Teratogenesis, i.e., the genesis of birth defects, is a complex process and one basic aspect of this process is teratogen-mediated cell death. More significant is the observation that teratogens kill certain cells in the embryo but spare others, i.e., teratogen exhibit cell-specific cytotoxicity. The long-term objectives of the proposed research are to understand the basis for cell specific cytotoxicity and its relationship to the overall process of teratogenesis. Using the technique of in vitro culture of mammalian (rat) embryos during the early phases of organogenesis, we have shown that the well-studied teratogen, cyclophosphamide (CP), induces a cytotoxic response in certain cells of the embryonic head. Cells of the developing heart, however, are completely resistant to the cytotoxic effects of CP. Building upon this information, we have developed a model system for studying the mechanisms underlying this differential sensitivity. Hopefully, information gained studying this model system will provide insights into the more general problem of why some cells within a developing tissue are sensitive to the cytotoxic effects of teratogen while others are resistant.
The specific aims of this continuation proposal are: (1) to continue to quantitate the kinetics of DNA-DNA and DNA- protein crosslinks in hearts and heads of embryos exposed to CP and/or metabolites/analogs using alkaline elution. (2) to assess the role of DNA repair in cell-specific cytotoxicity using alkaline elution, DNA adduct antibodies and mass spectrometry. (3) to assess the role of glutathione (GSH) and glutathione S- transferases (GST's) in cell-specific cytotoxicity. (4) to quantitate cell cycle times and cell cycle parameters for cells of different tissues of day 10 rat embryos as well as for cells within different regions of particular tissues using the technique of percent labeled mitosis (PLM).
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