Role of an N-ethylmaleimide-sensitive factor in the selective cellular uptake of low-density lipoprotein free cholesterol

Low-density lipoprotein (LDL) was the major contributor to an influx of free sterol from plasma which balances high-density lipoprotein (HDL)-mediated efflux from cultured skin fibroblasts. When HDL was absent, the uptake of LDL free cholesterol was associated with an increase in total cell cholesterol, due in part to accumulation of esterified cholesterol. This influx was mediated by a high-capacity, low-affinity pathway whose magnitude was similar in normal and LDL receptor-deficient cells. In the presence of HDL, some of the interiorized labeled LDL free cholesterol became available for HDL-mediated efflux and some was interiorized, as a result of a transport mechanism which was sensitive to N-ethylmaleimide (NEM) and nitrate ion but resistant to progesterone, azide, or vanadate. We suggest that normal free cholesterol homeostasis in these cells includes the initial binding of LDL followed by the selective transfer of free cholesterol to a compartment from which it is either returned to the membrane for efflux or internalized for storage or further metabolism within the cell. In the presence of NEM, LDL-derived free cholesterol remained mostly accessible for efflux from the cell surface. This free cholesterol pathway may function physiologically to stabilize plasma membrane cholesterol levels against the effect of varying concentrations of HDL and LDL.