Monocytes are a subset of white blood cells critical for normal immune function. They are important components of inflammatory responses and are believed to significantly contribute to atherosclerotic lesion pathology. In atherosclerotic lesions, monocyte/macrophages have been shown to express a lipid oxidizing enzyme, 15-lipoxygenase (15-LO), that produces potent inflammatory mediators. Unique among the products of this enzyme are oxidized linoleate which we have shown to be the predominant oxidized fatty acid in atherosclerotic lesions. Normal monocytes do not express 15-LO, but are induced to do so by exposure to the cytokine IL-13 and expression of this enzyme is believed to correlate with disease progression in animal models of atherosclerosis. The studies proposed in this application will explore the IL-13 triggered signal transduction pathways in primary human monocytes and relate them to the downstream expression of 15-LO. Studies in Aim I will characterize the IL-13 receptor complex in human monocytes. Next, in Aim 2, the association and activation of Jaks with these receptor components will be assessed and activated Stats will be identified. The activation of Ser/Thr kinases will be evaluated (in Aim 3), expanding on our preliminary data that MAP kinases are activated by IL-13. The role of Jaks in this activation process will be investigated. Studies in Aim 4 will explore the role of Ser/Thr phosphorylation in regulating Jak/Stat phosphorylation and function. The biologic relevance of each of the activated pathways to the expression of 15-lipoxygenase and the related enhancement of monocyte-mediated LDL oxidation is the topic to be examined in studies described in Aim 5. In this final aim, additional investigations will be conducted to determine the role of the identified, IL-13 responsive pathways to general monocyte function with the goal.of identifying signaling pathways that are selectively involved in 15-LO expression but not required for other essential monocyte functions such as phagocytosis and superoxide anion production. The novelty of these studies is derived from the unique approaches to be employed to specifically dissect the roles of these pathways in intact human monocytes by virtue of the fact that monocytes are very conducive to antisense oligodeoxyribonucleotide manipulation of specific protein expression. These studies will substantially enhance our understanding of the signaling mechanisms induced upon exposure of monocytes to IL-13 and may suggest specific and selective means for modulating this response.
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