? During the past funding period a new group of naturally occurring lipids, now known as the phospholipid growth factor (PLGF) family, has been isolated and characterized. We identified and synthesized several novel agonists and antagonists, clarified many of the receptor-mediated effects of PLGFs. PLGFs now emerge as important mediators of cell survival. The original objective continues in the renewal application, but the role in cell survival of PLGFs is the new focus of the investigation. The principal hypothesis of this proposal is that PLGFs through their specific receptors can regulate the survival of intestinal epithelium in response to injury caused by apoptosis-inducing treatments. The two aims for testing this hypothesis are: ? 1. Determine the mechanism by which LPA has antiapoptotic effects in IEC-6 intestinal epithelial cells. ? We will determine the structure-activity relationship among naturally occurring and synthetic lysophosphatidic acid (LPA) analogs to prevent the induction apoptosis after treatment with camptothecin and 7-irradiation, identify which LPA receptor subtypes are capable of mediating protection against apoptosis and determine the effects of LPA on the mitochondrial translocation and expression of Bcl-2-related proteins leading to cytochrome C release and ensuing activation of the caspase-9 --- caspase-3 pathway. ? 2. Optimize the protective effects of orally applied PLGF against radiation-induced apoptotic intestinal epithelial damage in vivo. ? We will determine the localization of LPA receptor subtypes in the villus-crypt unit and establish their role in the antiapoptotic effect using LPA1 and LPA2 receptor knockout mice, determine the pharmacokinetic properties of LPA to optimize the treatment regimen and evaluate the antiapoptotic efficacy amongst the three most effective PLGFs identified in aim 1. ? This is a revised application that has been considerably refocused in its scope. The experiments proposed will pave the way toward the therapeutic development of PLGFs for the treatment of the intestinal side effects of radiation and chemotherapy. These studies will have a twofold impact: first, advance our understanding of the physiological function of PLGF; and, second, provide new information concerning the regulation of apoptosis by G protein-coupled receptors. ? ?
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