COMPARISON OF CHOLESTEROL EGRESS FROM CULTURED-CELLS ENRICHED WITH CHOLESTEROL ESTER AFTER EXPOSURE TO CATIONIZED LDL OR TO LDL AND CHLOROQUINE

Human skin fibroblasts in culture were incubated with low density lipoprotein (LDL) and 50 μM chloroquine (Method 1) or with cationized LDL (Method 2) and a 20-50-fold increase in cholesterol ester was achieved. Incubation with cationized LDL resulted in an increase in cellular lipid droplets and in membrane bound vacuoles, containing the cationized 125I-labeled LDL as shown by radioautography. Large amounts of cationized 125I-labeled LDL were seen to adhere to cell surfaces both in cells fixed in the petri dish and in cells subjected to trypsinization and washing. Adhesion of cationized LDL to cells amounted to 30% of the material added during trypsinization as compared to less than 1% with native LDL. When the cells had been enriched with cholesterol ester by Method 1 and postincubated with high density apolipoprotein-sphingomyelin liposomes for 24-48 h, a 50-80% reduction in cellular cholesterol ester was found. Little or no reduction in cholesterol ester occurred when Method 2 was used to induce cholesterol accumulation. The slow egress of cholesterol from cells incubated with cationized LDL could be due to (a) over-loading of the lysosomal system, (b) overestimation of cellular cholesterol ester content owing to tenacious adherence of the cationized LDL to cell surfaces, and (c) reesterification of the free cholesterol liberated after intralysosomal hydrolysis, to cytoplasmic cholesterol ester. The present findings indicate that Method 2 for enrichment of cells with cholesterol ester is more complicated than the chloroquine LDL model, especially when the primary aim is quantitative determination of cholesterol egress from cells enriched in cholesterol ester. © 1979.