Atherosclerosis remains one of the major causes of morbidity and mortality in the US. Epidemiologic studies have shown a clear relationship between serum cholesterol levels and development and progression of atherosclerosis. As a result, major efforts have been instituted to reduce the serum cholesterol of individuals at risk, through lifestyle changes or drug therapies. Presumably, if reduction of serum cholesterol is to produce a reduction in existing atherosclerotic lesions an important mechanism would be to enhance removal of cholesterol from sites of accumulation, however, such mechanisms remain poorly understood. The long term goal of this proposal is to identify specific transport pathways involved in HDL mediated clearance of excess cholesterol from cells to gain further understanding of the processes involved in regression or formation of lipid laden foam cells as occur in atherosclerosis. We hypothesize that clearance of excess cellular cholesterol is an active process depending on stimulation by appropriate extracellular cholesterol acceptors that promote intracellular cholesterol transport to sites available for removal. This project aims to show that efflux of intracellular cholesterol requires an apolipoprotein dependent pathway mediated by active cellular processes distinct from non-specific aqueous diffusion of cholesterol and define cellular mechanism that facilitate transport of cholesterol from intracellular sites to sites available for removal by extracellular cholesterol acceptors. Cultured cells will be the experimental model to study cholesterol efflux pathways. Advantage will be made of somatic cell mutants with known defects in vesicular transport. Cholesterol efflux will be measured by changes in cell cholesterol mass and radioactivity, and changes of activities regulated by cell cholesterol levels. We will compare various extracellular cholesterol acceptor types to establish the contribution of apolipoprotein independent and dependent pathways for efficient cholesterol removal and establish if a link exists between efflux of excess cholesterol and phospholipids by the apolipoprotein dependent pathway. We will characterize cellular cholesterol transport pathways involving the Golgi apparatus and identify the contribution of proteins known to regulate vesicular transport. These studies will aid in delineating the cellular pathways involved in cholesterol transport through and out of the cell. Understanding these mechanisms will give greater knowledge of the atherosclerotic process and HDL function, and will lend insights into potential targets for pharmacological interventions to reduce the cholesterol contents of cells.
