Malformations of the heart comprise the most frequently occurring type of birth defect. Furthermore, greater than 50 percent of trisomy 21 children have endocardial cushion defects. To begin to understand why these malformations arise, it is necessary to determine the cellular and molecular mechanisms that lead to the development of normal cardiac tissues. To assess the importance of vitamins and other nutrients for normal embryonic development, research needs to be performed that analyzes their functions during cardiovascular development. This application will explore heart development during the earliest embryonic stages when nondifferentiated mesoderm gives rise to both the myocardium and endocardium. Specifically, they will focus on the functions of vitamins A, D and K, and calcium in promoting this developmental event. For these studies, they will use the novel model system for cardiac cell differentiation, the quail cell line QCE-6. This cell line is representative of the tissue from which it was derived, precardiac mesoderm. Moreover, these cells can be induced to differentiate into either myocardial or endocardial cell types. Experiments using serum- free chemically defined medium conditions have demonstrated that these vitamins, along with calcium, are essential requirements the differentiation of these cells.
The aims of this applications are to determine: 1) the molecular mechanism(s) by which vitamins A and D regulate cardiac cell differentiation: and 2) whether the requirement for calcium in promoting cardiac cell differentiation involves a signaling pathway that includes vitamins D, K, integrins and extracellular matrix proteins. This application will use cell culture, immunohistochemistry, RNA and protein immunoblots, n situ hybridization, cell transfection with foreign DNA, confocal microscopy, micro injections of cells into developing embryos, and cDNA library screening to examine gene expression and regulation, calcium fluxes, integrin expression and ligation, and the effects of altered vitamin responsiveness on cardiac cell differentiation. The potential medical benefits of these experiments will be in providing information on the importance of nutrient supplementation in the maternal, prenatal diet, thereby reducing the number of infants born with cardiac defects.
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