The research objective is to define the role and status of cellular constituents that function in cellular defense systems against reactive intermediates derived from toxic chemicals. The agents to be investigated are glutathione (GSH), vitamin E (alpha-tocopherol) and vitamin C (L-ascorbic acid). The studies are being conducted with rat hepatocytemodels that have been compromised in their cellular defense mechanisms against chemically induced cell injury and death. The models include cells that have either; 1) irreversibly inactivated glutathione reductase, 2) low levels of glutathione peroxidase due to being isolated from rats on a selenium deficient diet, 3) vitamin E deficiency due to being isolated from rats on a vitamin E deficient diet, 4) compromised by omission of calcium from the medium and 5) undergone depletion of GSH by GSH-depleting agents and placed in a medium that is free of sulfur amino acids. These hepatocyte models are being utilized to investigate the hypothesis that cells may have as a common pathway to injury and death from exposure to many toxic chemicals the loss of cellular defense systems which allows for endogenous oxidative stress to contribute to a significant degree to loss of cell viability. The mechanism proposed for endogenous oxidative stress and chemically induced oxidative stress is the alteration of the homeostasis of certain protein sulfhydryls leading to a loss of the thiol/disulfide redox balance needed for cell survival. A major effort will be devoted to the elucidation of the protein sulfhydryl mixed disulfides that are formed during acute cell injury. In addition, experiments are designed to determine if the GSH, vitamin E and vitamin C levels in hepatocytes during exposure to toxic agents including environmental agents are the result of these cellular protective agents having on electron antioxidant properties and thus possible """"""""sparing"""""""" effects. Efforts are being made to determine if an intracellular redox relationship exists between GSH, vitamins C and E that involves differential rates of electron and hydrogen transfers that may have a key role in the maintenance of cell viability during and after acute chemically induced cell injury. In vitro models with specific phospholipids are being utilized to provide the basis for the interpretation of results from the compromised hepatocyte model experiments. Model chemicals being used include ethyl methanesulfonate, bromobenzene, acetaminophen, A23187, Adriamycin, t-butyl hydroperoxide, methyl ethylketone peroxide and 1,3-bis-(20chlorethyl)-1-nitrosourea (BCNU).
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