The """"""""free radical theory of oxygen toxicity"""""""" links the deleterious pulmonary effects of hyperoxia, cellular oxygen metabolism and the respiratory burst of activated inflammatory cells to highly reactive metabolic products of oxygen. These reactive oxygen species can inactivate cellular enzymes, damage DNA and destroy lipid bilayers. To protect cells from these cytotoxic oxygen metabolites, a group of cooperative antioxidant systems have evolved. The primary defense being the superoxide dismutases. It has been shown repeatedly that the ability to respond to a free radical challenge with increased antioxidant defenses is strongly associated with the cell's tolerance to superoxide induced injury and often survival. Therefore, understanding the lung's normal mechanisms for stimulating endogenous antioxidant defenses may lead to logical steps in the development of therapeutic regimens that are effective in preventing and ameliorating free radical mediated pulmonary toxicity. To this end, the goals of this proposal are to understand the molecular mechanisms which control the acute phase activation of manganese superoxide dismutase in pulmonary epithelial and endothelial cells. We plan to identify and characterize promoter and enhancer elements, relative to their role in mediating the MnSOD gene's response to the inflammatory mediators: LPS, Il-1, TNF and the anti-inflammatory glucocorticoids. Using in vivo footprinting, we plan to delineate, at single nucleotide resolution, the position of cis-acting regulatory sequences which mediate the gene's response to these inflammatory mediators. The nucleotide contacts defined by in vivo footprinting will be utilized to identify and purify the appropriate stimulus-linked transacting factors and their coding sequences.
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