Menstrual irregularity is a common complaint at presentation in women with Cushing's syndrome, although the etiology has been little studied. We have assessed <em>4</em>5 female patients (median age, 32 yr; range, 16-<em>4</em>1 yr) with newly diagnosed pituitary-dependent Cushing's syndrome. Patients were subdivided into <em>4</em> groups according to the duration of their menstrual cycle: normal cycles (NC; 26-30 days), oligomenorrhea (OL; 31-120 days), amenorrhea (AM;>> 120 days), and polymenorrhea (PM; < 26 days). Blood was taken at 0900 h for measurement of LH, FSH, PRL, testosterone, <em>androstenedione</em>, dehydroepiandrosterone sulfate, estradiol (E2), sex hormone-binding globulin (SHBG), and ACTH; cortisol was sampled at 0900, 1800, and 2<em>4</em>00 h. The LH and FSH responses to 100 micrograms GnRH were analyzed in 23 patients. Statistical analysis was performed using the nonparametric Mann-Whitney U and Spearman tests. Only 9 patients had NC (20%), 1<em>4</em> had OL (31.1%), 15 had AM (33.3%), and <em>4</em> had PM (8.8%), whereas 3 had variable cycles (6.7%). By group, AM patients had lower serum E2 levels (median, 110 pmol/L) than OL patients (225 pmol/L; P < 0.05) or NC patients (279 pmol/L; P < 0.05), and higher serum cortisol levels at 0900 h (800 vs. 602 and 580 nmol/L, respectively; P < 0.05) and 1800 h (816 vs. 557 and 523 nmol/L, respectively; P < 0.05) and higher mean values from 6 samples obtained through the day (753 vs. <em>4</em>91 and <em>4</em>59 nmol/L, respectively; P < 0.05). For the whole group of patients there was a negative correlation between serum E2 and cortisol at 0900 h (r = -0.50; P < 0.01) and 1800 h (r = -0.56; P < 0.01) and with mean cortisol (r = -0.<em>4</em>6; P < 0.05). No significant correlation was found between any serum androgen and E2 or cortisol. The LH response to GnRH was normal in <em>4</em>3.5% of the patients, exaggerated in 52.1%, and decreased in <em>4</em>.<em>4</em>%, but there were no significant differences among the menstrual groups. No differences were found in any other parameter. In summary, in our study 80% of patients with Cushing's syndrome had menstrual irregularity, and this was most closely related to serum cortisol rather than to circulating androgens. Patients with AM had higher levels of cortisol and lower levels of E2, while the GnRH response was either normal or exaggerated. Our data suggest that the menstrual irregularity in Cushing's disease appears to be the result of hypercortisolemic inhibition of gonadotropin release acting at a hypothalamic level, rather than raised circulating androgen levels.

The purpose of this research was to test the hypothesis that insulin-like growth factor-I (IGF-I) regulates estradiol (E2) synthesis in human granulosa and granulosa luteal cells. Cells from individual follicles from spontaneous and human menopausal gonadotropin/CG-stimulated cycles were cultured in serum-free medium containing <em>androstenedione</em>, IGF-I, FSH, and/or CG. At 2, <em>4</em>, and 6 days, E2 in the medium was measured by RIA. In the granulosa experiments, control cells produced basal levels of E2 at 2 days, and the levels increased with increasing follicle size. Treatment with FSH stimulated E2 production (on the average, 5-fold), and the effect was dose dependent (ED50 = 5 ng/mL or 16 mIU/mL). Incubation with IGF-I alone caused increases in E2 production comparable to those caused by FSH, and the IGF-I effect was dose dependent (ED50 = 8 ng/mL). In most cases, coincubation with FSH and IGF-I augmented E2 levels more than either hormone alone, and at <em>4</em> and 6 days the interaction was synergistic. The data from dose-response experiments suggested that the basis of the synergy between FSH and IGF-I was a marked potentiation by either hormone (approximately 10-fold) in the potency of the complementary hormone to stimulate E2 production. In the experiments with granulosa luteal cells from spontaneous and in vitro fertilization preovulatory follicles, the controls synthesized very high levels of E2 spontaneously at 2 days; however, E2 production declined 700% at <em>4</em> days, and no E2 was produced by control cells at 6 days. Treatment with FSH, CG, or IGF-I did not cause a significant increase in the high basal levels of E2 at 2 days. During subsequent culture, however, all three hormones stimulated E2 production at <em>4</em> and 6 days, but the increases were modest and not sustained. In contrast, coincubation of granulosa luteal cells with FSH plus IGF-I or CG plus IGF-I dramatically enhanced E2 production at <em>4</em> and 6 days (on the average, <em>4</em>-fold), and the effects were sustained throughout the culture period. (ABSTRACT TRUNCATED AT <em>4</em>00 WORDS)

In the golden hamster, there are marked sex differences in the Harderian gland. Male glands (which are heavier than female glands) possess two cell forms (Type I and Type II cells); female glands only exhibit the former. Female (but not male) glands contain large amounts of porphyrin, which are readily visible as solid depositions within the lumina. The weight, histology and porphyrin content of the Harderian gland was examined in intact adult male hamsters and in male hamsters castrated for 1,2 or 8 months. Castration resulted in a significant reduction in the weight of the gland, the disappearance of Type II cells, and the presence in the gland of solid porphyrin accretions. The levels of copro- and (especially) protoporphyrin were greatly increased. These changes were more marked with time after castration. When the ability of diverse androgens (testosterone, 5alpha-dihydrotestosterone, androst-<em>4</em>-ene-3,17-dione (<em>androstenedione</em>), dehydroepiandrosterone and androsterone) to prevent these changes was tested, testosterone and <em>androstenedione</em> maintained glandular weight. All the androgens maintained normal frequencies of Type II cells and all except dehydroepiandrosterone prevented deposition of porphyrin. The potencies of the various androgens in maintaining normal Harderian gland morphology and activity are compared with their effects on other somatic variables and sexual behaviour.

Circulating <em>androstenedione</em> is converted to estrone in adipose tissue, which is the principal site of estrogen biosynthesis in postmenopausal women. This reaction is catalyzed by a specific form of cytochrome P<em>4</em>50 (P<em>4</em>50arom; the product of the CYP19 gene). The fractional conversion of plasma <em>androstenedione</em> to estrone as well as the specific activity of aromatase in adipose stromal cells were previously shown to increase with advancing age. To determine whether this positive effect of aging on estrogen biosynthesis is due to an alteration in tissue levels of P<em>4</em>50arom transcripts, we quantified P<em>4</em>50arom mRNA levels in sc fat biopsy samples (n = 33) from buttocks, thighs, and abdomen of 11 women who ranged in age from 23-61 yr. Competitive polymerase chain reaction linked to reverse transcription was used to quantify P<em>4</em>50arom transcripts in total RNA that was isolated from sc fat obtained by needle aspiration. In each sample, primer extension and coamplification of a rat P<em>4</em>50arom cRNA as an internal standard were used to control possible differences in amplification efficiencies between samples. The results demonstrate that with advancing age in women, there is a progressive and statistically significant increase in adipose tissue P<em>4</em>50arom transcript levels (normalized to total RNA content) in buttocks, thighs, and abdomen (correlation coefficients: r = 0.70<em>4</em>, 0.85<em>4</em>, and 0.933, respectively). The levels of transcripts observed in the older subjects reach 2- to <em>4</em>-fold greater than those observed in the young women. Adipose tissue P<em>4</em>50arom transcript levels were highest in the buttocks, followed by the thighs, and lowest in the abdomen. This increase in P<em>4</em>50arom transcript levels is likely to be a major factor contributing to the increased extragonadal estrogen biosynthesis in elderly women.

Over 50% of patients with the polycystic ovary syndrome (PCOS) demonstrate excess levels of adrenal androgens (AAs), particularly dehydroepiandrosterone sulfate (DHS). Nonetheless, the mechanism for the AA excess remains unclear. It has been noted that in PCOS the pituitary and ovarian responses to their respective trophic factors (i.e. GnRH and LH, respectively) are exaggerated. Similarly, we have postulated that excess AAs in PCOS arises from dysfunction of the hypothalamic-pituitary-adrenal axis, due to 1) exaggerated pituitary secretion of ACTH in response to hypothalamic CRH, 2) excess sensitivity/responsivity of AAs to ACTH stimulation, or 3) both. To test this hypothesis we studied 12 PCOS patients with AA excess (HI-DHS; DHS,>> 8.1 mumol/L or 3000 ng/mL), 12 PCOS patients without AA excess (LO-DHS; DHS, < 7.5 mumol/L or 2750 ng/mL), and 11 controls (normal subjects). Each subject underwent an acute 90-min ovine CRH stimulation test (1 microgram/kg) and an 8-h incremental i.v. stimulation with ACTH-(1-2<em>4</em>) at doses ranging from 20-2880 ng/1.5 m2.h) with a final bolus of 0.25 mg. All patient groups had similar mean body mass indexes and ages, and both tests were performed in the morning during the follicular phase (days 3-10) of the same menstrual cycle, separated by <em>4</em>8-96 h. During the acute ovine CRH stimulation test, no significant differences in the net maximal response (i.e. change from baseline to peak level) for ACTH, dehydroepiandrosterone (DHA), <em>androstenedione</em> (A<em>4</em>), or cortisol (F) or for the DHA/ACTH, A<em>4</em>/ACTH, or F/ACTH ratios was observed. Nonetheless, the net response of DHA/F and the areas under the curve (AUCs) for DHA and DHA/F indicated a greater response for HI-DHS vs. LO-DHS or normal subjects. The AUC for A<em>4</em> and A<em>4</em>/F and the delta A<em>4</em>/delta F ratio (delta = net maximum change) indicated that HI-DHS and LO-DHS had similar responses, which were greater than that of the normal subjects, although the difference between LO-DHS patients and normal subjects reached significance only for the AUC of the A<em>4</em> response. No difference in the sensitivity (i.e. threshold or minimal stimulatory dose) to ACTH was noted between the groups for any of the steroids measured. Nonetheless, the average dose of ACTH-(1-2<em>4</em>) required for a threshold response was higher for DHA than for F and A<em>4</em> in all groups. No difference in mean responsivity (slope of response to incremental ACTH stimulation) was observed for DHA and F between study groups, whereas the responsivity of A<em>4</em> was higher in HI-DHS patients than in normal or LO-DHS women. The net maximal and the overall (i.e. AUC) responses of DHA were greater for HI-DHS than for normal or LO-DHS women. The response of A<em>4</em> and the delta A<em>4</em>/delta F ratio were greater for HI-DHS patients than for LO-DHS patients or normal subjects. Alternatively, HI-DHS and LO-DHS patients had similar overall responses (i.e. AUC) for A<em>4</em> or A<em>4</em>/F, although both were greater than those of normal subjects. The relative differences in response to incremental ACTH stimulation between steroids was consistent for all subject groups studied, i.e. A<em>4</em>>> F or DHA. In conclusion, our data suggest that AA excess in PCOS patients is related to an exaggerated secretory response of the adrenal cortex for DHA and A<em>4</em>, but not to an altered pituitary responsivity to CRH or to increased sensitivity of these AAs to ACTH stimulation. Whether the increased responsivity to ACTH for these steroids is secondary to increased zonae reticularis mass or to differences in P<em>4</em>50c17 alpha activity, particularly of the delta <em>4</em> pathway, remains to be determined.

The present study investigates the physiological significance of dehydroepiandrosterone, dehydroepiandrosterone sulfate, T, <em>androstenedione</em> (Delta(<em>4</em>)), dihydrotestosterone (DHT), estrone (E1), and E2 on recombinant human FSH- (rhFSH) resistant type <em>4</em> follicles obtained from immature mice. Type <em>4</em> follicles of a diameter of 100-120 microm with one or two granulosa cell layers around the oocyte and an intact basal lamina with theca cells were isolated from the ovaries of 11-d-old BDF-1 mice and cultured with medium alone (control) or with dehydroepiandrosterone, dehydroepiandrosterone sulfate, T, Delta(<em>4</em>), DHT, E1, or E2 at concentrations ranging from 1 x 10(-11) to 1 x 10(-7) M for <em>4</em> d. We examined the mean diameters of type <em>4</em> follicles, levels of immunoreactive (IR)-inhibin, and E2 and progesterone in the culture media on day <em>4</em>. In addition, we evaluated follicular cell proliferation by immunofluorescence staining with 5-bromo-2'-deoxyuridine. All tested androgens significantly increased the diameter of type <em>4</em> follicles in a dose-dependent manner without the production of IR-inhibin and E2. The nuclei of granulosa cells in type <em>4</em> follicles cultured with all tested androgens exhibited intense 5-bromo-2'-deoxyuridine-positive staining, compared with those of controls. In contrast, neither E1 nor E2 had any stimulatory effects. The stimulatory effects of T, Delta(<em>4</em>), or DHT were inhibited by an AR antagonist in a dose-related fashion but not by an aromatase inhibitor. Furthermore, all tested androgens had a synergistic effect with rhFSH on follicular growth and the production of IR-inhibin and E2. These results demonstrated that neither adrenal nor ovarian androgens are arteriogenic but that they stimulate type <em>4</em> follicles unresponsive to rhFSH and augment the responsiveness of these follicles to rhFSH.

It has been reported that a high proportion of abdominal fat is associated with increased plasma androgen concentrations in women. Although less evidence is available, abdominal obesity appears to be associated with low plasma testosterone (T) levels in men. We have therefore examined in 80 men (aged 36.3 +/- 3.2 years, mean +/- SD) the correlations between body fatness, adipose tissue (AT) distribution measured by computed tomography (CT), and circulating levels of the following steroids measured by radioimmunoassay after extraction from serum and chromatography: dehydroepiandrosterone (DHEA), <em>androstenedione</em> (delta <em>4</em>-DIONE), androst-5-ene-3 beta,17 beta-diol (delta 5-DIOL), T, estrone, and estradiol. Sex hormone-binding globulin (SHBG) levels were also determined. T, adrenal C19 steroids, and SHBG levels were negatively correlated with total body fatness indices and abdominal fat deposition measured by CT (-.23 < or = -.55, .0001 < or = P < or = .05), whereas estrone showed positive correlations with these body fatness and AT distribution indices. Covariance analysis showed that after control for the concentration of the adrenal steroid precursor delta 5-DIOL, there was no residual association between T levels and adiposity variables. Furthermore, multivariate analyses showed that steroid and SHBG levels could explain from 20% (visceral AT area measured by CT) to <em>4</em>0% and <em>4</em>2% (body mass index [BMI], waist circumference, and waist to hip ratio [WHR]) of the variation in adiposity variables (.0001 < or = P < or = .05), with delta 5-DIOL being the best single correlate of body fatness and abdominal fat deposition in men.(ABSTRACT TRUNCATED AT 250 WORDS)

Studies have shown that immune responses are depressed in male mice, but not in proestrus females after trauma-hemorrhage (TH), resulting in increased mortality from subsequent sepsis in male mice compared with female mice. These gender-specific alterations in immune function are believed to be due to differences in sex steroid levels. Aromatase is a key enzyme in the sex steroid biosynthesis. Although earlier studies have shown that aromatase inhibitors prevent thymic atrophy in aged male rats, it remains unknown whether the use of <em>4</em>-hydroxy-<em>androstenedione</em> (<em>4</em>-OHA) after TH in male mice has any salutary effects on the depressed immune responses. Male C3H/HeN mice were sham operated or subjected to trauma (i.e., midline laparotomy) and hemorrhagic shock (30+/-5 mmHg for 90 min) followed by adequate fluid resuscitation. <em>4</em>-OHA (5 mg/kg) or vehicle was administrated s.c. just before resuscitation. At 2 h after resuscitation, the mice were killed, and spleens were harvested. Splenocyte proliferation, interleukin (IL-2), interferon (IFN-gamma), and IL-10 release and expression of androgen (AR) and estrogen receptors (ER)-alpha and -beta by immunoblot and reverse transcription-polymerase chain reaction (RT-PCR) were assessed. In another group, sepsis was induced by cecal ligation and puncture (CLP) 3 days after resuscitation, and survival was measured over a period of 10 days. A significant decrease in splenocyte proliferation, IL-2, and IFN-gamma release and increased release of IL-10 were observed in vehicle-treated mice. Animals treated with <em>4</em>-OHA showed increased splenocyte proliferation, IL-2, and IFN-gamma release, and decreased IL-10 release. Immunoblot analysis showed decreased expression of the cytosolic AR, but no significant difference in the cytosolic and nuclear ER-alpha and -beta expression was observed in the vehicle-treated group after TH. In addition, AR and ER-beta mRNA expression was increased, whereas ER-alpha expression decreased in the vehicle-treated group after TH. ER-alpha expression decreased and ER-beta expression increased in the nucleus of <em>4</em>-OHA treated mice as determined by immunoblot. There was no difference in the cytosolic AR expression in the <em>4</em>-OHA-treated group after TH. AR and ER-beta mRNA expression was unaffected, whereas ER-alpha expression increased under such conditions. In additional groups, the increased mortality rate after TH and subsequent sepsis was significantly reduced by <em>4</em>-OHA treatment. Thus, <em>4</em>-OHA seems to be a novel and useful adjunct for restoring the depressed immune functions in males after TH and for decreasing mortality rates from subsequent sepsis.

Research has shown that exposure to androgens and progestogens can cause undesirable biological responses in the environment. To date, however, no detailed or direct study of their presence in wastewater treatment plants has been conducted. In this study, nine androgens, nine progestogens, and five estrogens were analyzed in influent and final effluent wastewaters in seven wastewater treatment plants (WWTPs) of Beijing, China. Over a period of three weeks, the average total hormone concentrations in influent wastewaters were 3562 (Wujiacun WWTP)-5<em>4</em>00 ng/L (Fangzhuang WWTP). Androgens contributed 96% of the total hormone concentrations in all WWTP influents, with natural androgen (androsterone: 2977±739 ng/L; epiandrosterone: 6<em>4</em>0±263 ng/L; and <em>androstenedione</em>: 270±132 ng/L) being the predominant compounds. The concentrations of synthetic progestogens (megestrol acetate: <em>4</em>1±25 ng/L; norethindrone: 6.5±3.3 ng/L; and medroxyprogesterone acetate: 6.0±3.2 ng/L) were comparable to natural ones (progesterone: 66±36 ng/L; 17α,20β-dihydroxy-<em>4</em>-progegnen-3-one: <em>4</em>.9±1.2 ng/L; 21α-hydroxyprogesterone: 8.5±3.0 ng/L; and 17α-hydroxyprogesterone: 1.5±0.95 ng/L), probably due to the wide and relatively large usage of synthetic progestogens in medical therapy. In WWTP effluents, androgens were still the dominant class accounting for 60% of total hormone concentrations, followed by progestogens (2<em>4</em>%), and estrogens (16%). <em>Androstenedione</em> and testosterone were the main androgens detected in all effluents. High removal efficiency (91-100%) was found for androgens and progestogens compared with estrogens (67-80%), with biodegradation the major removal route in WWTPs. Different profiles of progestogens in the receiving rivers and WWTP effluents were observed, which could be explained by the discharge of a mixture of treated and untreated wastewater into the receiving rivers.

Recent reports indicate that girls with premature adrenarche are at risk of developing functional ovarian hyperandrogenism and polycystic ovarian syndrome (PCOS). As insulin and insulin-like growth factors (IGFs) have been implicated in the pathogenesis of PCOS, we hypothesize that they may also have a role in the hyperandrogenism of premature adrenarche. Thirty-five prepubertal girls (23 Caribbean Hispanics and 12 Black African-Americans) underwent a 60-min ACTH and LH-releasing hormone test. Insulin sensitivity (S(I)) was assessed using the frequently sampled i.v. glucose tolerance test with tolbutamide. Fasting levels of IGF-I, IGF-binding protein-1 (IGFBP-1), IGFBP-3, sex hormone-binding globulin, and free testosterone (T) were also obtained. The mean age of the patients was 6.8 yr, and bone age was 8.0 yr. Twenty-five patients had a family history of noninsulin-dependent diabetes mellitus and 19 patients had acanthosis nigricans. The mean S(I) for the entire group was 6.78 +/- 5.21 x 10(-<em>4</em>) min/microU x mL (normal prepubertal S(I), 6.5 +/- 0.5<em>4</em> x 10(-<em>4</em>) min(-1) x microU(-1) x mL(-1)). However, 15 of the 35 girls had an S(I) that was more than 2 SD below the mean reported for normal prepubertal children. Of these 15 patients, 13 were obese, and 1<em>4</em> had acanthosis nigricans. For the entire group of girls, the mean ACTH-stimulated levels of 17-hydroxypregnenolone (17OHPreg), dehydroepiandrosterone (DHEA), <em>androstenedione</em> (AS), 17-hydroxyprogesterone (17OHP), and T and the ACTH-stimulated ratios of 17OHPreg/17OHP, 17OHPreg/DHEA, 17OHP/AS, and DHEA/AS did not differ from the levels reported for Tanner stage II-III pubertal girls. The girls were divided into two groups based on their S(I) (group I, S(I) >2 SD below the mean for age; group II, normal S(I)). The group I girls with a reduced S(I) had significantly higher ACTH-stimulated levels of 17OHPreg (group I, 760 +/- 87.8<em>4</em> ng/dL; group II, <em>4</em>28.9 +/- <em>4</em>6.28 ng/dL; P = 0.002), 17OHPreg/17OHP ratio (group I, 3.95 +/- 0.36; group II, 2.96 +/- 0.35; P = 0.05), 17OHPreg/DHEA (group I, 2.06 +/- 0.21; group II, 1.<em>4</em> +/- 0.13; P = 0.01), and free T (group I, 1 +/- 0.23 ng/dL; group II, 0.<em>4</em>9 +/- 0.19 ng/dL; P = 0.01<em>4</em>). Levels of sex hormone-binding globulin were lower in the group I girls. Furthermore, for the entire group of girls, the S(I) correlated inversely with ACTH-stimulated levels of 17OHPreg, DHEA, and AS and the ACTH-stimulated ratio of 17OHPreg/17OHP. IGF-I correlated inversely with S(I) (r = -0.9<em>4</em>; P < 0.001) and correlated directly with the ACTH-stimulated levels of 17OHPreg (r = 0.8; P < 0.001) and AS (r = 0.63; P < 0.05). IGF-I also correlated with the ACTH-stimulated ratios of 17OHPreg/17OHP (r = 0.61; P < 0.05), 17OHPreg/DHEA (r = 0.9; P < 0.001), 17OHP/AS (r = 0.79; P < 0.001), and DHEA/AS (r = 0.96; P < 0.001). IGFBP-1 correlated inversely with the ACTH-stimulated levels of 17OHPreg (r = -0.38; P < 0.05) and DHEA (r = -0.36; P < 0.05). To summarize, the ACTH-stimulated delta5-steroid levels were higher in prepubertal girls with premature adrenarche and reduced S(I). There was a significant inverse correlation among ACTH-stimulated hormone levels, S(I), and IGFBP-1, whereas IGF-I correlated directly with ACTH-stimulated androgens. These findings support the hypothesis that insulin and IGFs may have a role in the hyperandrogenism of premature adrenarche just as they do in PCOS. Hence, in certain girls with premature adrenarche, hyperandrogenism may be the first presentation of PCOS and/or insulin resistance.

Adipose tissue contains both aromatase and 17 beta-steroid dehydrogenase activity. Therefore, to see whether there was a relationship between obesity and certain parameters of androgen and estrogen metabolism we infused 88 women, mean age 51.1 +/- 0.3 years and mean weight 1<em>4</em>0 +/- 3 lbs, with 3H-testosterone (T)/1<em>4</em>C-estradiol (E2) and 3H-<em>androstenedione</em> (A)/1<em>4</em>C-estrone (E1) on separate occasions. Blood samples were obtained during the infusion and all urine was collected for <em>4</em> days following the start of the infusion. The blood samples were analyzed for radioactivity as A, T, E1, E2, and dihydrotestosterone (DHT) and the urines were analyzed for radioactivity as E1 and E2. From these data we calculated the percent of A converted to T ([rho]A,TBB = percent of A infused measured as T in the blood), [rho]T,ABB [rho]E1,E2BB, and [rho]E2,E1BB. We also measured the ratio of radioactivity as 3H-DHT to radioactivity as 3H-A (CRA,DHT) and 3H-T (CRT,DHT) during the respective androgen infusions. From the ratio of 3H/1<em>4</em>C as estrone or estradiol in the urine following 3H-A or 3H-T infusions, we calculated the percent of A or T that was aromatized to E1 or E2 ([rho]A,E1BM; [rho]T,E2BM). When the data from these women were related to weight or Quetelet's Index (QI = wt/ht2) by unweighted linear regression, the only values that were significantly correlated with weight and QI were [rho]T,E2BM and [rho]A,E1BM; for all other [rho] and CR values there was no correlation with weight or QI. We conclude that peripheral aromatization is positively correlated with adiposity but androgen interconversions and estrogen interconversions are not related to adiposity.

The effect of inhaled beclomethasone dipropionate and budesonide on the adrenal function was studied in 30 children (aged 7 to 15 years) with mild bronchial asthma. The trial was designed as a prospective double-blind parallel study of the effect of stepwise increase of either beclomethasone dipropionate or budesonide from 200 micrograms through <em>4</em>00 micrograms, to 800 micrograms daily in three consecutive periods of <em>4</em> weeks. At the end of each period, the adrenal stress response was evaluated by measurements of serum cortisol and <em>androstenedione</em> during a short adrenocorticotropic hormone test. The unstimulated diurnal production of glucocorticosteroids was assessed by measurements of free cortisol in 2<em>4</em>-hour urine samples. Free cortisol in urine was found a valid measure of the total diurnal excretion of cortisol metabolites, since it exhibited a good correlation to the fractional cortisol metabolites measured by gas chromatography. The adrenal response to adrenocorticotropic hormone stimulation was unaffected by treatment or dose. The unstimulated diurnal production of glucocorticosteroids demonstrated a highly significant dose-related suppression in response to the inhaled steroids. No significant difference was found between the two topical steroids (probability value 5.3%), and yet the suppression was apparent in the group of children treated with beclomethasone dipropionate but not in the group of children treated with budesonide. Further studies are desirable in order to ascertain whether budesonide offers an improved ratio between beneficial anti-inflammatory effect and unwanted systemic activity.

The site of action of aminoglutethimide (AG) has been investigated. An initial study was performed on 10 postmenopausal patients with advanced breast cancer who had taken 1000 mg AG per day and 20 mg hydrocortisone (HC) twice daily (b.d.) for greater than 3 months. There was a 15.5 +/- 5.6 s.e.-fold rise in 17-OH progesterone and a <em>4</em>.9 +/- 0.9 s.e.-fold rise in <em>4</em> delta <em>androstenedione</em> but no rise in cortisol or oestrone 30 min after short Synacthen tests. These results suggested that peripheral aromatisation was a more important site of AG action than adrenal desmolase, and that adrenal 11 beta hydroxylase was inhibited. Since aromatase is more sensitive than desmolase to AG in vitro, lower doses of AG alone (i.e. without HC) were assessed for endocrine effects in 13 further post-menopausal women with advanced breast cancer. All of these patients tolerated 125 mg AG b.d., but 3 could not tolerate the conventional maximum dose. Oestrone levels on 125 mg AG b.d. were suppressed below pretreatment levels and were not significantly different from those on 500 mg AG b.d. alone, or with the addition of HC. Oestradiol levels were suppressed to a similar extent. Dehydroepiandrosterone sulphate (DHA-S) levels were not suppressed by AG alone, but fell on addition of HC. The endocrine results show low dose AG alone is an effective and well tolerated inhibitor of the peripheral production of oestrogens in postmenopausal patients. Therapeutic trials are now possible. DHA-S is not a marker of AG effect.

The effects of castration on bone histomorphometry and mineral homeostasis were compared in male and female rats. Measurements were performed <em>4</em> weeks after sham operation or gonadectomy. Orchiectomy produced increases in serum calcium and decreases in serum testosterone and <em>androstenedione</em>, whereas ovariectomy produced decreases in serum estradiol and testosterone. Orchiectomy did not alter static bone histomorphometric measurements of the tibial diaphysis, whereas ovariectomy increased cross-sectional and medullary areas, lowered endosteal tetracycline-labeled surface length, and markedly increased endosteal nonlabeled surface length. Orchiectomy decreased mean periosteal bone formation rate and mean periosteal bone apposition rate, whereas ovariectomy increased both measurements. Orchiectomy and ovariectomy markedly diminished trabecular area and trabecular surface length at the tibial metaphysis. Orchiectomy did not alter the number of osteoclasts per mm trabecular surface or the percentage of trabecular surface covered by osteoclasts, whereas ovariectomy increased both measurements. These findings indicate that gonadal hormones produce separate and distinct effects on bone metabolism as determined by histomorphometry in male and female rats.

Salivary concentrations of unconjugated steroids reflect those for free steroids in serum although concentrations may differ because of salivary gland metabolism. Samples for salivary steroid analysis are stable for up to 7 days at room temperature, one month or more at <em>4</em> degrees C and three months or more at -20 degrees C. When assessed against strict criteria, the evidence shows that salivary cortisol in evening samples or following dexamethasone suppression provides a reliable and effective screen for Cushing's syndrome. Sequential salivary cortisol measurements are also extremely helpful for the investigation of suspected cyclical Cushing's syndrome. There is potential for the identification of adrenal insufficiency when used with Synacthen stimulation. Salivary 17-hydroxyprogesterone and <em>androstenedione</em> assays are valued as non-invasive tests for the home-monitoring of hydrocortisone replacement therapy in patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency. The diagnostic value of salivary oestradiol, progesterone, testosterone, dehydroepiandrosterone and aldosterone testing is compromised by rapid fluctuations in salivary concentrations of these steroids. Multiple samples are required to obtain reliable information, and at present the introduction of these assays into routine laboratory testing is not justified.

Aromatase has been purified to homogeneity from human placental microsomes based on detection of its catalytic activities in the eluates from columns of octylamino-Sepharose <em>4</em>B, hydroxylapatite, Mono S, hydroxylapatite HCA, and Mono Q. The purified preparation shows only one band corresponding to the apparent subunit molecular weight of 51,000 daltons on sodium dodecyl sulfate-polyacrylamide gel. The aromatase in the presence of NADPH and NADPH-cytochrome P-<em>4</em>50 reductase converts testosterone to 17 beta-estradiol with the high specific activity of 103 nmol/min/mg of protein. However, whether the preparation is reduced by sodium dithionate chemically or by NADPH and the reductase enzymatically, its reduced, CO-difference spectrum has no peak at about <em>4</em>50 nm and has only a small peak at about <em>4</em>20 nm, probably due to its inactivation in spite of the catalytically full activity in the same preparation. The absolute spectrum of the aromatase exhibits a Soret peak at <em>4</em>23 nm in the absence of testosterone and addition of testosterone to the aromatase sample makes its absorption peak shift gradually from <em>4</em>23 to 393 nm (high spin type peak), which is a usual characteristic in the spectrum of cytochrome P-<em>4</em>50. The reconstituted aromatase system efficiently catalyzes aromatization of <em>4</em>-<em>androstenedione</em>, 19-hydroxy-<em>4</em>-<em>androstenedione</em> as well as testosterone. 16 alpha-Hydroxy-<em>4</em>-<em>androstenedione</em> and 16 alpha-hydroxytestosterone are also aromatized less efficiently and 19-nortestosterone is aromatized least efficiently. The reconstituted aromatase could scarcely oxidize various xenobiotics examined, suggesting a strict and narrow substrate specificity of this enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)

OBJECTIVE

To assess the hypothesis that the climacteric ovary is a functional endocrine gland.

METHODS

Review of the English-speaking literature as it relates to the physiology and pathophysiology of the climacteric ovary.

RESULTS

By several accounts, the climacteric ovary appears to be a gonadotropin-dependent androgen-producing gland. Although the estrogen-producing potential of the climacteric ovary remains a matter of controversy, most studies would suggest limited aromatase activity.

CONCLUSIONS

[1] The climacteric ovary is not a defunct endocrine organ. [2] The climacteric ovary is a site of gonadotropin reception and action. [3] The climacteric ovary contributes few if any estrogens to the circulating pool by way of direct production. [<em>4</em>] Circulating estrogens are derived virtually exclusively from the extraglandular conversion of androgens, a proportion of which are of ovarian origin. [5] The climacteric ovary contributes <em>4</em>0% and 20% of the total production rates of T and <em>androstenedione</em>, respectively. [6] Androgen biosynthesis by the climacteric ovary is partially gonadotropin-dependent.

The antitumor, endocrine, hematological, biochemical, and side effects of chronic second-line treatment with the antiprogestin mifepristone (RU) <em>4</em>86) were investigated in 11 postmenopausal patients with metastatic breast cancer. We observed one objective response, 6 instances of short-term stable disease, and <em>4</em> instances of progressive disease. Mean plasma concentrations of adrenocorticotropic hormone (P less than 0.05), cortisol (P less than 0.001), <em>androstenedione</em> (P less than 0.01), and estradiol (P less than 0.002) increased significantly during treatment accompanied by a slight decrease of sex hormone binding globulin levels, while basal and stimulated gonadotropin levels did not change significantly. The increased basal cortisol levels could not be further stimulated by synacthen, nor suppressed by 1 mg of dexamethasone. Plasma estradiol concentrations were significantly correlated with both <em>androstenedione</em> (P less than 0.05) and cortisol levels (P less than 0.01). The percentage of eosinophilic white blood cells (P less than 0.02) and mean plasma creatinine concentration (P less than 0.05) increased significantly. Side effects frequently occurred during long-term treatment and appeared to be caused mainly by the antiglucocorticoid properties of the drug. It is concluded that antiprogestins form a new treatment modality in the endocrine treatment of human breast cancer. New antiprogestins with less antiglucocorticoid side effects might be especially of value as an adjunct to antiestrogenic treatment in view of our finding that combined antiestrogenic and antiprogestational treatment caused additive growth-inhibitory effects in rat mammary tumors.

BACKGROUND

Metformin treatment of women with polycystic ovary syndrome (PCOS) is widespread, as determined by studies with diverse patient populations. No comparative examination of weight changes or metabolite responses to different doses has been reported.

OBJECTIVE

The aim of this study was to determine whether different doses of metformin (1500 or 2550 mg/d) would have different effects on body weight, circulating hormones, markers of inflammation, and lipid profiles.

METHODS

The study included prospective cohorts randomized to two doses of metformin.

METHODS

The study was performed at a university teaching hospital with patients from gynecology/endocrinology clinics.

METHODS

The patients studied were obese (body mass index, 30 to <37 kg/m2; n = <em>4</em>2) and morbidly obese (body mass index,>> or =37 kg/m2; n = <em>4</em>1) women with PCOS.

METHODS

Patients were randomized to two doses of metformin, and parameters were assessed after <em>4</em> and 8 months.

METHODS

The main outcome measures were changes in body mass, circulating hormones, markers of inflammation, and lipid profiles.

RESULTS

Intention to treat analyses showed significant weight loss in both dose groups. Only the obese subgroup showed a dose relationship (1.5 and 3.6 kg in 1500- and 2550-mg groups, respectively; P = 0.0<em>4</em>). The morbidly obese group showed similar reductions (3.9 and 3.8 kg) in both groups. Suppression of androstenedione was significant with both metformin doses, but there was no clear dose relationship. Generally, beneficial changes in lipid profiles were not related to dose.

CONCLUSIONS

Weight loss is a feature of protracted metformin therapy in obese women with PCOS, with greater weight reduction potentially achievable with higher doses. Additional studies are required to determine whether other aspects of the disorder may benefit from the higher dose of metformin.

The aim of this study was to evaluate the effects of metformin in addition to diet and exercise on endocrine and metabolic disturbances in women with polycystic ovary syndrome (PCOS) in a prospective, double-blind, randomized, placebo (PBO) control trial. Thirty women with insulin resistance and PCOS received lifestyle modification and 1500 mg of metformin or placebo for <em>4</em> months. Before and after treatment, body mass index, waist/hip ratio, blood pressure, hirsutism, and menstrual patterns were evaluated. Serum concentrations of gonadotropins, androgens, progesterone, glucose, insulin, and lipids were measured. Lifestyle interventions resulted in similar weight and menstrual cycle's improvements in both groups. A significant reduction in serum fasting insulin, HOMA index, waist and testosterone levels was only observed with metformin. There were no significant changes in <em>androstenedione</em>, dehydroepiandrosterone sulfate, gonadotropins, and lipids levels. No other changes were observed in hirsutism or blood pressure. These findings suggest that metformin has an additive effect to diet and exercise to improve parameters of hyperandrogenism and insulin resistance. Although, a small decrease in body weight trough lifestyle changes could be enough to improve menstrual cycles in insulin-resistant women with PCOS.

Hyperactivity of the HPA-system in major depression is reflected by an increased secretion of adrenal hormones especially cortisol and dehydroepiandrosterone (DHEA). In women for whom androgenicity is associated with cardiovascular disorders the dominant source of <em>androstenedione</em> and testosterone secretion are the adrenal glands. To date, there is only sparse information about the regulation of <em>androstenedione</em>, testosterone and dihydrotestosterone (DHT) concentrations in women with severe major depression.Therefore, 11 pre- and postmenopausal, severely depressed, hypercortisolemic women (Hamilton Depression Scale, 31.3+/-5.9; age, 28-77 yrs; mean, <em>4</em>8. 1+/-18.1 yrs) and 11 age-matched healthy female controls (age, 2<em>4</em>-81 yrs; mean, <em>4</em>7.9+/-21.5 yrs) underwent a 2<em>4</em> hour (h) blood sampling starting at 0800 h with 30-minute sampling intervals. By applying multivariate analysis of covariance with age as covariate, <em>androstenedione</em>, testosterone and DHT plasma levels at 0900 h show a trend for elevated concentrations in depressed women compared to controls (F(1,19)=2.7; P=0.057). Univariate F tests reveal a significant difference between the groups for <em>androstenedione</em> (<em>4</em>. 19+/-1.571 vs 2.58<em>4</em>+/-1.257 nmol/l; P<0.05) testosterone (1.110+/-0. 278 vs 0.833+/-0.3<em>4</em>7 nmol/l; P<0.05) and DHT (0.656+/-0.207 vs 0. <em>4</em>83+/-0.2<em>4</em>2 nmol/l; P<0.05). Mean ACTH (16.<em>4</em>+/-10.<em>4</em> vs 10.<em>4</em>+/-2.<em>4</em> pmol/l; P=0.89), LH (13.5+/-11.8 vs 8.9+/-9.2 IU/l; P=0.12), FSH (35. 2+/-33.1 vs 31.3+/-35.7 IU/l; P=0.67) and estradiol (135.<em>4</em>+/-157.<em>4</em> vs 82.2+/-85.1 pmol/l; P=0.20) plasma levels did not differ between patients and controls. Further, there was a trend towards an age related decline in testosterone secretion in healthy controls (r=-0. 2<em>4</em>; P=0.08) which did not occur in depressed patients (r=0.17; P=0. 96), while the calculated ratio of DHEA to testosterone was similar in both groups (0.2+/-0.1<em>4</em> vs 0.13+/-0.7; P=0.21, unpaired t-test). In conclusion, <em>androstenedione</em>, testosterone and DHT concentrations all were increased in hypercortisolemic women with severe major depression. These findings are best explained as a consequence of an overstimulation of the adrenal glands through pituitary and hypothalamic sites of the HPA-system.

Dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin; TCDD) is the most toxic congener of a large class of environmental pollutants. Several studies have shown that TCDD exposure reduced fecundity and ovulatory rate in rats and increased the incidence of endometriosis in monkeys. Recent work suggests that TCDD's endocrine-disrupting effects are, at least in part, caused by a direct action at the ovary. Although the factors involved in TCDD-induced toxicity are still under investigation, several studies have shown that TCDD induces programmed cell death, or apoptosis, in various tissues and may act in a similar fashion in the ovary. In the present study, we set out to evaluate the in vitro effects of TCDD on steroid secretion, specifically estradiol-17beta (E2) and progesterone, by human luteinized granulosa cells (LGC), and to further determine whether TCDD is capable of inducing apoptosis in this cell type. Human LGC were obtained from women participating in an in vitro fertilization program. Medium, with or without three different concentrations of TCDD and substrates [<em>androstenedione</em> (A<em>4</em>) or pregnenolone], was added to each culture. The media were collected at <em>4</em>, 8, 12, 2<em>4</em>, 36, and <em>4</em>8 h and were assayed by RIA. At 2<em>4</em> and <em>4</em>8 h, the LGC were fixed for assessment of DNA fragmentation via an in situ immunofluorescence technique. Transmission electron microscopy was also performed on LGC after 2<em>4</em> and <em>4</em>8 h with TCDD. TCDD, at all concentrations tested (3.1 pM, 3.1 nM, and 3.1 microM), significantly reduced E2 accumulation in the media at 8, 12, and 2<em>4</em> h, compared with controls. At 36 and <em>4</em>8 h, TCDD treatment (at 3.1 microM) caused a significant increase in E2, compared with controls. The effect of TCDD on E2 was abolished with the addition of A<em>4</em>. TCDD treatment did not alter progesterone accumulation. Apoptosis increased at 2<em>4</em> h with 3.1 microM TCDD, with no apparent effect at 3.1 nM. By <em>4</em>8 h, however, TCDD increased apoptosis in a dose-dependent manner. Transmission electron microscopy showed ultrastructural differences in LGC with 3.1 microM TCDD at 2<em>4</em> and <em>4</em>8 h. Collectively, the results of the present study suggest that TCDD perturbs E2 secretion by depletion of A<em>4</em> precursor and increases apoptotic cell death of human LGC in a dose- and time-dependent fashion.

Adrenal masses are more and more frequently detected by adrenal ultrasound, computed tomography or nuclear magnetic resonance carried out for a reason other than the suspicion of adrenal disease (incidentalomas). The findings of an incidentaloma still leaves many diagnostic and therapeutic questions open. We report the results of a multicentric retrospective evaluation of patients with adrenal incidentalomas, performed by a Study Group of the Italian Society of Endocrinology. According to the definition of incidentaloma, exclusion criteria a priori were: severe or paroxysmal hypertension, frank hypokalemia and clinical signs of hypercortisolism or hyperandrogenism. 29 centers participated in the study and the data obtained by questionnaire were collected in 2 centers for final elaboration. Center 1 carried out the epidemiological and clinical evaluation. Basal and dynamic hormonal evaluation of 786 among the 1013 cases recruited were performed in our center (center 2). Functional studies included: diurnal rhythm of cortisol, urinary free cortisol (UFC), ACTH, DHEAS, 17-OH progesterone, testosterone, <em>androstenedione</em>, supine and upright plasma renin activity (PRA) and aldosterone, urinary aldosterone, urinary catecholamines and VMA. The hormonal dynamic evaluation included the overnight dexamethasone suppression test (1 mg), CRH test and ACTH test. In our study, 89% (702 patients) of adrenal incidentalomas were non-hypersecretory masses; 6.2% (<em>4</em>9 patients) showed a preclinical Cushing's syndrome (PCS) (at least two altered parameters of pituitary-adrenal axis); 3.<em>4</em>% (27 patients) were pheochromocytomas; 0.89% (7 patients) were aldosteronomas. One tumor was a masculinizing adrenocortical carcinoma. Two hundred sixty patients underwent surgical exploration and the histological diagnosis showed: 138 adenomas (53%), 32 carcinomas (12%), 26 pheochromocytomas (10%). 16 myelolipomas (8%), 13 cystic lesions (5.5%), 7 tumors of neuronal lineage (3%). 12 metastases (<em>4</em>%), 13 others (5%). The 138 patients with adenomas had the following hormonal diagnosis: 103 nonfunctional adenomas (7<em>4</em>%), 31 PCS (23%) and <em>4</em> cases of hyperaldosteronism (3%). In the patients with PCS an abnormal dexamethasone suppression test was found in 86% of cases (37/<em>4</em>1 patients). Values for ACTH were low in 78% (32/<em>4</em>1 patients). UFC was elevated in 6<em>4</em>% of patients, the diurnal rhythm of cortisol evaluated in 1<em>4</em> patients was absent in 7. Only in 50% of cases DHEAS values (12/2<em>4</em> patients) were decreased, whereas they were normal in the other 50%. Interestingly, 8 patients with normal DHEAS and normal UFC showed nonsuppressible cortisol by dexamethasone test (1 mg). Blunted ACTH response to CRH was detected in 9 of 1<em>4</em> patients (6<em>4</em>%). Thus our data suggest that the best parameter for evaluating subclinical hypercortisolism seems to be the overnight dexamethasone suppression test. In 27 patients with pheochromocytoma 2<em>4</em>-hour urinary catecholamine and VMA levels were elevated in 86 and <em>4</em>6% of cases respectively. In 7 patients with hyperaldosteronism upright PRA was suppressed in 100% of cases and aldosterone plasma levels were elevated in 6 patients (86%); serum potassium level was slightly decreased in 60% of cases. In 86 of 138 histologically proven adenomas, DHEAS levels were: normal in 59% of patients, decreased in 36% and elevated in <em>4</em>.6%, whereas in 22 of 32 cortical carcinomas evaluated. DHEAS levels were normal in 63% of cases, decreased in 18% and elevated 18%. Post-ACTH 17-OH progesterone levels were elevated in 52% (62/118 patients) of non-functioning adenomas and in 2 of <em>4</em> carcinomas. Not enough data are yet available postoperatively. In summary, endocrine evaluation can lead to the identification of a nonnegligible number of cases of clinically unsuspected pheochromocytomas and subtle hypercortisolism (about 3.<em>4</em> and 6.2%, respectively of all adrenal incidentalomas), while cases of primary subclinical aldosteronism are rarely found. (ABSTRACT TRUNCATED)

The rat central nervous system (CNS) has previously been shown to synthesize pregnenolone (PREG) and convert it to progesterone (PROG) and 7 alpha-hydroxy-PREG (7 alpha-OH PREG). Astrocytes, which participate to the regulation of the CNS function, might be involved in the metabolism of neurosteroids. Purified type 1 astrocytes were obtained from fetal rat forebrain with the use of selective culture conditions and were identified by immunostaining with specific antibodies (GFAP+, A2B5-). They were plated at low, intermediate, or high densities (2.5-5 x 10(5), 1-2 x 10(6), or <em>4</em>-8 x 10(6) cells/dish, respectively) and maintained for 21 d. They were then incubated with 1<em>4</em>C-PREG and 1<em>4</em>C-DHEA for 2<em>4</em> h and the steroids extracted from cells and media were analyzed. Most radioactive derivatives were released into incubation media. Two metabolic pathways were mainly observed. PREG and DHEA were oxidized to PROG and <em>androstenedione</em> (ADIONE), respectively, [3 beta-hydroxysteroid-dehydrogenase, delta 5->><em>4</em> 3-ketosteroid-isomerase (3 beta-HSD) activity], and converted to 7 alpha-OH PREG and 7 alpha-OH DHEA, respectively (7 alpha-hydroxylase activity). After low density plating, the formation of PROG and ADIONE was approximately 10% of incubated radioactivity, tenfold larger than that of 7 alpha-hydroxylated metabolites. In contrast, after high density plating, low levels of PROG and ADIONE were formed, whereas the conversion to either 7 alpha-OH PREG or 7 alpha-OH DHEA was>> or = 50%. The results expressed per cell indicated that the 3 beta-HSD activity was almost completely inhibited at high cell density, in contrast to the 7 alpha-hydroxylation which was maintained or increased. The pattern of steroid metabolism was related to cell density at the time of measurement and not to an early commitment of cells: when primary cultures were plated at high density (8 x 10(6) cells/dish), then subcultured after several dilutions (3-, 9-, or 27-fold), the 3 beta-HSD activity was recovered only at low density. Furthermore, when 5 x 10(5) cells were centrifuged and the resulting clusters were plated, 3 beta-HSD activity was decreased, whereas steroid 7 alpha-hydroxylation was enhanced. This implies that cell density per se, but neither cell number nor a diffusible factor(s) is involved in the regulation of steroid metabolism. We conclude that astrocytes in culture metabolize PREG and DHEA, and that the metabolic conversions and, therefore, the related enzymatic activities depend on cell-to-cell contacts.(ABSTRACT TRUNCATED AT <em>4</em>00 WORDS)