Steroid metabolism by testicular homogenates of the Stanley-Gumbreck pseudohermaphrodite male rat. I. Increased formation of androsterone and androstanediol

Tritiated progesterone androstenedione and testosterone were incubated with testicular homogenates of 4- and 32-week-old Stanley-Gumbreck pseudohermaphrodite (Ps) and normal (N1) male littermate rats. In 15 and 180 minute incubations, both 4- and 32-week-old Ps testes converted all three substrates predominantly to androsterone and to a lesser extent androstanediol, while androstanediol in 4-week and testosterone in 32-week-old N1 testis were the major products. The addition of carrier testosterone (240 μg/g tissue) to 15 min incubations of testicular homogenates from 4- and 32-week-old N1 rats almost completely blocked the formation of androstanediol and markedly increased the accumulation of testosterone (47 and 41% from Progesterone-1,2-³ H; 66 and 92% from androstenedione-l,2-³H) indicating that androstanediol formed in the absence of carrier testosterone is, most likely, a product of testosterone reduction. When similar incubations were repeated using testicular homogenates from 4- and 32-week-old Ps rats, testosterone accumulation was not greatly increased (4–11%) by the addition of carrier testosterone, but androsterone formation was completely inhibited. However, when the incubations of Progesterone-1 ,2-³H with 4- and 32-week-old Ps and N1 testis in the presence of carrier testosterone were continued for 180 min, the major fraction of radioactivity from 32-week-old N1 testis was testosterone (79%) while that from 4-week-old N1 testis was androstanediol (60%) and from 4-and 32-week-old Ps testis was both androsterone (44–45%) and androstanediol (22–33%). The present data indicate that 4-week-old Ps testis, like the N1, has a high level of ring A reductase activity but forms androsterone rather than androstanediol as its major product. Unlike the normal mature male rat testis, in which ring A reductase activity diminishes allowing testosterone to become the major product, the 32-week-old Ps testis maintains a high level of reductase activity.     

Steroid metabolism in the gonads of a patient with testicular feminization. II. Metabolism of c19- and c18-steroids in vitro.

The metabolism of C19- and C18-steroids, in particular, the aromatization of androstenedione and testosterone, the interconversion of androgens to estrogens and the 5alpha-reductase activity of a right abdominal (r) and a left inguinal (l) testis of a patient with testicular feminization, are reported. Aromatization and 5alpha-reductase activity were also evaluated in tissue from the left ductus diferens (ld). The following results were obtained: 1. aromatization of androstenedione to estrone 2.52% (r), 0.02% (l), 0.94% (ld); 2. aromatization of testosterone to estradiol 0.58% (r), 2.88% (l); 3. conversion of androstenedione to testosterone 95.65% (r), 98.07% (l); 4. conversion of testosterone to androstenedione 33.14% (r), 53.65% (l); 5. conversion of estrone to estradiol85.29% (r), 100% (l), 6. conversion of estradiol to estrone 33.12% (r), 32.33% (l); 7.5alpha-reduction of testosterone to 5alpha-dihydrotestosterone 12.01% (r), 13.64% (l) and 4.10% (ld). A lack of 5alpha-reductase activity was not found in the tissues examined as stated in the literature. Estrogen production in these testes was demonstrated by the aromatization of androstenedione and testosterone to estrone and estradiol and is reflected in the difference of the estradiol concentration measured in spermatic and peripheral blood of the same patient (168 versus 33 pg/ml).     

The in vitro metabolism of progesterone, 4-androstene-3,17-dione, testosterone and 17 beta-hydroxy-5 alpha-androstan-3-one by fetal rhesus monkey testes.

Homogenates prepared from fetal rhesus monkey testes were incubated with progesterone, 4-androstene-3,17-dione, testosterone and 17 beta-hydroxy-5 alpha-androstan-3-one. The major progesterone metabolite was 17-hydroxy-4-pregnene-3,20-dione. Testosterone also accumulated in the progesterone incubations. 4-Androstene-3,17-dione was converted chiefly to testosterone. Testosterone was not actively metabolized by the fetal monkey testis. 17 beta-Hydroxy-5 alpha-androstan-3-one was actively converted primarily to 5 alpha-androstane-3 beta,17 beta-diol.     

Testicular steroidogenesis in the baboon Papio anubis.

We recently reported that the baboon testis converts pregnenolone to testosterone through the delta-4 pathway. The present studies were to determine the metabolism of intermediates of the delta-4 and delta-5 pathway by the baboon testis. Fragments (50 mg) were incubated for 3 hr with 10 muCi of the following tritium-labelled substrates: pregnenolone, progesterone, 17-hydroxypregnenolone, 17-hydroxyprogesterone, dehydroepiandrosterone, androstenedione, or testosterone. Pregnenolone was converted to testosterone primarily through the delta-4 pathway, with accumulation of progesterone, 17-hydroxyprogesterone and 20alpha-dihydroprogesterone as predominant intermediates. Similar results were obtained in progesterone incubations. 17-hydroxyprogesterone was not efficiently metabolized by the fragments, while 17-hydroxypregnenolone and dehydroepiandrosterone were efficiently converted into testosterone and androstenedione. Androstenedione was metabolized primarily to testosterone, while testosterone was not a suitable substrate. Some 5alpha-androstanediol was identified in each incubate. These results suggest that although testosterone is formed from pregnenolone through the delta-4 pathway, the delta-5 intermediates are more suitable substrates for testosterone synthesis in the baboon testis.     

Comparative ability of rat seminiferous tubules, interstitium, and whole testes to utilize cholesterol-1,5-3H as a substrate for testosterone synthesis and the intratesticular distribution of cholesterol side-chain cleavage enzyme.

Homogenates of rat seminiferous tubules, interstitium and intact testis tissues were assessed for their ability to convert cholesterol -1,2-3H to testosterone in vitro. While 3H-testosterone synthesis was observed in incubates of interstitial and whole testis homogenates, no synthesis was detectable in homogenates of seminiferous tubules. To determine whether cholesterol side-chain cleavage enzyme (CSCCE) was deficient or absent in tubules, mitochondria from tubules, interstitium and whole testes were analyzed for CSCCE activity by measuring conversion of cholesterol -26-14C to 14C-isocaproate (+pregnenolone). Interstitial mitochondrial preparations from each of six testes were found to be approximately 200 times more active in CSCCE than the corresponding tubule mitochondria, and 1600-1800 times more active on a specific activity basis. Although caution is required in extrapolation of in vitro data to the in vivo state, these findings suggest rat seminiferous tubules may be incapable of de novo testosterone biosynthesis and that this lack of synthetic ability may be due to a deficiency of CSCCE.     

Zymogen secretion and phospholipid metabolism in the pancreas. Phospholipids of the zymogen granule

1. The metabolism of [4-(14)C]pregnenolone to androst-16-enes has been studied in short-term incubations of boar testis tissue. With fresh tissue androsta-5,16-dien-3beta-ol (8%) and 5alpha-androst-16-en-3beta-ol (2%) were formed. Tissue that had been stored at -20 degrees C was still capable of metabolizing pregnenolone to androsta-5,16-dien-3beta-ol. 2. NADPH was essential for the formation of androsta-5,16-dien-3beta-ol from pregnenolone; NADH had less activity and ATP was not necessary for the reaction. 3. [4-(14)C]Androsta-5,16-dien-3beta-ol, prepared biosynthetically from [4-(14)C]pregnenolone, was shown to be converted by boar testis preparations into androsta-4,16-dien-3-one (31%) if NAD(+) was present or into 5alpha-androst-16-en-3beta-ol (4%) if NADPH was present. 4. 17alpha-Hydroxyandrost-4-en-3-one and 3beta,17alpha-dihydroxypregn-5-en-20-one were considered as possible precursors for androst-16-ene formation, but both were shown to be ineffective. 5. No radioactivity was incorporated into androst-5-en-3beta-ol used to trap any corresponding (14)C-labelled compound formed from [4-(14)C]pregnenolone.     

Steroid hormone biosynthesis by a sesterterpene pathway in the rat and rabbit testis

Rat and rabbit testis preparations were incubated with [4-14C]cholesterol and 23,24-dinor-[7 alpha-3H]5-cholen-3 beta-ol, the latter being a proposed intermediate in the sesterterpene pathway for steroid biosynthesis. Steroids were isolated, purified by thin-layer chromatography and crystallised to constant specific activity. It was found that rat and rabbit testis can utilise 23,24-dinor-5-cholen-3 beta-ol to produce testosterone. The tritium/carbon-14 ratios in the testosterone and androstenedione isolated indicated that these tissues differentiated between the two substrates. This finding is supported by the observation that, on stimulation with HCG, the tritium/carbon-14 ratios in the testosterone isolated were increased compared to the controls. The results of further experiments implied that, while the biosynthesis of testosterone from cholesterol occurred in the rat testis mitochondrial fraction, its biosynthesis from 23,24-dinor-5-cholen-3 beta-ol occurred in the microsomal fraction.     

The biosynthesis of some androst-16-enes from C21 and C19 steroids in boar testicular and adrenal tissue

1. The formation of androst-16-enes from [4-(14)C]progesterone has been investigated with long-term incubations and short-term kinetic studies. After 4hr., 1.7 and 10.3% respectively of 3alpha- and 3beta-hydroxy-5alpha-androst-16-enes were formed in boar testis minces, but much smaller yields were obtained in boar adrenal. Both tissues formed small quantities of androsta-4,16-dien-3-one. 2. The amounts of androst-4-ene-3,17-dione and testosterone isolated were small, suggesting that androst-16-ene formation may occur preferentially in the boar testis. 3. In the absence of tissue no radioactive androst-16-enes were formed. 4. Incubation of both [4-(14)C]pregnenolone and [7alpha-(3)H]progesterone resulted in 3alpha- and 3beta-hydroxy-5alpha-androst-16-enes containing (3)H/(14)C ratios of near unity and confirmed that both C(21) steroids were precursors. A similar incubation with 17alpha-hydroxy[4-(14)C]-progesterone and [7alpha-(3)H]progesterone gave the same Delta(16)-alcohols, but they contained only (3)H, indicating that side-chain cleavage of pregnenolone and progesterone occurred before 17alpha-hydroxylation. 5. Dehydroepiandrosterone, testosterone, testosterone acetate and 16-dehydroprogesterone were not found to be precursors of Delta(16)-steroids. 6. A pathway is proposed for the biosynthesis of 3alpha- and 3beta-hydroxy-5alpha-androst-16-enes from pregnenolone and progesterone; this may involve androsta-4,16-dien-3-one as an intermediate, but excludes 17alpha-hydroxyprogesterone, testosterone and dehydroepiandrosterone.     

Cell-free transfer of sterols from dictyosome-like structures to plasma membrane vesicles of guinea pig testes

Summary Transfer of radiolabeled lipids from dictyosome-like structures (DLS) from testis tubules of the guinea pig as donor to unlabeled plasma membrane from testis tubules immobilized on nitrocellulose as acceptor was studied in a completely cell-free system. As a general label for lipids of the donor DLS, isolated testis tubules were incubated with [¹⁴C]acetate. Time- and temperature-dependent transfer of [¹⁴C]acetate labeled constituents was observed in the cellfree system. However, despite the fact that phospholipids and other constituents were highly labeled in the donor fraction, primarily radioactive sterols were transferred to the plasma membrane acceptor vesicles. Transfer at 37°C represented 0.4 to 0.7% of the total radiolabeled cholesterol at 37°C but little or no transfer occurred at 4°C. The sterols transferred exhibited Chromatographic mobilities corresponding to those of cholesterol and lanosterol. Similar results were obtained with [¹⁴C]mevalonic acid. In subsequent experiments, cholesterol transfer from DLS to plasma membrane was demonstrated by incubation of DLS with [³H]squalene which was converted into sterol or with [¹⁴C]cholesterol. Transfer of sterols required ATP, but not cytosol, and was both time- and temperature-dependent. DLS were more effective than either endoplasmic reticulum or plasma membrane as the donor fraction. The results from the cell-free analysis suggest a possible functional role of the DLS in sterol biogenesis and transfer to the plasma membrane during spermatid development.Abbreviations DLS dictyosome-like structure(s) - PBS phosphatebuffered saline - HEPES 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid - BSA bovine serum albumin     

The metabolic route by which oleate is converted into cholesterol in rat hepatocytes.

The effect of (-)-hydroxycitrate on the conversion of [1-14C]oleate into cholesterol was dependent on the time of day at which the cells were prepared and on the extracellular oleate concentration. In hepatocytes prepared during the light phase of the diurnal cycle (L2-hepatocytes), (-)-hydroxycitrate inhibited the conversion of L-[U-14C]lactate (2 mM) and of 0.13 mM-[1-14C]oleate into cholesterol. However, when [1-14C]oleate was present at 1.3 mM, most of the sterol carbon was derived from this source, and under these conditions (-)-hydroxycitrate had no inhibitory effect on [14C]cholesterol formation. In these cells, non-radioactive acetoacetate blocked the conversion of 1.3 mM-[1-14C]oleate, but not of 0.13 mM-[1-14C]oleate, into cholesterol. In cells prepared during the dark phase of the diurnal cycle (D6-hepatocytes), irrespective of the concentration of [1-14C]oleate, (-)-hydroxycitrate decreased its conversion into cholesterol. In both types of cell preparation, the inhibitory effect of (-)-hydroxycitrate on the conversion of L-[U-14C]lactate into cholesterol was greater than that on the overall rate of cholesterol production from all endogenous sources. These results provide evidence for the following. (1) The major metabolic route by which oleate is converted into cholesterol is dependent on its extracellular concentration. (2) When oleate is the major source of hepatic sterol carbon, the flux of substrate through citrate into cholesterol is dependent on the nutritional state of the animal. (3) When endogenous substrates are the sole source of sterol carbon, a substantial proportion of the carbon enters the cholesterol pathway through routes not involving citrate cleavage.     

Metabolism of cholesteryl palmitate by rat brain in vitro; formation of cholesterol epoxides and cholestane-3beta,5alpha,6beta-triol

Incubation of [4-(14)C]cholesteryl palmitate with the 12,000 g supernatant fraction of adult rat brain fortified with an NADPH-generating system and beta-mercaptoethylamine resulted in formation (2-5%) of more polar metabolites characterized as a mixture of cholesterol-5,6-epoxides. Under extended incubation conditions, cholestane-3beta-5alpha-6beta-triol was isolated as the major end product of the incubations. Free [4-(14)C]cholesterol incubated under similar conditions was not oxidized, whereas oxidation of [4-(14)C]cholesteryl palmitate appeared to be dependent upon hydrolysis of the ester by the rat brain microsomal subcellular fraction. Elimination of the NADPH-generating system or the addition of EDTA to the incubation mixture inhibited epoxide formation, suggesting that the products are derived from an NADPH-dependent enzymatic lipoperoxidation mechanism. The in vitro conversion of [4-(14)C]cholesterol-5alpha,6alpha-epoxide to cholestane-3beta,5alpha,6beta-triol was also demonstrated in rat brain subcellular fractions in the absence of added cofactors.     

Seasonal variation in steroid biosynthesis by the testis of the lizard Tiliqua Rugosa.

1. Seasonal change in the biosynthesis of androgens by testicular tissue from a scincid lizard Tiliqua rugosa has been studied in vitro using [7-3H]pregnenolone and [4-14C]progesterone as substrates. 2. Evidence is presented for the synthesis of epitestosterone, testosterone, androstenedione, 17 alpha-hydroxyprogesterone, 20 beta-hydroxypregn-4-en-3-one, and 17 alpha, 20 beta-dihydroxypregn-4-en-3-one. 3. Epitestosterone was the major conversion product. Its yield was highest (68%) during summer and lowest (14%) during spring. The yield of testosterone was maximal (10%) during spring (mating season). The production of 20 beta-hydroxypregn-4-en-3-one was highest during spring and undetectable during summer. 4. Histological evidence and changes in testicular weight indicated that spermatogenic activity was greatest during spring. Pronounced regression of the testis occurred immediately after the breeding season and lasted until autumn.     

21-Hydroxylation of C-11-oxygenated and C-11-deoxysteroids by perfused dog adrenals.

The left adrenal of two female dogs were perfused with either 3 muCi [4-14C]cpd. S (2) and 45 muCi [1, 2-3H] 21-deoxycortisone (Dog I), or with 1 muCi [4-14C] 17-hydroxyprogesterone and 65 muCi [1,2-3H] 21-deoxycortisone (Dog II). In Dog I, 40% of the perfused 21-deoxycortisone was converted to cortisone and 23% of cpd. S was converted to cortisol. In Dog II the percent conversion of 21-deoxycortisone to cortisone and of 17- hydroxyprogesterone to cortisol was 24% and 15% respectively. The results demonstrate that the dog adrenal has the capability of hydroxylating an 11-oxygenated steroid at C-21.     

Pathway of testosterone biosynthesis in the testis of the marmoset Saguinus oedipus

These studies were undertaken to determine the principal pathway of androgen biosynthesis by the testis of the marmoset $\text{Saguinus oedipus}$. Testicular fragments (25 mg) were incubated at 37°C in Krebs-Ringer bicarbonate buffer, pH 7.4, containing pregnenolone-7-³H (3β-hydroxy-5-pregnen-20-one) or progesterone-7-³H. Duplicate fragments were incubated with each substrate for 30 min, one hr, three hr, or five hr, for a total of 16 separate incubations. Metabolites were separated by paper and thin-layer chromatography, with identity established by recrystallization to constant specific activities and ³H/¹⁴C ratios. Pregnenolone was readily metabolized to progesterone, 17α-hydroxyprogesterone, androstenedione (4-androstene-3, 17-dione) and testosterone. Progesterone was converted to 17α-hydroxyprogesterone, androstenedione and testosterone. 17α-hydroxyprogesterone was the predominant metabolite obtained from both substrates at one, three and five hrs of incubation. Neither 17α-hydroxypregnenolone (3β-17-dihydroxy-5-pregnen-20-one) nor dehydroepiandrosterone (3β-hydroxy-5-androsten17-one) was detected in the incubates. These data suggest a predominant Δ⁴ pathway with accumulation of 17α-hydroxyprogesterone in the testis of this primate specie.     

Intestinal and hepatic microsomal metabolism of testosterone and progesterone by a 3 alpha-hydroxysteroid dehydrogenase to the 3 alpha-hydroxy derivatives in the channel catfish,Ictalurus punctatus

Intestinal or hepatic microsomes from channel catfish converted [4-14C]-testosterone to three major metabolites: 6 beta-hydroxytestosterone, androstenedione and a third metabolite. Formation of the unknown metabolite required NADPH as cofactor. When incubated with 200 microM testosterone, the rate of formation of the unknown metabolite was 265+/-158 pmol/(min mg) protein (mean+/-S.D.) in microsomes from the proximal intestine, 515+/-93 pmol/(min mg) protein in distal intestine and 226+/-42 pmol/(min mg) protein in hepatic microsomes. Comparison of the chromatographic and spectral properties of the unknown metabolite with those of authentic testosterone derivatives showed that this metabolite corresponded to 4-androstene-3 alpha,17 beta-diol. No 3 alpha-reduced metabolite was formed in incubations of testosterone with catfish intestinal cytosol. Testosterone was reduced to 5 alpha-dihydrotestosterone primarily in the cytosolic fraction and not in microsomes. Incubation of progesterone with intestinal microsomes resulted in the formation of a metabolite with properties similar to that of the 3 alpha-reduced testosterone, and this metabolite was identified by co-chromatography with authentic standard as 3 alpha-reduced progesterone. Thus, 3 alpha-hydroxysteroid dehydrogenase is an important pathway in intestinal microsomes of the channel catfish.     

Androgenic modulation of progesterone metabolism by rat granulosa cells in culture

Effects of androgens on progesterone accumulation, utilization of exogenous progesterone and accumulation of [4-14C]progesterone metabolites by rat granulosa cells in culture were studied. Androgen increased progesterone accumulation in cultures without exogenous progesterone and slowed the overall decline of progesterone concentration in cultures supplemented with exogenous progesterone. Both aromatizable testosterone and nonaromatizable 5 alpha-dihydrotestosterone decreased [4-14C]progesterone utilization by granulosa cells by 12 to 30%. This effect was observed irrespective of whether the cells were continuously exposed to androgens or only pre-exposed. In he same experiments, androgens decreased conversion of radiolabeled progesterone to 20 alpha-hydroxy-4-pregnen-3-one by 11 to 50% and to 5 alpha-pregnane-3 alpha, 20 alpha-diol by 26 to 49%. Accumulation of 3 alpha-hydroxy-5 alpha-pregnan-20-one was not altered in 3 h incubations and was increased by up to 43% in 24 h incubations by androgen treatment. It is suggested that androgens alter progesterone catabolism by granulosa cells by decreasing 20 alpha-hydroxysteroid dehydrogenase activity and that this effect may contribute to overall stimulatory action of androgens on progesterone accumulation.     

Stimulation of liver cholesterol synthesis by actinomycin D

1. An eightfold increase in the incorporation of [2-(14)C]acetate into liver cholesterol in vivo was observed 24hr. after starved rats had been given actinomycin D (0.5mg./kg. of body wt.). Liver cholesterol radioactivity declined faster in the treated animals, suggesting a greater rate of cholesterol turnover. 2. Liver slices from treated animals showed a tenfold increase in the incorporation of [2-(14)C]acetate into cholesterol; conversion into CO(2) and into fatty acids was less markedly increased, and conversion into ketone bodies was not significantly affected. 3. The patterns of conversion into liver cholesterol in vivo of the lactone and the sodium salt of mevalonic acid differed markedly. The former was converted at a faster rate and to a greater extent than the latter. Treatment with actinomycin D increased the conversion of both forms of mevalonic acid into liver cholesterol, but only to a small extent. 4. Stimulation of the incorporation of acetate into cholesterol occurred at 4hr. after the administration of actinomycin D but not at 2hr. The response was abolished by the simultaneous administration of dl-ethionine or puromycin. 5. Pre-feeding with a cholesterol-rich diet greatly diminished the stimulation of conversion of acetate into cholesterol caused by actinomycin D, though it did not completely suppress it. Adrenalectomized animals responded to the drug, but much less markedly. 6. It is concluded that actinomycin D stimulates the synthesis of cholesterol in the liver at a stage in the pathway before mevalonic acid, by a mechanism that probably requires protein synthesis. A likely site would be the beta-hydroxy-beta-methylglutaryl-CoA reductase, the rate-limiting enzyme in cholesterol biosynthesis. Some possible mechanisms by which the drug may lead to increased activity of this enzyme are considered.     

Inhibition of the conversion of cholesterol to pregnenolone in bovine adrenocortical preparations.

The inhibition of the conversion of [4-14C] cholesterol to [4-14C] pregnenolone by a number of steroids has been studied in bovine adrenocortical mitochondrial acetonedried preparations. At equimolar substrate and inhibitor concentrations (3.3 muM) the most potent inhibitors were cholesterol derivatives containing a nitrogen function at c-22, followed by derivatives containing oxygen functions at c-22 or c-20 or both. The presence of a hydroxyl group at c-17 or the replacement of the 3beta-hydroxyl group by fluorine reduced the inhibitory efficacy. In the presence of inhibitors that were also relatively good substrates of the cholesterol side-chain cleavage system, such as some cholesterol derivatives hydroxylated in the side-chain,the rate of [4-14C] pregnenolone formation increased with time as the inhibitor was consumed. (20S)-20,21-Dihydroxycholesterol exerted such an effect on the kinetics of [4-14C]pregnenolone formation, and yielded 21-hydroxypregnenolone which was identified by gas chromatography-mass spectrometry. The synthesis of (20R)-22-ketocholesterol, of (20R,22R)-22hydroxycholesterol, (20R,22S)-hydroxycholesterol, and of (20S)-desmosterol is described.     

Inhibition of testicular testosterone biosynthesis following experimental varicocele in rats

In an attempt to determine whether defective testicular testosterone (T) biosynthesis may be associated with a varicocele, an experimental study was performed in adult rats whereby a unilateral left varicocele was surgically created. At 2, 4, 8, and 12 wk following the creation of the varicocele, intratesticular T as well as the activities of three (17 alpha-hydroxylase, 17,20-desmolase, and 17 beta-hydroxysteroid dehydrogenase) of the five enzymes in the delta 4 pathway of testicular T biosynthesis were measured. Intratesticular T (ng/g testis +/- SEM) in the left testis decreased significantly from 121 +/- 21 in the control group to 59 +/- 8 in the two-wk varicocele group (p less than 0.01), and remained significantly suppressed throughout the experimental period. The T concentrations in the right testis paralleled those in the left in both the control and varicocele animals. At 2 wk following the creation of the varicocele, the activity (nmol/min/testis +/- SEM) of the 17,20-desmolase enzyme decreased significantly, from 115 +/- 8 in the left testis of control rats to 87 +/- 6 in the left testis of the varicocele animals (p less than 0.025), and remained low throughout the 12 weeks of the study. The activity of the 17 alpha-hydroxylase enzyme was significantly decreased at the 8th and 12th weeks of the study, while the 17 beta-hydroxysteroid dehydrogenase activity did not show any significant change during the study period. The enzyme activities in the right testis paralleled those in the left testis.(ABSTRACT TRUNCATED AT 250 WORDS)     

Biosynthesis of steroids from cholesterol-14C by human adrenal carcinoma tissue

Slices prepared from human adrenal carcinoma tissue obtained froa a 51 year old female with virilism were incubated with cholesterol-4-¹⁴C (l) in the presence and absence of ACTH or cyclic AMP. The tissue homogenates were analyzed for steroids by the reverse isotope dilution method. Purification and identification of the radio-active metabolites were achieved by column, paper and thin-layer chromatography, derivative formation and crystallization to constant specific activity. Cholesterol-¹⁴C was converted to pregnenolone-¹⁴C (0.75%), progesterone- ¹⁴C (0.17%) and dehydroepiandrosterone-¹⁴C(0.15%). No evidence was found for the formation of labeled 17-hydroxyprogesterone, androstenedione, testosterone, and estrogens. Addition of ACTH to the incubations resulted in a two-fold inoreas of radioactive pregnenolone-¹⁴C and or dehydroepiandrosterone-¹⁴C. A two-fold increase of progesterone-¹⁴C synthesis was found in cyclic AMP-stimulated incubations.