Isoeicosanoids& Biologically Active Oxidized Phospholipi

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Lipid peroxidation is a naturally occurring process that in part, protects cells and cellular macromolecules from reactive oxygen species. It is now becoming widely appreciated that lipid peroxidation also leads to biologically active products that mediate cellular response to free radical processes. Also, reactive chemical entities are formed during lipid peroxidation that are sufficiently electrophilic to covalently modify proteins and other biopolymers. The overall goal of the proposed research is to explore the initial chemical event taking place during the free radical oxidation of polyunsaturated fatty acids present as phospholipid esters in the membranes of cells. A major focus of the individual studies will center around the oxidation of arachidonic acid as a particularly susceptible target for peroxidative events. Mass spectrometry will be used as the primary tool for structural characterization of oxidized phospholipids. Structural studies of covalent adducts of electrophilic lipid free radical products and target proteins will be carried to determine the site of adduct formation and techniques developed to determine the structure of the lipid portion of the covalent adduct. A second major focus will be developed to determine the structure of the lipid portion of the covalent adduct. A second major focus will be testing the alternative pathway of leukotriene A4. The structural characterization of biologically active phospholipid products of phospholipid oxidation will be a focus of a collaborative project to characterize those oxidized phospholipid products that may induce the anti-inflammatory macrophage phenotype. A second collaborative study will involve the characterization of oxidized phospholipid products that stimulate cyclooxygenase gene induction in endothelial cells. Finally, a method to collect lipid peroxidation products present in aerosols of expired breath in human subjects will be investigated. This novel, non-invasive method will be used to assess lipid peroxidation events in the lung that can be quantitated through measurement of markers using mass spectrometric techniques. Studies of human subjects with upper airway, lower airway, and alveolar lung disease will be undertaken using these techniques of collecting non- volatile molecules in expired breath and quantitation of specific products of arachidonate peroxidation.