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Purpose: Alcohol consumption is an established breast cancer risk factor, though further research is needed to advance our understanding of the mechanism underlying the association. We used global metabolomics profiling to identify serum metabolites and metabolic pathways that could potentially mediate the alcohol-breast cancer association., Methods: A cross-sectional analysis of reported alcohol consumption and serum metabolite concentrations was conducted among 211 healthy women 25-29 years old who participated in the Dietary Intervention Study in Children 2006 Follow-Up Study (DISC06). Alcohol-metabolite associations were evaluated using multivariable linear mixed-effects regression., Results: Alcohol was significantly (FDR p < 0.05) associated with several serum metabolites after adjustment for diet composition and other potential confounders. The amino acid sarcosine, the omega-3 fatty acid eicosapentaenoate, and the steroid 4-androsten-3beta,17beta-diol monosulfate were positively associated with alcohol intake, while the gamma-tocopherol metabolite gamma-carboxyethyl hydroxychroman (CEHC) was inversely associated. Positive associations of alcohol with 2-methylcitrate and 4-androsten-3beta,17beta-diol disulfate were borderline significant (FDR p < 0.10). Metabolite set enrichment analysis identified steroids and the glycine pathway as having more members associated with alcohol consumption than expected by chance., Conclusions: Most of the metabolites associated with alcohol in the current analysis participate in pathways hypothesized to mediate the alcohol-breast cancer association including hormonal, one-carbon metabolism, and oxidative stress pathways, but they could also affect risk via alternative pathways. Independent replication of alcohol-metabolite associations and prospective evaluation of confirmed associations with breast cancer risk are needed.

Study Objective: Hirsutism has negative impact on adolescent psychosocial development for both cosmetic and endocrine reasons. This study evaluated the effectiveness of a new intermittent, low-dose finasteride regimen consisting of 2.5 mg of drug given every 3 days (1 day of treatment, 2 days of drug withdrawal) for 6 months in girls with hirsutism by polycystic ovarian syndrome (PCOS) or idiopathic hirsutism (IH)., Design and Participants: Twenty-eight girls (15-19 y old) with hirsutism were randomly assigned to 2 treatment groups and treated for 6 months. Fourteen patients (7 with IH, 7 with PCOS) received finasteride; fourteen patients (7 with IH, 7 with PCOS) received placebo. Hirsutism score (HS), clinical, and hormonal effects were compared between the 2 groups., Results: In patients treated with finasteride, the HS value at 6 months was 52.9% lower than that observed at baseline in girls with IH, and 52.8% lower in girls with PCOS (P < .0001 for both). Similarly, the 3α-17 β-androstenediol glucuronide serum levels were decreased by 34.8% in patients with IH, and by 47.5% in patients with PCOS (P < .0001, respectively). Finasteride treatment was well tolerated and did not alter values of BMI, serum levels of sexual hormones, metabolic parameters related to liver and kidney function as well as glycemic and lipidic asset., Conclusions: A low-dose of finasteride, given every 3 days, reduces the HS in young patients affected by PCOS or IH. Compared with conventional continuous finasteride administration, the intermittent low-dose regimen has similar efficacy with the advantage to be safer and less expensive., (Copyright © 2014 North American Society for Pediatric and Adolescent Gynecology. Published by Elsevier Inc. All rights reserved.)

The widely expressed steroid hydroxylase CYP7B1 is involved in metabolism of a number of steroids reported to influence estrogen and androgen signaling. Several studies by us and other investigators have linked this enzyme to effects on estrogen receptor activation. In a previous report we examined the effect of CYP7B1-mediated hormone metabolism for estrogen-mediated response in kidney-derived HEK293 cells. In the current study we used an androgen response element (ARE) reporter system to examine androgen-dependent response of some CYP7B1 substrates and CYP7B1-formed metabolites in several cell lines derived from different tissues. The results indicate significantly lower androgen receptor activation by CYP7B1-formed steroid metabolites than by the corresponding steroid substrates, suggesting that CYP7B1-mediated catalysis may decrease some androgenic responses. Thus, CYP7B1-dependent metabolism may be of importance not only for estrogenic signaling but also for androgenic. This finding, that CYP7B1 activity may be a regulator of androgenic signaling by converting AR ligands into less active metabolites, is also supported by real-time RT-PCR experiment where a CYP7B1 substrate, but not the corresponding product, was able to stimulate known androgen-sensitive genes. Furthermore, our data indicate that the effects of some steroids on hormone response element reporter systems are cell line-specific. For instance, despite transfection of the same reporter systems, 5-androstene-3β,17β-diol strongly activates an androgen-dependent response element in prostate cancer cells whereas it elicits only ER-dependent responses in kidney HEK293 cells. Potential roles of cell-specific metabolism or comodulator expression for the observed differences are discussed., (Copyright © 2012 Elsevier B.V. All rights reserved.)

A series of 19-substituted androstenediol derivatives was prepared. Some of the novel analogs were surprisingly potent and selective ligands for ER-beta.

The use of anabolic agents in food producing animals is prohibited within the EU since 1988 (96/22/EC directive). The control of the illegal use of natural steroid hormones in cattle is still an exciting analytical challenge as far as no definitive method and non-ambiguous analytical criteria are available. The ability of gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) to demonstrate the administration of 17beta-estradiol to bovine has been investigated in this paper. By comparison of 13C/12C isotopic ratio of main urinary estradiol metabolite, i.e. 17alpha-estradiol, with two endogenous reference compounds (ERCs), i.e. dehydroepiandrosterone (DHEA) and 5-androstene-3beta,17alpha-diol, the differentiation of estradiol metabolite origin, either endogenous or exogenous, has been proved to be achievable. After treatment, the delta(13)C(VPDB)-values of 17alpha-estradiol reached -27 per thousand to -29 per thousand, whereas delta13CVPDB-values of DHEA remained between -13 per thousand and -20 per thousand depending on the diet, maize and grass, respectively. A significant difference of delta13CVPDB between ERCs and 17alpha-estradiol was measurable over a period of 2 weeks after estradiol ester administration to the animal.

7-Hydroxy-metabolites of dehydroepiandrosterone (DHEA) and 3beta,17beta-androstenediol (AD) possess immunomodulatory and neuroprotective properties; therefore, the measurement of these steroids in patients with autoimmune diseases or disturbances in the CNS may be of interest. A gas chromatography-mass spectrometry (GC-MS) method for the determination of 7-hydroxy-metabolites of pregnenolone, DHEA, AD, and testosterone including the parent steroids was applied to serum samples from 12 adult men (27-66 years), 13 male adolescents (13-20 years), 5 boys (6-10 years), 15 women in the follicular phase of the menstrual cycle (22-45 years), 17 women in the luteal phase (22-45 years), and 4 girls (6-10 years). The steroids were age and sex dependent, but independent of the menstrual cycle. The ratio of the 7alpha-hydroxy-metabolites to their parent steroids were age dependent, exhibiting an increasing trend (p < 0.0001, ANOVA) from pregnenolone (5%) to AD (20%). The ratio of 7beta- to 7alpha-metabolites ranged from 0.6 to 1. These results are consistent with models suggesting 7alpha-hydroxylation of the parent steroid, conversion to a 7-oxo-steroid and finally to the 7beta-hydroxylated-metabolite. Partial correlations suggested that 7-hydroxylation might reduce the concentration of circulating androgens. Despite the three times lower concentration of AD-metabolites, their antiglucocorticoid, immunomodulatory, and neuroprotective effects may be comparable to that of DHEA based on their reported greater biological activity.

5-Androsten-3beta, 17beta-diol (HE2100), and a synthetic analogue HE3204 are regarded as immune-regulating hormones, because both induce changes in the reporter antigen-popliteal lymph node assay (RA-PLNA). Mice were injected in the footpad with either HE2100 or HE3204 (0.01-3 mg), and a nonsensitizing dose of trinitrophenyl ovalbumin (TNP-OVA) was used as bystander reporter antigen. Seven days later, nodes were removed and numbers of cells (CD3, CD4, CD8, CD19; flow cytometry), TNP-specific IgM, IgG1, and IgG2a antibody-forming cells (AFCs; ELISPOT assay), and cytokines (interleukin-4 [IL-4], interferon-gamma [IFN-gamma]; ELISA) were measured. HE2100 and HE3204 increased cell numbers in a dose-dependent fashion. T (helper and suppressor) cells and B cells were increased (>5-fold). HE3204 was apparently twice as potent as HE2100. Both increased the B/T ratio (fivefold), increased TNP-specific IgM and IgG1 ( approximately 50-fold), and induced IgG2a AFCs. Both increased IL-4 and IFN-gamma secretion (up to threefold). Both displayed anti-inflammatory activity in the murine model of carrageenan-induced pleurisy, as evidenced by reduced neutrophil numbers and exudate volumes. Our observations suggest that both HE2100 and HE3204 are immune-regulating steroid hormones that exhibit anti-inflammatory properties. HE2100 (1 mg/mouse per day) provided significant benefit when given at disease onset in the SJL/J female mouse model of experimental autoimmune encephalomyelitis. These compounds and their analogues are candidates for further testing in autoimmune diseases.

We compared in vivo radioprotective efficacy of 5-androstenediol (5-AED) to that of ten other steroids: 17alpha-androstenediol, dehydroepiandrosterone, 5-androstenetriol (AET), 4-androstenedione (AND), testosterone, estradiol, fluasterone, 16alpha-bromoepiandrosterone, 16alpha-fluoro-androst-5-en-17alpha-ol (alpha-fluorohydrin, AFH), and 16alpha-fluoro-androst-5-en-17beta-ol (beta-fluorohydrin). Steroids were administered 24 or 48 hr before, or 1 hr after, whole-body gamma-irradiation. Two days after irradiation at 3 Gy, blood elements were counted. In addition, after irradiation at 9-12.5 Gy, survival was recorded for 30 days. The results showed radioprotective efficacy was specific for 5-AED. One other steroid, AFH, demonstrated appreciable survival effects but was less efficacious than 5-AED. AND and AET produced slight enhancement of survival in some experiments. This is the first demonstration that the prophylactic window for survival enhancement by 1 subcutaneous (s.c.) injection of 5-AED is as long as 48 hr in mice. Moreover, the results indicate that 1 s.c. injection of 5-AED 1 hr after irradiation is much less effective than 1 injection 24-48 hr before irradiation. Comparing the molecular features of steroids with radioprotective efficacy leads to the following conclusions: 1) these effects are due to interaction with specific receptors, since s.c. injection of extremely similar molecules with the same physicochemical properties as 5-AED were not radioprotective; 2) the 17-hydroxyl group is essential; 3) this group must be in the beta configuration in the absence of nearby side groups; 4) a halogen atom at 16 changes the 17-hydroxyl specificity to alpha; 5) the 3beta-hydroxyl group is not essential; 6) addition of a 7beta-hydroxyl group is deleterious; and 7) the effects are not due to activation of sex steroid receptors.

Since it is known that androstenediol (ADIOL) has potent immunoregulatory effects, changes in ADIOL levels during and after pregnancy might affect the maternal immune system. We examined serum concentrations of ADIOL and androstenediol 3-sulfate (ADIOLS) together with IFN-gamma and IL-4 production levels during pregnancy and after delivery up to 10-11 months postpartum. The subjects were 73 normal pregnant, 76 normal postpartum, and 28 normal non-pregnant women. ADIOL and ADIOLS were measured using EIA and GC/MS, respectively. The cytokine levels in the supernatant of whole-blood cultures stimulated with phorbol 12-myristate 13-acetate and ionomycin were measured using ELISA. ADIOL levels significantly decreased compared to non-pregnant levels in the first trimester (P < 0.05) and were reversed in the third trimester (P < 0.05). After pregnancy, ADIOL levels gradually declined, and a significant decrease was observed at 10-11 months postpartum (P < 0.05). ADIOLS levels were significantly lower in the third trimester (P < 0.05) and significantly higher at the first month postpartum (P < 0.001) compared to non-pregnant women. IFN-gamma and IL-4 levels decreased during pregnancy and subsequently increased postpartum. On the other hand, we found significant negative correlations between ADIOL concentrations and production levels of IFN-gamma (P < 0.05) or IL-4 (P < 0.05). These findings suggest that ADIOL may be involved in modifying the maternal immune response during and after pregnancy.

Androstenediol (5-androsten-3beta, 17beta-diol, ADIOL) and androstenediol 3-sulfate (ADIOLS) are active metabolites of dehydroepiandrosterone (DHEA) and DHEA sulfate (DHEAS), respectively, and have estrogenic activity and immunoregulatory function. We examined serum concentrations of ADIOL, ADIOLS, DHEA, DHEAS and pregnenolone sulfate (5-pregnen-3beta-ol-20-one sulfate, PREGS) in patients with Graves' thyrotoxicosis (male/female 9/14), hypothyroidism (11/20) and in normal controls (14/29). In hypothyroidism serum levels of all these steroids were significantly decreased in both genders. In hyperthyroidism, in contrast, serum levels of ADIOLS (male 1.49 +/- 0.69, female 0.64 +/- 0.31 micromol/l), DHEAS (male 7.43 +/- 3.91, female 5.13 +/- 2.03 micromol/l), and PREGS (male 1.13 +/- 0.58, female 1.07 +/- 0.85 micromol/l) were markedly increased, but serum concentrations of ADIOL and DEHA were not significantly different from controls (ADIOLS male 0.36 +/- 0.33, female 0.14 +/- 0.09 micromol/l; DHEAS male 2.88 +/- 1.70, female 1.86 +/- l1.03pmol/l; PREGS male 0.18 +/- 0.12, female 0.11 +/- 0.08 micromol/l; ADIOL male 3.76 +/- 1.35, female 1.91 +/- 1.17 nmol/l; DHEA male 9.23 +/- 3.49, female 13.5 +/- 10.8nmol/l). Serum concentrations of all these steroids correlated with the serum concentration of the thyroid hormones in these patients. Serum albumin and sex hormone-binding globulin concentrations were not related to these changes in the concentrations of steroids. These findings indicate that serum concentrations of ADIOLS, ADIOL, DHEAS, DHEA and PREGS were decreased in hypothyroidism, whereas serum ADIOLS, DHEAS and PREGS concentrations were increased but ADIOL and DHEA were normal in hyperthyroidism. Thyroid hormone may stimulate the synthesis of these steroids and sulfotransferase is speculated to be increased in hyperthyroidism. Increased ADIOLS might contribute to menstrual disturbances and gynecomastia in hyperthyroidism.

Studies of urinary steroids were performed in males after oral administration of 5-androsten-3,17-dione; 5-androsten-3beta,17beta-diol; dehydroepiandrosterone; and 19-nor-5-androsten-3,17-dione. 5-Androsten-3,17-dione; 5-androsten-3beta,17beta-diol; and dehydroepiandrosterone amplify most endogenous steroids, but to a lesser extent than their delta4 analogues do. Especially affected are androsterone, etiocholanolone, dehydroandrosterone, dehydroepiandrosterone, and isomeric 5-androstendiols. 5-Androsten-3,17-dione; 5-androsten-3beta,17beta-diol; and dehydroepiandrosterone elevate the urinary testosterone to epitestosterone (T/E) ratio by a factor of 2-3 a few hours after administration. This may cause a positive T/E test (> 6) for individuals with normal T/E ratios higher than 2. Most of the steroids return to their original concentrations in less than 24 h. Etiocholanolone and 5beta-androstan-3alpha,17beta-diol remain elevated for several days. A reduced androsterone to etiocholanolone (A/E) ratio may be an indication of delta5 steroids abuse. 19-Nor-5-androsten-3,17-dione has a similar effect, except that all metabolites in urine are 19-nor exogenous steroids. Identification criteria for 19-nor-5-androsten-3,17-dione may be the same as nandrolone, that is, detection of 19-norandrosterone and 19-noretiocholanolone. Specific abundant metabolites of 19-nor-5-androsten-3,17-dione are 19-nordehydroandrosterone and 19-nordehydroepiandrosterone. In the later stages of excretion, higher concentration of 1 9-noreticholanolone relative to 19-norandrosterone specifically indicates administration of 19-nor delta5 steroids.

Objectives: We evaluated the role of delta-5-androstenediol (adiol) and its sulphates in health and endocrine diseases., Design: Serum and urine samples from healthy adult men and pre and post-menopausal women were analysed by gas chromatography-mass spectrometry to establish reference values. In patients who were either evaluated or treated for endocrine diseases, sequential serum samples were collected and analysed., Patients: Reference values were obtained from 24 healthy male, 23 premenopausal and 30 post-menopausal female volunteers. Adiol and delta-5-androstenediol-3-sulphate (adiol-3S) concentrations were determined in combination with other relevant steroids in patients with either pituitary (n = 5), adrenal (n = 2) or gonadal dysfunction (n = 1), or testicular carcinoma (n = 19)., Measurements: After addition of deuterated adiol as internal standard, serum and urine samples were extracted. Steroid sulphates were hydrolysed. The extracts and hydrolysates were purified on HPLC, adiol was derivatized and finally quantified by gas chromatography-mass spectrometry., Results: The calculated reference ranges for adiol and adiol-3S concentrations in serum are respectively: in men 1.78-7.24 and 123-579 nmol/l, in premenopausal women 0.65-6.93 and 21.2-298 nmol/l and in post-menopausal women 0.29-2.90 and 6.1-184 nmol/l. Urinary values varied considerably. In the population with endocrine abnormalities serum adiol and adiol-3S concentrations were compared with other relevant steroids., Conclusions: The wide concentration range of adiol and adiol-3S in urine makes analysis of these steroids in urine of little clinical value. Serum concentrations of adiol and adiol-3S are higher in men than in women. Premenopausal values are higher than post-menopausal. Adiol and adiol-3S in serum are significantly correlated in pre and post-menopausal women, r = 0.51 and r = 0.69 respectively, but not in men. In endocrine patients the serum concentrations of adiol show an ACTH or LH dependency in women; adiol correlates with cortisol, dehydroepiandrosterone or androstenedione and, in males, additionally with testosterone. However, in several situations adiol correlates with none of these steroids. Although adiol secretion can be stimulated by ACTH and LH, the level of serum adiol is also determined by other factors. Finally, in adrenal carcinoma serum adiol and adiol-3S may be used as tumour markers.

Objectives: We developed an assay for delta-5-androstenediol (adiol) and delta-5-androstenediol-3-sulphate (adiol-3S) in serum and adiol and total adiol sulphate (adiol-S) in urine., Design: An analytical procedure using HPLC and gas chromatography-mass spectrometry was devised and tested for its reliability., Measurements: After addition of deuterated androstenediol as internal standard, serum and urine samples were extracted. Steroid sulphates were hydrolysed. The extracts and hydrolysates were purified on HPLC, adiol was derivatized using heptafluorobutyric anhydride and finally quantified by gas chromatography-mass spectrometry., Results: The assay is accurate and reproducible. The coefficient of variation (CV) for the determination of adiol in serum samples is 4% (intra-assay) and 9% (interassay) and for urine samples 3 and 8% respectively. The intra-assay CV for the adiol-3S analyses is 5% for serum and 2% for urine samples while the interassay CV values for adiol-3S are 10% for serum and 7% for urine samples. The recovery of adiol and adiol-3S from serum and urine samples is 97%., Conclusions: The developed assay meets the analytical demands needed for clinical applications.

On solvolysis of Westphalen-type steroids with a leaving group in the position 6 beta (e.g., 2), products of elimination (followed by rearrangement and fragmentation of the steroid skeleton) were prepared (e.g., 4 and 5). These products were subsequently converted to suitable analogs of the compound, which has been reported to promote hair growth (1). Compounds 11 to 13 exhibited strong antiandrogenic activity in vivo; however, this activity could not be interpreted either in terms of inhibition of 5 alpha-reductase or by strong binding to an androgen receptor.

A commercially available antidihydrotestosterone antiserum was used for the direct radioimmunoassay of androstenediol-3-sulfate (ADS) in human serum. Aliquots of 1 or 2 microliter male serum (mean age of 40 subjects, 38.2 +/- 5.0 years) were diluted and extracted with ethanol for assay. The tracer, [7-3H]ADS, was prepared by sodium borohydride reduction of [7-3H]dehydroepiandrosterone sulfate (DS). Significantly cross-reacting steroids were testosterone, DS, androsterone sulfate, and epiandrosterone sulfate, which combined to produce a mean overestimation of ADS of 4.3 micrograms/dl in male serum. Mean serum ADS was 23.6 +/- 10.0 micrograms/dl (SD) in 20 fresh-frozen sera versus 28.4 +/- 9.7 micrograms/dl (SD) in 20 long-term (24.4 +/- 1.2 years) frozen specimens, showing stability on long-term frozen storage. Androstenediol-3-sulfate also showed a strong correlation with serum DS (r = 0.75). The possible physiologic significance of ADS is discussed, particularly in terms of the known estrogenicity of unconjugated androstenediol.

The secretion of 17 beta-estradiol (E2) and the effect of exogenous E2 on progesterone (P) production by granulosa-luteal cells from 18 women attending an in vitro fertilization (IVF) program were studied. The cells were separated from follicular fluid and precultured in medium containing fetal calf serum. On the third day of culture E2 (0.25-2.0 micrograms/ml) was added onto the cells and its effect on both hCG-stimulated and basal P production was measured. E2 inhibited both basal and hCG-stimulated P production. 1 microgram/ml of E2 caused mean decrements of 55 and 56% in basal and hCG-stimulated P production, respectively. The maximal inhibition by E2 occurred at 6 hours of incubation, but when the cells were allowed to react with E2 for longer periods of time the effect became less significant and more variable. At 48 h no inhibition was observed. At 6 h E2 (1.0 microgram/ml) increased both basal and hCG-stimulated pregnenolone production by approx. 10-fold, suggesting that the suppression of P production was due to inhibition of 3 beta-hydroxysteroid dehydrogenase. Exogenous androgen, 5-Androsten-3 beta-ol-17-one sulfate, dehydroepiandrosterone sulfate (DHEAS), in a dose-dependent manner increased granulosa-luteal cell E2 production. The maximal response was about 1000-fold above the E2 production of unstimulated cells and was not affected by hCG. However, the maximal amount of E2 produced was minor in comparison to exogenous doses required for the suppression of P production and did not have the inhibitory effect. It is concluded that the production of E2 by granulosa-luteal cells is mainly regulated by the availability of androgen substrate, and that E2 functions as a modulator of luteal P production.

Female pattern androgenic alopecia (AA) is a relatively common endocrine abnormality in premenopausal women. However, unlike hirsutism, little is known about the androgen metabolism and plasma C19 steroid sulphate profiles in this disorder. We have therefore measured the plasma levels of dehydroepiandrosterone sulphate (DHEA-S), 5-androstene-3 beta,17 beta-diol sulphate (5-ADIOL-S), 5 alpha-androstane-3 alpha, 17 beta-diol sulphate (3 alpha-DIOL-S), androstenedione (AD), total testosterone (T), free testosterone (FT), sex hormone binding globulin (SHBG), non-SHBG bound T, luteinizing hormone (LH) and follicle stimulating hormone (FSH), and have calculated the free androgen index (FAI): 100 x T (nmol/l) divided by SHBG (nmol/l), in premenopausal women with AA (n = 25-45) and in normal premenopausal women (n = 17-73). While mean plasma concentrations of DHEA-S and T were not significantly different from controls, mean SHBG concentrations were significantly lower (47 +/- 3 vs 64 +/- 3 nmol/l) and the mean free androgen index (4.4 +/- 0.4 vs 2.4 +/- 0.2), and mean concentrations of free testosterone (45 +/- 5 vs 26 +/- 1.4 pmol/l), non-SHBG bound T (0.9 +/- 0.2 vs 0.6 +/- 0.1 nmol/l) and androstenedione (4.3 +/- 0.3 vs 3.4 +/- 0.2 nmol/l) were significantly elevated in women with AA. Furthermore, mean plasma concentrations of 5-ADIOL-S (512 +/- 42 nmol/l) and 3 alpha-DIOL-S (76 +/- 7 nmol/l) were significantly higher than levels found in normal women (272 +/- 12 nmol/l and 52 +/- 2 nmol/l respectively). The nature of the hyperandrogenism associated with AA may thus only be revealed by a comprehensive plasma androgen and androgen sulphate profile, which may explain apparently aberrant data for a given patient. In addition, 5-ADIOL-S and 3 alpha-DIOL-S may serve as excellent plasma markers of both the existence of the disorder and the efficacy of its treatment.

Plasma levels of cortisol, dehydroepiandrosterone (DHA), dehydroepiandrosterone sulphate (DHAS), delta 5-androstenediol (delta 5-DIOL), delta 5-androstenediol sulphate (delta 5-DIOL-S) and testosterone were determined every 2 h from 10.00 to 20.00 h in 8 normal women and 10 with anorexia nervosa. Plasma levels of cortisol. DHA and delta 5-DIOL were significantly (P less than 0.001) higher while DHAS levels were significantly (P less than 0.001) lower in the anorexic women. The levels of delta 5-DIOL-S and testosterone were similar in both groups of women. In the normal women there were significant (P less than 0.001) diurnal fluctuations in the levels of cortisol, DHA and DHAS with high levels in the morning and a nadir in the evening; however, there were significant P less than 0.001) 'reverse' diurnal fluctuations in the levels of delta 5-DIOL, delta 5-DIOL-S and testosterone with low levels in the morning and elevated levels in the evening. In the anorexia nervosa women there was a loss of the diurnal variation in the levels of cortisol, DHA and DHAS and delta 5-DIOL-S; the diurnal variations of delta 5-DIOL and testosterone levels in the anorexic women were similar to those in the normal women. In general, these findings support the suggestion of a disturbance in the mechanisms regulating hypothalamic-pituitary-adrenal function resulting from a primary hypothalamic defect and/or abnormal alterations in steroid metabolism associated with the malnutrition in anorexia nervosa.

Human pituitary LH (1200 i.u.) was infused for 4 h (from 10.00 to 14.00 h) into six women with anorexia nervosa and into five women with polycystic ovarian disease (PCO). Plasma dehydroepiandrosterone sulphate (DHAS), androstenediol sulphate, dehydroepiandrosterone (DHA), androstenediol and testosterone were estimated by gas-liquid chromatography in blood samples taken every 2 h from 10.00 to 20.00 h. The values were compared with those obtained at the same times on the previous control day. There were no significant changes in the plasma levels of DHAS and androstenediol sulphate in response to LH at any of the sampling times in either the anorexia nervosa or the PCO patients. In the anorexia nervosa women, plasma DHA levels were significantly increased at 16.00 (P less than 0.001), 18.00 (P less than 0.001) and 20.00 h (P less than 0.05) after LH infusion. In the PCO women, DHA levels increased significantly at 14.00 (P less than 0.01), 16.00 (P less than 0.001), 18.00 (P less than 0.001) and 20.00 h (P less than 0.001) in response to LH infusion. Plasma androstenediol levels increased significantly in the anorexia nervosa patients at 12.00 (P less than 0.001), 14.00 (P less than 0.01) and 16.00 h (P less than 0.01) in response to LH. Similar increases were also found in the PCO patients at 12.00 (P less than 0.01), 14.00 (P less than 0.001) and 16.00 h (P less than 0.01). Plasma testosterone decreased progressively in the anorexic women in response to LH, becoming significant at 16.00 (P less than 0.05), 18.00 (P less than 0.05) and 20.00 h (P less than 0.01). A similar progressive decrease in plasma testosterone was seen in the PCO women, the levels being significantly lower than controls at 16.00 (P less than 0.05), 18.00 (P less than 0.05) and 20.00 h (P less than 0.05). The results represent the first experimental evidence for a direct regulatory role for LH on androgen secretion in women. In addition, the data have a significant bearing on the pathogenesis of the PCO syndrome and the development of hirsutism which may be directly related to the high androgen levels in PCO women in whom the levels of LH are normally raised. The data may also offer an explanation for the mechanisms responsible for the low androgen levels in anorexia nervosa patients in whom there is a gonadotrophin deficiency.

Long-chain fatty acid esters of the adrenal-derived estrogen 5-androstene-3 beta, 17 beta-diol (ADIOL) were found to accumulate in four human mammary cancer cell lines (MCF-7, ZR-75-1, MDA-MB-231 and MDA-MB-330) when explosed to 10-30 nM ADIOL for variable time periods. At each time point examined, the monoester fraction, which represented the major component of the total lipoidal fraction, contained fatty acids linked to either the 3 beta- or 17 beta-positions. However, there was considerable variation in the ratio of 3 beta- to 17 beta-monoesters in the four cell lines. By means of reverse phase HPLC and referral to authentic synthesized compounds, each monoester fraction was found to contain a number of long-chain fatty acid components whose composition resembled that previously determined for the fatty acid esters formed from 17 beta-estradiol. A specific and measurable turnover of a subfraction of ADIOL-17 beta-monoesters composed of essential fatty acids (22:6, 20:4, 18:3) occurred in MCF-7 cells, and to a lesser extent in ZR-75-1 cells. No changes were observed with time in any of the components of the 3 beta- or 17 beta-monoester fractions in MDA-MB-231 and MDA-MB-330 cells. These results, coupled with other studies, now suggest that a very rapid turnover of some components of these lipoidal derivatives may be occurring. If so, it is possible that the system of acylation-deacylation may be involved in a transport mechanism for estrogens and perhaps other steroid hormones.

Microsomal preparations derived from bovine placenta cotyledons, previously investigated as a convenient source of fatty acyl coenzyme A: estradiol-17 beta-acyl transferase, have been shown to acylate other steroids bearing 3 beta- or 17 beta-hydroxyl groups. In the presence of 0.1 mM oleoyl CoA, the apparent Km values for dehydroepiandrosterone, testosterone, and 5-androstene-3 beta,17 beta-diol (delta 5-DIOL) were 45, 67, and 20 microM, respectively. Acylation of delta 5-DIOL occurred at either the 3 beta- or 17 beta-positions to give monoesters. Testosterone, estradiol-17 beta, and delta 5-DIOL acted as competitive inhibitors for the acylation of the 3 beta-hydroxyl group of dehydroepiandrosterone (Ki values 71, 75, and 41 microM, respectively). Such data indicate that a single enzyme of wide substrate specificity may be involved in these acylation reactions. When estrogen receptor (ER) positive and negative human mammary cancer cell lines were incubated with 10 nM [3H]delta 5-DIOL, intracellular accumulation of delta 5-DIOL long-chain fatty acid esters occurred; rates being higher (p less than 0.001) in ER negative cells (MDA-MB-231 and MDA-MB-330) compared to MCF-7 cells (ER positive), and higher (P less than 0.005) in MDA-MB-231 cells compared to ZR-75-1 cells (ER positive). After exposure to 10 nM [3H]delta 5-DIOL for 16 h, the total labeled steroid fatty acid fraction was composed predominantly of delta 5-DIOL-3 beta- and 17 beta-monoesters (approximately 85%), the remainder containing approximately equal amounts of delta 5-DIOL-diesters and dehydroepiandrosterone-3 beta-esters. Subsequent transfer to medium lacking delta 5-DIOL was accompanied by a breakdown of the labeled esters, which was more rapid in the ER positive cell lines. During this period, intracellular free delta 5-DIOL levels rapidly declined in MDA-MB-330 cells but were maintained in MCF-7 cells, presumably by binding to ER. This behavior parallels that of estradiol-17 beta previously observed in these cell lines and further emphasizes the potential importance of the adrenal-derived estrogen delta 5-DIOL in consideration of a hormone-based etiology of human breast cancer.

The sulphoconjugates of delta 5-C19O2 steroids were determined in 32 women with idiopathic hirsutism (IH). The plasma and urinary levels of 5-androsten-3 beta, 17 beta-diol (delta 5, 3 beta-diol) were 82 +/- 24 ng/ml and 169 +/- 170 micrograms/24 h respectively for the monosulphate ester and 193 +/- 101 ng/ml and 180 +/- 108 micrograms/24 h for the disulphate ester. These levels are statistically higher (p less than 0,001) as compared with the levels observed in women without IH (30 +/- 20 ng/ml and 23 +/- 13 micrograms/24 h for the monosulphate, 78 +/- 40 ng/ml and 48,0 +/- 23,7 micrograms/24 h for the disulphate). There is a correlation between the urinary levels of the monosulphate of delta 5, 3 beta-diol (y) and of the dehydroepiandrosterone sulphate (S-DHA) (x) : y (microgram/24 h) = 0,124x + 16,8; r = 0,93 (p less than 0,01) and n = 32. The plasma levels of S-DHA ranged form 1,250 ng/ml to 7,000 ng/ml in the abnormal population and from 1,000 ng/ml to 2,400 ng/ml in the normal population. A concomitant determination of plasma free androgens showed that 60 % of the dehydroepiandrosterone values were above the upper limits of normal values. Conversely, the testosterone and androstenedione levels were only higher in 20 % and 27 % of cases respectively. These results suggest that the possible importance of the sulphoconjugates of delta 5-C19O2 steroids in the IH pathogeny should not be overlooked.

Beside dehydroepiandrosterone sulphate and androsterone sulphate, 5-androstene-3 beta, 17 beta-diol-3-sulphate and testosterone sulphate were isolated and identified from the axillary sweat of sexually mature women and men. The quantities of these four C19-steroid sulphates were determined individually in the sweat of eight healthy, sexually mature women and men: dehydroepiandrosterone sulphate was found in the highest amount, it was followed by 5-androstene-3 beta, 17 beta-diol-3-sulphate and androsterone sulphate. Testosterone sulphate occurred in the lowest quantity, its excretion amounting to about one-thousandth of that of dehydroepiandrosterone sulphate. Study of the C19-steroid sulphates excreted by the apocrine sweat glands permitted an insight into the steroid metabolism in human skin, which presumably is, closely connected with the steroid hormones of peripheral blood.