TOXIC AND THERAPEUTIC EFFECTS OF OXYGEN

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This application is for a Program Project Grant to study relationships at the cellular level between molecular oxygen, reactive oxygen species (ROS) and nitric oxide (NO.) under physiological and pathological conditions. Factors that determine the fate of oxygen and regulate its interactions with NO. and tissue thiol pools is the investigative theme for the four projects that comprise the proposal. Project 1, a study of brain metabolism during and after carbon monoxide (CO) hypoxia, proposes that mitochondrial ROS generation in different brain regions is a significant source of neuronal injury. The investigators will relate ROS production to changes in excitotoxicity, NO. production and O2 availability. ROS production including NO. production will be correlated with changes in neuropathology and function to better understand their roles in neuronal injury. The next project studies the role of nitric oxide in the cellular biochemistry of the airways of the lungs. It addresses a major paradox in the lung, i.e., that the biological activity of NO. is in conflict with its established toxicity in an oxygen-rich environment. Answers to this apparent paradox will be sought in the capacity of NO. to undergo oxidation-reduction reactions that determine the formation of adducts with protein thiol moieties located both extracellularly and intracellularly. Protein thiol NO. adducts, or S-nitrosothiols, which are resistant to reactions with molecular oxygen, possess potent biological activity. Project 3 will evaluate responses of isolated endothelial cells to nitric oxide. The mechanisms of action of NO., which interacts with a variety of cellular targets, will be investigated via the glycolytic enzyme, GAPDH. Efforts also will focus on mechanisms of NO. formation in endothelial cells, regulation of NO. formation, and the effects of prolonged exposure of cells to NO. with emphasis on changes in cellular protein thiol pools. The last project will evaluate the role of extracellular superoxide dismutase (EC-SOD) in modulating lung airway and vascular function. It is based on the important finding that EC-SOD, which is highly constitutively expressed in the lungs, has been localized in the extracellular spaces around smooth muscle cells in airways and in pulmonary vasculature. The hypothesis will be tested that EC-SOD is a key bioregulator of NO. activity. Both EC-SOD and NOS. will be colocalized in several tissues and the functional relationships of EC-SOD, O2 and NO. will be explored in both airway and vascular structures.