The objectives of this proposal are to determine if GPIHBP1 is particularly important for the lipolytic processing of apo-B48-containing lipoproteins, to understand the atherosclerosis in Gpihbp1-deficient (Gpihbp1-/-) mice, and to create and analyze knock-in mouse models for naturally occurring GPIHBP1 mutations in humans. We recently found that Gpihbp1-/- mice have chylomicronemia, even on chow diet, with plasma triglyceride levels as high as 8000 mg/dl and plasma cholesterol levels as high as 800 mg/dl. Because GPIHBP1 is located on the luminal surface of capillaries of """"""""lipolytic tissues"""""""" (e.g., heart, muscle, fat) and because GPIHBP1 binds both lipoprotein lipase (LPL) and chylomicrons, we suspect that GPIHBP1 serves as a platform for the metabolic processing of triglyceride-rich lipoproteins. Of note, GPIHBP1 contains a strongly negatively charged amino-terminal domain that is important for binding both LPL and lipoproteins. Gpihbp1-/- mice have elevated levels of apo-B48-containing lipoproteins in their plasma, but normal levels of apo-B100-containing lipoproteins. I hypothesize that GPIHBP1 may have a particularly important role in the processing of apo-B48-containing liporoteins. To explore this hypothesis, I will examine the phenotypes of Gpihbp1-/- mice that are homozygous for the """"""""apo-B100-only"""""""" or """"""""apo-B48-only"""""""" mutations in Apob. Large lipoproteins such as chylomicrons are generally assumed to be nonatherogenic, but I have demonstrated that chow-fed Gpihbp1-/- mice develop spontaneous atheroscerotic lesions. To determine if the atherosclerosis in Gpihbp1-/- mice is driven by the cholesterol content of the """"""""triglyceride-rich"""""""" lipoproteins, I will examine the impact of ezetimibe and Npc1l1 deficiency on the susceptibility of Gpihbp1-/- mice to atherosclerosis. Recently, homozygous G56R and Q115P mutations in GPIHBP1 were identified in patients with chylomicronemia. Both mutations occurred in highly conserved residues. I found that the Q115P mutation abolishes the ability of GPIHBP1 to bind to LPL or chylomicrons, but the G56R mutation had no detectable effect-as judged by in vitro assays. These findings left me with uncertainty regarding whether the in vitro assays of GPIHBP1 were truly reliable indicators of GPIHBP1 function in vivo. To explore this issue, I will generate and characterize knock-in mouse models for the G56R and the Q115P mutations. The three specific aims of this proposal are: (1) to determine if GPIHBP1 is particularly important for the clearance of apo-B48-containing lipoproteins;(2) to determine if atherosclerotic lesions in chow-fed Gpihbp1- /- mice are driven by the cholesterol content of """"""""triglyceride-rich"""""""" lipoproteins;and (3) to create knock-in mouse models for the G56R and Q115P mutations in GPIHBP1.
TO PUBLIC HEALTH: Understanding lipoprotein metabolism is important because plasma lipoproteins deliver triglyceride fuel to vital tissues and because lipoproteins are causal factors in atherogenesis. A key goal of this project is to understand the role of GPIHBP1 in the lipoprotein lipase-mediated processing of triglyceride-rich lipoproteins. The proposed studies are likely to provide new information on how lipid nutrients are delivered to vital tissues;my experiments will also explore mechanisms for atherosclerosis in the setting of Gpihbp1 deficiency and define the in vivo relevance of specific human GPIHBP1 missense mutations.
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