Regulation of sterol synthesis in L cells: Steady-state and transitional responses

The patterns of flux, content, and synthesis of sterol were studied in L cells cultured in media containing various concentrations of phospholipid and sterol, finding that cellular desmosterol and the rate of acetate incorporation into sterol varied inversely with exogenous cholesterol.

Abstract

The patterns of flux, content, and synthesis of sterol were studied in L cells cultured in media containing various concentrations of phospholipid and sterol. L cells do not synthesize cholesterol and, when grown in sterol-free medium, they contained 7.5 μg of desmosterol per mg cellular protein. When the medium was supplemented with cholesterol (40 μg/ml) and lecithin (20 μg/ml), the cells contained 19 μg cholesterol per mg cellular protein, almost no desmosterol, and the rates of incorporation of [14C]acetate and 3H2O into digitonin-precipitable material were only 20% of the rates of cells grown in sterol-free medium. When cells were grown with various concentrations of exogenous cholesterol (0–40 μg/ml) the cellular cholesterol varied directly with concentration of exogenous cholesterol, whereas, cellular desmosterol and the rate of acetate incorporation into sterol varied inversely with exogenous cholesterol. When increasing concentrations of exogenous lecithin were added to the cholesterol (40 μg/ml) medium, the L cells contained less cholesterol, more desmosterol, and showed increasing rates of incorporation of acetate into sterol. Upon the addition of cholesterol to cells initially growing in sterol-free medium, the following transitions occurred: An immediate influx of cholesterol, a gradual decrease in desmosterol, an exponential decay of acetate incorporation into sterol (half-time 7 h), and a doubling of total cellular sterol by 20 h. Upon the removal of cholesterol from the culture medium the cellular cholesterol decreased (33% decrease by 10 h, 57% decrease by 20 h), while the rate of acetate incorporation increased (150% by 5 h, and 450% by 20 h). After 5 h, synthesized desmosterol compensated for the reduction in cellular cholesterol; total sterol remained at 85 % of initial value from 5 to 25 h.