OF WORK This research area involves the study of embryonic stem cells prior to and during differentiation to cardiomyocytes. For these studies, we employ embryonic stem (ES) cells (R1, D3), embryonic germ cells and embryonic carcinoma cells. We have established one of the most efficient systems available for the generation of cardiomyocytes from ES cells in vitro. Immunofluorescence analyses of the differentiating ES-derived cardiac cells using monoclonal and polyclonal antibodies have been successfully performed for sarcomeric actins, troponin T. We have established when ryanodine receptor, SR CaATPase, phospholamban and dihydropyridine receptor are first expressed with in vitro differentiation, permitting studies of EC coupling with differentiation. Selection protocols have permitted the isolation of subpopulations of cardiomyocytes. Overall, the research is aimed at generating cardiac-lineage specific cells to understand the role of regulatory proteins in the formation of cardiomyocytes in vitro. Additionally, we have used genomic techniques like serial analysis of gene expression or microarrays to identify transcripts that may be involved in 1) pluripotentiality, 2) differentiation regulation, and 3) the viablility of cardiomyocytes in vitro. We are currently examining a number of these genes, which may be implicated in embryonic pluripotentiality and/or self-renewal. Several target genes are also being modified in the stem cells to determine there role in cardiomyocyte differentiation, and more recently, we have begun isolating (and targeting) cells to select sub-populations of cardiac progenitor cells. By studying the basic biology of embryonic stem cells, we hope to delineate mechanisms responsible for cardiomyocyte development and renewal. The goal in the coming year is to take some of our findings from mouse ES cells and test their functionin human
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