Macrophage 3-hydroxy-3-methylglutaryl coenzyme a reductase activity in sitosterolemia: effects of increased cellular cholesterol and sitosterol concentrations

Sitosterolemia is a rare, recessively inherited disease characterized clinically by accelerated atherosclerosis and xanthomas and biochemically by hyperabsorption and retention of sitosterol and other plant sterols in tissues. Decreased cholesterol biosynthesis and inhibition of 3-hydroxy-3-methylgluratyl coenzyme A (HMG-CoA) reductase and other enzymes in the biosynthetic pathway have been associated with enhanced low-density lipoprotein (LDL) receptor function. We examined the effects of cholesterol and sitosterol on sterol concentrations and composition and HMG-CoA reductase activity in monocyte-derived macrophages (MDM) from 12 control and 3 homozygous sitosterolemic subjects. The cells were cultured up to 7 days in media devoid of plant sterols, but containing increasing amounts of serum cholesterol. Before culture, MDM from the homozygous sitosterolemic subjects contained 22% more total sterols than cells from control subjects. Plant sterols and stanols represented 15.6% of MDM total sterols in sitosterolemic cells, but only 3.8% in control cells. After 7 days of culture in 10% delipidated serum (DLS) (20 [mu ]g/mL cholesterol, no sitosterol), all plant sterols were eliminated so that cells from both phenotypes contained only cholesterol. When DLS was replaced with fetal bovine serum (FBS) (300 [mu ]g/mL cholesterol), with and without addition of 200 [mu ]g/mL LDL, cholesterol levels in MDM from sitosterolemic subjects increased 108% (P [lt ] .05) compared with a 65% increase (P [lt ] .04) in control MDM cultured similarly. MDM HMG-CoA reductase activity from the 3 sitosterolemic subjects, which was significantly lower than controls at baseline (24 [plusmn] 3 v 60 [plusmn] 10 pmol/mg/min, P [lt ] .05), was not downregulated by increased cellular cholesterol levels, as observed in control cells. Control MDM were also cultured in medium that contained 10% DLS and was supplemented with 100 [mu ]g/mL cholesterol or sitosterol dissolved in ethanol or the ethanol vehicle alone. In contrast to cellular cholesterol accumulation, which significantly downregulated HMG-CoA reductase activity ([minus ]53%, P [lt ] .05), the increase in cellular sitosterol up to 25.1% of total sterols did not change MDM HMG-CoA reductase activity. Evidence of a normal HMG-CoA reductase protein in sitosterolemic cells, which was not derepressed upon removal of cellular sitosterol, and the failure of cellular sitosterol to inhibit normal HMG-CoA reductase activity argue against feedback inhibition by sitosterol as a mechanism for low reductase activity in this disease. The larger accumulation of sterols and inadequate downregulation of HMG-CoA reductase in MDM may be mechanisms for foam cell formation and explain, in part, the increased risk of atherosclerosis in sitosterolemia.