LIPID METABOLISM: DYNAMICS OF INTRACELLULAR TRANSFER

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One mechanism of atherosclerosis, at a cellular level, involves imbalances between cholesterol biosynthesis, uptake and efflux, as one important factor in this disease process. Reduction in cardiovascular risk by elevated HDL levels has been attributed to increased cholesterol efflux from peripherial tissues promoted by HDL for transport to the liver. An alternate view is that HDL may be simply a marker in persons with reduced cardiovascular risk. Current ideas about intracellular regulation of cholesterol synthesis have been formulated with little information about rate limiting processes, the cellular location of rate limiting steps and the relative rates of competing processes of cholesterol and fatty acid transport. Lipid distributions that are determined by thermodynamics have not been distinguished from those that exist as the result of kinetic barriers that prevent rapid lipid movement by passive transfer and cellular transport mechanisms. The objective is to define the kinetics and mechanisms of cellular uptake, intracellular transfer, and efflux of cholesterol and fatty acids in cultured murine 3T3L1 preadipocytes and human fibroblasts. A combination of digital fluorescence imaging and biochemical experiments are proposed to answer the following questions. 1. Is the role of fatty acid binding protein that of an intracellular buffer or a transport vesicle between membranes? 2. Which intracellular membranes have continuous lipid domains and which membranes create kinetic barriers to passive transfer of lipids? 3. Does redistribution of cholesterol between cellular compartments increase synthesis of 3-hydroxy-3-methyglutaryl-CoA reductase? 4. Is the control of transcription and translation of FABP and lipoprotein lipase influenced primarily by fatty acid flux or by hormone action?