The atherogenicity of oxidatively low density lipoprotein (LDL) is being investigated by numerous groups throughout the world. Studies addressing the atherogenic effects of oxidized LDL are aimed at providing new insights into mechanisms by which progression of this important disease takes place, enabling development of alternate or complimentary measures for identifying risk factors in humans as well as new therapeutic approaches. The relationship between oxidant stress and atherogenic effects of oxidized LDL will be studied in this project in terms of the pro-oxidant properties of LDL, effects apparently associated with lipid peroxides. This project draws upon the recent isolation of an oxidatively modified LDL from human plasma which is enriched in lipid peroxidation products as well as modifications to athe apoprotein. This oxidized lipoprotein (referred to as LDL)-) exists in a state approaching the threshold of modification characteristic of in vitro forms of oxidized LDL that exert atherogenic effects on vascular cells, largely described in model and cell culture systems. A primary goal of the proposed studies will be to d attermine the extent to which LDL- exhibits atherogenic properties analogous to the in vitro forms of oxidatively modified LDL's its potential for further oxidation, and the relationship between its extent of oxidation and """"""""atherogenicity"""""""". We propose that the atherogenic properties of oxidized LDL, described in cell culture systems via cytotoxicity, uptake by non- regulated alternate receptors rather than the regulated normal LDL receptor, and accumulation of cholesterol esters (intracellular lipid loading) are linked to its pro-oxidant properties. This hypothesis will be tested in endothelial and macrophage cell systems by treatments with LDL- before and after in vitro oxidation using well characterized oxidizing systems. The oxidant stress produced on these cells will be determined by measurements of cell lipid peroxidation and stress protein/antioxidant responses. Studies focus on heme oxygenase (HO) which may exert an antioxidant effect by degrading heme released during oxidative attack of cellular hemoproteins. Concurrent induction of certain cytokines also takes place and this project will focus on MCP-1 in terms of its relationship to HO induction ad specific atherogenic reactions described for endothelial cells and monocyte macrophages treated with oxidized LDL preparations. The ability to reduce the pro-oxidant responses with antioxidants or by pre-adaptation of cells to oxidant stress will serve as parameters for determining the LDL oxidant stress capacity. Concurrently, the ability of LDL- to induce non-LDL-receptor-mediated uptake and cholesterol ester accumulation, together with inhibition of these effects by antioxidants or pre-adaptation to oxidant stress, will be key parameters of investigation.
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