Selection of ventricular-like cardiomyocytes from ES cells in vitro

Ischemic disorders of the heart can cause an irreversible loss of cardiomyo cytes resulting cause an irreversible loss of cardiomyocytes resulting in a substantial decrease of cardiac output. The therapy of choice is heart tra nsplantation, a technique that is hampered by the low number of donor organ s. in the present study, we describe the specific labeling, rapid but gentl e purification and characterization of cardiomyocytes derived from mouse pl uripotent embryonic stem (ES) cells. To isolate the subpopulation of ventri cular-like cardiomyocytes, ES cells were stable transfected with the enhanc ed green fluorescent protein (EGFP) under transcriptional control of the ve ntricular-specific 2.1 kb myosin light chain-2v (MLC-2v) promoter and the 0 .5 kb enhancer element of the cytomegalovirus (CMVenh.). First fluorescent cells were detected at day 6 + 8 of differentiation within EBs. Four weeks after initiation of differentiation 25% of the cardiomyocyte population dis played fluorescence. Immunohistochemistry revealed the exclusive cardiomyog enic nature of EGFP-positive cells. This was further corroborated by electr ophysiological studies where preferentially ventricular phenotypes, but no pacemaker-like cardiomyocytes, were detected among the EGFP-positive popula tion. The enzymatic digestion of EBs, followed by Percoll gradient centrifu gation and fluorescence-activated cell sorting, resulted in a 97% pure popu lation of cardiomyocytes. Based on this study, ventricular-like cardiomyocy tes can be generated in vitro from EBs and labeled using CMVenh./MLC-2V-dri ven marker genes facilitating an efficient purification. This method may be come an important tool for future cell replacement therapy of ischemic card iomyopathy especially after the proof of somatic differentiation of human E S cells in vitro.