increased blood vessel density in group-3 at 40x ( p <0.01 vs group-2). Conclusions: iPS cell-derived PCs comprise a multipotent precursor population capable of repopulating the infarcted heart to preserve global cardiac function.
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