Twice-daily treatments with subovulatory doses of hCG result in the development of large ovarian cysts that possess the capacity to produce preovulatory amounts of estradiol (E2) in the presence of exogenous substrate (Biol Reprod 1991; 45:34-42). To determine the effects of prolonged stimulation by subovulatory doses of LH-like activity on the ability of ovarian follicles to produce aromatizable androgens and their metabolites and on the capacity of similar follicles to metabolize exogenous androstenedione (A4) and testosterone, pregnant rats were treated with either 0 (control), 1, or 3 IU hCG twice daily for 9 days, beginning on Day 13 of pregnancy. The largest follicles or cysts in the ovaries of these animals on Days 15, <em>17</em>, 19, and 22 were incubated for 4 h in the presence of 1) medium alone, 2) 1 mM cAMP, 3) 800 ng/ml A4 with or without cAMP, or 4) 800 ng/ml testosterone with or without cAMP. In the presence or absence of cAMP, follicular incubates from controls displayed limited amounts of A4, testosterone, E2, estrone (E1), and 5alpha-reduced androgen accumulation compared to incubates from rats treated with hCG. By Day 22, follicular incubates from rats treated with 3 IU hCG contained more A4 than incubates from animals treated with 1 IU hCG. However, on each day tested, incubates from rats treated with 1 IU hCG displayed at least as much, and usually more, testosterone, E2, E1, and 5alpha-reduced androgens as did incubates from rats treated with 3 IU hCG. Follicles from rats treated with 3 IU hCG lost their ability to respond to cAMP with increased steroidogenesis. The capacity of follicles from controls and from rats treated with 3 IU hCG to metabolize exogenous A4 to testosterone, E2, and 5alpha-reduced androgens was maintained between 32 and 37 ng of steroid/4 h from Day 15 to Day 22, whereas the capacity of follicles from rats treated with 1 IU hCG to metabolize A4 ranged from 68 to 92 ng of steroid/4 h. Similar patterns for E2 and 5alpha-reduced steroid production were observed when exogenous testosterone was used as substrate. Follicles from all in vivo treatment groups displayed similar capacities to reverse-metabolize exogenous testosterone to A4 and E1 on Day 15 of pregnancy. This capacity increased dramatically, from 29 to 75 ng of steroid/4 h, between Days 15 and <em>17</em> for follicles from rats treated with 3 IU hCG. A similar increase was not observed for follicular incubates from rats treated with 1 IU hCG until Day 22 of pregnancy, and was never observed for follicles from controls. In summary, prolonged exposure to stimulation by subovulatory doses of LH-like activity differentially affects the forward and reverse metabolism of aromatizable androgens by ovarian cysts that are developing in the pregnant rat. The limited ability of large, hCG-induced cysts to forward-metabolize A4 and the apparent increased capacity of these follicles to reverse-metabolize testosterone to A4 indirectly support the notion that follicular <em>17</em>-<em>ketosteroid</em> reductase and <em>17</em>beta-hydroxysteroid dehydrogenase activities may be differentially regulated during the induction of ovarian cysts in response to prolonged stimulation by gonadotropins in the anovulatory environment of the pregnant rat.

A 42 year old (46 XY) subject with <em>17</em>-<em>ketosteroid</em> reductase deficiency was investigated. The patient reared as female, has developed masculine features (facial hair, male distribution of body hair, male body habitus, acne and clitoridomegaly) at about 15 years of age but never consulted. She married at 22 years and for 20 years thought to have a "normal" female sex life. Only when her 14 year old "niece" was investigated (1) and treated for similar problems she realized hers. She had a small phallus with perineal urethra, vaginal pouch absence of labia minora and undescended testis, no breast development. Baseline peripheral studies showed plasma testosterone (T) in the range of Tanner II stade of puberty (150 ng/dl), elevated delta 4-androstenedione (delta 4) (930 ng/dl) and estrone (E1) (33,5 NG/DL) LEVELS 6--8 times above normal, but subnormal estradiol levels. Increased basal gonadotropins showed an hyper-response to LHRH stimulation. Dynamic tests (ACTH test, Dexamethasone suppression, and hCG stimulation) showed that abnormal delta 4 and E1 were not of adrenal origin. In the spermatic veins delta 4 levels were extremely high (239 micrograms/dl) but T levels low (11.4 micrograms/dl). delta 4/T ratio in the spermatic vein was much higher than in the peripheral blood suggesting intact peripheral conversion of delta 4 to T. After castration all hormone levels returned to the range usually seen in agonadic male or female adults.

We studied the testicular function and some androgen-mediated events in 22 males (16-30 years of age) with male pattern baldness that was treated with finasteride (10 mg once daily) for 2 years. Patients were evaluated every 3 months. Prostatic volume was determined in six subjects by endorectal ultrasound scans. Serum gonadotropin, prostate-specific antigen (PSA), and sex hormone levels were determined basally and periodically during the treatment period. Fourteen subjects underwent gonadal stimulation with human chorionic gonadotropin (hCG), and the gonadotropin response to gonadotropin releasing hormone (GnRH) was determined in eight subjects, prior to and after 2 years of therapy. Finasteride treatment resulted in an improvement in the male pattern baldness and prostatic shrinkage that was associated with an increase in serum testosterone levels (<em>17</em>.2 +/- 2.5 vs. 26.3 +/- 1.7 nmol/L) and a decrease in dihydrotestosterone (DHT) levels (1.45 +/- 0.41 vs. 0.38 +/- 0.10 nmol/L), causing a marked increase in that testosterone/DHT ratio. A significant increase in the serum levels of androstenedione (3.67 +/- 0.49 vs. 7.05 +/- 0.70 nmol/L) and estradiol (132 +/- 44 vs. 187 +/- 26 pmol/L) was also noted, whereas androstanediol glucoronide (33.3 +/- 6.4 vs. 10.7 +/- 4.5 pmol) and PSA (1.6 +/- 0.6 vs. 0.4 +/- 0.1 ng/ml) were significantly decreased. No changes in basal or stimulated levels of gonadotropin were observed. There was a significant increase in the testosterone response to hCG during finasteride therapy (delta: 16.7 vs. 35.5 nmol/L) that could be explained, at least in part, by the reduction of testosterone metabolism resulting from the blockage induced by finasteride. The decrease in the androstenedione to testosterone and estrone to estradiol ratios observed after hCG treatment, however, strongly suggests increased activity of the <em>17</em>-<em>ketosteroid</em> reductase enzyme and an improvement of the testicular capacity for testosterone production.

Nowadays there are increasing experimental and clinical data indicating an important role of an endocrine system (especially its neuroendocrine part and sex hormones) in the pathogenesis of epilepsy. The relationships between patomechanisms of epilepsy and activity of hypothalamo-pituitary-ovarian axis in animals and humans are quite well recognized but the role of male sex hormones, i.e androgens, in seizure susceptibility processes is less known. Epidemiological data clearly show that adrogens-related disorders occur more frequently in epileptic men than in general male population. Usually, they appear in the form of hypogonadism associated with low levels of plasma free testosterone and with low excretion of its <em>17</em>-<em>ketosteroid</em> metabolites in the urine. Reproductive and sexual disorders can be attributed to hypogonadism. Androgen abnormalities in epileptics men are often affected by chronically used anti-epileptic drugs. Antiepileptic drugs, particularly classical ones, substantially modify bioavailability of androgens and can inhibit the activity of hypothalamo-pituitary-testicular axis, and--in a consequence--aggravate hypogonadism. Since neuroactive androgens cross the blood-brain barrier and modify seizure susceptibility, changes in their plasma concentrations can affect the course and clinical outcome of epilepsy. Effects of testosterone on seizures seem to depend on its different metabolic pathways. Aromatization of testosterone leads to formation of <em>17</em>beta-estradiol that is believed to have proconvulsive activity. Activation of 5alpha-reductase pathway leads to formation of <em>ketosteroid</em> metabolites, primarily andosterone and etiocholanolone that demonstrate the ability to prevent convulsions in majority of animal studies. Recently, it has been shown that androsterone enhances the antiepileptic activity of phenobarbital, carbamazepine, and gabapentin in animal model of epilepsy. Antiepileptic activity of testosterone and its metabolites encourage further investigation of androgens as promising candidates for treatment of epilepsy in men with androgens-related disorders.

The patient, diagnosed as a case of testicular feminisation in infancy, was examined at the age of 15 years because of severe symptoms of virilising puberty with poor breast development. Plasma steroid analyses revealed a 10-fold elevated androstenedione concentration (A: 1562 ng/100 ml). Testosterone (T: 266 ng/100 ml) was in the male pubertal range. Thus the A/T-ratio was far above normal. The oestrone/oestradiol ratio was also elevated (Oe1/Oe2: 10.2/2.2 ng/100 ml). A, T, Oe1 and Oe2 could not be suppressed by dexamethasone, but reacted promptly to fluoxymesterone (A: 781 ng/100 ml). hCG caused a further increase of the A/T-radio (2220/246 ng/100 ml); ACTH did not alter the A-concentration. These findings together with simular investigations after gonadectomy suggest that the failure to convert A to T and Oe1 to Oe2 is essentially located in the testes. In vitro incubations of testicular tissue showed reduced <em>17</em>-<em>ketosteroid</em> reductase activity in tissue slices and in the subcellular fractions microsomes and cytosole. This form of male pseudohermaphroditism can easily be detected already in infancy, if steroid analyses and stimulation tests are performed. In case of female sex assignment patients should be submitted to early orchidectomy in order to avoid virilisation in puberty.

OBJECTIVE

The aim of this study was to investigate the possible effect of prolactin on the metabolism of androgens and estrogens in patients with prolactinoma. To accomplish this, prolactin, urinary androgen, and estrogen metabolite levels were determined. In order to indirectly evaluate the possible involvement of enzymes, the concentration ratios of precursor metabolite to product metabolite were also compared with controls.

METHODS

Urine samples were obtained from 27 female patients with prolactinoma (macro, micro, and idiopathic) and from 31 age-matched normal female subjects. Urinary metabolites of 21 androgens and corticoids and 20 estrogens were analyzed by a gas chromatography-mass spectrometry system.

RESULTS

In patients with prolactinoma, urinary <em>17</em>-<em>ketosteroids</em>, and all estrogen metabolite concentrations were elevated. The ratios of delta 5/delta 4-steroids and 5 beta/5 alpha-hydrogensteroids were higher in the patients with prolactinoma than in normal female controls. In addition, no significant differences between patients and controls were observed in the precursor metabolite to product metabolite ratios relating to estrogen metabolism.

CONCLUSIONS

Our data suggests that enhanced PRL levels have a direct effect on urinary steroid secretion and metabolism, probably due to lowered activities of 5 alpha-reductase and 3 beta-hydroxysteroid dehydrogenase in the patients with prolactinoma.

The aldo-keto reductase (AKR) superfamily comprises NAD(P)H-dependent enzymes that catalyze the reduction of a variety of carbonyl compounds. AKRs are classified in families and subfamilies. Humans exhibit three members of the AKR1B subfamily: AKR1B1 (aldose reductase, participates in diabetes complications), AKR1B10 (overexpressed in several cancer types), and the recently described AKR1B15. AKR1B10 and AKR1B15 share 92% sequence identity, as well as the capability of being active towards retinaldehyde. However, AKR1B10 and AKR1B15 exhibit strong differences in substrate specificity and inhibitor selectivity. Remarkably, their substrate-binding sites are the most divergent parts between them. Out of 27 residue substitutions, six are changes to Phe residues in AKR1B15. To investigate the participation of these structural changes, especially the Phe substitutions, in the functional features of each enzyme, we prepared two AKR1B10 mutants. The AKR1B10 m mutant carries a segment of six AKR1B15 residues (299-304, including three Phe residues) in the respective AKR1B10 region. An additional substitution (Val48Phe) was incorporated in the second mutant, AKR1B10mF48. This resulted in structures with smaller and more hydrophobic binding pockets, more similar to that of AKR1B15. In general, the AKR1B10 mutants mirrored well the specific functional features of AKR1B15, i.e., the different preferences towards the retinaldehyde isomers, the much higher activity with steroids and ketones, and the unique behavior with inhibitors. It can be concluded that the Phe residues of loop C (299-304) contouring the substrate-binding site, in addition to Phe at position 48, strongly contribute to a narrower and more hydrophobic site in AKR1B15, which would account for its functional uniqueness. In addition, we have investigated the AKR1B10 and AKR1B15 activity toward steroids. While AKR1B10 only exhibits residual activity, AKR1B15 is an efficient <em>17</em>-<em>ketosteroid</em> reductase. Finally, the functional role of AKR1B15 in steroid and retinaldehyde metabolism is discussed.

In our studies on patients with head injury, it was noted that there are some correlations between their clinical courses and the urinary excretion of creatine (cr), creatinine (Crn), <em>17</em>-<em>ketosteroid</em> and <em>17</em>-hydroxycorticosteroid. We observed the high urinary excretion of Cr in patients with severe head injury while almost negative in a mild case. We reported those facts in 1974. Also noted in patients with head injury is the relationship between the enzyme-activities (GOT, GPT, LDH and CPK) in the cerebrospinal fluid and their clinical courses. In this paper, we reported 34 cases of head injured patients (simple type: 2, concussion: 9, contusion: 8, acute intracranial hematoma: 7 and chronic intra-cranial hematoma: 8). The control values of CSF enzyme-activities were determined in these 14 cases (simple head injury, whip-lash injury and osteoma of the skull) as GOT less that 15, GPT less than 7, LDH less than 12 and CPK less than 8 units. In the moderate cases, a slight increase in activities of 4 enzymes in CSF were observed, while in severe or comatose cases, the enzyme-activities (especially LDH and CPK) were greater than in the controls. In the dead cases these values were five times as high as the normal case. In the patients recovering from a serious stage, these activities decreased to normal. High CSF enzyme-levels tend to indicate a poor prognosis and low levels a favorable progrosis. In the patients with a significant elevation of CSF enzymes, a high urinary excretion of Cr [normal range: 0-150 (ca. 50)mg/day] was often observed. There was no apparent correlation between the enzyme level in CSF and that in serum and the increase or decrease of these 4 enzymes are not always proprotionate with each other. As reported by Green (1958) and Lending (1961), cerebral cell necrosis and increased permeability of BLB, BBB or cerebral cell membrane can be related to the increase of enzymeactivities. With these observations, it can be considered that severe head injury gives influence on metabolic function in the hypothalamus and may cause in the levels of CSF enzymes and/or the urinary excretions of Cr, Crn and corticosteroids. And the examinations of enzyme activities in the patients with head injury may become a useful aid to make an outlook of their clinical coure and prognosis.

1. Urine samples were collected from four men before and during test cold exposures in Melbourne, Australia, and Mawson, Antarctica. Changes in the response of body temperature to the test exposures showed that the men had acclimatized to cold at Mawson.2. Excretion rates of <em>17</em>-hydroxycorticosteroids and <em>17</em>-<em>ketosteroids</em> were significantly greater at Mawson than in Melbourne, in both the pre-exposure and exposure periods.3. Excretion rates of noradrenaline, adrenaline, sodium, potassium and creatinine did not differ significantly between Mawson and Melbourne, nor did urine flow rates.4. During the cold exposure significant increases occurred, to the same extent at Mawson as in Melbourne, in urine flow rate and in all measured urinary constituents except creatinine.

Eleven analytes were presented in the Urine Chemistry Survey Program during the year 1977. These included amylase, calcium, phosphorus, sodium, potassium, creatinine, osmolality, <em>17</em>-<em>ketosteroids</em>, total protein, glucose, and urea nitrogen. More than 450 laboratories participated. Deficiencies in assay performance were found for amylase, <em>17</em>-<em>ketosteroids</em>, and total protein. Sharp shifts in precision performance were seen in urea nitrogen analysis. Severe biases are apparent in glucose analysis using neocuproine and ferricyanide methods.

A pilot survey program for urinary chemistry was implemented by the College of American Pathologists during the year 1976. The nine analytes selected for the program were amylase, calcium, phosphorus, sodium, potassium, creatinine, osmolality, <em>17</em>-<em>ketosteroids</em> and total protein. Data obtained from more than 230 enrolled laboratories indicate deficiencies in assay performance for amylase, <em>17</em>-<em>ketosteroids</em> and total protein as measured by consensus evaluation. Acceptable performance was apparent for the remaining analytes.

The zonal testis of the dogfish (Squalus acanthias) has proven advantageous to study biochemical changes in relation to stage of spermatogenesis, including information on steroidogenic enzymes and steroid receptors. To investigate whether sulfotransferase is part of a mechanism regulating the availability of biologically active hormone in close proximity to receptors, we measured in vitro conversion of [3H]estrone (E1) to sulfoconjugated metabolites in cytosolic subfractions of testes grossly dissected according to germ cell composition (premeiotic-PrM, meiotic-M, and postmeiotic-PoM stages). Assays were carried out in the presence of adenosine 3'-phosphate 5'-phosphosulfate (PAPS) at 22 degrees C and optimized for time (60 min) and protein (500 micrograms/ml). Michaelis-Menten kinetics and saturation analysis gave the following reaction constants for [3H]E1: Km = 0.33 microM, Vmax = 2.5 pmol/min/mg; and for PAPS: Km = 33 microM, Vmax = 1.1 pmol/min/mg; competition studies carried out in the absence or presence of 1- or 5-fold excess radioinert steroids indicated that estrogen (E2>> E1) as well as androgens (T = DHEA>> 5 alpha dihydrotestosterone, DHT) were effective inhibitors. Sulfotransferase activity was found to be stage-related, being highest in PoM regions (2.31 +/- 0.24 pmol/min/mg protein) when compared to M and PrM regions (1.22 +/- 0.22 and 1.28 +/- 0.21 pmol/min/mg protein, respectively). Sulfoconjugation and the intratesticular distribution of steroid sulfates were also measured in vivo by perfusion of the intact testis with [3H]androgen or -estrogen. The pathway of blood flow via the genital artery was epigonal organ->>PoM->>M->>PrM (mature->>immature). Perfused [3H]E2, T, and DHT were all extensively metabolized in a one-pass, 1 hr perfusion, less than 10% of perfused [3H] steroid being recovered from testicular tissues as unchanged steroid. In general, recovery of polar metabolites was greater than non-polar metabolites from all three substrates. Sequential hydrolysis with glucuronidase and glusulase indicated that sulfoconjugation is a minor component (< 20%) of several "inactivating" pathways, which include glucuronide conjugation, <em>17</em>-<em>ketosteroid</em> synthesis, and pathways leading to unidentified polar metabolites. No consistent stage-related distribution patterns were observed for any of the metabolite subfractions; however, total recovered radioactive steroid (polar plus non-polar) formed a decreasing concentration gradient from point of entry of perfusate (PoM region) to point of exit (PrM region). These data support the conclusion that access to receptors by steroid ligands may be controlled by a balance between activating and inactivating pathways.(ABSTRACT TRUNCATED AT 400 WORDS)

Cytochrome P450IIC5 (rabbit liver 21-hydroxylase) is unusual among hepatic forms of cytochromes P450 because it catalyzes the conversion of one active steroid hormone (progesterone) to another active hormone (deoxycorticosterone). Another interesting aspect of this steroid-hydroxylating enzyme is the ability to convert delta 5-3 beta-hydroxysteroids to the delta 4-3-<em>ketosteroid</em> configuration. The delta 5-3-beta-hydroxysteroid, pregnenolone, was readily 21-hydroxylated, and this product was further metabolized to the delta 4-3-<em>ketosteroid</em>, deoxycorticosterone. It is suggested that the mechanism of this cytochrome P450-mediated, 3 beta-hydroxysteroid dehydrogenase/delta 5----4 isomerase-like reaction is through a gem-diol formation. In this study, COS-1 cells were transfected with the plasmid encoding cytochrome P450IIC5 to express a functional enzyme within the cell milieu. Transfected COS cells preferentially metabolize pregnenolone compared with all other steroids tested. Progesterone and <em>17</em> alpha-hydroxypregnenolone are also 21-hydroxylated, whereas <em>17</em> alpha-hydroxyprogesterone is a poor substrate. Substrate preference of this 21-hydroxylase differs from that seen with bovine adrenal P450XXIA1 (formerly P450C21) hydroxylase. Additionally, this study demonstrated that C19 steroids, like dehydroepiandrosterone and androstenedione, are hydroxylated at the 16 alpha position. Contrary to previous reports, no metabolite of estradiol-<em>17</em> beta was detected, presumably due to the unstable nature of catechol estrogens (2-hydroxyestradiol).

The properties of 5-ene-3 beta hydroxysteroid oxidoreductase (3 beta-HSD) from human placental homogenates were studied in vitro. The apparent Michaelis constants for 3 beta-HSD with the substrates pregnenolone (delta 5P) and dehydroepiandrosterone (DHA) were <em>17</em>0 nM and nM respectively. The optimal pH for both these substrates was between 10 and 12. With NAD as the substrate, the Km for the pregnenolone was 20 microM and for DHA, <em>17</em> microM. The activity of 3 beta-HSD was inhibited by various steroids. Competitive inhibitors (pregnenolone substrate) included: ethynylestradiol (inhibition constant Ki=7.3 nM), DHA (Ki=46 nM), estradiol-<em>17</em> beta (Ki=46 nM), cholesterol (Ki=0.68 microM) and 16 alpha-hydroxydehydroepiandrosterone (16 alphaOHDHA) (Ki=2.2 microM). When the substrate was DHA, competitive inhibition occurred with the following steroids: ethynylestradiol (Ki=6.4 nM), estradiol-<em>17</em> beta (Ki=69 nM), pregnenolone (Ki=91 nM), cholesterol (Ki=1.3 microM) and 16 alphaOHDHA (Ki=1.9 microM). 4-Ene-3-<em>ketosteroids</em> such as androstenedione, progesterone (delta 4P), norethindrone and chlormadinone acetate acted as noncompetitive inhibitors towards both substrates.

Alternative metabolic pathways for the biosynthesis of testosterone from pregnenolone exist in mammalian testes. The following experiments were designed to identify the preferred testosterone biosynthetic pathway in rat and rabbit testes. The experimental protocol included the infusion of steroidogenic reaction inhibitors and testosterone biosynthetic intermediates into testes perfused in vitro. Under these conditions, the testicular steroid secretions were a measure of specific reaction activities. Infusion of medrogestone (6,<em>17</em>-dimethyl-4,6-pregnadiene-3,20-dione, Ayerst), an inhibitor of delta5-4isomerization, permitted the reactions converting pregnenolone to androstenediol to be studied separately from those converting progesterone to testosterone. For example, the activity of the pregnenolone vector <em>17</em>alpha-hydroxypregnenolone reaction was measured as the total of the <em>17</em>alpha-hydroxypregnenolone, dehydroepiandrosterone, and androstenediol secreted by medrogestone-inhibited testes. The reactions convering delta5-3beta-hydroxysteroids to delta4-3-<em>ketosteroids</em> were studied in testes infused with SU-10603 (7-chloro-3,4-dihydro-2-[3-pyridyl]-1(2H)-naphthalenone. Ciba-Geigy), an inhibitor of <em>17</em>alpha-hydroxylation and C-<em>17</em>, C-20 cleavage reactions. The results indicated that preferred testosterone biosynthetic pathways are present and different in rat and rabbit testes.

Plasma concentrations of free 5 alpha-androstane-3 alpha, <em>17</em> beta-diol (3 alpha-diol), 5 alpha-androstane-3 beta, <em>17</em> beta-diol (3 beta-diol), 5 alpha-dihydrotestosterone (DHT) and of testosterone (T) were determined in female rats undergoing spontaneous sexual maturation or pregnant mare's serum gonadotropin (PMSG)-induced precocious ovulation. In the prepubertal period, 3 alpha-diol levels showed an inverse relationship with uterine weights and a significant drop between Days 21 and 26 of life. Postpubertally, 3 alpha-diol concentrations tended to rise again, to a lesser degree, in parallel with increments in plasma T. Concentrations of 3 alpha-diol were increased tenfold 48 h after the administration of PMSG, i.e., at a time when T and estradiol-<em>17</em> beta also peaked. Concentrations of 3 beta-diol and DHT remained low throughout both experiments. In earlier studies, it was found that exogenous 3 alpha-diol suppresses reproductive function in the immature rat, but the high concentrations of endogenous 3 alpha-diol reported here for the PMSG model clearly do not interfere with ovulation. The observed constellations of plasma steroids are compatible with the postulate of a peripubertal shift in ovarian progesterone metabolism from 5 alpha-reduced C21 steroids to delta 4-3-<em>ketosteroids</em>.

The inhibition of human testicular testosterone production in vitro by oestrogen was studied by measuring effects of oestradiol-<em>17</em> beta and ethinyl oestradiol on the 3 beta-hydroxysteroid dehydrogenase/isomerase and the delta 4-pathway of testosterone synthesis. Progesterone, <em>17</em> alpha-hydroxyprogesterone, androstenedione and testosterone concentrations of incubation media were all depressed after oestrogen addition. To minimize differences in substrate amounts between incubations, 3H-labelled pregnenolone, progesterone, <em>17</em> alpha-hydroxyprogesterone or androstenedione was added to the incubation media. The conversion of pregnenolone, progesterone and <em>17</em> alpha-hydroxyprogesterone was inhibited by oestrogen. It is concluded that oestrogen inhibits 3 beta-hydroxysteroid dehydrogenase/isomerase, <em>17</em> alpha-hydroxylase and C<em>17</em>-20-lyase enzyme activities of human testicular tissue in vitro while <em>17</em> beta-<em>ketosteroid</em> reductase activity is unaffected.

Ovarian homogenates from 10-, 23- 128- and 60-day-old golden hamsters were incubated with [14C]-4-androstene-3,<em>17</em>-dione or [7-3H]-progesterone in the presence of NADPH and enzyme activities and metabolism of progesterone were estimated. A rapid increase in uterine weight was found around 28 days of age. The activity of 5 alpha-reductase was very high in the ovaries of 23-day-old hamsters (647 +/- 1<em>17</em> (SD) nmol/g tissue/h), high in those of 28-day-old hamsters (135 +/- 4) and low in those of 10- and 60-day-old animals (20 +/- 16, 39 +/- 11). However, the activity of 5 beta-reductase was high in all ovaries of golden hamsters at different stages of development (84-132 nmol/g tissue/h). The major C-21-<em>17</em>-hydroxysteroids and C19-steroids formed from progesterone by the ovaries of 23-day-old hamsters were 5 alpha-steroids such as 3 alpha, <em>17</em>-dihydroxy-5 alpha-pregnan-20-one and androsterone, whereas those by the ovaries of 28- and 60-day-old hamsters were 4-ene-3-<em>ketosteroids</em> and 5 beta- and 5 alpha-steroids such as <em>17</em>-hydroxy-4-pregnene-3,20-dione. 3 alpha, <em>17</em>-dihydroxy-5 alpha- and 5 beta-pregnan-20-one, 4-androstene-3,<em>17</em>-dione, testosterone and androsterone. The formation of oestradiol-<em>17</em> beta and oestrone from progesterone was found only in the ovaries of 38- and 60-day-old hamsters. These results show that similarly high levels of 5 beta-reductase are present in all ovaries from suckling, immature and adult golden hamsters and that high levels of 5 alpha-reductase are formed only in ovaries from immature hamsters, especially those with small uterus. The active 5 alpha-reduction of 4-ene-3-<em>ketosteroids</em> may be responsible for the decrease in the formation of oestrogens in immature hamster ovaries.

A 46, XY phenotypically female infant with <em>17</em>-<em>ketosteroid</em> reductase (<em>17</em>-KSR) showed normal plasma androgens for chromosomal sex shortly after birth, but did not show the physiologic testosterone rise. One intramuscular injection with human chorionic gonadotropin resulted in high ratios between androstenedione/testosterone and dehydroepiandrosterone/delta 5-androstenediol, confirming the diagnosis. In spermatic vein plasma similarly elevated ratios were found. A urinary steroid profile revealed elevated levels of metabolites of <em>17</em>-OH-progesterone and androstenedione. In vitro studies in testicular tissue showed a decreased capacity of <em>17</em>-<em>ketosteroid</em> reductase, the reduction capacity being more affected than the oxidation capacity. The activity of 3 beta-hydroxysteroid-dehydrogenase was slightly increased. The serial analysis of plasma androgens provides more insight in the natural history of <em>17</em>-<em>ketosteroid</em> reductase.

Abstract The narcotically active progesterone metabolite 3alpha-hydroxy-5alpha-pregnan-20-one (HPO) modulates gamma-aminobutyric acid (GABA) neurotransmission by direct actions on the GABA(A)-receptor complex. In the present investigation, the formation of HPO was quantified in man. Twenty-four h urine samples were collected during the night (2300 to 0700 h) and day (0700 to 2300 h) from 11 healthy subjects (31 +/- 5 years) for two consecutive days. The concentration of HPO was measured after enzymatic hydrolysis of conjugated HPO and multiple chromatographic separation steps by gas chromatography-mass spectrometry. The mean excretion rates of HPO were 14.8 +/- 11.8 mug/24 h with high inter-individual variations. There were no significant differences in the production of HPO during the night and day. It seemed unlikely that HPO was primarily formed in the adrenals or gonads since the excretion rates of HPO poorly correlated with the formation of <em>17</em>-<em>ketosteroids</em> or <em>17</em>-hydroxycorticosteroids that are widely used as indices of adrenal and gonadal steroid production. The data showed for the first time that HPO is produced in male subjects in concentrations similar to that of classical steroid hormones.

A simple three-step synthetic method is reported on the conversion of delta 4-3-<em>ketosteroids</em> to the corresponding 3 beta-hydroxy-delta 5-steroid analogues. <em>17</em> alpha-Hydroxy-4-pregnen-3,20-dione (10a) was used as a model to develop a method for the synthesis of 3 beta, <em>17</em> alpha-dihydroxy-5-pregnen-20-one (16). The major problem being the synthesis of 3,<em>17</em> alpha-diacetoxy-3,5-pregnadien-20-one (14) was solved by acetylating using a mixture of acetic anhydride and perchloric acid. The conversion of 15 beta, <em>17</em> alpha-dihydroxy-4-pregnen-3, 20-dione (8), product of Penicillium citrinum fermentation, to the desired 3 beta,15 beta,<em>17</em> alpha-trihydroxy-5-pregnen-20-one (1), is described using a modification of this method. Reaction of 8 with acetic anhydride and perchloric acid in ethyl acetate gave 3,15 beta,<em>17</em> alpha-triacetoxy-3,5-pregnadien-20-one (<em>17</em>) which on reduction with sodium borohydride gave 5-pregnen-3 beta,15 beta,<em>17</em> alpha, 20(S + R)-tetrols (18a and 18b); however, reduction of <em>17</em> with a mixture of sodium borohydride and potassium bicarbonate gave after basic hydrolysis with methanolic sodium hydroxide the desired product 3 beta,15 beta,<em>17</em> alpha-trihydroxy-5-pregnen-20-one (1) in good yield (54%).

Adreno cortical function was carried out in 43 cases of leprosy. These cases were further divided into tuberculoid, borderline, lepromatous and Lepra reaction. Serum and urinary electrolyte, urinary <em>17</em>-<em>Ketosteroid</em> and <em>17</em>-Ketogenic steroid and plasma cortisol levels were measured to assess the adrenocortical status in these different forms of leprosy. It was observed that these parameters were within normal limit in tuberculoid leprosy except low value of urinary <em>17</em>-Ketogenic steroid. The borderline and Lepromatous leprosy cases revealed low values of urinary sodium, potassium and <em>17</em>-Ketogenic steroid and high level of serum potassium. However, the cases of lepra reaction revealed low value of serum and urinary sodium and potassium, urinary <em>17</em>-Ketogenic steroid. The basal plasma cortisol level was high in this group but it was statistically insignificant.

Thirteen analytes were presented in the Urine Chemistry Survey Program during the year 1978. These included amylase, calcium, phosphorus, sodium, potassium, creatinine, osmolality, total protein, glucose, and urea nitrogen as regular constituents. Optional analytes included <em>17</em>-<em>ketosteroids</em>, <em>17</em>-ketogenic steroids, and vanillylmandelic acid. Certain samples elevated sodium, potassium, calcium, and phosphorus concentration to stimulate human disease states and to test consensus results in ranges heretofore not possible. Serious loss of precision performance was not noted although some methods are noticeably affected. Improvement in interlaboratory precision has sllowed as compared with previous years.