title: Effects of cyclosporin on cholesterol 27-hydroxylation and LDL receptor activity in HepG2 cells.
abstract: The hypothesis that mitochondrial sterol 27-hydroxylase plays a role in the sterol-mediated down-regulation of LDL receptor activity was evaluated in HepG2 cells. 27-Hydroxycholesterol was found to be more potent at suppressing LDL receptor activity than cholesterol (IC50 values of 8 mu M and 142 mu M for 27-hydroxycholesterol and cholesterol, respectively) when the sterols were delivered to cells from 2-hydroxypropyl-beta-cyclodextrin (beta-CD)-solubilized solutions. Cyclosporin, an immunosuppressant which has been shown to inhibit the 27-hydroxylation of sterols, was used to assess whether the formation of endogenous 27-hydroxycholesterol was required for the cholesterol-induced suppression of LDL receptor activity. Cyclosporin dose-dependently inhibited the 27-hydroxylation of cholesterol by HepG2 mitochondria (Ki = 0.25 mu M) and HepG2 cell cultures (IC50 = 1 mu M). At 1 mu M, cyclosporin had no effect on LDL receptor activity, and did not prevent the suppression of LDL receptor activity caused by: 1) the addition of beta-CD-solubilized cholesterol, 2) the receptor-mediated uptake of beta-VLDL, or 3) the inhibition of cholesterol esterification. In contrast, 10 mu M cyclosporin was found to inhibit the esterification of cholesterol and to increase the cellular level of free cholesterol resulting in suppression of LDL receptor activity. These results suggest that if mitochondrial sterol 27-hydroxylase plays a role in the regulation of LDL receptor activity, it is not through the formation of potent regulatory oxysterols, but through its effects on the availability and/or size of the free cholesterol pool regulating LDL receptor activity.
authors.affiliation: Division of Pharmacokinetics and Drug Metabolism, Glaxo Wellcome Inc., Research Triangle Park, NC 27709, USA.
authors.forename: D A
authors.initials: DA
authors.lastname: Winegar
authors.forename: J A
authors.initials: JA
authors.lastname: Salisbury
authors.forename: S S
authors.initials: SS
authors.lastname: Sundseth
authors.forename: R L
authors.initials: RL
authors.lastname: Hawke
mesh_terms: D001692:Biological Transport / Q000187:drug effects; D053493:Cholestanetriol 26-Monooxygenase; D016572:Cyclosporine / Q000494:pharmacology*; D065607:Cytochrome P-450 Enzyme Inhibitors; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D008077:Lipoproteins, LDL / Q000378:metabolism*; D008099:Liver / Q000378:metabolism*; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D011973:Receptors, LDL / Q000187:drug effects*; D012995:Solubility; D013250:Steroid Hydroxylases / Q000037:antagonists & inhibitors*; D014407:Tumor Cells, Cultured
chemical_list: D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D008077:Lipoproteins, LDL; D011973:Receptors, LDL; D016572:Cyclosporine; D013250:Steroid Hydroxylases; C506831:CYP27A1 protein, human; D053493:Cholestanetriol 26-Monooxygenase