AJP - Gastrointestinal and Liver Physiology, Vol 270, Issue 3 401-G410, Copyright © 1996 by American Physiological Society

ARTICLES

Hep G2 cells: a model for studies on regulation of human cholesterol 7alpha-hydroxylase at the molecular level

W. M. Pandak, R. T. Stravitz, V. Lucas, D. M. Heuman and J. Y. Chiang
Department of Medicine, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298-0711.

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

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