11β-HSD1 Activity Research Articles

Diabetes and Insulin Injection Modalities: Effects on Hepatic and Hippocampal Expression of 11β-Hydroxysteroid Dehydrogenase Type 1 in Juvenile Diabetic Male Rats.

Dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis is often encountered in diabetes, leading to several clinical complications. Our recent results showing an elevated tetrahydrocortisol/tetrahydrocorticosterone ratio in morning urine of diabetic children compared to that of controls suggest an increased nocturnal activity of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) in the former. We hypothesized that these observations could be explained by a reduced inhibition of hepatic 11β-HSD1 activity by exogenous insulin owing to its subcutaneous (SC) administration and absence of first hepatic passage. Additionally, we hypothesized that hippocampal 11β-HSD1 activity might also be impaired by diabetes. We therefore measured HPA axis activity and 11β-HSD1 expression and activity in liver and hippocampus in streptozotocin-induced diabetic juvenile rats treated with SC or intraperitoneal (IP) insulin. Plasma corticosterone levels were elevated in untreated diabetic rats during the resting phase and restored by both types of insulin treatment. The mRNA expression and activity of 11β-HSD1 were increased in the untreated diabetic group in liver. Although diabetes was controlled equally whatever the route of insulin administration, liver 11β-HSD1 gene expression and activity was decreased only in the IP group, suggesting that a first hepatic pass is needed for 11β-HSD1 hepatic inhibition. In hippocampus, 11β-HSD1 activity was elevated in the untreated diabetic group but restored by both types of insulin treatment. Thus, these data extend our findings in diabetic children by showing impairment of hippocampal 11β-HSD1 in diabetes and by demonstrating that IP is preferable to SC insulin administration to restore 11β-HSD1 activity in liver.

Mechanisms for the species difference between mouse and pig oocytes in their sensitivity to glucorticoids.

Although in vitro exposure to physiological concentrations of glucorticoids did not affect maturation of mouse oocytes, it significantly inhibited nuclear maturation of pig oocytes. Studies on this species difference in oocyte sensitivity to glucocorticoids will contribute to our understanding of how stress/glucocorticoids affect oocytes. We showed that glucorticoid receptors (NR3C1) were expressed in both oocytes and cumulus cells (CCs) of both pigs and mice; however, while cortisol inhibition of oocyte maturation was overcome by NR3C1 inhibitor RU486 in pigs, it could not be relieved by RU486 in mice. The mRNA level of 11β-hydroxysteroid dehydrogenase 1 (HSD11B1) was significantly higher than that of HSD11B2 in pig cumulus-oocyte complexes (COCs), whereas HSD11B2 was exclusively expressed in mouse COCs. Pig and mouse cumulus-denuded oocytes (DOs) expressed HSD11B2 predominantly and exclusively, respectively. In the presence of cortisol, although inhibiting HSD11B2 decreased maturation rates of COCs in both species, inhibiting HSD11B1 improved maturation of pig COCs while having no effect on mouse COCs. Cortisol-cortisone interconversion observation confirmed high HSD11B1 activities in pig oocytes but none in mouse oocytes, a higher HSD11B2 activity in mouse than in pig oocytes, and a rapid cortisol-cortisone interconversion in pig COCs catalyzed by HSD11B1 from CCs and HSD11B2 from DOs. In conclusion, the species difference in glucocorticoid sensitivity between pig and mouse oocytes is caused by their different contents/ratios of HSD11B1 and HSD11B2, which maintain different concentrations of active glucocorticoids. While cortisol inhibited pig oocytes by interacting with NR3C1, glucocorticoid suppression of mouse oocytes was apparently not mediated by NR3C1.

The relationship between dioxins exposure and risk of prostate cancer with steroid hormone and age in Vietnamese men

Although Vietnam's massive herbicide exposure in 1960s and 1970s was clearly injurious to health, not all causal relationships have been clarified. We therefore explored associations among dioxins, steroid hormones, age and prostate cancer risk in men. We compared serum levels of dioxin, steroid hormones and prostate specific antigen (PSA) in men aged 56–81years from herbicide-exposed hotspots (n=50) with those from non-sprayed regions (n=48). Mean serum levels of dioxin congeners in the hotspot group were 1.5–11.3 times higher than the non-sprayed group depending on specific compound. Levels of testosterone, estradiol and 3β-hydroxysteroid dehydrogenase (3β-HSD) activity in the hotspot group were also significantly higher than in non-sprayed group. Estradiol levels were significantly related to levels of several specific dioxin derivatives in both group. Significant positive correlations were also found between DHT and 1234678-HpCDD or 1234678-HpCDF; and between 3β-HSD activity and 123678-HxCDD, 123478-HxCDF, 123678-HxCDF, or HxCB#169. After adjusting for age, body mass index, and tobacco use, multiple linear regressions showed levels of dihydrotestosterone (DHT), estradiol, testosterone and 3β-HSD activity were not associated with dioxins in the two groups; however, levels of DHT, testosterone and 3β-HSD activity increased significantly with age in the hotspot group. The hotspot and non-sprayed groups did not significantly differ in PSA levels. But six of the hotspot subjects had PSA levels >3ng/mL, 3 of whom were suspected to have prostate cancer (PC) after digital rectal examination. Our findings suggest that dioxin exposure can lead to increased levels of several sex steroid hormones with age. The correlation of dioxin with steroid hormone levels and prostate cancer risk should be studied further.

11 Targeting 11β-HSD1 to promote angiogenesis – consequences for solid tumour growth

11β-Hydroxysteroid dehydrogenase type 1 (11β-HSD1), which generates active glucocorticoids from inactive precursors, is expressed in glucocorticoid target tissues, including the arterial wall. Since active glucocorticoids are anti-angiogenic 11β-HSD1 inhibitors enhance angiogenesis and may have therapeutic potential in ischaemia. However increased angiogenesis may be detrimental in tumours. This investigation tested the hypothesis that 11β-HSD1 inhibition promotes angiogenesis and tumour growth in mouse models of squamous cell carcinoma (SCC) and pancreatic ductal adenocarcinoma (PDA). Murine tumour cells (1 × 106) were injected subcutaneously into mice (female, 10–12 weeks, FVB/C57Bl6/J) receiving RM-1 diet with, or without (Control), the 11β-HSD1 inhibitor UE2316 (175 mg/kg). Tumour size was measured every 2–3 days for 2–3 weeks. Sections of tumours were stained for vascular markers (CD31, alpha-smooth muscle actin) for quantification of vessel density. Steroid/drug levels were measured in plasma/tissues using liquid chromatography tandem mass spectrometry whilst 11β-HSD1 activity was assayed in tissue homogenates by high performance liquid chromatography. The effects of 11β-HSD1 inhibition on angiogenesis were examined ex vivo using an aortic ring assay. Data are mean±SEM. 11β-HSD1 inhibition increased SCC tumour growth in FVB mice (p 11β-HSD1 inhibition does not promote angiogenesis in SCC or PDA tumours, but may increase SCC growth through a mechanism involving reduced fibrosis. Whether 11β-HSD1 inhibition increases angiogenesis in glucocorticoid-sensitive tumours remains to be established.

11β-HSD1 Modulates the Set Point of Brown Adipose Tissue Response to Glucocorticoids in Male Mice

Glucocorticoids (GCs) are potent regulators of energy metabolism. Chronic GC exposure suppresses brown adipose tissue (BAT) thermogenic capacity in mice, with evidence for a similar effect in humans. Intracellular GC levels are regulated by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) activity, which can amplify circulating GC concentrations. Therefore, 11β-HSD1 could modulate the impact of GCs on BAT function. This study investigated how 11β-HSD1 regulates the molecular architecture of BAT in the context of GC excess and aging. Circulating GC excess was induced in 11β-HSD1 knockout (KO) and wild-type mice by supplementing drinking water with 100 μg/mL corticosterone, and the effects on molecular markers of BAT function and mitochondrial activity were assessed. Brown adipocyte primary cultures were used to examine cell autonomous consequences of 11β-HSD1 deficiency. Molecular markers of BAT function were also examined in aged 11β-HSD1 KO mice to model lifetime GC exposure. BAT 11β-HSD1 expression and activity were elevated in response to GC excess and with aging. 11β-HSD1 KO BAT resisted the suppression of uncoupling protein 1 (UCP1) and mitochondrial respiratory chain subunit proteins normally imposed by GC excess. Furthermore, brown adipocytes from 11β-HSD1 KO mice had elevated basal mitochondrial function and were able to resist GC-mediated repression of activity. BAT from aged 11β-HSD1 KO mice showed elevated UCP1 protein and mitochondrial content, and a favorable profile of BAT function. These data reveal a novel mechanism in which increased 11β-HSD1 expression, in the context of GC excess and aging, impairs the molecular and metabolic function of BAT.

Modulation of steroidogenesis by vitamin D3 in granulosa cells of the mouse model of polycystic ovarian syndrome

ABSTRACTPolycystic ovarian syndrome (PCOS) is the most common endocrine disorder of women of reproductive age characterized by polycystic ovarian morphology, anovulation or oligomenorrhea, and hyperandrogenism. It is shown that disruption in the steroidogenesis pathway caused by excess androgen in PCOS is a critical element of abnormal folliculogenesis and failure in dominant follicle selection. Vitamin D plays an important role in the regulation of ovulatory dysfunction and can influence genes involved in steroidogenesis in granulosa cells. In the present study, we investigated the effects of vitamin D3 on steroidogenic enzyme expression and activities in granulosa cell using a PCOS mouse model. In our study, the PCOS mouse model was developed by the injection of dehydroepiandrosterone (DHEA) for 20 days. The mRNA and protein expression levels of genes involved in steroidogenesis in granulosa cells were compared between polycystic and normal ovaries using real-time PCR and Western blotting assays. Granulosa cells of DHEA-induced PCOS mice were then cultured with and without vitamin D3 and mRNA and protein expression levels of steroidogenic enzymes and serum 17beta-estradiol and progesterone levels were investigated using qRT-PCR, western blot, and radioimmunoassay, respectively. Steroidogenic enzymes including Cyp11a1, StAR, Cyp19a1, and 3β-HSD were upregulated in granulosa cells of PCOS mice when compared to normal mice. Treatment with vitamin D3 decreased mRNA and protein expression levels of steroidogenic enzymes in cultured granulosa cells. Vitamin D3 also decreased aromatase and 3β-HSD activity that leads to decreased 17beta-estradiol and progesterone release. This study suggests that vitamin D3 could modulate the steroidogenesis pathway in granulosa cells of PCOS mice that may lead to improving follicular development and maturation. This is a step towards a possible conceivable treatment for PCOS.Abbreviations: AMHR-II: anti-müllerian hormone receptor-II; 3β-HSD: 3β-hydroxysteroid dehydrogenase; Cyp11a1: Cytochrome P450 Family 11 Subfamily A Member 1; Cyp19a1: cytochrome P450 aromatase; DHEA: dehydroepiandrosterone; FSH: follicle stimulating hormone; FSHR: follicle stimulating hormone receptor; IVF: in vitro fertilization; 25OHD: 25-hydroxy vitamin D; OHSS: ovarian hyperstimulation syndrome; PCOS: polycystic ovarian syndrome; P450scc: P450 side-chain cleavage enzyme; StAR: steroidogenic acute regulatory protein; VDRs: vitamin D receptors

Regulation of 11β-hydroxysteroid dehydrogenase type 1 following caloric restriction and re-feeding is species dependent

Evidence in the current literature suggests that expression and activity of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), a key regulatory enzyme in glucocorticoid metabolism, is elevated in the liver and reduced in visceral adipose tissue and skeletal muscle following caloric restriction (CR). In order to investigate the influence of CR on 11β-HSD1 in more detail, we assessed expression and activity of 11β-HSD1 in several tissues in two independent CR and re-feeding animal models. Levels and activity of 11β-HSD1 after CR and re-feeding were measured [mouse liver and pig liver, pig visceral adipose tissue and pig skeletal muscle] using semi-quantitative RT-PCR, Western Blot analysis, and HPLC.After CR, no significant difference on mRNA levels was detected in mouse liver. But 11β-HSD1 mRNA expression was upregulated after subsequent re-feeding. In contrast, 11β-HSD1 protein expression after CR was significantly up-regulated, while no difference was detected after re-feeding. Interestingly, upregulation of protein after CR (1.4-fold) was lower than the increase in enzymatic activity (2.6-fold). Furthermore, while no difference was observed in protein levels after two weeks re-feeding, 11β-HSD1 activity increased 2.5-fold. In pig tissues neither 11β-HSD1 mRNA levels, protein expression or enzyme activity were influenced after CR and re-feeding.Overall, the results demonstrate species-dependent differences in regulation of 11β-HSD1 following CR and suggest the presence of an additional regulation step for 11β-HSD1 activity in mouse liver.

11-Beta Dehydrogenase Type 2 Activity Is Not Reduced in Treatment Resistant Hypertension.

Decreased renal 11-beta dehydrogenase type 2 (11β-HSD2) activity, as reflected by an increased urinary free cortisol to cortisone ratio (UFF/UFE), is associated with having hypertension (HTN). The current study was conducted to determine if reduced 11β-HSD2 activity is also associated with having resistant HTN. We evaluated 55 consecutive patients with RHTN, defined as blood pressure (BP) ≥140/90 mm Hg despite using ≥3 antihypertensive medications including a diuretic, and 38 patients whose BP was controlled on ≤3 medications to serve as a non-RHTN comparator group. All patients underwent biochemical evaluation, including measurement of 24-hour urinary UFF/UFE. The 2 study groups had similar demographic characteristics. Systolic, diastolic BP, and number of antihypertensive medications were greater in patients with uncontrolled RHTN vs. the control group (167.5 ± 28.2/91.2 ± 18.8 vs. 126.6 ± 11.4/77.8 ± 8.65 mm Hg and 4.31 ± 1.23 vs. 2.74 ± 0.6, respectively). The 24-hour UFF was 13.6 ± 11.8 vs. 14.3 ± 10.7 µg/24 h and UFE was 64.9 ± 36.3 vs. 76.1 ± 44 µg/24 h such that the UFF/UFE was 0.22 ± 0.16 vs. 0.19 ± 0.09 in RHTN vs. the control group. This ratio was not associated to age, race, gender, and body mass index. An elevated UFF/UFE was not present in this large cohort of patients with uncontrolled RHTN. This suggests that reduced conversion of cortisol to cortisone does not contribute to the development of RHTN.

FSH Stimulation promotes progesterone synthesis and output from human granulosa cells without luteinization.

Can granulosa cells produce progesterone (P) in response to FSH stimulation? FSH actively promotes P synthesis and output from granulosa cells without luteinization by up-regulating the expression and increasing enzymatic activity of 3β-hydroxysteriod dehydrogenoase (3β-HSD), which converts pregnenolone to P. Serum P level may rise prematurely prior to ovulation trigger in stimulated IVF cycles and adversely affect implantation and clinical pregnancy rates by impairing endometrial receptivity. A translational research study. Human ovarian cortical samples (n=15) and non-luteinizing FSH-responsive human mitotic granulosa cell line (HGrC1) were stimulated with rec-FSH at 12.5, 25 and 50 mIU/ml concentrations for 24 and 48 h. FSH receptor expression was knocked-down and up-regulated in the granulosa cells using short hairpin RNA (shRNA) technology and activin-A administration, respectively. The expressions of the steroidogenic enzymes were analyzed at mRNA level by real-time quantitative RT-PCR, and protein level by western blot and immunoprecipitation assay. The enzymatic activity of 3β-HSD was measured using a spectrophotometric method. In vitro estradiol (E2) and P productions of the cells before and after FSH stimulation were measured by electro-chemiluminescence immunoassay method. Stimulation of the HGrC1 cells with FSH resulted in a dose-dependent increase in the mRNA and protein level of 3β-HSD. Overall, when all time points and FSH doses were analyzed collectively, FSH significantly up-regulated the mRNA expression of its own receptor (3.73±0.06-fold, P<0.001), steroidogenic acute regulatory protein (stAR, 1.7±0.03-fold, P<0.01), side-chain cleavage enzyme (SCC, 1.75±0.03-fold, P<0.01), aromatase (4.49±0.08-fold, P<0.001), 3β-HSD (1.68±0.02-fold, P<0.01) and 17β-hydroxy steroid dehydrogenase (17β-HSD, 2.16±0.02-fold, P<0.01) in the granulosa cells. Expression of 17α-hydroxylase (17α-OH, 1.03±0.01-fold P>0.05) did not significantly change. Similar changes were observed in the protein expression analysis of these enzymes on western blotting after FSH stimulation. FSH significantly increased 3β-HSD, 17β-HSD and aromatase in a dose-dependent manner but did not affect 17α-OH. Protein expression of P was increased along with 3β-HSD after FSH stimulation, which was further evidenced by immunoprecipitation assay. Enzymatic activity of 3β-HSD was significantly enhanced by FSH administration in the HGrC1 cells in a dose-dependent manner. In line with these findings P output (1.05±0.3 vs. 0.2±0.1 ng/ml, respectively, P<0.001) from the samples stimulated with FSH were significantly increased along with E2 (1918±203 vs. 932±102 pg/ml, respectively, P<0.001) compared to unstimulated controls. FSH-induced increase in 3β-HSD expression was amplified and reversed in the HGrC1 cells when FSH receptor expression was up-regulated by activin-A and down-regulated with shRNA, respectively. As only the effect of FSH was studied we cannot extrapolate our findings to the potential effects of HMG and recombinant LH. This data provides a molecular explanation for the largely unexplained phenomenon of P rise during the follicular phase of gonadotropin stimulated IVF cycles. Our findings may progress the research to uncover potential mechanisms for preventing premature P rise that appears to be associated with inferior outcomes in women undergoing IVF. Funded by the School of Medicine and the Graduate School of Health Sciences of Koc University. All authors declare no conflict of interest. None.

Novel 11β-hydroxysteroid dehydrogenase 1 inhibitors reduce cortisol levels in keratinocytes and improve dermal collagen content in human ex vivo skin after exposure to cortisone and UV.

Activity and selectivity assessment of new bi-aryl amide 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) inhibitors, prepared in a modular manner via Suzuki cross-coupling, are described. Several compounds inhibiting 11β-HSD1 at nanomolar concentrations were identified. Compounds 2b, 3e, 7b and 12e were shown to selectively inhibit 11β-HSD1 over 11β-HSD2, 17β-HSD1 and 17β-HSD2. These inhibitors also potently inhibited 11β-HSD1 activity in intact HEK-293 cells expressing the recombinant enzyme and in intact primary human keratinocytes expressing endogenous 11β-HSD1. Moreover, compounds 2b, 3e and 12e were tested for their activity in human skin biopsies. They were able to prevent, at least in part, both the cortisone- and the UV-mediated decreases in collagen content. Thus, inhibition of 11β-HSD1 by these compounds can be further investigated to delay or prevent UV-mediated skin damage and skin aging.

Maternal salivary cortisone to cortisol ratio in late pregnancy: An improved method for predicting offspring birth weight

Overexposure to maternal cortisol in utero has been associated with lower birth weight of offspring. In order to regulate fetal exposure to this hormone, placental expression of 11-β-hydroxysteroid dehydrogenase-2, an enzyme that converts active cortisol to inactive cortisone, increases across pregnancy. Because of this increase in 11-βHSD2 activity, measuring maternal cortisol in isolation may not reflect actual fetal exposure to the hormone. Previous work by Hellgren et al. (2016) has shown that maternal serum cortisone:cortisol ratio was a better predictor of offspring birth weight than cortisol measured in isolation. This paper sought to replicate these results when examining maternal salivary cortisone:cortisol ratio. Data come from 55 pregnant women from Auckland, New Zealand. Cortisol and cortisone were measured in saliva samples collected at waking and prior to going to sleep on two consecutive weekdays between 34 and 36 weeks of gestation. We found that salivary cortisol and cortisone followed the expected diurnal rhythm and that cortisone was higher than cortisol at both times of day. Maternal bedtime cortisone:cortisol ratio was significantly and inversely related to offspring birth weight. However, waking and bedtime cortisol, as well as waking cortisone:cortisol ratio, were unrelated to birth weight. These results show that maternal salivary cortisone:cortisol ratio, like serum cortisone:cortisol ratio, is a more sensitive biomarker for predicting infant birth weight than cortisol measured in isolation. This ratio could be a valuable, minimally-invasive measurement for future studies interested in understanding the relationship between maternal HPA-axis function and offspring birth weight.

Effects of Cellular 11β-hydroxysteroid Dehydrogenase 1 on LPS-induced Inflammatory Responses in Synovial Cell Line, SW982.

11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) catalyzes the conversion of inactive cortisone into active cortisol, which has pleiotropic roles in various biological conditions, such as immunological and metabolic homeostasis. Cortisol is mainly produced in the adrenal gland, but can be locally regenerated in the liver, fat, and muscle. Its diverse actions are primarily mediated by binding to the glucocorticoid receptor. SW982, a human synovial cell line, expresses 11β-HSD type 1, but not type 2, that catalyzes the conversion of cortisone to cortisol. In this study, therefore, we investigated the control of lipopolysaccharide (LPS)-induced inflammatory responses by prereceptor regulation-mediated maintenance of cortisol levels. Preliminarily, cell seeding density and incubation period were optimized for analyzing the catalytic activity of SW982. Additionally, cellular 11β-HSD1 still remained active irrespective of monolayer or spheroid culture conditions. Inflammatory stimulants, such as interleukin (IL)-1β, tumor necrosis factor (TNF)α, and LPS, did not affect the catalytic activity of 11β-HSD1, although a high dose of LPS significantly decreased its activity. Additionally, autocrine effects of cortisol on inflammatory responses were investigated in LPS-stimulated SW982 cells. LPS upregulated pro-inflammatory cytokines, including IL-6 and IL-1β, in SW982 cells, while cortisol production, catalyzed by cellular 11β-HSD1, downregulated LPS-stimulated cytokines. Furthermore, suppression of NFκB activation-mediated pro-inflammatory responses by cortisol was revealed. In conclusion, the activity of cellular 11β-HSD1 was closely correlated with suppression of LPS-induced inflammation. Therefore, these results partly support the notion that prereceptor regulation of locally regenerated cortisol could be taken into consideration for treatment of inflammation-associated diseases, including arthritis.

Effects of 11β-Hydroxysteroid Dehydrogenase Type 1 Enzyme in the Liver in Fructose Induced Metabolic Syndrome Rat Model

Excessive expression and activity of 11β-hydroxysteroid dehydrogenase enzyme type 1 (11β-HSD1) enzyme in mature adipocyte leads to obesity and metabolic syndrome. Fructose in drinking water was proven to induce metabolic syndrome in male Wistar rat. Hence, the aim of the study was to assess the effects of expression and activity of 11β-HSD1 enzyme in the liver in an established metabolic syndrome rat model induced by fructose drinking water. Twelve male Wistar rats were randomly divided into two groups: Control group, C (n=6) and Fructose drinking water 20%, F20 (n=6). The food and fluid intake were given as ad libitum for eight weeks. At the end of the experiment, the expression of 11β-HSD1 enzyme in the liver was measured by immunohistochemical staining method. The score was given according to the intensity of the staining of the granules in the hepatocyte cytoplasm which was measured using double-blinded method. Meanwhile, the activity of 11β-HSD1 enzyme in the liver was measured using ELISA technique. Following eight weeks of consumption of fructose drinking water, the F20 group showed an increased in both expression and activity of 11β-HSD1 in the liver. The obtained data clearly suggest that 11β-HSD1 enzymes in the liver may play a role in the development of metabolic syndrome and its complications in male Wistar rat.

Alternations in miRNA Expression in Chronic Stress-Induced Ageing of Leydig Cells

Purpose: Chronic stress represents a critical influence on ageing of Leydig cells in testis. We aimed to investigate expression changes of microRNA (miRNA) associated with stress-induced ageing in Leydig cells.Methods: Brown Norway rats were separated into three groups, young control group, stress group and age group. Physiological changes, including serum corticosterone and testosterone levels, 11β-HSD (11β- hydroxysteroiddehydrogenase) activity, GSH/GSSG (glutathione/glutathione disulfide) ratio and DNA damage after chronic stress treatment were tested by radioimmunoassay and Immunohistochemistry. Differentially expressed miRNAs in Leydig cells between young control group and stress group were identified by miRNA microarray analysis, followed by target prediction of miRNAs. Functional annotation of predicted targets of miRNA was carried out using GeneSpring software and miRNA-target gene interaction network was built by String software.Results: Under chronic stress, increased serum corticosterone levels and reduced testosterone levels were observed. Meanwhile, decline of 11β-HSD activity and GSH/GSSG ratio, increase of DNA damage were also shown in stress group. After performed miRNA microarray analysis, three differentially expressed miRNAs were screened out, including rno-miR-31a-5p, rno-miR-192-5p and rno-miR-191a-3p. Targets of them were mainly involved in apoptosis, cell cycle and transport. In miRNA-target gene interaction network, targets of miR-31a-5p (Tp53, Ywhae and Ndel1), and targets of miR-192-5p (Cdk2, Rac2, Stk3) were with higher degree.Conclusion: These data suggest a central role of miRNA-regulated gene expression in response to chronic stress, especially dys-regulation of rno-miR-31a-5p and rno-miR-192-5p play a vital role in ageing of Leydig cells in testis.

Neuromodulatory effect of oestradiol in the metabolism of ovarian progesterone and oestradiol during dioestrus II: participation of the superior mesenteric ganglion.

The aims of the present study were to determine: (1) whether oestradiol (E2) in the superior mesenteric ganglion (SMG) modifies the release of ovarian progesterone (P4), androstenedione (A2) and E2, the activity and gene expression of 3β-hydroxysteroid dehydrogenase (3β-HSD) and 20α-HSD and the expression of P450 aromatase (Cyp19a1) and (2) whether any such modifications are related to changes in ovarian nitric oxide (NO) and noradrenaline (NA) levels during dioestrus II. Using an ex vivo SMG-ovarian nervous plexus-ovary system, ovarian P4 release was measured following the addition E2 plus tamoxifen (Txf) (10-6M) to the ganglion, whereas A2, E2, NA and NO were measured following the addition of E2 alone. Steroids were measured by radioimmunoassay, NA concentrations were determined by HPLC and gene expression was evaluated using reverse transcription-polymerase chain reaction. Oestradiol in the ganglion decreased ovarian P4, E2 and NA release, as well as 3β-HSD activity, but increased the release of A2 and nitrites, as well as the 20α-HSD expression and its activity. No changes were observed in Cyp19a1 gene expression. The addition of E2 plus Txf to the ganglion reversed the effects of E2 alone. The action of oestradiol in SMG favours the beginning of functional luteolysis, due to an increase in NO release and a decrease in NA in the ovary. These results may help elucidate the role of E2 in hormone-dependent pathologies in women.

Lindane Induces Spermatotoxicity and inhibits Steroidogenesis in Adult Rats

This study was conducted to investigate the influence of lindane on spermatogenesis and testicular biochemical parameters and to get insight into its mechanism. Adult male albino rats were treated orally with lindane at doses of 0, 1, 2 or 4 mg/kg/day for 30 consecutive days. Testes weight and sperm count and motility were significantly decreased. Testicular activities of 3β-HSD, 17β-HSD were significantly inhibited. Testicular cholesterol level was significantly increased while glycogen and sialic acid content were significantly decreased. Testicular activities of LDH-X, γ-GT, β-glucuronidase and acid phosphatase (ACP) were significantly decreased. Aldo-ketoreductase activity was significantly decreased in response to 2 or 4 mg/kg/day of lindane while, protein carbonyl contentswere significantly increased. Hydrogen peroxide and hydroxyl radical generation and LPO were significantly increased. The enzymatic antioxidants SOD, CAT and GPx and the non-enzymatic antioxidants GSH and Vit C were significantly decreased. Lindane at doses 2 and 4 mg/kg/day showed a significant reduction in Vit E, while at a dose of 1 mg/kg/day did not show any significant change. In conclusion, lindane inhibits spermatogenesis, steroidogenesis and suppresses Sertoli cell marker enzymes which may be due, partly, to oxidative stress. Lindane enhances ROS generation and LPO and depletes antioxidant enzymes and non-enzymatic antioxidants.

11β-Hydroxysteroid Dehydrogenase Type 1 Inhibition Attenuates the Adverse Effects of Glucocorticoids on Dermal Papilla Cells.

PurposeGlucocorticoids, stress-related hormones, inhibit hair growth. Intracellular glucocorticoid availability is regulated by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). 11β-HSD1 was recently detected in keratinocytes and fibroblasts. However, the expression of 11β-HSD1 in human hair follicles remains unknown. We aimed to examine 11β-HSD1 expression in human dermal papilla cells (DPCs) and to investigate whether modulation of 11β-HSD1 activity can regulate the negative effects of glucocorticoids on DPCs.Materials and Methods11β-HSD1 expression in normal human scalp skin was examined by immunohistochemistry. 11β-HSD1 protein was detected in Western blots of human DPCs. Cultured human DPCs were treated with cortisol with or without a selective 11β-HSD1 inhibitor and subsequently stained for Ki-67 antibody. Expression levels of 11β-HSD1, Wnt5a, alkaline phosphatase (ALP), and vascular endothelial growth factor (VEGF) were analyzed by Western blotting.Results11β-HSD1 was detected in dermal papilla in human scalp skin by immunohistochemistry. Human DPCs expressed 11β-HSD1 protein in vitro. Furthermore, cortisol stimulated the expression of 11β-HSD1 in DPCs. Glucocorticoids decreased cellular proliferation and the expression of Wnt5a, ALP, and VEGF in DPCs. A specific 11β-HSD1 inhibitor significantly attenuated the anti-proliferative effects of cortisol and reversed the cortisol-induced suppression of Wnt5a, ALP, and VEGF expression in DPCs.ConclusionOur data demonstrated the expression of 11β-HSD1 in human DPCs and revealed that inhibition of 11β-HSD1 activity can partially prevent the negative effect of glucocorticoids on DPCs, suggesting the possible application of 11β-HSD1 inhibitors for stress-related hair loss.

Visualizing the Interrenal Steroidogenic Tissue and Its Vascular Microenvironment in Zebrafish.

This protocol introduces how to detect differentiated interrenal steroidogenic cells through a simple whole-mount enzymatic activity assay. Identifying differentiated steroidogenic tissues through chromogenic histochemical staining of 3-β-Hydroxysteroid dehydrogenase /Δ5-4 isomerase (3β-Hsd) activity-positive cells is critical for monitoring the morphology and differentiation of adrenocortical and interrenal tissues in mammals and teleosts, respectively. In the zebrafish model, the optical transparency and tissue permeability of the developing embryos and larvae allow for whole-mount staining of 3β-Hsd activity. This staining protocol, as performed on transgenic fluorescent reporter lines marking the developing pronephric and endothelial cells, enables the detection of the steroidogenic interrenal tissue in addition to the kidney and neighboring vasculature. In combination with vibratome sectioning, immunohistochemistry, and confocal microscopy, we can visualize and assay the vascular microenvironment of interrenal steroidogenic tissues. The 3β-Hsd activity assay is essential for studying the cell biology of the zebrafish interrenal gland because to date, no suitable antibody is available for labeling zebrafish steroidogenic cells. Furthermore, this assay is rapid and simple, thus providing a powerful tool for mutant screens targeting adrenal (interrenal) genetic disorders as well as for determining disruption effects of chemicals on steroidogenesis in pharmaceutical or toxicological studies.

Soy isoflavones exert beneficial effects on letrozole-induced rat polycystic ovary syndrome (PCOS) model through anti-androgenic mechanism

Context: Soy is the main source of phytoestrogens, which has long been used as traditional food. One major subtype of phytoestrogens includes isoflavones and they are scientifically validated for their beneficial actions on many hormone-dependent conditions. Objective: The present study examines the effect of soy isoflavones on letrozole-induced polycystic ovary syndrome (PCOS) rat model. Materials and methods: PCOS was induced in Sprague–Dawley rats with of 1 mg/kg letrozole, p.o. once daily for 21 consecutive days. Soy isoflavones (50 and 100 mg/kg) was administered for 14 days after PCOS induction. Physical parameters (body weight, oestrous cycle determination, ovary and uterus weight) metabolic parameters (oral glucose tolerance test, total cholesterol), steroidal hormone profile (testosterone and 17β-oestradiol), steroidogenic enzymes (3β-hydroxy steroid dehydrogenase (HSD) and 17β-HSD), oxidative stress and histopathology of ovary were studied. Results: Soy isoflavones (100 mg/kg) treatment significantly altered the letrozole-induced PCOS symptoms as observed by decreased body weight gain (p < 0.05), percentage diestrous phase (p < 0.001), testosterone (p < 0.001), 3β-HSD (p < 0.01) and 17β-HSD (p < 0.001) enzyme activity and oxidative stress. Histological results reveal that soy isoflavones treatment in PCOS rats resulted in well-developed antral follicles and normal granulosa cell layer in rat ovary. Discussion: Treatment with soy isoflavones exerts beneficial effects in PCOS rats (with decreased aromatase activity) which might be due to their ability to decrease testosterone concentration in the peripheral blood. Conclusion: Analysis of physical, biochemical and histological evidences shows that soy isoflavones may be beneficial in PCOS.

Getting to the heart of intracellular glucocorticoid regeneration: 11β-HSD1 in the myocardium.

Corticosteroids influence the development and function of the heart and its response to injury and pressure overload via actions on glucocorticoid (GR) and mineralocorticoid (MR) receptors. Systemic corticosteroid concentration depends largely on the activity of the hypothalamic–pituitary–adrenal (HPA) axis, but glucocorticoid can also be regenerated from intrinsically inert metabolites by the enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), selectively increasing glucocorticoid levels within cells and tissues. Extensive studies have revealed the roles for glucocorticoid regeneration by 11β-HSD1 in liver, adipose, brain and other tissues, but until recently, there has been little focus on the heart. This article reviews the evidence for glucocorticoid metabolism by 11β-HSD1 in the heart and for a role of 11β-HSD1 activity in determining the myocardial growth and physiological function. We also consider the potential of 11β-HSD1 as a therapeutic target to enhance repair after myocardial infarction and to prevent the development of cardiac remodelling and heart failure.