Surgery, trauma, and sepsis all induce tissue damage and inflammation. The cellular production of radical oxygen intermediates (ROI) [including superoxide (O2-), hydrogen peroxide (H2O2) and hydroxyl radical (OH)] play a major role in disturbances to physiologic homeostasis in all these insults. It has been found that administration of protective agents against these reactive species can, under some but not all conditions, afford protection against cell and tissue damage (oxidative injury). Nitrogen radicals are also formed under inflammatory conditions. The present proposal focuses on the role in oxidative tissue injury of the interaction between ROI and nitric oxide (NO). Our laboratory was the first to discover NO production in the liver, and we have published extensive data delineating the regulation of its production by cytokines. Recently we have shown that endogenous NO production in the isolated hepatocyte inhibits a protective mechanism against oxidative damage (catalase) and increases oxygen radical production; stimulation of oxygen radical production in these isolated cells also decreases NO formation and modifies its oxidation products. In the isolated heart, we have also shown endogenous NO production and found that this NO protects against ischemia/reperfusion injury. We have already shown previously that endogenous production of NO protects against oxygen radical-mediated hepatic injury in sepsis, a model for end-stage organ failure. We therefore know that NO/oxygen radical interactions occur in vitro and in vivo. In this proposal we will systematically and serially vary the presence of oxygen and nitrogen radicals. We will add them exogenously or induce cells and tissues to produce them endogenously. Our approach will be to determine the effects on oxidative injury (lipid peroxidation, cell lysis) of the simultaneous presence of these two types of radicals, because the chemistry of their interaction predicts that under some circumstances they will neutralize one another, and, under other circumstances, toxic hydroxyl radicals will be generated. We will therefore determine the effects of nitrogen radicals on key biochemical events associated with oxygen radical-induced cell and organ damage. We then experimentally manipulate protective mechanisms known to be effective against oxygen radicals and determine the effects when nitrogen radicals are also present. Although the focus will be on isolated hepatocytes in vitro where conditions can be precisely controlled, a limited number of experiments are planned with isolated perfused liver and with liver in vivo. A single experiment with the isolated perfused heart will provide comparative data in another organ. Considerable information should be generated concerning the protective and possibly injurious role of nitric oxide under conditions of stress.
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