Lipid bodies, non-membrane bound cytoplasmic inclusions, have newly identified roles in the formation of arachidonate-derived mediators by leukocytes and other cells involved in inflammation. Lipid bodies are prominent in cells in vivo associated with inflammation, and the formation of lipid bodies represents a cellular response mediated by protein kinase C activation which leads to a coalescence of lipids and proteins into discrete intracellular domains. Lipid bodies are both stores of arachidonyl-phospholipids and sites at which specific enzymes involved in arachidonate metabolism localize. The proposed studies will evaluate the roles of lipid bodies in the metabolism of arachidonic acid. Three aspects of lipid body formation and function will be studied. First, the cellular events that lead to the formation of arachidonate-containing lipid bodies will be investigated in leukocytes and other cells. Second, the mechanisms that regulate the mobilization of arachidonic acid from within glycerolipids of lipid bodies will be evaluated, examining the activities and regulation of phospholipases active on classes of lipid body phospholipids. Third, the mechanisms by which liberated arachidonic acid is utilized to form eicosanoid derivatives will be explored. These studies will assess the means by which lipoxygenase and cyclooxygenase enzymes associate with lipid bodies in order to utilize lipid body stores of arachidonic acid. These three aspects of lipid body function will be investigated using lipid bodies isolated by subcellular fractionation as well as methods of immunolocalization to evaluate proteins associated with lipid bodies within cells.
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