title: Modulation of regulatory oxysterol formation and low density lipoprotein suppression of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity by ketoconazole. A role for cytochrome P-450 in the regulation of HMG-CoA reductase in rat intestinal epithelial cells.
abstract: The effects of ketoconazole, a lanosterol demethylase and cytochrome P450 inhibitor, on the regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase (EC 1.1.1.34, reductase) activity and sterol biosynthesis were studied in rat intestinal epithelial cell cultures (IEC-6). Incubation of cells with 0.15-2 microM ketoconazole resulted in a concentration-dependent inhibition of reductase activity. As the drug concentration approached 15 microM, the reductase activity returned to control values, and at 30 microM ketoconazole, a stimulation of enzyme activity was observed. The drug had no effect on reductase activity in homogenates of IEC-6 cells. Ketoconazole (0.15-30 microM) caused a concentration-dependent inhibition of the incorporation of [3H] mevalonolactone into cholesterol with a concomitant accumulation of radioactivity in methyl sterols; e.g. lanosterol and 24,25-epoxylanosterol. Interestingly, the incorporation of radioactivity into polar sterols showed a biphasic response which was inversely proportional to the biphasic response of reductase activity. Thus, incorporation of [3H]mevalonolactone into polar sterols increased at low concentrations of ketoconazole (0.15-2 microM) and decreased to control values at high concentrations of the drug. Treatment of cells with ketoconazole (30 microM) and [3H]mevalonolactone followed by removal of the drug and radiolabel resulted in an inhibition of reductase activity and a redistribution of radioactivity from lanosterol and 24,25-epoxylanosterol to cholesterol and polar sterols. These results suggested that the inhibition of reductase activity at low concentrations of ketoconazole (less than 2 microM) was due to a formation of regulatory polar sterols generated from the methyl sterols. At high concentrations of ketoconazole (30 microM) where no suppression in reductase activity was observed, the conversion of exogenously added [3H]24(S),25-epoxylanosterol to polar sterols was prevented. Exogenously added 24,25-epoxylanosterol inhibited reductase activity in a dose-dependent fashion, and ketoconazole (30 microM) prevented the inhibition caused by low concentrations of epoxylanosterol. The drug, however, was unable to prevent the dose-dependent suppression of reductase activity by 25-hydroxylanosterol, a reduced form of 24,25-epoxylanosterol. These results indicated that 24,25-epoxylanosterol per se was not an inhibitor of reductase activity but could be metabolized to regulatory polar sterols through a cytochrome P-450 dependent reaction which was sensitive to ketoconazole. Treatment of cells with ketoconazole totally abolished the inhibition of reductase activity by low density lipoprotein (LDL).(ABSTRACT TRUNCATED AT 400 WORDS)
mesh_terms: D000818:Animals; D002784:Cholesterol / Q000031:analogs & derivatives / Q000378:metabolism; D002788:Cholesterol Esters / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004848:Epithelium / Q000201:enzymology; D006903:Hydroxymethylglutaryl CoA Reductases / Q000378:metabolism*; D007422:Intestines / Q000201:enzymology*; D007654:Ketoconazole / Q000494:pharmacology*; D008077:Lipoproteins, LDL / Q000494:pharmacology*; D008798:Mevalonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D008956:Models, Chemical; D019301:Oleic Acid; D009829:Oleic Acids / Q000378:metabolism; D051381:Rats; D002785:Sterol O-Acyltransferase / Q000037:antagonists & inhibitors; D013261:Sterols / Q000096:biosynthesis*
chemical_list: D002788:Cholesterol Esters; D008077:Lipoproteins, LDL; D009829:Oleic Acids; D013261:Sterols; D019301:Oleic Acid; C015367:mevalonolactone; C028358:24,25-epoxycholesterol; D003577:Cytochrome P-450 Enzyme System; D002784:Cholesterol; D006903:Hydroxymethylglutaryl CoA Reductases; D002785:Sterol O-Acyltransferase; D007654:Ketoconazole; D008798:Mevalonic Acid