Identification of novel molecules involved in cardiac differentiation of embryonic stem cells

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The aims of this project are to clarify the molecular mechanisms involved in in vitro differentiation of embryonic stem (ES) cells into cardiac myocytes, to improve the efficiency of cardiac differentiation, and to develop the isolation method of ES cell-derived cardiac myocytes. For those purposes, the multiple approaches have been taken. First, we analyzed the role of Smads and their specific ligands during cardiac differentiation of ES cells, and found that Smad2 was activated bimodally in the early and late phases of cardiac differentiation. Smad2 is activated by Nodal/Cripto in the early phase, which is indispensable for endodermal and mesodermal induction and subsequent cardiac differentiation. TGF-β and Activin are responsible for the late Smad2 activation, which negatively regulates cardiomyogenesis by modulating proliferation and differentiation of cardiac myocytes. Second, we performed signal sequence trap on cardiac myocytes, and identified several transmembrane molecules, which have not been recognized as being expressed in cardiac myocytes. Four of these are induced to be expressed during cardiac differentiation of ES cells and Pl9CL6 cells, and the role of these molecules in cardiac differentiation is now being analyzed. We also identified RNA aptamers, which specifically bind to cardiac myocytes, by systematic evolution of ligands by exponential enrichment (SELEX). The isolation method of cardiac myocytes from differentiated ES cells are now being developed with the identified aptamers.