Three cases of gingival overgrowth induced by cyclosporin and/or nifedipine have been reported. Patient A was under medication with cyclosporin plus nifedipine, patient B with nifedipine only and patient C with cyclosporin only. The significance of androgen metabolism in gingival tissue with respect to hyperplastic changes has been studied by several workers. Hence, we have investigated whether gingival tissue from the above patients showed significant metabolism of the androgen, testosterone, to its biologically-active form, 5 alpha-dihydrotestosterone (5 alpha-DHT). Radical gingivectomy was carried out in all 3 cases to remove the hyperplastic tissue. The excised tissue was incubated with labelled testosterone in order to study the extent of androgen metabolism. Healthy gingivae from males and females produced 5 alpha-DHT (22.<em>4</em> +/- 7.7, s.e.m., n = 8). Very significantly higher values (p less than 0.001) were recorded for patients A, B and C (1139, 5<em>4</em>2 and 99<em>4</em> fmol/mg, respectively). These represented increases of 51-, 2<em>4</em>- and <em>4</em><em>4</em>.<em>4</em>-fold, respectively over control values. Corresponding production of <em>4</em>-<em>androstenedione</em> from testosterone was 28 +/- 8.3, s.e.m., n = 8, fmol/mg. In patients A, B and C, <em>4</em>-<em>androstenedione</em> production was elevated: 85, 901 and 113 fmol/mg, respectively, representing increases of 3-, 32- and <em>4</em>-fold. Even the lower values of 85 and 113 fmol/mg were very highly significant (p less than 0.001) compared with control values. Although healthy female gingival tissue does not metabolize testosterone significantly, in the presence of inflammation the extent of 5 alpha-DHT formation is comparable to that of male samples.(ABSTRACT TRUNCATED AT 250 WORDS)

BACKGROUND

The aim of the present study was to investigate the steroidogenic response pattern to HCG in obese women with polycystic ovary syndrome (PCOS) and the possible effects of metformin treatment on it.

METHODS

A single injection of human chorionic gonadotrophin (HCG, 5000 IU) was given to 12 obese [body mass index (BMI)>> or = 27 kg/m2] women with PCOS and to 27 control women. Blood samples for assays of 17alpha-hydroxyprogesterone (17-OHP), <em>androstenedione</em>, testosterone and oestradiol were collected at baseline and 1, 2 and <em>4</em> days after the injection. Responses to HCG were also assessed in the PCOS women after 2-month treatment with metformin (500 mg x 3 daily).

RESULTS

Serum 17-OHP and oestradiol concentrations peaked at 2<em>4</em> h in the PCOS women and preceded the maximum testosterone concentration, which was seen at <em>4</em>8 h. In the control women the maximum concentrations of all these steroids were reached 96 h after HCG. After metformin treatment, the basal serum testosterone concentration and the areas under the <em>androstenedione</em> (AUC(A)) and testosterone (AUC(T)) response curves after HCG decreased significantly.

CONCLUSIONS

The results demonstrate that obese PCOS women have a male-type steroidogenic response pattern to a single injection of HCG and a higher androgen secretory capacity than control women, which may be explained by the increased thecal cell activity in the polycystic ovary. The slight alleviation of hyperandrogenism brought about by metformin therapy appears to be due to its effect on ovarian steroidogenesis possibly mediated by decreased insulin action.

Acne is known to be one of the features of hyperandrogenism. The aim of the present work was to study women with persistent acne and without other evidence of hyperandrogenism, such as hirsutism, alopecia, or irregular menses. Among 87 female patients with acne and/or hirsutism, we defined three groups: group 1 (n = 29), patients having treatment-resistant acne without menstrual disturbance, alopecia, or hirsutism; group 2 (n = 27), patients with acne and hirsutism; and group 3 (n = 31), patients with hirsutism alone. Clinical chemistry criteria for hyperandrogenism were based on elevated values of one or more of the following parameters: plasma testosterone, delta-<em>4</em>-<em>androstenedione</em>, dehydroepiandrosterone, urinary 5 alpha-androstane 3 alpha-17 beta-diol, and 17-ketosteroids (with chromatography). Plasma and urine samples were drawn between the 18th and 25th days of the cycle. Among group 1 patients, we found 25 subjects (86%) with hyperandrogenism, according to these laboratory criteria. The etiologies were: polycystic ovary syndrome (36%), adrenal hypersecretion (<em>4</em>0%, of which 12% showed secondary polycystic ovaries), isolated increase in 5 alpha-androstane 3 alpha-17 beta-diol (20%), and hyperandrogenism without diagnosis (<em>4</em>%). The parameters were found to be more elevated in these patients than in a control group of 30 normal volunteer women. In groups 2 and 3, the findings were essentially the same as in group 1, except for increased levels of testosterone and the testosterone/SHBG ratio. Furthermore, it was evident that persistent acne may be an isolated sign of hyperandrogenism.

Megestrol acetate (Megace), an antiandrogen, was administered in a dosage of 80 mg daily to 6 patients with benign prostatic hypertrophy (BPH) for <em>4</em> to 25 days prior to transurethral resection of the prostate (TURP). Surgical tissue from drug-treated patients was compared to untreated controls in regard to: 1) the enzymatic reduction of testosterone (T) and dihydrotestosterone (DHT); 2) DHT binding to a cytosol receptor protein; 3) tissue levels of endogenous dihydrotestosterone and androstanediols (diols). When minced prostate was incubated with 3H-T and 1<em>4</em>C-<em>androstenedione</em> for 1 h at 37 C, prostate 5alpha-reductase activity, measured as reduced products formed from substrate, decreased to 31% and 39%, respectively, of the control values. Prostate 3-oxido-reductase enzyme activity, measured as diols formed from 3H-DHT, was decreased to neglible values in Megace-treated patients compared to an 8.7% conversion to diols in controls. No 3H-DHT binding to a cytosol receptor protein could be demonstrated in <em>4</em> out of 5 prostates from Megace-treated patients, whereas the presence of such a receptor was noted in 1<em>4</em> out of 17 untreated controls. Endogenous DHT levels in Megace-treated patients averaged 1.1 ng/g (SE = 0.26), significantly less than the average of 3.9 ng/g (SE = 0.<em>4</em>9) found in controls (P less than 0.001). No significant difference was noted in endogenous diols. In addition to these effects on tissue, Megace significantly decreased plasma levels of T, LH, and FSH at the end of the <em>4</em>- to 25-day period; plasma prolactin levels did not change. Continued studies of Megace for the possible treatment of benign prostatic hypertrophy may be warranted since the drug appears to block several important biochemical steps which mediate the effects of androgen on the human prostate.

Infusion of dexamethasone into chronically catheterized foetal kids induced delivery in <em>4</em>1--65 h. Changes in the concentrations of placental and ovarian steroids in the maternal circulation at dexamethasone-induced delivery mimicked those preceding spontaneous kidding at term; in both instances the peripheral concentration of progesterone fell and the concentration of oestradiol-17beta rose. The concentration of cortisol in the foetus was low at dexamethasone-induced delivery. Metabolism of pregnenolone, progesterone, 17alpha-hydroxyprogesterone and androst-<em>4</em>-ene-3,17-dione by extracts of foetal placenta was investigated in late pregnancy, after premature parturition induced with dexamethasone or prostaglandins and after spontaneous parturition at term. In placenta obtained before the onset of labour (or from animals induced to kid by administration of prostaglandins), the main product of progesterone metabolism was a 5beta-pregnane-3,20-diol. In contrast, placentae from animals in which the foetal level of glucocorticoid had been raised (after spontaneous parturition or by administration of dexamethasone to the foetus) were able to 17alpha-hydroxylate and progesterone was metabolized to 5beta-pregnane-3alpha/3beta,17alpha,20alpha-triols and 17alpha,20alpha-dihydroxypregn-<em>4</em>-en-3-one. The appearance of placental 17alpha-hydroxylase was correlated with raised maternal concentrations of 17alpha,20alpha-dihydroxypregn-<em>4</em>-en-3-one and <em>androstenedione</em>. The induction or activation of placental 17alpha-hydroxylase may represent the mechanism by which foetal glucocorticoid controls the onset of labour in the goat.

We previously reported that tributyltin chloride (TBT) and triphenyltin chloride (TPT) powerfully suppressed human chorionic gonadotropin- and 8-bromo-cAMP-stimulated testosterone production in pig Leydig cells at concentrations that were not cytotoxic [Nakajima Y, Sato Q, Ohno S, Nakajin S. Organotin compounds suppress testosterone production in Leydig cells from neonatal pig testes. J Health Sci 2003;<em>4</em>9:51<em>4</em>-9]. This study investigated the effects of these organotin compounds on the activity of enzymes involved in testosterone biosynthesis in pig testis. At relatively low concentrations of TPT, 17beta-hydroxysteroid dehydrogenase (17beta-HSD; IC(50)=2.6microM) and cytochrome P<em>4</em>50 17alpha-hydroxylase/C(17-20) lyase (IC(50)=117microM) activities were inhibited, whereas cholesterol side-chain cleavage cytochrome P<em>4</em>50 and 3beta-HSD/Delta(<em>4</em>)-Delta(5) isomerase activities were less sensitive. Overall, TPT was more effective than TBT. TPT also inhibited both ferredoxin reductase and P<em>4</em>50 reductase activities at concentrations over 30microM; however, TBT had no effect, even at 100microM. The IC(50) values of TPT were estimated to be 25.7 and 22.8microM for ferredoxin reductase and P<em>4</em>50 reductase, respectively. The inhibitory effect of TPT (30microM) on microsomal 17beta-HSD activity from pig testis was eliminated by pretreatment with the reducing agents dithiothreitol (1mM) and dithioerythritol (1mM). On the other hand, TPT (0.03microM) or TBT (0.1microM) exposure suppressed the testosterone production from <em>androstenedione</em> in pig Leydig cells indicating that these organotins inhibit 17beta-HSD activity in vivo as well as in vitro, and the IC(50) values of TPT and TBT for 17beta-HSD activity were estimated to be <em>4</em>8 and 11<em>4</em>nM, respectively. Based on these results, it appears possible that the effects of TBT and TPT are largely due to direct inhibition of 17beta-HSD activity in vivo.

The objective of this work was to evaluate the effects of testosterone (T) and 17beta-estradiol (E(2)) on coronary microvascular endothelial cells (CMECs) of male and female rats. To analyze the short-term effects of such sex steroid hormones on intracellular Ca(2+) concentration ([Ca(2+)](i)) kinetics, we used the chelating agent fura-2 acetoxymethyl ester. We also explored the possibility of testosterone aromatization by using selective inhibitors of the aromatase enzyme cytochrome P-<em>4</em>50 aromatase (P<em>4</em>50(arom)), aminoglutethimide (<em>4</em> microM), and <em>4</em>-hydroxy<em>androstenedione</em> (<em>4</em> microM). The presence of P<em>4</em>50(arom) was investigated by immunocytochemical and immunoblot assays using peptide-generated polyclonal antibodies raised against a 20-amino acid synthetic fragment of rat P<em>4</em>50(arom) and by in situ hybridization to locate the aromatase mRNA in such cells. The activity of P<em>4</em>50(arom) was demonstrated by the stereospecific loss of the tritium atom of [1beta-(3)H]<em>androstenedione</em>. Our results indicate that both T and E(2) induced a rapid increase in [Ca(2+)](i). The fact that the effects of E(2) and T were carried out within milliseconds suggests that they were exerted at the membrane level and not through intracellular receptors. The possibility of involvement of PLC-beta in these effects is suggested because U-73122 (a PLC inhibitor) blocked the effects of both T and E(2). Immunocytochemical assays indicated the expression of androgenic and estrogenic receptors in these cells. The effects of T were blocked by the selective aromatase inhibitors. We also demonstrated membrane association of P<em>4</em>50(arom), expression of the ovary-specific mRNA after in situ hybridization, and E(2) formation resulting from a significant activity of P<em>4</em>50(arom) in CMECs. There were no gender-based differences.

Cortisol and steroids with progestational or androgenic activity were studied to determine the effects of these steroids on the conversion of <em>androstenedione</em> (A) to estrone (E1) in human cultured breast carcinoma cells. Cortisol (10(-6) M) stimulated aromatase activity in two estrogen unresponsive cell lines (MD, DM) and in an estrogen responsive cell line (MCF7) with the maximum stimulation occurring during confluence. Cortisol inhibited the replication of MCF7 cells but not MD and DM. Dihydrotestosterone, androsterone and 5 alpha-androstanedione (10(-6) M) inhibited the conversion of A to E1 by greater than 90% under basal and cortisol stimulated conditions. Progesterone (10(-6) M) had no effect on aromatase activity while the progestational agent R5020 (10(-6) M) produced a 30% inhibition. The anabolic steroids 19-nortestosterone and 19-nor<em>androstenedione</em> which also have progestational activity inhibited the conversion of A to E1 in a dose dependent manner with 90% inhibition at 10(-6) M. Danazol (10(-6) M) a drug with both androgenic and progestational activity inhibited E1 formation by 30%. Under the same conditions, the known inhibitor of aromatase, <em>4</em>-hydroxy<em>androstenedione</em> (10(-6) M) decreased E1 formation by more than 90% and aminoglutethimide (10(-6) M) caused only 25% inhibition. These studies demonstrate that endogenous and exogenous steroids may have significant effects in modulating the local formation of estrogens from androgen precursors in cultured breast carcinoma cells. This effect on estrogen formation may be a factor in the biological response of breast tissue.

It is well known that the larynx is a target organ for androgens and the cancer of larynx is more frequent in male subjects. We have evaluated the androgen receptors (AR) in the cytosol (ACR) and in salt extractable (ANR) and salt resistant nuclear fraction (AMR) in a group of 2<em>4</em> male patients with cancer of the larynx surgically removed. In addition specimens obtained from the normal mucosa of the same subjects were analyzed. In 5 patients estrogen (ER) and progesterone (PgR) receptors were also assayed. In all subjects blood samples were taken before surgery for the assay of the following hormones: LH, FSH, estradiol, testosterone, dihydrotestosterone, delta <em>4</em>-<em>androstenedione</em>, dehydroepiandrosterone sulfate, cortisol. The results observed showed that 18 out of 2<em>4</em> normal larynx mucosa specimens and 17 out of 2<em>4</em> larynx cancer specimens were positive for ACR or ANR or AMR. The 5 samples of normal and cancer tissues analyzed for ER and PgR were negative. In conclusion there is no significant correlation between AR positivity from one size, histology, degree of differentiation and invasivity of the cancer, age of patients and hormonal blood levels from the other. The high ANR and AMR positivity (normal hormonal translocation and binding on DNA acceptors) confirm that the normal and cancer larynx are target tissue for androgens and establish the hormone dependence of this cancer. Hormonal therapy could be envisaged as an alternative or a complementary therapy for this type of cancer at least in the cases in which the analysis of hormone receptors will prove to be positive.

OBJECTIVE

To describe the presentation and fertility sparing treatment of a young woman found to have a steroid cell tumor not otherwise specified (NOS) and her spontaneous pregnancy and delivery shortly after surgery.

METHODS

A 20-year-old Hispanic female presented with hirsuitism, virilization, and elevated androgen levels (testosterone 328 ng/dL) and was wrongly diagnosed with polycystic ovarian syndrome. Four months later she sought a second opinion. Her androgens were as follows: testosterone level 485 ng/dL, androstenedione 1,738 ng/dL and DHEA 1,459 ng/dL. She had normal levels of progesterone, estradiol, and DHEA-SO4. On transvaginal ultrasound she had a solid-appearing right ovarian mass. She underwent fertility sparing surgery with a laparoscopic right oophorectomy.

RESULTS

Gross and histological pathology confirmed a benign steroid cell tumor NOS. She had rapid normalization of all androgens 13 days after surgery. She had spontaneous resumption of menses 4 months later. She conceived despite using emergency contraception approximately 9 months following surgery and delivered a healthy boy at term without complication.

CONCLUSIONS

Prompt evaluation for an androgen producing tumor should be performed when testosterone levels are greater than 200 ng/dL. Pregnancy following removal of this rare tumor has not previously been reported.

BACKGROUND

Differences in circulating steroid hormone levels have been hypothesized to explain ethnic differences in steroid-related diseases. The aim of this study was to determine the serum levels of a wide panel of steroid hormones, both androgens and estrogens, in healthy middle-aged African-Caribbean and European men.

METHODS

Serum steroid hormone levels were determined in men participating in a systematic public health study funded by the French National Health Insurance system. Blood was collected in the morning from 30<em>4</em> healthy African-Caribbean and European men aged between <em>4</em>0 and 69 years. Serum steroids were measured by mass spectrometry-gas chromatography, except for DHEAS and sex hormone-binding globulin, which were determined by RIA. Data were analyzed in 10-year age intervals by analysis of covariance, with adjustment for age, body mass index, waist-to-hip ratio, tobacco and alcohol consumption, and season of sampling.

RESULTS

Compared with Europeans, African-Caribbean men presented significantly higher serum levels of measured bioavailable testosterone, <em>4</em>-<em>androstenedione</em> (<em>4</em>-dione), and estrone (E1) regardless of the age group, of 5-androstenediol (5-diol) in those aged <em>4</em>0-<em>4</em>9 and 50-59 years, and of testosterone (TT) and dihydrotestosterone in those aged <em>4</em>0-<em>4</em>9 years. In contrast, European men aged <em>4</em>0-69 years showed significantly higher serum levels of DHEA and DHEAS.

CONCLUSIONS

Significant differences in serum steroid hormone levels were observed in middle-aged African-Caribbean and European men. Whether such differences could contribute to ethnic differences in disease risk in adult men remains to be investigated. Some steroids, such as bioavailable TT, <em>4</em>-dione, 5-diol, and E1, deserve particular attention.

The role of 5 alpha-reduction of testosterone in the inhibition of LH secretion was investigated in rat anterior pituitary cell cultures. Pituitary cells were preincubated with testosterone or dihydrotestosterone (17 beta-hydroxy-5 alpha-androstan-3-one) for 17 h and then with LHRH for an additional <em>4</em> h. Dihydrotestosterone was 6-fold more potent than testosterone in the inhibition of LHRH-induced LH release. Basal LH secretion was not affected by either androgen. The inhibition curves of testosterone and dihydrotestosterone were not shifted by the presence of the 5 alpha-reductase inhibitors 17 beta-N,N-diethylcarbamoyl-<em>4</em>-methyl-<em>4</em>-aza-5 alpha-androstan-3-one (<em>4</em>-MA) and 17 beta-N,N-diisopropylcarbamoyl-<em>4</em>-aza-androstan-3-one (DIPA). Neither <em>4</em>-MA nor DIPA alone had an effect on either basal or LHRH-induced LH release. When pituitary cells were incubated with [3H]testosterone for 17 h, the radioactivities were found to be unmetabolized testosterone (66.9 +/- 2.<em>4</em>%), dihydrotestosterone (13.3 +/- 0.5%), <em>androstenedione</em> (15.9 +/- 1.3%), 5 alpha-androstane-3,17-dione (2.8 +/- 0.3%), and 3 alpha (beta), 17 beta-androstanediol (less than 1%). In the presence of <em>4</em>-MA or DIPA, 5 alpha-reduction of testosterone was completely inhibited; <em>androstenedione</em> was the only metabolite. <em>Androstenedione</em> was only 12% as potent as testosterone in the inhibition of LHRH stimulation of LH release, and conversion of [3H]<em>androstenedione</em> to testosterone and dihydrotestosterone did occur in these cells. When [3H]dihydrotestosterone was incubated with pituitary cells, the radioactivities were dihydrotestosterone (6<em>4</em>.<em>4</em> +/- 0%), 5 alpha-androstanedione (19.3 +/- 1%), 3 alpha (beta), 17 beta-androstanediol (7.7 +/- 1.7%), and unknown polar metabolites. <em>4</em>-MA and DIPA had no effect on the metabolism of dihydrotestosterone. These results indicate that both testosterone and dihydrotestosterone inhibit LHRH-induced LH release and that this activity of testosterone does not depend on its 5 alpha-reduction.

OBJECTIVE

The origin of oestrogens in men is only partly understood. From infusion studies with radioactively labelled hormones, we know that oestradiol (E2) and oestrone (E1) are either directly secreted by the testes and adrenal glands or peripherally produced from testicular or adrenal androgens.

METHODS

We determined E2, E1, androstenedione, testosterone and dehydroepiandroster-one sulphate (DHEAS) in 292 elderly men and 367 postmenopausal women. We considered post-menopausal women as men without testes, assuming that the postmenopausal ovary is not endocrinologically active and that the testes do not contribute to circulating levels of DHEAS. Subjects were stratified by DHEAS levels to adjust for differences in DHEAS levels between sexes. For men and women separately, mean levels of E2, E1, androstenedione and testosterone were calculated per DHEAS stratum. The relative direct and indirect contributions of the testes to steroid levels in men were calculated by the formula [(C(m) - C(f))/C(m)] x 100%, in which C(m) and C(f) represent the mean concentrations of the steroid in men and women respectively.

RESULTS

The relative contributions (%) of the testes to hormone levels per DHEAS stratum (<2, 2-4, 4-6 and >6 micromol/l) respectively were, for E2, 72%, 60%, 52% and 44%; for E1, 54%, 47%, 35% and 34%; for androstenedione, 14%, 4%, 12% and 0%; and, for testosterone, 88%, 88%, 87% and 83%.

CONCLUSIONS

We conclude that in elderly men dependent on DHEAS levels, 44-72% of E2 and 34-54% of E1 originate directly or indirectly from the testes.

Several studies have now documented the existence of IGFBPs in follicular fluid and their correlation with the health of the follicle. In particular, increased levels of IGFBP-<em>4</em> have been reported in androgen-dominant atretic follicles and those from polycystic ovaries. The aim of this study was to elucidate the role of IGFBP-<em>4</em> in ovarian steroidogenesis. Granulosa cells and theca tissue were incubated with or without LH or FSH in the presence or absence of IGFBP-<em>4</em> (0.5-50 ng/ml). Inhibition by IGFBP-<em>4</em> of estradiol production in the presence of testosterone alone was seen in three of four experiments. IGFBP-<em>4</em> completely inhibited FSH-stimulated estradiol production in three experiments and caused 67% inhibition in a fourth. Similar results were obtained for theca, in which concurrent incubation with IGFBP-<em>4</em> completely negated the stimulatory effects of LH on <em>androstenedione</em> production. The mechanism by which IGFBP-<em>4</em> exerts these potent effects and the possibility that this may by IGF-independent are currently being investigated.

The possible existence of correlations between bone mineral content (BMC), age and serum levels of steroid hormones was investigated. It was found that dehydroepiandrosterone sulphate (DHEA-S), oestradiol (E2) and delta-<em>4</em>-<em>androstenedione</em> (A) were correlated with BMC, whereas oestrone (E1) and testosterone (T) were not. Partial correlations after adjustment for age were significant (P less than 0.05) only between E2 and DHEA-S and BMC at the L2-L<em>4</em> lumbar site (BMC-1) and between DHEA-S (P less than 0.01) and BMC at the midradius site (BMC-r). Stepwise multiple regression analysis showed that, apart from age, E2 was the only factor to fit (P less than 0.05) into the mathematical model with BMC-1 as the dependent variable, while DHEA-S was the only factor to fit (P less than 0.01) with BMC-r as the dependent variable. These data suggest that different hormonal influences are related to BMC at different sites, namely E2 to lumbar trabecular bone (L2-L<em>4</em>) and DHEA-S to cortical bone (midradius).

OBJECTIVE

Peripheral aromatization of testosterone and androstenedione is the principal source for circulating oestrogens in men and in castrated and post-menopausal women. Since human bone is a target organ for androgens and oestrogens, aromatase activity was assessed in human spongiosa obtained from patients who were undergoing orthopaedic surgery.

METHODS

In initial experiments for assessing aromatization, oestrogen formation from 1,2,6,7-3H-androstenedione was compared with the release of tritiated water from 1 beta-3H-androstenedione. Since the rates of enzyme activity were similar with the two methods, rates of oestrogen formation were determined under standardized conditions with the tritiated water generation technique in bone specimens obtained from 4 men and 11 post-menopausal women.

RESULTS

The apparent Km of the aromatase ranged between 6 and 50 nM (20.4 +/- 3.9; mean +/- SEM), values in the range of those reported for human placental microsomes. The maximum velocity (Vmax) of the aromatase activity ranged between 0.14 and 1.23 nmol/g DNA/h.

CONCLUSIONS

Oestrogens formed in human bone may play a physiological role in steroid hormone action in this tissue.

BACKGROUND

Dehydroepiandrosterone (DHEA) mainly exerts indirect action via downstream conversion toward sex steroids within peripheral target cells including immune cells. In vitro DHEA has been shown to enhance IL-2 release from T lymphocytes, whereas it inhibits IL-6 secretion. Conversely, aging is associated with a decline in both DHEA and IL-2, whereas IL-6 increases.

OBJECTIVE

The objective of the study was to investigate age-related differences in expression and functional activity of steroidogenic enzymes involved in downstream conversion of DHEA in peripheral blood mononuclear cells (PBMCs).

METHODS

This study was cross-sectional.

METHODS

Healthy young men (n = 8; age range, 23-29 yr) and healthy middle-aged men (n = 8; age range, 52-66 yr) were studied in an academic setting.

METHODS

mRNA expression of steroidogenic enzymes in PBMCs was measured by qualitative and quantitative RT-PCR analysis and enzyme activity assays after incubation of PBMCs with radiolabeled DHEA, <em>4</em>-androstene-3,17-dione (<em>androstenedione</em>), and testosterone.

RESULTS

RT-PCR analysis showed expression of all enzymes required for DHEA conversion toward active androgens and to the immune-stimulatory metabolite androstenediol. Steroid conversion patterns indicated a particularly increased activity of 17beta-hydroxysteroid dehydrogenase type 5 (17beta-HSD5) in the older men, demonstrated by significantly higher conversion rates of DHEA to androstenediol and of androstenedione to testosterone (all P < 0.05). By contrast, conversion of DHEA to androstenedione via 3beta-HSD occurred at a similar rate. Quantitative RT-PCR analysis revealed increased expression of 17beta-HSD 5 mRNA in PBMCs from the older men.

CONCLUSIONS

Our results provide evidence for significant changes in sex steroid metabolism by human PBMCs with aging, which may represent an endocrine link to immune senescence.

A female newborn infant with ambiguous genitalia was found to have hypertension (121/82 mm Hg) immediately after birth. The plasma testosterone (T) (0.73 nmol/l), delta <em>4</em>-<em>androstenedione</em> (delta <em>4</em>-A) (5.9 nmol/l), dehydroepiandrosterone (DHEA) (8.9 nmol/l), as well as 17 OH-hydroxyprogesterone (17 OHP) (152 nmol/l) were elevated. The diagnosis of 11 beta-hydroxylase deficiency was finally established on the basis of elevated plasma eleven-deoxycortisol (compound S) (greater than 0.6 mumol/l) and confirmed by the normalisation of the blood pressure during hydrocortisone therapy. Our case is probably the youngest patient with 11 beta-hydroxylase deficiency in whom the hypertension was found at birth.

To assess the influence of endogenous opioids on human gonadotropin secretion, integrated concentrations of gonadotropins during 2<em>4</em>-h naloxone infusion (2.08 mg/h) were examined in a group of six normal men. Naloxone significantly stimulated LH secretion in all subjects, whereas serum FSH levels were similar during both saline and naloxone infusion. Serum testosterone, dihydrotestosterone, and 17 alpha-hydroxyprogesterone levels increased during the infusion of naloxone, but the delta <em>4</em>-<em>androstenedione</em> concentration was not modified. The frequency and amplitude of LH secretory episodes were clearly increased when an equal amount of the drug was given and LH concentrations were measured every 15 min for 6 h. The results suggest the endogenous opioids inhibit the hypothalamic-pituitary-gonadal axis in normal men.

Dehydroepiandrosterone (DHEA) is a C19 adrenal steroid synthesized in the human adrenal cortex and serving as a biosynthetic precursor to testosterone and 17beta-estradiol. Despite the fact that it is one of the most abundant steroid hormones in circulation, the physiological role of DHEA in humans remains unclear. The action of DHEA itself, such as its interactions with receptors and nuclear transcription factors, is not well understood, and a specific DHEA receptor has yet to be identified. Although the activity of DHEA can be due to its metabolism into androgens and estrogens, DHEA has been shown to interact with the androgen receptor and the estrogen receptor (ER) in vitro. We demonstrate in this study that DHEA (3beta-Hydroxy-5alpha-androstan-17-one) inhibits 17beta-estradiol (E2) binding to its receptor in vivo in yeast. DHEA stimulates human ER dimerization in yeast, as determined by ER fusion protein interactions, GAL<em>4</em> reconstitution and subsequent measurement of increased beta-galactosidase activity. DHEA causes an increase in estrogen response element-dependent beta-galactosidase activity, demonstrating that the ER dimer induced by DHEA is transcriptionally active, but at a concentration of DHEA about 1000 times greater than E2. Inclusion of the nuclear receptor co-activator RIP1<em>4</em>0 in the yeast enhances ER transactivation by DHEA or E2 in a ligand-dependent manner; moreover, only in the presence of RIP1<em>4</em>0 is DHEA able to stimulate beta-galactosidase activity to levels similar to those achieved by E2. Ligand-receptor interaction for other C19-steroids was also examined. While 5-androstene-3beta, 17beta-diol (ADIOL) displayed estrogenic activity in this system, <em>4</em>-androstene-17-dione (<em>androstenedione</em>) and <em>4</em>-androstene-17beta-ol,3-one (testosterone) did not. We have investigated whether DHEA can interact with the human ER in vivo. Our findings demonstrate a mechanism by which DHEA interacts directly with estrogen signaling systems; however, because DHEA is several orders of magnitude less potent than E2 in this system, we conclude that it essentially is not an estrogen agonist.

At the time of surgery, women were infused with [3H]dehydroepiandrosterone sulfate ([3H]DS)/[1<em>4</em>C]testosterone ([1<em>4</em>C]T) for 6 h; blood samples were obtained from an artery the ovarian veins, and a peripheral vein; and fluid was obtained from ovarian follicles. Both blood and follicular fluid samples were analyzed for radioactivity as DS, dehydroepiandrosterone (D), <em>androstenedione</em> (delta <em>4</em>A). T, and dihydrotestosterone (DHT), and the blood was also analyzed for the concentration of nonisotopic DS by RIA. In other subjects the concentrations of D and DS were measured in paired samples of blood and follicular fluid. From these data, values of 13.6 +/- 0.69 L/day four (mean +/- SE; n = <em>4</em>) for MCRDS, 607 +/- 90 L/day (n = 3) for MCRT, and 0.0190 +/- 0.0089 (n = 3) for [p]DS-T (fraction of plasma DS metabolized to plasma T) were obtained. The ratio of the concentration of the tracer-labeled steroid in the follicular fluid to the concentration in the arterial plasma sample was elevated significantly above 1 for three 3H-labeled and three [1<em>4</em>C-labeled metabolites: [3H]D (21-fold; P less than 0.001), [3H]T (81-fold; P less than 0.001), [3H]DHT (19-fold; P less than 0.001), [1<em>4</em>C]T (<em>4</em>-fold; P less than 0.025), [1<em>4</em>C]DHT (21-fold; P less than 0.01), and [1<em>4</em>C]delta <em>4</em>A (50-fold; P less than 0.001). The estimated concentrations of steroids in follicular fluid derived from DS based on specific activity calculations were as follows: [geometric mean (95% confidence limits; n)]: DS, 5600 (<em>4</em>800-6500 nmol/L; 12); D, 370 (88-1500 nmol/L; 10); delta <em>4</em> A, 120 (67-220 nmol/L; 12); T, 130 (39-<em>4</em>50 nmol/L; 10); and DHT, 6<em>4</em> (35-120 nmol/L; 8). Comparison of these data to known follicular fluid steroid concentrations shows that DS from the intravascular pool can be used as an ovarian prehormone.

To explore the abnormal steroidogenesis in subjects with primary testicular failure, we measured serum levels of testosterone (T) and its precursors <em>androstenedione</em> (delta <em>4</em>A) and 17-hydroxyprogesterone (17-OHP) in the basal state and after stimulation by human chorionic gonadotropin (hCG) for <em>4</em> hours (acute reserve) or 72 hours (chronic reserve). Subjects with primary testicular failure had decreased mean basal serum T, decreased mean chronic T reserve, and absent mean acute T reserve. In contrast, these subjects had normal mean basal serum 17-OHP and delta <em>4</em>A, normal chronic 17-OHP reserve, and non-zero mean acute 17-OHP and delta <em>4</em>A reserve, although mean chronic delta <em>4</em>A reserve was reduced. Heterogeneity of biochemical abnormalities depending on the etiology of testicular failure was apparent. In primary testicular failure, the testicular reserves of the T precursors 17-OHP and delta <em>4</em>A are better maintained than is the reserve of T itself, raising the possibility that this disorder might be associated with biochemical blocks in conversion of T precursors to T.

Testosterone and <em>androstenedione</em> were measured in testicular and epididymal tissue of 37 previously healthy infants between 1 and 2<em>4</em> months of age who died suddenly. In half of the patients elevated plasma levels of cortisol and <em>androstenedione</em> suggested preterminal stress. Plasma testosterone levels, however, did not differ from those in healthy infants. Testicular testosterone concentrations were maximal in boys from 1-3 months of age (median, 36.6 ng/g; range, 7-380 ng/g) with peak values similar to those found in pubertal or even adult testes. Thereafter testicular testosterone concentrations decreased and after the age of 6 months all values were below 12.5 ng/g, which corresponds to the low normal range of older prepubertal boys. Plasma testosterone and testicular testosterone correlated significantly (P less than 0.001). On average the testicular concentrations were 36.<em>4</em> times higher than the corresponding plasma concentrations. Testicular <em>androstenedione</em> was low but correlated significantly with testicular testosterone (P less than 0.001). Epididymal testosterone concentrations were surprisingly high (1-3 months: median, 10.3 ng/g; range, <em>4</em>-<em>4</em>2.7 ng/g) and averaged 30% of the testicular testosterone concentration. Thus, epididymal testosterone concentrations were significantly higher than the circulating plasma testosterone levels, indicating the capacity of the infant epididymis to accumulate androgens. These findings suggest that high local testosterone concentrations during early infancy are important not only for the testis itself but particularly for the developing epididymis.

A purified rat hepatic monooxygenase system containing cytochrome P-<em>4</em>50b oxidizes testosterone to <em>androstenedione</em> and 16 alpha- and 16 beta-hydroxytestosterone at approximately equal rates. The metabolism of epitestosterone by the same system is characterized by a marked stereoselectivity in favor of 16 beta-hydroxylation (<em>4</em>- to 5-fold relative to 16 alpha-hydroxylation), formation of 15 alpha-hydroxyepitestosterone, and a rate of <em>androstenedione</em> formation which is three to five times higher than that observed with testosterone. Apparent Km values for 16 alpha- and 16 beta-hydroxylation and <em>androstenedione</em> formation are 20-30 microM with either substrate. Mass spectral analysis of the <em>androstenedione</em> formed from [16,16-2H2]testosterone and [16,16-2H2] epitestosterone indicates essentially complete retention of deuterium, thereby ruling out a mechanism of <em>androstenedione</em> formation via C-16 hydroxylation followed by loss of water and rearrangement. Mass spectral analysis of the C-16 hydroxylation products from incubations of testosterone or epitestosterone in 18O2 shows essentially complete incorporation of 18O (greater than 95%). <em>Androstenedione</em> formed from testosterone is enriched in 18O only 2-fold (5-8%) over background, while the <em>androstenedione</em> formed from epitestosterone shows 8<em>4</em>% enrichment. Kinetic experiments utilizing [17-2H]testosterone and [17-2H]epitestosterone as substrates indicate that cleavage of the C-17 carbon-hydrogen bond is involved in a rate-limiting step in the formation of <em>androstenedione</em> from both substrates. Taken together, our results indicate that <em>androstenedione</em> formation from epitestosterone proceeds exclusively through the gem-diol pathway, while <em>androstenedione</em> formation from testosterone may proceed through a combination of gem-diol and dual hydrogen abstraction pathways.