Metabolism of androgens in the seminal vesicles and the different lobes of the prostate in young mature rats

Oxidation and reduction of 4-androstene-3,17-dione (androstenedione), 17 beta-hydroxy-4-androsten-3-one (testosterone), 17 beta-hydroxy-5 alpha-androstan-3-one (DHT), 5 alpha-androstan-3 alpha,17 beta-diol (3 alpha-A'diol) and 5 alpha-androstane-3 beta,17 beta-diol (3 beta-A'diol) were measured in homogenates from ventral (VP), dorsal (DP) and lateral prostate (LP), the coagulating gland (CG) and seminal vesicle (SV) of the intact sexually mature rat using NAD(H) or NADP(H) as cofactors. The specific activity of the various enzymes varied significantly between the different organs. 5 alpha-Reductase activity was highest in the DP and the CG, and undetectable in the LP. 17 beta-Hydroxysteroid oxidoreductase (17 beta-HSOR) activity was mainly confined to the LP. 3 alpha-Hydroxysteroid oxidoreductase (3 alpha-HSOR) activity was also highest in the LP. In the VP the highest 3 alpha-HSOR activity was recorded using NAD(H) as cofactor. In the other organs, similar or higher enzymatic activities were measured using NADP(H) as added cofactor. 3 beta-Hydroxysteroid oxidoreductase (3 beta-HSOR) activity was high in the LP and low or undetectable in the other tissues. Our results indicate that isoenzymes of 3 alpha-HSOR, 3 beta-HSOR and 17 beta-HSOR are present in the accessory sex organs of the rat.     

DHT formation and degradation in cultured human skin fibroblasts: DHT accumulation in the genital skin

The conversion of testosterone to dihydrotestosterone (DHT) by 5 alpha-reductase and the interconversion between DHT and 5 alpha-androstane-3 alpha, 17 beta-diol (3 alpha-diol) by 3 alpha-hydroxy-steroid oxidoreductase (3 alpha-HSOR) were studied in fibroblasts derived from the genital skin of 22 males and 6 females, and from the nongenital skin of 19 males and 9 females with normal gonadal function. The formation of DHT from testosterone (5 alpha-reduction) was significantly greater in fibroblasts from genital skin than in those from nongenital skin in both males (2.15 +/- 1.43 vs 0.81 +/- 0.46 pmol/mg protein/h, mean +/- SD, P less than 0.001) and females (2.52 +/- 1.99 vs 0.69 +/- 0.18, P less than 0.01). Furthermore, DHT formation from 3 alpha-diol (3 alpha-HSOR oxidation) was also significantly greater in genital skin fibroblasts than in nongenital skin fibroblasts of males (5.47 +/- 3.37 vs 2.52 +/- 1.74 pmol/mg protein/h, P less than 0.01). However, the degradation of DHT to 3 alpha- and/or 3 beta-diol (3 alpha- and/or 3 beta-HSOR reductions) was not different between genital and nongenital skin fibroblasts of either males or females. Respective ratios of DHT formation to DHT degradation (5 alpha-reduction/3 alpha-HSOR reduction, 3 alpha-HSOR oxidation/3 alpha-HSOR reduction) were also significantly greater (P less than 0.002) in genital skin fibroblasts than in nongenital skin fibroblasts of males. On the other hand, both DHT formation and degradation were not different between male and female genital skin fibroblasts. These results suggest that the increased production of DHT in genital compared to nongenital skin results from increased 5 alpha-reduction and 3 alpha-HSOR oxidation.     

The identification and characterization of a new C19O3 steroid metabolite in the rat ventral prostate: 5 alpha-androstane-3 beta, 6 alpha, 17 beta-triol.

This study represents the first report of the formation of 5 alpha-androstane-3 beta, 6 alpha, 17 beta-triol (6 alpha-triol) by prostatic tissue. The 6 alpha-triol has been identified by rigorous methods and a chemical synthesis of this triol has been accomplished. This 6 alpha-triol is the major metabolite of 5 alpha-androstane-3 beta, 17 beta-diol (3 beta-diol) in the rat ventral prostate. A minor metabolite of 3 beta-diol has been identified as 5 alpha-androstane-3 beta, 7 alpha, 17 beta-triol (7 alpha-triol). Using a variety of C19 androstane substrates, the 6 alpha- and 7 alpha-triols were always found as the major components of the total 3 beta-hydroxy-5 alpha-androstane metabolites produced by the ventral prostate. Following intraperitoneal injection of 3H-3 beta-diol, both 6 alpha- and 7 alpha-triol were formed in vivo by the ventral prostate and found in the blood. The 6 alpha- and 7 alpha-triols were found to possess no androgenic activity when tested by the ventral prostatic growth bioassay in the castrate rat.     

Age dependent changes in androgen metabolism in the rat prostate

Oxidation and reduction of androstenedione, testosterone, dihydrotestosterone (DHT), 5 alpha-androstan-3 alpha,17 beta-diol and 5 alpha-androstane-3 beta,17 beta-diol (3 alpha- and 3 beta-A'diol) were measured in homogenates from the ventral prostate (VP), dorsal prostate (DP), lateral prostate (LP), the coagulating gland (CG) and seminal vesicles (SV) in intact rats of different ages from young mature (3-6 months) to senescent rats (20-30 months). Some very old intact rats (30-32 months) were treated with testosterone in order to rule out the effect of this hormone on androgen metabolism. The enzymatic activities for young mature rats were significantly altered by increasing age, both with regard to differences between the various organs as well as differences in cofactor requirement. With increasing age, the specific activity of most enzymes gradually decreased. With testosterone as substrate, 5 alpha-reductase activity was significantly reduced in the old rats in all tissues studied and was undetectable in the oldest animals in the VP and the SV. On the other hand, 5 alpha-reductase could not be recorded in any tissue in any tissue in old rats when androstenedione was the substrate. 3 alpha-Hydroxysteroid oxidoreductase (3 alpha-HSOR) in the VP was the only enzyme which did not decrease in activity by increasing age. In the other lobes this enzyme activity decreased similar to 3 beta-hydroxysteroid oxidoreductase (3 beta-HSOR) and the 17 beta-hydroxysteroid oxidoreductase (17 beta-HSOR) activity. Administration of testosterone to old rats increased the specific activity of most of the enzymes studied.     

Prostatic cytosol binding of 5 alpha-androstane-3 beta, 17 beta-diol and estradiol-17 beta

The goal of the present research was characterization of the interaction of 5 alpha-androstane-3 beta, 17 beta-diol (3 beta-diol) with prostatic estradiol-17 beta(E2) binding sites to address the role of this 5 alpha-dihydrotestosterone(DHT)a metabolite in prostatic regulation. Using dextran-charcoal assay we demonstrated specific 3 beta-diol and E2 binding sites in rat ventral prostate cytosol (RVPC) and dog prostate cytosol (DPC). In both cytosols, E2 binding is of high affinity (Ka congruent to 10(9) M-1; RVPC:68 fmol/mg protein), DPC:170 fmol/mg protein), and 3 beta-diol binding is of moderate affinity (Ka congruent to 10(8) M-1; RVPC:62 fmol/mg protein, DPC:165 fmol/mg protein). Unlabeled 3 beta-diol competes effectively for cytosolic 3H-E2 binding sites, whereas unlabeled DHT, 5 alpha-androstane-3 alpha, 17 beta-diol (3 alpha-diol) and testosterone (T) are poor competitors for 3H-E2 binding sites. Using DNA-cellulose column chromatography, we separated prostatic androgen and estrogen binding activities. The E2 binding activity which adhered to DNA-cellulose was displaced by 100-fold excess 3 beta-diol but not by DHT. Thus data from two assay procedures show competition of 3 beta-diol for 3H-E2 binding sites in rat and dog prostate.     

Hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol by rat prostate microsomes: effects of antibodies and chemical inhibitors of cytochrome P450 enzymes.

The purpose of the present study was to test the hypothesis that rat prostate microsomes contain a single cytochrome P450 enzyme responsible for the conversion of 5 alpha-androstane-3 beta,17 beta-diol to a series of trihydroxylated products. The three major metabolites formed by in vitro incubation of 5 alpha-[3H]androstane-3 beta,17 beta-diol with rat prostate microsomes were apparently 5 alpha-androstane-3 beta,6 alpha,17 beta-triol, 5 alpha-androstane-3 beta,7 alpha,17 beta-triol, and 5 alpha-androstane-3 beta,7 beta,17 beta-triol, which were resolved and quantified by reverse-phase HPLC with a flow through radioactivity detector. The ratio of the three metabolites remained constant as a function of incubation time, microsomal protein concentration, ionic strength, and substrate concentration. The ratio of the three metabolites was dependent on pH, apparently because the hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol shifted from the 6 alpha- to the 7 alpha-position with increasing pH (6.8-8.0). The V(max) values were 380, 160, and 60 pmol/mg microsomal protein/min for the rate of 6 alpha-, 7 alpha-, and 7 beta-hydroxylation, respectively. Similar Km values (0.5-0.7 microM) were measured for enzymatic formation of all three metabolites, which suggests that formation of all three metabolites was catalyzed by a single, high-affinity enzyme. Testosterone, 5 alpha-dihydrotestosterone, and 5 alpha-androstane-3 alpha,17 beta-diol did not appreciably inhibit the hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol, suggesting that this enzyme exhibits a high degree of substrate specificity. Formation of all three metabolites was inhibited by antibody against rat liver NADPH-cytochrome P450 reductase (85%) and by a 9:1 mixture of carbon monoxide and oxygen (60%). Several chemical inhibitors of cytochrome P450 enzymes, especially the antimycotic drug clotrimazole, also inhibited the formation of all three metabolites. Polyclonal antibodies that recognize liver cytochrome P450 1A, 2A, 2B, 2C, and 3A enzymes did not inhibit 5 alpha-androstane-3 beta,17 beta-diol hydroxylase activity. Overall, these results are consistent with the hypothesis that the 6 alpha-, 7 alpha-, and 7 beta-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol by rat prostate microsomes is catalyzed by a single, high-affinity P450 enzyme. This cytochrome P450 enzyme appears to be structurally distinct from those in the 1A, 2A, 2B, 2C, and 3A gene families.     

Age-related changes in conversion of 5 alpha-androstan-17 beta-ol-3-one to 5 alpha-androstane-3 alpha,17 beta-diol and 5 alpha-androstane-3 beta,17 beta-diol by rat testicular cells in vitro

Conversion of labelled 5 alpha-androstane-17 beta-ol-3-one (DHT) by isolated testicular cells from rats of different ages was examined under saturating substrate conditions in vitro (5--10 micrograms DHT/ml in a 24 h incubation). Two detectable metabolites of DHT were produced by testicular cells in vitro. 5 alpha-androstane-3 alpha, 17 beta-diol (3 alpha-diol) and 5 alpha-androstane-3 beta, 17 beta-diol (3 beta-diol). Production of these diols during a 24 h period was linear, and the amounts formed were directly related to the cell number. The amount of 3 alpha- and 3 beta-diols formed by testicular cells of rats of different ages increased from Day 10 to Day 25, then declined. Testicular cells from rats 10 to 20 days of age converted DHT mainly to 3 alpha-diol, but thereafter 3 beta-diol was the predominant testicular metabolite of DHT.     

Isolation and catalytic activity of cytochrome P-450 from ventral prostate of control rats

5 alpha-Androstane-3 beta, 17 beta-diol hydroxylase (3 beta-diol hydroxylase), a form of cytochrome P-450, was purified from rat ventral prostate, and its regulation as a function of age and 5 alpha-dihydrotestosterone (DHT) treatment was examined. Cytochrome P-450 could be quantitated by its CO difference spectrum only after partial purification from the microsomal membrane, and this was achieved by chromatography on p-chloroamphetamine-coupled Sepharose. Further purification of prostate microsomal P-450 by anion exchange chromatography yielded a preparation with a P-450 content of 8-10 nmol/mg of protein, which upon sodium dodecyl sulfate electrophoresis showed, in the molecular weight region between 50,000 and 60,000 where P-450 is expected to migrate, a single protein band of Mr 54,000. This preparation upon reconstitution with cytochrome P-450 reductase and microsomal lipid catalyzed the formation of three triols, 5 alpha-androstane-3 beta, 7 beta, 17 beta-triol, 5 alpha-androstane-3 beta, 6 alpha, 17 beta-triol, and 5 alpha-androstane-3 beta, 7 alpha, 17 beta-triol from 3 beta-diol in the ratio 1:7:3. Both turnover number and the ratio of the three products in the reconstituted system were similar to that found in prostate microsomes. These data indicate that a single form of P-450 catalyzes the formation of all three triols and that 3 beta-diol hydroxylase is the major, if not the only, form of P-450 in the prostate microsomes of untreated rats. The yield of P-450 from prostate microsomes varied as a function of age from a high level of 0.05 nmol/mg of microsomal protein in 6-week-old rats to 0.002 nmol/mg of microsomal protein in rats 11 weeks or older. 3 beta-Diol hydroxylase activity followed a similar age-related pattern varying between 2,000 and 4,000 nmol of triols formed/g of tissue/h in 6-week-old rats to 100 nmol of triols formed/g of tissue/h in 11-week-old rats. Treatment of 6-week-old rats with DHT did not prevent the age-related decrease in 3 beta-diol hydroxylase activity. However, DHT does play a role in the regulation of this enzyme since castration resulted in a loss of catalytic activity from the prostate and treatment of castrated rats with DHT caused an induction of the enzyme.     

Pituitary metabolism of 5alpha-androstane-3beta-17beta-diol: intense and rapid conversion into 5alpha-androstane-3beta,6alpha,17beta-triol and 5alpha-androstane-3beta,7alpha, 17beta-triol.

In the male rat pituitary, 5alpha-androstane-3beta, 17beta-diol (3beta-diol) is extensively metabolized into polar steroids. They were identified as 5alpha-androstane-3beta, 6alpha-17beta-triol (6alpha-triol) and 5alpha-androstane-3beta, 7alpha, 17beta-triol (7alpha-triol). 6-alpha-Triol represents 53% and 7alpha-Triol 28% of the total 3beta-diol metabolites. The remaining percentage is related to 6beta and 7beta isomers. The biological role of triols is still unknown.     

Possible indices for the detection of the administration of dihydrotestosterone to athletes.

Dihydrotestosterone (DHT) can be used by an athlete as an anabolic steroid to evade the current International Olympic Committee approved drug tests. To investigate the possibility of a method for its detection, the heptanoate ester of DHT was administered to two male subjects (150 mg i.m.). Urine samples, collected before and after the injection, were subjected to enzymatic hydrolysis and the excretion rates of DHT, 5 alpha-androstane-3 alpha,17 beta-diol (3 alpha-diol) and testosterone (T) were determined by radioimmunoassay. Relative changes in the excretion of DHT, 3 alpha-diol, 5 alpha-androstane-3 beta,17 beta-diol (3 beta-diol), 5 beta-androstane-3 alpha, 17 beta-diol (5 beta-diol), T and epitestosterone (17 alpha hydroxyandrost-4-en-3-one; Epi-T) were determined by gas chromatography-mass spectrometry (GC-MS). Following administration of DHT, the urinary excretion rates of DHT, 3 alpha-diol and 3 beta-diol increased when compared to those of T, Epi-T, 5 beta-diol and luteinizing hormone (LH). Concentrations of DHT in the plasma increased whereas those of T, LH and follicle stimulating hormone decreased. The changes following such modest doses of DHT suggest that these ratios of urinary hormones may be used for the detection of doping with DHT.     

Androgen metabolism and actions in rat ventral prostate epithelial and stromal cell cultures

The rat ventral prostate requires androgens for normal development, growth, and function. To investigate the relationship between androgen metabolism and its effects in the prostate and to examine differences between the epithelial and stromal cells, we have established a system of primary cell cultures of immature rat ventral prostate cells. Cultures of both cell types after reaching confluency (6-7 days) actively metabolized 3H-labelled testosterone (T), 5 alpha-dihydrotestosterone (5 alpha-DHT), 5 alpha-androstane-3 alpha,17 beta-diol, and 5 alpha-androstane-3 beta,17 beta-diol. The epithelial cells actively reduced T to 5 alpha-DHT and formed significant amounts of 5 alpha-androstane-3,17-dione from T, 5 alpha-DHT, and 5 alpha-androstane-3 alpha,17 beta-diol. All substrates were converted to significant amounts of C19O3 metabolites. The stromal cells also metabolized all substrates, but very little 5 alpha-androstane-3,17-dione was formed. The metabolism studies indicate that both cell types have delta 4-5 alpha-reductase, 3 alpha- and 3 beta-hydroxysteroid oxidoreductase and hydroxylase activities. The epithelial cells have significant 17 beta-hydroxysteroid oxidoreductase activity. The epithelial cells cultures grown in the presence of T have higher acid phosphatase (AP) contents (demonstrated histochemically and by biochemical assay). Tartrate inhibition studies indicate that the epithelial cells grown in the presence of T are making secretory AP. Stromal cell AP is not influenced by T. The results indicate that the cultured cells maintain differentiated prostatic functions: ability to metabolize androgens and, in the case of the epithelial cells, synthesize secretory AP.     

Uptake and metabolism of androgens by bovine spermatozoa

Spermatozoa from bovine ejaculates and cauda epiditymidis were incubated with either tritiated 17 beta-hydroxy-5 alpha-androstane-3-one (DHT) or 5 alpha-androstane-3 alpha, 17 beta-diol (3 alpha-diol). Examination of the medium incubations demonstrated metabolic conversion of both DHT and 3 alpha-diol when these steriods were incubated with ejaculated sperm. In addition to this interconversion, the following metabolities were identified: 5 alpha-androstane-3 beta, 17 beta-diol, (3 beta-diol), androsterone and 5 alpha-androstane-3, 17-dione (5 alpha-A-dione). Incubations with cauda spermatozoa showed similar metabolic patterns. Androgen binding was exhibited by both sperm types. Examination of the washed cauda sperm pellet, following incubations with 3 alpha-diol showed that the incubated steroid was the most abundantly bound. DHT and 5 alpha-androst-16-en-3 alpha-ol (delta 16-3 alpha-ol1 were also detected. The major part of the radioactivity bound in the sperm pellet was identified as DHT when this steroid was used as the substrate; the remaining radioactivity consisted of 3 alpha-diol and delta 16-3 alpha-ol. Investigations of ejaculated sperm pellets gave similar results apart from the additional identification of 5 alpha-androst-16-en-3 one (delta 16-3-one) and 5 alpha-androst-16-en-3 beta-ol (delta 16-3 beta-ol (delta 16-3 beta-ol).     

Hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol by rat prostate microsomes: potent inhibition by imidazole-type antimycotic drugs and lack of inhibition by steroid 5 alpha-reductase inhibitors.

5 alpha-Dihydrotestosterone, the principal androgen mediating prostate growth and function in the rat, is formed from testosterone by steroid 5 alpha-reductase. The inactivation of 5 alpha-dihydrotestosterone involves reversible reduction to 5 alpha-androstane-3 beta,17 beta-diol by 3 beta-hydroxysteroid oxidoreductase followed by 6 alpha-, 7 alpha-, or 7 beta-hydroxylation. 5 alpha-Androstane-3 beta,17 beta-diol hydroxylation represents the ultimate inactivation step of dihydrotestosterone in rat prostate and is apparently catalyzed by a single, high-affinity (Km approximately 0.5 microM) microsomal cytochrome P450 enzyme. The present studies were designed to determine if 5 alpha-androstane-3 beta,17 beta-diol hydroxylation by rat prostate microsomes is inhibited by agents that are known inhibitors of androgen-metabolizing enzymes. Inhibitors of steroid 5 alpha-reductase (4-azasteroid analogs; 10 microM) or inhibitors of 3 beta-hydroxysteroid oxidoreductase (trilostane, azastene, and cyanoketone; 10 microM) had no appreciable effect on the 6 alpha-, 7 alpha-, or 7 beta-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol (10 microM) by rat prostate microsomes. Imidazole-type antimycotic drugs (ketoconazole, clotrimazole, and miconazole; 0.1-10 microM) all markedly inhibited 5 alpha-androstane-3 beta,17 beta-diol hydroxylation in a concentration-dependent manner, whereas triazole-type antimycotic drugs (fluconazole and itraconazole; 0.1-10 microM) had no inhibitory effect. The rank order of inhibitory potency of the imidazole-type antimycotic drugs was miconazole greater than clotrimazole greater than ketoconazole. In the case of clotrimazole, the inhibition was shown to be competitive in nature, with a Ki of 0.03 microM. The imidazole-type antimycotic drugs inhibited all three pathways of 5 alpha-androstane-3 beta,17 beta-diol hydroxylation to the same extent, which provides further evidence that, in rat prostate microsomes, a single cytochrome P450 enzyme catalyzes the 6 alpha-, 7 alpha-, and 7 beta-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol. These studies demonstrate that certain imidazole-type compounds are potent, competitive inhibitors of 5 alpha-androstane-3 beta,17 beta-diol hydroxylation by rat prostate microsomes, which is consistent with the effect of these antimycotic drugs on cytochrome P450 enzymes involved in the metabolism of other androgens and steroids.     

Metabolism of 17 alpha-methyltestosterone in the rabbit: C-6 and C-16 hydroxylated metabolites

17 alpha-Methyltestosterone and the reduced metabolites, 17 alpha-methyl-5 alpha-androstane-3 alpha, 17 beta-diol, 17 alpha-methyl-5 alpha-androstane-3 beta, 17 beta-diol and 17 alpha-methyl-5 beta-androstane-3 alpha, 17 beta-diol, together with three hydroxylated metabolites, 17 alpha-methyl-5 beta-androstane-3 alpha, 16 alpha, 17 beta-triol, 17 alpha-methyl-5 beta-androstane-3 alpha, 16 beta, 17 beta-triol and a new metabolite, 17 alpha-methyl-5 alpha-androstane-3 beta, 6 alpha, 17 beta-triol, were isolated and identified in the urine of rabbits dosed with 17 alpha-methyltestosterone. No hydroxylated 5 alpha-metabolite of 17 alpha-methyltestosterone has been identified previously. No of 17 alpha-methyltestosterone has been identified previously. No evidence for epimerization at the C-17 position was observed.     

Species differences in 5 alpha-androstane-3 beta,17 beta-diol hydroxylation by rat, monkey, and human prostate microsomes.

The 6 alpha-, 7 alpha-, and 7 beta-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol by rat prostate microsomes appears to be catalyzed by a single, high-affinity cytochrome P450 enzyme. In the present study we have examined the hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol by prostate microsomes from cynomolgus monkeys and from normal subjects and patients with benign prostatic hyperplasia. Our results suggest that although rat, monkey, and human prostate microsomes catalyze the 6 alpha-, 7 alpha-, and 7 beta-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol, these pathways of oxidation in monkeys and humans are not catalyzed by a single cytochrome P450 enzyme. The ratio of the three metabolites was not uniform among prostate microsomal samples from individual humans or monkeys. The 6 alpha-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol varied independently of both the 7 alpha- and 7 beta-hydroxylation, which varied in unison. The 6 alpha-, 7 alpha-, and 7 beta-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol by monkey prostate microsomes appeared to be differentially affected by in vivo treatment of monkeys with beta-naphthoflavone or dexamethasone. Treatment of a monkey with dexamethasone appeared to cause a 2.5-fold increase in both the 7 alpha- and the 7 beta-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol without increasing the 6 alpha-hydroxylation. The 7 alpha- and 7 beta-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol by human and monkey prostate microsomes, but not the 6 alpha-hydroxylation, was inhibited by antibody against rat liver NADPH-cytochrome P450 reductase. Similarly, the 7 alpha- and 7 beta-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol by human prostate microsomes, but not the 6 alpha-hydroxylation, was markedly inhibited (greater than 85%) by equimolar concentrations of the imidazole-containing antimycotic drugs ketoconazole, clotrimazole, and miconazole. These results suggest that the 7 alpha- and 7 beta-hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol by monkey and human prostate microsomes is catalyzed by a cytochrome P450 enzyme, whereas the 6 alpha-hydroxylation is catalyzed by a different enzyme which may or may not be a cytochrome P450 monooxygenase. The hydroxylation of 5 alpha-androstane-3 beta,17 beta-diol by prostate microsomes from normal human subjects was quantitatively and qualitatively similar to its hydroxylation by prostate microsomes from patients with benign prostatic hyperplasia.(ABSTRACT TRUNCATED AT 400 WORDS)     

The effect of short term treatment with cyproterone acetate or flutamide on the metabolism of 5 alpha-dihydrotestosterone in human testicular tissue

Testicular tissue obtained from ten patients orchiectomized for prostatic cancer was incubated with [3H]5 alpha-dihydrotestosterone (DHT) in order to study the metabolic transformation into 5 alpha-androstane-3 alpha,17 beta-diol (3 alpha-diol) and 5 alpha-androstane-3 beta,17 beta-diol (3 beta-diol). Throughout 5 days before surgery four subjects were treated with cyproterone acetate (CA). To three patients flutamide (F) was administered for the same period of time. Three subjects remained untreated. Compared to the control group the administration of CA decreased the formation of 3 beta-diol whereas that of 3 alpha-diol increased. Treatment with F lead to an elevated formation of both diols. However, the 3 alpha/3 beta ratio did not change. As 3 beta-diol is considered to be an index of tubular function in the human testis it is concluded that CA has a direct inhibitory effect upon this testicular compartment whereas F has none.     

Steroid metabolism by rat nasal mucosa: studies on progesterone and testosterone

The metabolism of [4-14C]progesterone and [4-14C]testosterone by slices of the nasal mucosa from rats was studied. As shown by gas chromatography-mass spectrometry there was a preferential formation of reduced progesterone-metabolites (5 alpha-pregnane-3,20-dione, 3 alpha- and 3 beta-hydroxy-5 alpha-pregnane-20-one, 20 alpha- and 20 beta-hydroxypregn-4-en-3-one, 2 alpha,3 alpha-dihydroxy-5 alpha-pregnane-20-one, 3 alpha,16 alpha-dihydroxy-5 alpha-pregnane-20-one) and reduced testosterone-metabolites (4-androstene-3,17-dione, 5 alpha-dihydrotestosterone, 3 alpha-hydroxy-5 alpha-androstane-17-one, and 5 alpha-androstane-3 alpha, 17 beta-diol, 2 alpha-hydroxy-5 alpha-dihydrotestosterone, 5 alpha-androstane-2 alpha,3 alpha, 17 beta-triol) indicating the presence of 5 alpha-reductase, 3 alpha-, 3 beta-, 17 beta-, 20 alpha- and 20 beta-hydroxysteroid oxidoreductase activities in this tissue. Progesterone-metabolites hydroxylated at positions 2 alpha, 6 alpha, 6 beta, 15 alpha and 16 alpha and testosterone-metabolites hydroxylated at positions 1 beta, 2 alpha, 6 beta, 15 beta and 16 alpha were also identified, indicating the presence of several steroid hydroxylases in the nasal mucosa. Autoradiography of the nasal region of rats injected with [4-14C]progesterone or [4-14C]testosterone showed a selective localization of radioactivity in the mucosa covering the olfactory region of the nasal cavity.     

Metabolism and binding in vitro of 5 alpha-androstane-3 beta, 17 beta-diol and of 5 alpha-androstane-3 alpha, 17 beta-diol in cell fractions of rat ventral prostate and liver.

Rat ventral prostate and liver were investigated for the binding in vitro to particulate fractions and for the metabolism of 5 alpha-androstane-3 beta, 17 beta-diol. Comparative investigations were carried out on the metabolism of 5 alpha-androstane-3 alpha, 17 beta-diol. Preparations of the liver were investigated in order to establish the organ specificity of the method. In the prostate, the bulk of the metabolites of 5 alpha-androstane-3 beta, 17 beta-diol was present as steroids of high polarity. Of the less polar metabolites, 17 beta-hydroxy-5 alpha-androstan-3-one, 3 beta-hydroxy-5 alpha-androstan, 17-one and 5 alpha-androstane-3 alpha, 17 beta-diol were detectable. The binding of a 5 alpha-androstane-3 beta, 17 beta-diol to mitochondria and microsomes was unspecific. In the liver, among the less polar metabolites, 3 beta-hydroxy-5 alpha-androstan-17-one was the main metabolite, and the binding was unspecific. The main metabolite in the prostate homogenate of 5 alpha-androstane-3 alpha, 17 beta-diol was 17 beta-hydroxy-5 alpha-androstan-3-one. The portion of highly polar steroids was very low. The portion of unmetabolized hormone was distributed almost equally among the different cell preparations except the nuclei, in which 17 beta-hydroxy-5 alpha-androstan-3-one was higher and 5 alpha-androstane-3 alpha, 17 beta-diol was lower than in the remaining cell fractions.     

Androgen metabolism in rat L6 myoblast cells; high formation of 5 alpha-androstane-3 alpha,17 beta-diol from testosterone

We have studied androgen metabolism in L6 rat myoblasts. 4-androstene-3,17-dione (Adione), testosterone, 5 alpha-dihydrotestosterone (DHT), and 5 alpha-androstane-3 alpha, 17 beta-diol (3 alpha-diol) were used for substrates and the amounts of metabolites formed from the respective substrates in the medium were determined. Conversion of Adione to testosterone was dominant over the reverse conversion. DHT formation from testosterone was low and did not change with the duration of incubation, whereas 3 alpha-diol formation increased in a time-dependent manner. Major metabolite of testosterone was not DHT but 3 alpha-diol. A large amount of 3 alpha-diol was formed from DHT, however, DHT formation from 3 alpha-diol was very low. These data indicate that L6 cells have high 5 alpha-reductase activity and suggest that DHT formed from testosterone is rapidly metabolized to 3 alpha-diol in these cells.     

Determination of 5 alpha-androstane-3 alpha, 17 beta-diol and 5 alpha-androstane-3 beta,17 beta-diol in human plasma by radioimmunoassay

5 alpha-Androstane-3 alpha,17 beta-diol (3 alpha-diol) and 5 alpha-androstane-3 beta,17 beta-diol (3 beta-diol) were measured in human peripheral plasma by radioimmunoassay using celite microcolumn purification. The antisera used for the assay were obtained by immunization of rabbits with 3 alpha,17 beta-dihydroxy-5 alpha-androstane-6-(O-carboxymethyl) oxime: BSA for 3 alpha-diol and 3 beta,17 beta-dihydroxy-5 alpha-androstane-15 alpha-carboxymethyl: BSA for 3 beta-diol. The concentrations (pg/ml +/- SD) of the two diols in normal male and female plasma are respectively: 216 +/- 51 and 49 +/- 32 for 3 alpha-diol, 239 +/- 76 and 82 +/- 45 for 3 beta-diol. Comparison of these results with published ones shows that 3 beta diol concentrations were significantly lower. The high specificity of the assay is due to chromatography on celite microcolumns, allowing elimination of 5-androstene-3 beta,17 beta-diol from the plasma sample.