Synthesis and secretion of apoE in thioglycolate-elicited mouse peritoneal macrophages: Effect of cholesterol efflux

ApoE synthesis and secretion, as a function of cellular cholesterol content and cholesterol efflux, was studied in thioglycolate-elicited mouse peritoneal macrophages. As expected, loading elicited macrophages with cholesterol induced a 5-fold increase in apoE secretion and a 2.5-fold increase in cellular apoE content over a 5-h period. Treatment of cholesterol-loaded cells with HDL₃ further increased apoE secretion 1.7-fold and decreased cellular cholesterol by 20%. Treatment of cholesterol-loaded cells with HDL₃ and SAH 58.035 (an ACAT inhibitor) increased apoE secretion 2.4-fold and decreased cellular cholesterol content by 35%. Treatment of the cells with the ACAT inhibitor alone suppressed apoE secretion by 40% but did not change cellular cholesterol content. Northern blot analysis of RNA indicated that cholesterol loading increased apoE mRNA 2-fold. ApoE mRNA levels were not further affected by treatment with HDL₃ and/or the ACAT inhibitor. Cholesterol-loaded cells, in the absence of HDL₃, secreted apoE into the media in two fractions as determined by column chromatography: a large molecular weight complex, (larger than HDL), and an essentially lipid-free protein. In the presence of HDL₃, the cells secreted apoE in three fractions: a large molecular weight complex, an essentially lipid-free protein, and over 50% of apoE associated with HDL. In the process, HDL₃ became larger and eluted in a position identical to that of HDL₂. A small amount of HDL₃-derived material was also transformed to an LDL-size particle. Incubation of HDL₃ in the absence of cholesterol-loaded cells did not produce these changes. It is concluded that cholesterol-loading increases apoE mRNA content and apoE synthesis. ApoE synthesis and secretion, but not apoE mRNA levels are further increased by HDL-mediated cholesterol efflux.