Abstract 305: CREG Induces Cardiac Differentiation From Embryonic Stem Cells
The use of embryonic stem (ES)-derived cardiomyocytes to repair damaged heart is a new therapeutic paradigm for heart failure due to myocardial infarction. In order to generate a large number of cardiomyocytes at their appropriate differentiation stage for transplantation and to ensure their integration and maturation in the diseased heart, it is necessary to understand the mechanisms of cardiogenesis in embryonic development and ES cell differentiation. In this study we have developed a model system in which ES cells are first differentiated into cystic embryoid bodies (EBs) that contain visceral and parietal endoderm as well as epiblast. By selective ablation of specific germ layers, we show that the visceral endoderm plays an instructive role in cardiac differentiation whereas the parietal endoderm has inhibitory effects. We further show that the cellular repressor of E1A-activated genes (CREG), a secreted glycoprotein, is highly expressed together with its putative receptor insulin-like growth factor 2 receptor in the visceral endoderm and developing cardiomyocytes during mouse embryogenesis and EB differentiation. Stable overexpression of CREG in EBs augments cardiomyocyte differentiation and maturation as evidenced by increased number of beating EBs, enlarged beating area and accelerated beating frequency. Of note, the CREG-overexpressing EBs often differentiate to form distinct beating spherules with well developed adherens junctions. Moreover, our immunofluorescence data demonstrate that the expression of the gap junction protein connexin43 is significantly increased in a punctate pattern at cardiomyocyte junctions as compared to the control EBs. These results suggest that CREG is an endoderm-derived factor that promotes the differentiation and maturation of cardiomyocytes from ES cells.