Peroxidation of membrane lipids is intrinsic to many pathological processes. Elaborate antioxidant defenses have evolved to prevent or minimize lipid peroxidation suggesting that its control is critical to maintaining cell integrity and survival. Nevertheless, persistent oxidant stress can compromise cellular defenses as detected through oxidatively damaged cell components. This points to the need for secondary defense systems to handle damaged membrane components that escaped primary antioxidant defenses. Free radical induced membrane lipid peroxidation is known to activate phospholipase A2 with excision of peroxidized fatty acids. Although this hydrolytic activity is directed largely to oxidized lipids, there is evidence that considerable amounts of intact fatty acids are also released. Analysis of lipids from ischemic tissues, as well as tissues subjected to other forms of oxidative injury, indicates that sizable amounts of phospholipids are degraded by phospholipase A2. Degradation of membrane phospholipids under conditions of severe oxidant stress is thought to contribute to injury by physical disruption of the membrane as well as by formation of inflammatory mediators derived from the eicosanoid pathway. The proposed studies will examine mechanisms responsible for phospholipase A2 activation following membrane lipid peroxidation to test the hypothesis that peroxidation creates alterations in membrane phospholipid structure that are susceptible to hydrolysis. The proposed studies will examine relationships between preferred release of arachidonic acid along with oxidized fatty acids since recent findings suggest that peroxidation may specifically mediate the release of arachidonic acid. Liposomes, isolated cell membranes and cultured cells will be used to examine relationships between induction of membrane microheterogeneity and phospholipase A2-mediated release of specific fatty acyl components following lipid peroxidation. Using specific labeling techniques, we will also examine the effect of peroxidation on phospholipase A2-mediated arachidonic acid release and subsequent eicosanoid synthesis in cultured cells. These studies are aimed at determining the influence of low level lipid peroxidation on eicosanoid synthesis which is postulated to occur via membrane structural modulation of phospholipase A2 activity.
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