Initial studies have suggested that transplantation of embryonic stem (ES) cells following myocardial infarction (MI) in animal models is beneficial; however, the mechanism of benefit is largely unknown. The present study investigated the fate of mouse ES cells transplanted post-MI to determine if the ES cells give rise to the range of major cell types present in the native myocardium. MI was produced by coronary artery ligation in C57BL/6 mice. Two different mouse ES cell lines, expressing eGFP and beta-galactosidase, respectively, were tested. Post-MI intramyocardial injection of 3 x 10(4) ES cells was compared to injection of media alone. Histochemistry and immunofluorescence were used to track the transplanted ES cells and identify the resulting cell types. Echocardiography assessed the cardiac size and function in a blinded fashion. Two weeks post-MI, engraftment of the transplanted ES cells was demonstrated by eGFP or beta-galactosidase-positive cells in the infarct region without evidence for tumor formation. Co-immunolabeling demonstrated that the transplanted ES cells had become cardiomyocytes, vascular smooth muscle, and endothelial cells. Echocardiographic analysis showed that ES cell transplantation resulted in reduced post-MI remodeling of the heart and improved cardiac function. In conclusion, transplanted mouse ES cells can regenerate infarcted myocardium in part by becoming cardiomyocytes, vascular smooth muscle, and endothelial cells that result in an improvement in cardiac structure and function. Therefore, ES cells hold promise for myocardial cellular therapy.