HEP G2 CELLS - A MODEL FOR STUDIES ON REGULATION OF HUMAN CHOLESTEROL7-ALPHA-HYDROXYLASE AT THE MOLECULAR-LEVEL

The present study examines the feedback control governing human choles terol 7 alpha-hydroxylase mRNA expression in the human hepatoblastoma cell line, Hep G2. Glycochenodeoxycholate (GCDC) and glycodeoxycholate , hydrophobic bile salts, decreased cholesterol 7 alpha-hydroxylase mR NA levels and bile acid synthesis in a concentration-dependent (76 +/- 8%, P < 0.001, and 48 +/- 3%, P < 0.01, respectively) and time-depend ent manner. Cholesterol 7 alpha-hydroxylase mRNA levels were repressed with a half-maximal inhibitory concentration of < 12.5 mu M by GCDC a nd a half-life of 30 min by 100 mu M of the bile acid. The addition of actinomycin D (10 mu g/ml) alone or in combination with GCDC (100 mu M) led to similar concentration- and time-dependent suppression of cho lesterol 7 alpha-hydroxylase mRNA. Glycocholate (100 mu M), not intern alized based on lack of uptake of a fluorescent cholate analogue, had no effect on cholesterol 7 alpha-hydroxylase mRNA or total bile acid s ynthesis. In cultures transfected with a rat cholesterol 7 alpha-hydro xylase promoter construct, reporter gene activity was decreased (31%, P < 0.01) by GCDC (100 alpha M). Hep G2 cells maintain the intracellul ar machinery to express and rapidly regulate human cholesterol 7 alpha -hydroxylase by hydrophobic bile acids. These data suggest that Hep G2 cells will support functional studies of the human cholesterol 7 alph a-hydroxylase gene.