This project proposes to determine factors that regulate glutathione synthesis in the lung and lung cells under normoxic conditions and conditions of oxidant exposure. We have obtained preliminary data which suggest that the level of GSH in pulmonary artery endothelial cells in culture is correlated with the rate of uptake (transport) of glutamic acid and cystine. Our first studies will include further characterization of this amino acid transport system and determination of the relationship between GSH levels and endothelial cell uptake of cystine and glutamate during different exposures to oxidant stresses, growth conditions and phases of the cell cycle. We will then investigate other lung cells including fibroblasts, smooth muscle cells, macrophages, and alveolar type II epithelial cells to see if GSH levels in these cells are also controlled by availability of amino acid precursors, or if other mechanisms of control such as direct uptake of intact GSH, transport of gamma-glutamyldipeptides or regulation of the feedback inhibition of glutathione are predominant control factors in these cells. We have succeeded in increasing GSH levels up to five-fold in cultured endothelial cells by treatment with low concentrations of diethylmaleate (DEM). We will attempt to produce similar increases in GSH in vivo by low doses of DEM over a 24 hr period. We will also use DEM and other treatments to increase cell levels of GSH and determine which types of oxidant stresses can be protected against by increases in GSH levels and during what time period GSH availability is most critical. Finally, we will attempt to determine whether blood or erythrocyte GSH levels reflect either the tissue GSH status or the general state of oxidant stress first in rats and then in patients undergoing oxygen therapy for treatments of disease conditions. The results of these studies should provide new insights into the development of strategies to protect against oxidant injury.
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