出版社：American Society for Biochemistry and Molecular Biology
摘要：Exchange and net mass efflux of cholesterol were investigated in [3H]cholesterol-labeled or cholesteryl ester-loaded murine peritoneal macrophages, respectively. Macrophages were subjected to mild proteolysis prior to measurements of mass efflux or exchange to assess whether plasma membrane proteins participated in either process. Cholesterol exchange and mass efflux were inhibited up to 70% following trypsinization. The inhibitory effect was reversible as cells regained normal efflux and exchange 6-8 hr following treatment. Incubation of trypsinized cells with cycloheximide prevented recovery, indicating that protein synthesis was necessary for restoration of normal cholesterol efflux. Studies with peptide and nonpeptide inhibitors of proteolysis suggested that active catalytic activity of trypsin was necessary for the inhibitory effect to be expressed. The degree of inhibition for both cholesterol exchange and mass efflux was dependent in a quantitatively similar manner on the time of incubation and the concentration of trypsin, suggesting that the mechanism of cholesterol exchange and mass efflux were similar at the level of the plasma membrane. Two other serine-proteases, thrombin and elastase, were also capable of inhibiting cholesterol removal in a similar manner. No cell death was observed by altered morphology, detachment, changes in DNA or protein content, or trypan blue exclusion even under the most severe proteolytic conditions. These studies suggest that protease-sensitive plasma membrane proteins play a role in cholesterol efflux in macrophages.
