Generation of regulatory oxysterols: 26-hydroxylation of cholesterol by ovarian mitochondria.

TitleGeneration of regulatory oxysterols: 26-hydroxylation of cholesterol by ovarian mitochondria.
Publication TypeJournal Article
Year of Publication1990
AuthorsRennert H, Fischer RT, Alvarez JG, Trzaskos JM, Strauss JF
Date Published1990 Aug
KeywordsAminoglutethimide, Animals, Cells, Cultured, Cholestanetriol 26-Monooxygenase, Cholesterol, Chorionic Gonadotropin, Female, Gas Chromatography-Mass Spectrometry, Gonadotropins, Equine, Granulosa Cells, Humans, Hydroxycholesterols, Hydroxylation, Kinetics, Mitochondria, Models, Biological, Pregnenolone, Progesterone, Rats, Steroid Hydroxylases, Sterols, Superovulation

De novo synthesis of cholesterol and low-density lipoprotein (LDL) receptor levels are suppressed in the presence of cholesterol. Recent evidence suggests that a cholesterol metabolite (possibly a hydroxysterol), not cholesterol per se, is the effector that inhibits transcription of genes encoding enzymes involved in sterol synthesis and LDL receptors. We found that 26-hydroxycholesterol inhibits human ovarian cell sterol synthesis, and that luteinized human granulosa cells contain 26-hydroxylase messenger RNA (mRNA). We proceeded to characterize the enzyme generating 26-hydroxycholesterol in the rat ovary. Mitochondria derived from ovaries of PMSG-human CG (hCG) primed immature rats (day 3 post-hCG) metabolized [3H] cholesterol into [3H]26-hydroxycholesterol in the presence of nicotinamide adenine dinucleotide phosphate and aminoglutethimide (100 micrograms/ml), added to inhibit metabolism of sterols by the cholesterol side-chain cleavage system. The identity of the product was confirmed by chromatography in several systems; recrystallization to constant specific activity and mass spectrometry. Negligible 26-hydroxylase activity was detected in other ovarian subcellular fractions. 26-Hydroxycholesterol formation progressed at a linear rate for up to 40 min and was linearly related to mitochondrial protein added to the incubation mixture. 26-Hydroxylase was markedly stimulated (5-fold) by calcium (0.2 mM). Maximal rates of 26-hydroxycholesterol formation observed were 1 pmol/ protein. This activity is substantially lower than cholesterol side-chain cleavage measured in the absence of aminoglutethimide. Ketoconazole (1-100 microM) inhibited 26-hydroxylase in a dose-dependent manner. Pregnenolone (1-1000 microM) and progesterone (1-100 microM) inhibited 26-hydroxylase in a dose-dependent manner, with appreciable inhibitory effects in the 1-10 microM range. We suggest that 26-hydroxycholesterol is an intracrine regulator that controls cellular sterol metabolism. Formation of 26-hydroxcholesterol in ovarian cells may be regulated by steroidogenic activity in such a way as to ensure availability of steroid hormone precursors. When steroidogenesis is active, 26-hydroxylase is inhibited by products of the side-chain cleavage system, allowing increased de novo sterol synthesis and LDL uptake. With reduced steroidogenic activity and less demand for cholesterol, 26-hydroxylase is not blocked, permitting formation of 26-hydroxycholesterol with attendant reduction in sterol synthesis and LDL receptor gene expression.

Alternate JournalEndocrinology
PubMed ID2373053
Grant ListHD-06274 / HD / NICHD NIH HHS / United States
HD-17301 / HD / NICHD NIH HHS / United States
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