HDL and apoA-I plasma levels are inversely related to coronary artery disease. Two major subclasses of HDL are those containing only apoA-I (LpAI) and those containing both apoA-I and apoA-II (LpAI/AII). Whereas apoA-1 is considered to be generally anti-atherogenic, evidence points to apoA-II being pro-atherogenic. The HDL receptor, SR-BI, plays an important role in the metabolism in HDL. SR-BI mediates the selective uptake of cholesterol esters from HDL into cells and enhances the efflux of cellular free cholesterol to HDL. Although SR-BI is considered to exhibit a broad lipoprotein binding specificity, the group of the PI has found in unpublished studies that LpAI/AII is markedly deficient in its ability to bind SR-BI. The central hypothesis to be examined in this proposal is that apolipoprotein AII exerts a regulatory function by inhibiting HDL binding to SR-BI and consequently SR-BI-mediated lipid flux between HDL and cells. Such a regulatory function of apoA-II would have major implications for HDL metabolism, including its role in reverse cholesterol transport. This proposal therefore seeks to examine the influence of apoA-II on SR-BI-mediated HDL metabolism and on cholesterol flux between HDL and cells.
The specific aims are: 1) to determine the effect of apoA-II on SR-BI-mediated cholesterol flux between HDL and cells. This will be accomplished by studying the influence of human apoA-II on the binding and metabolism of human HDL by SR-BI expressed in transfected CHO cells. ApoA-I only and apoAI/apoAII-containing particles will be compared and the effect of increasing apoA-II levels in reconstituted particles studied, 2) to assess the effect of apoA-II on HDL levels and on hepatic selective cholesterol ester uptake in mice. This will be achieved by determining relative rates of hepatic selective cholesterol ester uptake from LpAI and LpAI/AII in apoA-1-/- mice and the effect of adenovirus-mediated expression of increasing plasma levels of human or mouse apoA-II on HDL levels in human apoA-I transgenic mice or C57BL/6 mice and 3) to investigate the structural requirements of apoA-II necessary for the modulation of apoA-I as an SR-BI ligand. This will be accomplished by studying both in vivo and in cultured cells, the effects of a series of structural variants and mutations of apoA-II on HDL binding to SR-BI and on selective cholesterol ester uptake.
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