Hydroxylation of 5α-androstane-3β,17β-diol by rat prostate microsomes: Effects of antibodies and chemical inhibitors of cytochrome P450 enzymes

Gemzik, Brian; Green, Justin; Parkinson, Andrew

doi:10.1016/0003-9861(92)90585-k

Cited by 9 publications

(4 citation statements)

References 34 publications

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“…Because this is the first study on CYP7B1‐mediated 3β‐Adiol metabolism in the pig, GC/MS analysis was carried out to determine the structure of the 3β‐Adiol hydroxymetabolite formed. Previous studies report the formation of both 6‐ and 7‐hydroxymetabolites from 3β‐Adiol [17–19]. The GC/MS analysis carried out in the present study showed that the main product formed from 3β‐Adiol in pig liver is 5α‐androstane‐3β,7α,17β‐triol (for GC/MS chromatogram, see Supplementary material).…”

Section: Resultssupporting

confidence: 62%

CYP7B1‐mediated metabolism of dehydroepiandrosterone and 5α‐androstane‐3β,17β‐diol – potential role(s) for estrogen signaling

et al. 2008

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CYP7B1, a cytochrome P450 enzyme, metabolizes several steroids involved in hormonal signaling including 5alpha-androstane-3beta,17beta-diol (3beta-Adiol), an estrogen receptor agonist, and dehydroepiandrosterone, a precursor for sex hormones. Previous studies have suggested that CYP7B1-dependent metabolism involving dehydroepiandrosterone or 3beta-Adiol may play an important role for estrogen receptor beta-mediated signaling. However, conflicting data are reported regarding the influence of different CYP7B1-related steroids on estrogen receptor beta activation. In the present study, we investigated CYP7B1-mediated conversions of dehydroepiandrosterone and 3beta-Adiol in porcine microsomes and human kidney cells. As part of these studies, we compared the effects of 3beta-Adiol (a CYP7B1 substrate) and 7alpha-hydroxy-dehydroepiandrosterone (a CYP7B1 product) on estrogen receptor beta activation. The data obtained indicated that 3beta-Adiol is a more efficient activator, thus lending support to the notion that CYP7B1 catalysis may decrease estrogen receptor beta activation. Our data on metabolism indicate that the efficiencies of CYP7B1-mediated hydroxylations of dehydroepiandrosterone and 3beta-Adiol are very similar. The enzyme catalyzed both reactions at a similar rate and the K(cat)/K(m) values were in the same order of magnitude. A high dehydroepiandrosterone/3beta-Adiol ratio in the incubation mixtures, similar to the ratio of these steroids in many human tissues, strongly suppressed CYP7B1-mediated 3beta-Adiol metabolism. As the efficiencies of CYP7B1-mediated hydroxylation of dehydroepiandrosterone and 3beta-Adiol are similar, we propose that varying steroid concentrations may be the most important factor determining the rate of CYP7B1-mediated metabolism of dehydroepiandrosterone or 3beta-Adiol. Consequently, tissue-specific steroid concentrations may have a strong impact on CYP7B1-dependent catalysis and thus on the levels of different CYP7B1-related steroids that can influence estrogen receptor beta signaling.

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Section: Resultssupporting

confidence: 62%

CYP7B1‐mediated metabolism of dehydroepiandrosterone and 5α‐androstane‐3β,17β‐diol – potential role(s) for estrogen signaling

et al. 2008

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“…We think this unlikely. First, substrate specificity and stereochemistry for these conversions can vary according to reaction conditions (26,31). Second, the hydroxylation profiles of DHEA and A/anediol by brain and recombinant CYP7B enzyme are indistinguishable.…”

Section: Brain Expression; Reporter Activity Is Robustly and Selectivmentioning

confidence: 99%

Neurosteroid Hydroxylase CYP7B

Rose

Allan

Gauldie

et al. 2001

Journal of Biological Chemistry

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The major adrenal steroid dehydroepiandrosterone (DHEA) enhances memory and immune function but has no known dedicated receptor; local metabolism may govern its activity. We described a cytochrome P450 expressed in brain and other tissues, CYP7B, that catalyzes the 7␣-hydroxylation of oxysterols and 3␤-hydroxysteroids including DHEA. We report here that CYP7B mRNA and 7␣-hydroxylation activity are widespread in rat tissues. However, steroids related to DHEA are reported to be modified at positions other than 7␣, exemplified by prominent 6␣-hydroxylation of 5␣-androstane-3␤,17␤-diol (A/anediol) in some rodent tissues including brain. To determine whether CYP7B is responsible for these and other activities we disrupted the mouse Cyp7b gene by targeted insertion of an IRES-lacZ reporter cassette, placing reporter enzyme activity (␤-galactosidase) under Cyp7b promoter control. In heterozygous mouse brain, chromogenic detection of reporter activity was strikingly restricted to the dentate gyrus. Staining did not exactly reproduce the in situ hybridization expression pattern; post-transcriptional control is inferred. Lower level staining was detected in cerebellum, liver, and kidney, and which largely paralleled mRNA distribution. Liver and kidney expression was sexually dimorphic. Mice homozygous for the insertion are viable and superficially normal, but ex vivo metabolism of DHEA to 7␣-hydroxy-DHEA was abolished in brain, spleen, thymus, heart, lung, prostate, uterus, and mammary gland; lower abundance metabolites were also eliminated. 7␣-Hydroxylation of 25-hydroxycholesterol and related substrates was also abolished, as was presumed 6␣-hydroxylation of A/anediol. These different enzyme activities therefore derive from the Cyp7b gene. CYP7B is thus a major extrahepatic steroid and oxysterol hydroxylase and provides the predominant route for local metabolism of DHEA and related molecules in brain and other tissues.Brain function is subject to hormonal control, notably by steroids synthesized from the adrenal glands and gonads. Accumulating evidence also points to local steroid synthesis and metabolism in brain; a growing field of investigation focuses on the biological role of brain-active steroids, or "neurosteroids" (1-5). Attention has focused on the major adrenal steroid in primates, dehydroepiandrosterone (DHEA), 1 in view of a possible link with cognitive aging and immunosenescence. DHEA and related steroids, including pregnenolone, have memoryenhancing properties in rodents (6 -8) as well as immunostimulatory effects (9 -11). In primates, levels of DHEA and its sulfate (DHEAS) decline asymptotically with age (12-14). A causal relationship with age-related physiological impairments has been debated (15, 16). Because DHEA replacement therapy has brought mixed results (15,(17)(18)(19), and no dedicated receptor has been described for DHEA, its bioactivity may require local metabolism.B-ring hydroxylation is a major metabolic route for 3␤-hydroxysteroids, including DHEA and pregnenolone, in diverse tissues includ...

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“…However, this does not mean that cyp7b1 is the sole enzyme with 7α-hydroxylase activity in the prostate. An enzyme possessing 7α-hydroxylase activity has previously been described in both rat and human prostates [14,26,27], which appears to be specific for A\anediol and fails to metabolize DHEA. Interestingly, DHEA and A\enediol can inhibit the 7α-hydroxylation of A\anediol in the prostate [14].…”

Section: Discussionmentioning

confidence: 99%

“…In the prostate, 7α-hydroxylation of 3β-hydroxysteroids has also been described, but the enzyme responsible for the 7αhydroxylation of DHEA and related molecules has not been identified [14,26,27]. In the present study, we used a combination of RNA analysis [reverse-transcription (RT) with PCR, Northern blotting and in situ hybridization] and biochemical techniques to investigate the 7α-hydroxylase enzyme in rat prostate.…”

Section: Introductionmentioning

confidence: 99%

cyp7b1 catalyses the 7α-hydroxylation of dehydroepiandrosterone and25-hydroxycholesterol in rat prostate

Martin

Bean

Rose

et al. 2001

Biochemical Journal

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Dehydroepiandrosterone (DHEA) is the most prominent circulating steroid in humans, and it is a precursor for sex-steroid synthesis in peripheral tissues, including the prostate. Recently, enzyme-mediated pre-receptor metabolism has been recognized as a key step in determining steroid action in vivo. Hydroxylation of 3β-steroids at the 7α-position has been reported in rat and human prostate to be a major inhibitory pathway to sex-steroid synthesis/action. However, the molecular identity of the enzyme responsible is so far unknown. We recently described a novel cytochrome P450 enzyme, cyp7b1, strongly expressed in the hippocampus of rodent brain, which catalyses the metabolism of DHEA, pregnenolone and 25-hydroxycholesterol to 7α-hydroxy products. In the light of this new enzyme, we have examined its possible role in 7α-hydroxylation conversion in rat prostate. NADPH-dependent 7α-hydroxylation was confirmed for 3β-hydroxysteroids including DHEA and androstenediol, as well as 25-hydroxycholesterol. Kinetic analysis yielded an apparent Km of 14±1µM for 7α-hydroxylation of DHEA in the prostate gland, a value similar to that recorded for recombinant cyp7b1 enzyme [13.6µM; Rose, Stapleton, Dott, Kieny, Best, Schwarz, Russell, Bjoorkheim, Seckl and Lathe (1997) Proc. Natl. Acad. Sci. U.S.A. 94, 4925-4930]. The Vmax value of the prostate was 46±2pmol/min per mg, and this activity was inhibited by clotrimazole, a P450-enzyme blocker. Moreover, RNA analysis (reverse-transcription PCR, Northern blotting and in situ hybridization) revealed a high expression of cyp7b1 mRNA in the rat prostate, restricted to the epithelium, suggesting that cyp7b1 catalyses oxysterol 7α-hydroxylation in the prostate gland.

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Hydroxylation of 5α-androstane-3β,17β-diol by rat prostate microsomes: Effects of antibodies and chemical inhibitors of cytochrome P450 enzymes

Cited by 9 publications

References 34 publications

CYP7B1‐mediated metabolism of dehydroepiandrosterone and 5α‐androstane‐3β,17β‐diol – potential role(s) for estrogen signaling

CYP7B1‐mediated metabolism of dehydroepiandrosterone and 5α‐androstane‐3β,17β‐diol – potential role(s) for estrogen signaling

Neurosteroid Hydroxylase CYP7B

cyp7b1 catalyses the 7α-hydroxylation of dehydroepiandrosterone and25-hydroxycholesterol in rat prostate

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