Lipid storage and secretion. Studies on VLDL assembly and foam cell formation

Abstract: This thesis deals with formation of two factors of upmost importance for the development of atherosclerosis: the triglyceride rich very low density lipoproteins 1 (VLDL1) and the lipid loaded macrophages ("foam cells"- characteristic features of the atherosclerotic lesion). Both these processes depend on the cellular accumulation of lipids. There exist two forms of VLDL; VLDL1 and VLDL2. VLDL2 is the precursor to VLDL1. The formation of VLDL1, but not of VLDL2, was dependent on the ADP-ribosylation factor 1 (ARF1). ARF1 is required for the correct intracellular transfer of secretory proteins, and the loss of VLDL1 assembly coincided with loss of anterograde transport from the endoplasmic reticulum (ER) to the Golgi apparatus. Moreover the VLDL1 assembly was completely abolished at 15º C, which blocks the anterograde transport between the ER-Golgi intermediate compartment and cis-Golgi. We therefore propose that the conversion of VLDL2 to VLDL1 occurs in a compartment distal to ER, most likely the Golgi apparatus. The assembly of VLDL1 is highly dependent on the levels of triglycerides in the liver. We therefore investigate the influence of ADRP (adipocyte differentiation-related protein; a major protein on cytosolic lipid droplets) on the assembly of VLDL1. When the cellular levels of ADRP were increased, the pool of cytosolic lipid droplets (and triglycerides) increased but the formation of VLDL1 decreased. ADRP promoted the incorporation of fatty acids into cytosolic triglycerides and their storage in lipid droplets, thereby diverting them from the VLDL assembly pathway. When the levels of ADRP decreased (knockdown by siRNA), the pool of lipid droplets (and triglycerides) decreased; but the effect on VLDL1 was small. Instead there was a more than two fold increase in the oxidation of fatty acids. These results demonstrate a complex relation between storage, oxidation and secretion of fatty acids, in which the levels of ADRP have an important role. ADRP also has an important role in the accumulation of lipids in macrophages, and triglycerides accounts for a substantial portion of the lipids in macrophages from the atherosclerotic lesions. Since the oxygen tension in the lesions is low, we investigated if such a hypoxia could influence macrophages to store triglycerides. When isolated human macrophages were cultured in the presence of 1 % O2, the pool of cytosolic lipid droplets (and triglycerides) increased. This increase could be explained by an increased expression of ADRP. Moreover there was a down regulation of enzymes in the beta-oxidation and a subsequent decrease in this oxidation, providing more fatty acids for lipid biosynthesis. Moreover, the triglyceride biosynthesis increased. Together our results indicate that the hypoxia could contribute the formation of foam cells in the atherosclerotic lesion and to the accumulation of triglycerides in such lesions.