<strong class="sub-title"> Background: </strong> Patients with steroid <em>5α</em>-reductase 2 deficiency (<em>5α</em>-RD) caused by SRD5A2 (OMIM #607306) variants present variable genotypes and phenotypes. The genotype-phenotype correlations remain unclear.

Methods: We investigated genotype-phenotype correlations of SRD5A2 variants in a large Chinese single-center cohort. Phenotypes were categorized using the external masculinization score (EMS), urethral meatus and gonad position, and penile length-standard deviation score.

Results: Of the 130 included patients, 113 had hypospadias, and 17 had a normal urethral meatus position. Testosterone/dihydrotestosterone (T/DHT) values were not significantly associated with phenotypic severity (p = 0.539-0.989). Of the 31 SRD5A2 variants, including 10 novel variants, p.R227Q was the most prevalent (39.62%), followed by p.Q6* (16.92%), p.R246Q (13.46%), and p.G203S (10.38%). Compared to biallelic missense mutations, biallelic nonsense mutations were associated with a lower EMS and urethral meatus score (p = 0.009 and p = 0.024, respectively). Patients homozygous for p.R227Q exhibited mild and variable phenotypes, while those homozygous for p.Q6*, p.R246Q, or p.G203S showed consistently severe phenotypes. The phenotypes were variable and milder in patients with compound heterozygosity for p.R227Q and these mutations.

Conclusion: T/DHT does not predict phenotype severity. The most prevalent SRD5A2 variant in Han Chinese is p.R227Q, which is associated with milder phenotypes and greater phenotypic variability. SRD5A2 variants may significantly influence phenotypic variation.

<strong class="sub-title"> Keywords: </strong> SRD5A2 gene; disorders of sex development; genotype-phenotype correlation; steroid <em>5α</em>-reductase type 2 deficiency.

Purpose: Polycystic ovary syndrome (PCOS) is a complex reproductive disorder often associated with obesity, insulin resistance, and dyslipidemia. Hormonal changes in PCOS may also include altered glucocorticoid signaling. Our aim was to examine whether alterations in hepatic glucocorticoid signaling are associated with disturbances of glucose and lipid metabolism in animal model of PCOS.

<strong class="sub-title"> Methods: </strong> Female rats, 3 weeks old, were subcutaneously implanted with <em>5α</em>-dihydrotestosterone (<em>DHT</em>) or placebo pellets for 90 days to induce PCOS. Expression of 11β-hydroxysteroid dehydrogenase 1 (11βHSD1) and A-ring reductases (<em>5α</em> and 5β), as well as intracellular distribution of glucocorticoid receptor (GR) and expression of its regulated genes were examined in the liver. Proteins of hepatic lipid and carbohydrate metabolism and markers of inflammation were also assessed.

<strong class="sub-title"> Results: </strong> <em>DHT</em> treatment induced increase in body and liver mass, as well as in triglycerides and free fatty acids levels in plasma. Elevation of 11βHSD1 and reduction of <em>5α</em>-reductase expression was observed together with increased hepatic corticosterone concentration and nuclear GR activation. Induced expression of Krüppel-like factor 15 and decreased expression of genes for proinflammatory cytokines and de novo lipogenesis (DNL) were detected in the liver of <em>DHT</em>-treated rats, while DNL regulators and proinflammatory markers were not changed. However, increased mRNA levels of stearoyl-CoA desaturase and apolipoprotein B were observed in <em>DHT</em> animals.

Conclusions: DHT treatment stimulated hepatic glucocorticoid prereceptor metabolism through increased corticosterone availability which is associated with enhanced GR activation. This does not affect gluconeogenesis and DNL, but could be linked to stimulated triglyceride synthesis and hypertriglyceridemia.

<strong class="sub-title"> Keywords: </strong> <em>5α</em>-Dihydrotestosterone; Glucocorticoids; Liver; Polycystic ovary syndrome; Triglycerides.

Kit ligand (KITL) is an important granulosa cell-derived growth factor in ovarian folliculogenesis, but its expression and function in human granulosa cells are currently poorly understood. Based on studies performed in animal models, it was hypothesised that KITL gene expression in human granulosa cells is regulated by androgens and/or growth differentiation factor 9 (GDF9). We utilised two models of human granulosa cells, the KGN granulosa tumour cell line and cumulus granulosa cells obtained from preovulatory follicles of women undergoing assisted reproduction. Cells were treated with combinations of <em>5α</em>-dihydrotestosterone (<em>DHT</em>), recombinant mouse GDF9, and the ALK4/5/7 inhibitor SB431542. KITL mRNA levels were measured by quantitative real-time PCR. No change in KITL mRNA expression was observed after <em>DHT</em> treatment under any experimental conditions, but GDF9 treatment resulted in a significant decrease in KITL mRNA levels in both KGN and cumulus cells. The effect of GDF9 was abolished by the addition of SB431542. These results indicate that KITL is not directly regulated by androgen signalling in human granulosa cells. Moreover, this study provides the first evidence that GDF9 negatively regulates KITL gene expression in human granulosa cells providing new information on the regulation of these important growth factors in the human ovary.

Sebaceous gland cells (sebocytes) differentiate to intracellularly accumulate lipid droplets - a phenomenon similar to that found in adipocytes. In the present study, we examined whether the regulation of lipogenesis in sebocytes is the same as that in preadipocytes. When sebocytes and preadipocytes, prepared from auricle and subcutaneous adipose tissues from the inguinal region of hamsters, respectively, were treated with a common differentiation inducer, insulin, intracellular lipid-droplet formation and triacyglycerol (TG) production were dose- and time-dependently augmented in both. Insulin increased the production of perilipin, a differentiation marker in both sebocytes and adipocytes. Insulin-like growth factor 1 (IGF-1) augmented the intracellular level of TG in sebocytes and preadipocytes. In addition, the action of 1α,25-dihydroxyvitamin D₃ [1,25(OH₂)D₃] on TG production was the opposite between sebocytes and preadipocytes. Furthermore, <em>5α</em>-dihydrotestosterone (<em>5α</em>-<em>DHT</em>) augmented the TG level in sebocytes, whereas it did not alter TG production in preadipocytes. Moreover, insulin-augmented TG production in sebocytes was enhanced by IGF-1 and <em>5α</em>-<em>DHT</em>, while diminished by 1,25(OH₂)D₃. In preadipocytes, the insulin-augmented production of TG was decreased by IGF-1, 1,25(OH₂)D₃, and <em>5α</em>-<em>DHT</em>. These results suggest that sebocytic lipogenesis is partially similar to but substantially different from adipocyte lipogenesis due to the forementioned hormones and growth factors in the skin under physiological conditions.

Our previous observations proposed Pelophylax nigromaculatus as a model species for studying the masculinizing effects of androgenic EDCs in amphibians. To better develop this model species, we studied the process of the gonadal differentiation/development and the sensitive stage to androgens. We found that the earliest sexual dimorphism in gonads at morphological and histological levels occurred at stages 38-40 and stage 36 respectively. Further examination of molecular markers for testicular and ovarian differentiation during development revealed that the cyp17 and cyp19 expressions were sexually dimorphic from stage 32 and stage 36 respectively. Further, we investigated the sex-reversal induced by 100 ng/L <em>5α</em>-dihydrotestosterone (<em>DHT</em>) when exposures were initiated at stages 24, 26 and 28. We found that when exposed from stage 24, <em>DHT</em> resulted masculinization of all tadpoles with no typical ovaries, whereas exposures from stage 26 or 28 dramatically reduced the effect of <em>DHT</em>. Our findings show that gonads of P. nigromaculatus are bipotential at stage 24, in the process of differentiation at stage 26 and determined to become either testis or ovary at stage 28. Altogether, exposure of P. nigromaculatus should begin at stage 24 in order to sensitively detect masculinizing effects of EDCs. Present study provides useful information about the gonadal differentiation and development in P. nigromaculatus for effectively evaluating masculinizing effects of EDCs on gonads.

Background: Hypoandrogenic men showed a higher prevalence of major depressive disorder (MDD), which could be ascribed to overlapping symptoms such as sexual dysfunction, or additionally to core emotional symptoms such as sadness and anhedonia. We examined whether androgen levels 1) differ between men with and without MDD cross-sectionally, 2) are associated with an elevated risk for onset of MDD prospectively, and 3) associate with all individual MDD symptoms, or only with hypogonadism overlapping symptoms.

<strong class="sub-title"> Methods: </strong> In 823 men (mean age 43.5 years), baseline plasma levels of total testosterone, <em>5α</em>-dihydrotestosterone (<em>5α</em>-<em>DHT</em>), and androstenedione were determined with liquid chromatography-tandem mass spectrometry, and dehydroepiandrosterone-sulphate (DHEAS) and sex hormone binding globulin with radioimmunoassay, whereas free testosterone was calculated. MDD status was assessed at baseline and after two years using structured interviews and individual MDD symptoms were self-rated at baseline, and after one and two years.

<strong class="sub-title"> Results: </strong> None of the androgen levels were associated with current or onset (incidence or recurrence) of MDD. Free testosterone was only inversely associated with interest in sex. Also, androstenedione and DHEAS were positively associated with some individual MDD symptoms, and <em>5α</em>-<em>DHT</em> levels showed non-linear associations (both with low and high levels) with MDD symptom severity and several individual MDD symptoms.

<strong class="sub-title"> Conclusions: </strong> These results support the idea that circulating androgens synthesised by the testes are of limited clinical relevance to MDD in adult men, but levels of androstenedione, DHEAS and <em>5α</em>-<em>DHT</em> may be associated with some individual MDD symptoms.

Keywords: Androgens; Depression; Depressive disorder, major; Hypogonadism; Men; Testosterone.

Four major kallikreins (mK1, mK22, mK9, and mK13) were identified in the mouse submandibular gland (SMG). mK1, a true tissue kallikrein, was used as a protein marker to identify different types of SMG granular convoluted tubule (GCT) cells along with epidermal growth factor (EGF), nerve growth factor (NGF), and renin. Kallikrein mK1 was localized in a very small number (~5%) of GCT cells, which were scattered throughout the GCT, indicating that the majority of GCT cells are mK1-negative. Among mK1-positive cells, particularly strong signals were observed in a small number of narrow cells, recognized as slender granular cells (SG cells, Type IV), in the GCT. After postnatal development of the SMG, GCT cells are no longer uniform based on the bioactive substances (mK1, EGF, NGF, and renin) that they produce and secrete. GCT cells were classified into four subtypes, Types I-IV, and it became clear that these subtypes are complicatedly and reversibly converted by the endocrine hormones <em>5α</em>-dihydrotestosterone (<em>DHT</em>) and triiodothyronine (T₃). Duct segments with similar morphology or hormone dependency were recognized in the sublingual and parotid glands. The presence of duct cells with such characteristics is therefore a common feature of the three major salivary glands of rodents.

Previous studies have demonstrated that some amphibian species can be sex-reversed by high concentrations of androgens. Little attention has focused on the effects of androgenic endocrine-disrupting chemicals (EDCs) on amphibians. The present study aimed to investigate the effects of lower concentrations of the androgenic EDC <em>5α</em>-dihydrotestosterone (<em>DHT</em>) on gonadal differentiation and development in Pelophylax nigromaculatus, a true frog distributed widely in East Asia. Tadpoles at Gosner stage 24/25 were exposed to nominal concentrations of 40 ng/L, 400 ng/L, and 4000 ng/L <em>DHT</em> to complete metamorphosis. In all <em>DHT</em> treatment groups, males and ambiguous sexes were identified based on gonadal morphology, whereas no females were found; thus, all treatment groups exhibited male-skewed ratios compared with the control group. Gonadal histological examination revealed that ambiguous sexes displayed overall testicular structure with certain ovarian characteristics, demonstrating that <em>DHT</em>-induced sex-ambiguous gonads were incomplete ovary-to-testis reversals (IOTTRs). The expression levels of some ovary-biased genes in the IOTTRs were significantly higher than in the control testes but lower than in the control ovaries. These results show that low concentrations of <em>DHT</em> induced complete or incomplete female-to-male sex reversal in P. nigromaculatus, and incomplete sex reversal retained certain ovarian characteristics not only at gonadal morphological and histological levels but also at the molecular level. They present study highlights potential risks of <em>DHT</em> and other androgenic EDCs for P. nigromaculatus.

BACKGROUND

Persistent organic pollutants (POPs) are persistent in the environment after release from industrial compounds, combustion productions or pesticides. The exposure of POPs has been related to various reproductive disturbances, such as reduced semen quality, testicular cancer, and imbalanced sex ratio. Among POPs, dichlorodiphenyldichloroethylene (4,4'-DDE) and polychlorinated biphenyls (PCBs) are the most widespread and well-studied compounds. Recent studies have revealed that 4,4'-DDE is an antagonist of androgen receptor (AR). However, the mechanism of the inhibition remains elusive. CB-153 is the most common congener of PCBs, while the action of CB-153 on AR is still under debate.

RESULTS

Molecular docking and molecular dynamics (MD) approaches have been employed to study binding modes and inhibition mechanism of 4,4'-DDE and CB-153 against AR ligand binding domain (LBD). Several potential binding sites have been detected and analyzed. One possible binding site is the same binding site of AR natural ligand androgen <em>5α</em>-dihydrotestosterone (<em>DHT</em>). Another one is on the ligand-dependent transcriptional activation function (AF2) region, which is crucial for the co-activators recruitment. Besides, a novel possible binding site was observed for POPs with low binding free energy with the receptor. Detailed interactions between ligands and the receptor have been represented. The disrupting mechanism of POPs against AR has also been discussed.

CONCLUSIONS

POPs disrupt the function of AR through binding to three possible biding sites on AR/LBD. One of them shares the same binding site of natural ligand of AR. Another one is on AF2 region. The third one is in a cleft near N-terminal of the receptor. Significantly, values of binding free energy of POPs with AR/LBD are comparable to that of natural ligand androgen DHT.

Excess androgen-induced obesity has become a public health problem, and its prevalence has increased substantially in recent years. Chemokine-like receptor 1 (CMKLR1), a receptor of Chemerin secreted by adipose tissue, is linked to adipocyte differentiation, adipose tissue development and obesity. However, the effect of CMKLR1 signaling on androgen-mediated adiposity <i>in vivo</i> remains unclear. Here, using pharmacological method, orchidectomized model and CMKLR1 knockout mice, we demonstrated that androgen excess in female mice resulted in a larger cell size in white adipose tissue (WAT) and brown adipose tissue (BAT), whereas androgen deprivation of male mice induced a smaller cell size. Both effects relating to the adipocytes size were both attenuated in CMKLR1 knockout mice. CMKLR1 deficiency influenced the effect of androgen on adipose tissue by regulating the mRNA expression of androgen receptor (AR) and adipocyte markers (such as Fabp4 and Cidea). Moreover, <i>in vivo</i> suppression of CMKLR1 by its antagonist α-NETA can also weaken the enlargement of the adipocyte cell size caused by <em>5α</em>-dihydrotestosterone (<em>DHT</em>). Furthermore, using <i>in vitro</i> BAT explants culture and PI3K signaling antagonist wortmannin, we found <em>DHT</em> could reduce the phosphorylated ERK (pERK) in BAT, whereas CMKLR1 inactivation inhibited this effect caused by <em>DHT</em> through PI3K signaling pathway. Taken together, these findings reveal an anti-obesity role of CMKLR1 deficiency in regulating lipid accumulation, highlighting the scientific importance for further development of small molecule CMKLR1 antagonists as fundamental scientific tools or a potential drug for the treatment of adiposity.

OBJECTIVE

Benign Prostatic Hyperplasia (BPH) is a form of benign tumor that occurs in humans mainly with ageing. It affects more than 50% of over 50 years old males and it is characterized by an increased synthesis of dihydrotestosterone (<em>DHT</em>), due to the <em>5α</em>-reductase activity. The BPH therapeutic approach mainly uses <em>5α</em>-reductase inhibitors, such as the active compounds present in the extracts deriving from species Serenoa repens. Many lipidosterolic extracts are available on the market, which are obtained with different solvents, among them ethanol is recognized as non-toxic and has less handling risks than hexane. The purpose of the present experimental study was to investigate in-vitro the potency of an ethanol extract of S. repens comparing it with an n-hexane one.

METHODS

Two different lipido-sterolic extracts of S. repens have been tested: ethanol extract and n-hexane extract, two batches for each one. The inhibitory action of the extract was evaluated estimating in-vitro the activity of enzyme <em>5α</em>-reductase type I (<em>5α</em>-RI), which was mainly active under the experimental condition of pH 7.5. <em>DHT</em> amount, synthesized from testosterone (1 μM), was evaluated in a co-culture model of epithelial cells and fibroblasts resulting from prostatic biopsy of a patient with BPH.

RESULTS

The analysis of the resulting dose-response curves showed that the entire S. repens extracts inhibited the <em>5α</em>-RI showing no difference between the two kinds of extract or between the batches. The resulting IC50 values were the following: 8.809 (95% CI = 5.133-15.56) and 9.464 (95% CI = 5.094- 18.27) for ethanol extracts; 11.08 (95% CI = 6.389-19.98) and 12.72 (95% CI = 7.758-21.53) for n-hexane extracts.

CONCLUSIONS

The potency of ethanol extracts of S. repens was comparable with the one of n-hexane extracts.

This study aimed to investigate the beneficial effects of <i>A. melanocarpa</i> on testosterone propionate (TP)-induced benign prostatic hyperplasia (BPH) in Wistar rats. Moreover, the bioactive constituents in the extract were determined using LC/MS and HPLC analyses. The dried fruits of <i>A. melanocarpa</i> were extracted using accelerated solvent extraction (ASE) under different extract conditions (temperature, 30 C or 100 C; extract solvent, 60% or 100% ethanol) to yield four extracts (T1~T4). Of the four <i>A. melanocarpa</i> extracts, T1 extracted under the condition of 100% ethanol/low temperature (30 C) exhibited the greatest inhibitory activity on TP-induced prostatic hyperplasia in rats. The administration of T1 (100 mg/kg body weight, p.o.) for six weeks attenuated TP-induced prostate enlargement and reduced the levels of dihydrotestosterone (<em>DHT</em>) and <em>5α</em>-reductase in both serum and prostate tissue. The suppression of PCNA mRNA expression in prostate tissue was remarkable in T1-treated rats. In LC/MS analysis, the levels of main anthocyanins and phenolics were significantly higher in T1 than in the other extracts. Furthermore, the quantitative study showed that the contents of cyanidin-3-glucose and cyanidin-3-xylose in T1 exhibited 1.27~1.67 and 1.10~1.26 folds higher compared to those in the other extracts. These findings demonstrated that <i>A. melanocarpa</i> extract containing anthocyanins as bioactive constituents attenuated the development of testosterone-induced prostatic hyperplasia, and suggested that this extract has therapeutic potential to treat prostate enlargement and BPH.

Keywords: 5-alpha-reductase; Aronia melanocarpa; androgen receptor; benign prostatic hyperplasia; constituents; testosterone.

<AbstractText>Chinese Skullcap (Scutellaria baicalensis Georgi), which is part of the 50 fundamental herbs of Traditional Chinese Medicine, has been extensively used in the several East Asian countries to treat pyrexia, micturition disorder and inflammation. Although skullcap has effective properties on various diseases, the effects and molecular mechanism of Chinese Skullcap on BPH are still needed for better understanding.</AbstractText><AbstractText>In present study, we aimed to demonstrate the efficacy of Chinese Skullcap root extract (SRE) in testosterone-induced BPH rats and investigate the exact regulatory mechanism involved.</AbstractText><AbstractText>We followed a protocol of testosterone-induced BPH. Rats were allocated into five groups: Group 1, control; Group 2, BPH-induced rats; Group 3, BPH-induced rats administrated with finasteride; Group 4, BPH-induced rats administrated with SRE 100 mg/kg/day; Group 5 - BPH-induced rats administrated with SRE 200 mg/kg/day. We measured the weight of prostate, and thickness of prostate using H&E staining. Western blotting, immunostaining and real-time PCR were used to measure proliferation- and inflammation-relative markers. To confirm the effects of SRE on apoptotic events in BPH-induced tissues, we performed the TUNEL assay.</AbstractText><AbstractText>Compared with the untreated group, the SRE administration group suppressed pathological alterations, such as prostate growth and increase in serum <em>DHT</em> and <em>5α</em>-reductase levels. Furthermore, SRE significantly obliterated the expression of AR and PCNA. SRE also restored Bax/Bcl-2 balance, inducing apoptosis in rats with BPH. These effect of SRE was more prevalent than commercial <em>5α</em>-reductase inhibitor, finasteride.</AbstractText><AbstractText>Taken together, we propose that SRE suppresses abnormal androgen events in prostate tissue and inhibits the development of BPH by targeting inflammation- and apoptosis-related markers. These finding strengthens that SRE could be used as plant-based <em>5α</em>-reductase inhibitory alternative.</AbstractText>

3α-Hydroxysteroid dehydrogenase (3α-HSD) is an enzyme that is essential in the regulation of the concentration of <em>5α</em>-dihydrotestosterone (<em>5α</em>-<em>DHT</em>) in the prostate. It catalyzes the hydride reduction of <em>5α</em>-<em>DHT</em> to 3α-androstanediol, which activates androgen receptors. Elucidating details about the hydride reduction of <em>5α</em>-<em>DHT</em> by 3α-HSD and the environment around the active site of the enzyme could lead to the development of effective drugs for the treatment of prostate cancer. In this study, the X-ray crystal structure of human 3α-HSD type 3 was comprehensively evaluated. Moreover, molecular dynamics (MD) simulations and hybrid ONIOM-type quantum mechanics/molecular mechanics (QM/MM) calculations were performed using a large QM region (maximum 232 atoms). It was determined that the reaction proceeded in a single step without the formation of an alkoxide ion owing to the direct hydride reduction of the substrate by nicotinamide adenine dinucleotide phosphate (NADPH) and concerted proton transfer by Tyr55 and Lys84. Noncovalent interaction (NCI) analysis highlighted the roles of Tyr216 and Trp227 in 3α-HSD. Specifically, Tyr216 assisted the reaction by π/π interactions with the neighboring nicotinamide ring of NADP(H), whereas Trp227 played an important role in recognition of the size of the substrate by CH/π interactions.

<b>Background:</b> Hepatocellular carcinoma (HCC) is a male-predominant cancer. However, the relationship between <em>5α</em>-dihydrotestosterone (<em>DHT</em>), the active form of testosterone, and HCC risk has not been established yet. <b>Methods:</b> We performed a serum epidemiological study in the Chinese population. From 2010 to 2012, 106 male HCC patients and 318 age-matched controls were detected for their serum <em>DHT</em> and estradiol (E2). The odds ratios (ORs) and 95% confidence interval (CI) were estimated by logistic regression analysis with adjustment for potential risk factors. Bivariate Pearson correlations between hormone concentrations and liver function index were investigated. <b>Results:</b> Serum <em>DHT</em> levels were lower (to 1/3 of control), and E2 levels were higher (to 1.5-fold of control) in HCC patients. Compared with the low <em>DHT</em> level, men with a medium level had an adjusted multiple OR of 0.15 (95% CI 0.05-0.43, <i>p</i> trend < 0.01), and men with a high level had an OR of 0.05 (95% CI 0.01-0.21, <i>p</i> trend < 0.01). Notably, <em>DHT</em> concentration, but not E2, is correlated with liver injury. <b>Conclusion:</b> The data suggest that serum <em>DHT</em> is closely associated with HCC risk, providing a reference in order to accurately predict liver cancer and study the pathogenesis of this disease.

Keywords: dihydrotestosterone; estradiol; hepatocellular carcinoma (HCC); liver function; liver injury.

Androgens' metabolism and activity are gaining a more and more important role in human physiology particularly referring to aging and to neurodegenerative diseases. Androgen treatment is often required for long-lasting disorders. In order to improve their duration and effects, androgens can be administered as esters of carboxylic acids. The novelty of our research is the use of esters of androgens with specific unsaturated fatty acids, in order to reduce possible side effects particularly related to chronic pathologies with altered lipid homeostasis such as X-linked adrenoleukodystrophy and cardiovascular disorders. Thus the esters of the main androgenic substances testosterone, dihydrotestosterone (<em>DHT</em>) and their metabolite <em>5α</em>-androstan-3α,17β-diol were chemically obtained by coupling with different unsaturated fatty acids. To this aim, fatty acids with various degree of unsaturation and belonging to different series were selected. Specifically, oleic acid (18:1, n-9), linoleic acid (18:2, n-6), and the n-3 fatty acids, α-linolenic acid (18:3), eicosapentaenoic acid (EPA, 20:5), and docosahexaenoic acid (DHA, 22:6) were used obtaining corresponding esters with acceptable yields and good degree of purity. All the synthesized compounds were tested for their cytotoxic activities in mouse NIH3T3 and human astrocyte cell lines. The esters demonstrated good tolerability and no in vitro cytotoxic effect in both cell cultures. After these promising preliminary results, the esters will be suitable for in vivo studies in order to ascertain their pharmacokinetic characteristics and their biological effects.

Trenbolone acetate (TBA) is a synthetic anabolic steroidal growth factor that is used for rapid muscle development in cattle. The absorbed TBA is hydrolyzed to the active form, 17β-trenbolone (17 TB; 17β-hydroxy-estra-4,9,11-trien-3-one) in meat and milk products, which can cause adverse health effects in humans. Similar to <em>5α</em>-dihydrotestosterone (<em>DHT</em>), 17 TB was reported to exhibit endocrine disrupting effects on animals and humans due to its androgenic effect via binding to the androgen receptor. The purpose of this study is to investigate the molecular mechanism of cell proliferation in prostate cancer (PCa) cells treated with 17 TB. We found that 17 TB induces AR-dependent cell proliferation in the human prostate cancer cell line, 22Rv1 in a concentration dependent manner. Treatment with 17 TB increased the expression of cell cycle regulatory proteins, cyclin D2/CDK-4 and cyclin E/CDK-2, whereas the expression of p27 was down-regulated. Furthermore, phosphorylation of Rb and activation of E2F were also induced, which suggests the activation of cyclin D2/CDK-4 and cyclin E/CDK-2 in the cells. When 22Rv1 cells were exposed to 30 pM of 17 TB, which is the effective concentration (EC50) value required to observe proliferative effects on 22Rv1 cells, the expression levels of the phosphorylated forms of Akt and GSK3β were increased. This study demonstrates that 17 TB induces AR-dependent proliferation through the modulation of cell cycle-related proteins in the Akt signaling pathway. The present study provides an effective methodology for identifying cell proliferation signaling of veterinary drugs that exert AR agonistic effects.

Bovine herpesvirus 1 (BoHV-1), a significant viral pathogen, establishes latency in sensory neurons. The viral genome contains more than 100 consensus glucocorticoid receptor (GR) regulatory elements (GREs): consequently, stress stimulates viral replication and reactivation from latency. The immediate early transcription unit 1 (IEtu1) and bICP0 early promoters are transactivated by GR and synthetic corticosteroid dexamethasone. The androgen receptor (AR), like GR, is a Type 1 nuclear hormone receptor that binds and stimulates certain promoters containing GREs. Consequently, we hypothesized AR and <em>5α</em>-Dihydrotestosterone (<em>DHT</em>) stimulate productive infection and key viral promoters. New studies demonstrated AR, <em>DHT</em>, and Krüppel like transcription factor 4 (KLF4) cooperatively stimulated productive infection and bICP0 E promoter activity in mouse neuroblastoma cells (Neuro-2A). KLF15 also cooperated with AR and <em>DHT</em> to stimulate IEtu1 promoter activity. We suggest AR and testosterone increase the prevalence of virus in semen by stimulating viral gene expression and replication.

BACKGROUND

Three decades after the first nerve-sparing radical prostatectomy, postoperative erectile dysfunction (ED) remains a challenging and common problem. Despite considerable advances and improvements in surgical techniques, full recovery of erectile function remains elusive even for young, potent men. This suggests, ipso facto, that factors other than surgical technique must be important to recovery of erectile function.

OBJECTIVE

This study aims to review evidence that the prostate is an endocrine gland with contributions to local and systemic concentrations of <em>5α</em>-dihydrotestosterone (<em>5α</em>-<em>DHT</em>), a potent androgen shown to be critical to penile physiology.

METHODS

Literature review of human and animal studies related to endocrine role of prostate and postoperative recovery of erectile function.

METHODS

Effect of <em>5α</em>-<em>DHT</em> on erectile function and recovery after surgical injury.

RESULTS

We advance the following hypothesis: "Loss of endocrine function of the prostate, specifically reduced local <em>5α</em>-<em>DHT</em> concentration plays a major role in the failure of full recovery of erectile function following anatomic nerve-sparing radical prostatectomy."

CONCLUSIONS

We propose two separate, yet interrelated, mechanisms whereby the loss of <em>5α</em>-<em>DHT</em> interferes with postoperative recovery of erectile function: (i) <em>5α</em>-<em>DHT</em> contributes to cavernous nerve integrity and its ability to recover from surgical insult. (ii) <em>5α</em>-<em>DHT</em> is important to the structural/functional integrity of penile tissues and erectile physiology. Kacker R, Morgentaler A, and Traish A. Medical hypothesis: Loss of the endocrine function of the prostate is important to the pathophysiology of postprostatectomy erectile dysfunction.

The novel A-YAS assay for the detection of androgenic activity in liquid samples such as urine has been developed and assessed. The assay is based on transgenic Arxula adeninivorans yeast cells as the bio-component. The cells were engineered to co-express the human androgen receptor (hAR) gene and the inducible phytase reporter gene (phyK, derived from Klebsiella sp. ASR1), under the control of an Arxula derived glucoamylase (GAA) promoter, which had been modified by the insertion of hormone-responsive elements (HREs). The Arxula transformation/expression platform Xplor®2 was used to select stable mitotic resistance marker free transformants and the most suitable cells were characterized for performance as a sensor bio-component. The assay is easy-to-use, fast (6-25 h) and is currently the most sensitive yeast-based androgen screen with an EC50, limit of detection and of quantification values for <em>5α</em>-dihydrotestosterone (<em>DHT</em>) of 277.1±53.0, 56.5±4.1 and 76.5±6.7 ng L(-1), respectively. Furthermore, the assay allows the determination of androgenic and anti-androgenic activity of various compounds such as naturally occurring androgens and estrogens, pharmaceuticals and biocides. The robustness of the A-YAS assay enables it to be used for analysis of complex samples such as urine. The results of the analysis of a number of cattle urine samples achieved by the A-YAS assay correlate well with GC-MS analysis of the same samples.

Knowledge of how and why secondary sexual characters are associated with sex hormones is important to understand their signalling function. Such a link can occur if i) testosterone participates in the elaboration of sex-traits, ii) the display of an ornament triggers behavioural response in conspecifics that induce a rise in testosterone, or iii) genes implicated in the elaboration of a sex-trait pleiotropically regulate testosterone physiology. To evaluate the origin of the co-variation between melanism and testosterone, we measured this hormone and the expression of enzymes involved in its metabolism in feathers of barn owl (Tyto alba) nestlings at the time of melanogenesis and in adults outside the period of melanogenesis. Male nestlings displaying smaller black feather spots had higher levels of circulating testosterone, potentially suggesting that testosterone could block the production of eumelanin pigments, or that genes involved in the production of small spots pleiotropically regulate testosterone production. In contrast, the enzyme <em>5α</em>-reductase, that metabolizes testosterone to <em>DHT</em>, was more expressed in feathers of reddish-brown than light-reddish nestlings. This is consistent with the hypothesis that testosterone might be involved in the expression of reddish-brown pheomelanic pigments. In breeding adults, male barn owls displaying smaller black spots had higher levels of circulating testosterone, whereas in females the opposite result was detected during the rearing period, but not during incubation. The observed sex- and age-specific co-variations between black spottiness and testosterone in nestling and adult barn owls may not result from testosterone-dependent melanogenesis, but from melanogenic genes pleiotropically regulating testosterone, or from colour-specific life history strategies that influence testosterone levels.

Treatment of cultured testicular cells from adult rats with 5 alpha-dihydrotestosterone (<em>DHT</em>; 10(-6) M) or the synthetic androgen methyltrienolone (R1881; 10(-6) M) inhibited Leydig cell 3 beta-hydroxysteroid dehydrogenase/delta 5-4 isomerase (3 beta-HSD) enzyme activity, whereas no effect of both androgens on cultured cells derived from neonatal animals could be observed. The inhibitory effect of <em>DHT</em> or R1881 on Leydig cell 3 beta-HSD enzyme activity, however, was abolished when adult cells were cultured in the presence of the anti-androgen cyproterone acetate (CPA; 10(-6) M) or the protein synthesis inhibitor cycloheximide (CX; 1 microgram/ml). Testicular cells from adult animals were also cultured in the presence of the different treatments described above, and the spent media was collected and thereafter used as conditioned culture medium (CCM) in subsequent experiments performed with neonatal cells. Dispersed testicular cells from neonatal rats were cultured for 12 days in McCoy's <em>5a</em> medium of in CCM derived from R1881-treated adult cells, and fresh culture medium or CCM was replaced every 2 days. The human chorionic gonadotropin (hCG)-stimulated testosterone production of neonatal cells was abolished in the presence of CCM derived from R1881-treated adult cells. Nevertheless, the steroidogenic response to hCG recovered when neonatal cells were cultured for two additional days in McCoy's <em>5a</em> medium. Treatment of neonatal cells with increasing concentrations of hCG (0.1-10 ng/ml) resulted in a dose-dependent augmentation in Leydig cell 3 beta-HSD enzyme activity and testosterone production.(ABSTRACT TRUNCATED AT 250 WORDS)

The target of the current study is to formulate letrozole loaded nanoemulsion (LET-NE) for the direct nose to brain delivery to reduce peripheral effects of letrozole (LET). LET-NE is compared against intraperitoneally administered free LET in kainic acid (KA) induced status epilepticus (SE) in mice. LET loaded nanoemulsion (LET-NE) was prepared by aqueous microtitration method using Triacetin, Tween 80 and PEG-400 as the oil phase, surfactant, and co-surfactant. Nanoemulsion was studied for droplet size, polydispersity index (PDI), zeta potential, percentage transmittance, drug content, surface morphology. TEM images of developed formulation demonstrated spherical droplets with a mean diameter of 95.59 ± 2.34 nm, PDI of 0.162 ± 0.012 and zeta potential of -7.12 ± 0.12 mV respectively. In in-vitro and ex-vivo drug release, LET-NE showed prolonged drug release profile as compared to suspension. SE was induced by KA (10 mg/kg, i.p.) in Swiss albino mice. Behavioral seizure monitoring, biochemical estimations, and histopathological examination were performed. The onset time of SE was significantly enhanced and % incidence of SE was reduced by intranasal administration of LET-NE as compared to KA and LET administered intraperitoneally. Biochemical estimations revealed that LET-NE effectively decreased levels of 17-β estradiol while the levels of <em>5α</em>-Dihydrotestosterone (<em>5α</em>-<em>DHT</em>) and 3α-androstanediol (3α-Diol) were significantly increased in the hippocampus. In cresyl violet staining LET-NE showed better protection of the hippocampus from neurotoxicity induced by KA as compared to LET. Also, in gamma scintigraphy of mouse brain, intranasal administration of nanoemulsion exhibited the presence of high concentration of LET. The study demonstrates the anticonvulsant and neuroprotective effect of LET-NE probably by inhibition of aromatization of testosterone into 17-β estradiol, proconvulsant, and diverting the pathway into the synthesis of testosterone metabolites, 3α-Diol with known anticonvulsant and neuroprotective action. Brain targeting of LET-NE showed better anticonvulsant and neuroprotective action than LET.

Summary In human beings, androgen metabolism plays an important role in mediating the action of male hormones upon target structures of the skin. First, human skin is capable of transforming inactive steroids supplied through the blood, such as androstenedione and dehydroisoandrosterone, into the active androgen testosterone. Second, human skin is able to reduce testosterone to <em>5a</em>lpha-<em>DHT</em>, an essential prerequisite, during embryogenesis, for the male differentiation of target structures derived from urogenital sinus. At puberty, hair growth in sexual areas of skin also requires the transformation of testosterone to <em>DHT</em>. Regulation of <em>5a</em>lpha-reductase activity varies according to the anatomical site of the enzyme. In foetuses, <em>5a</em>lpha-reductase activity present in tissues derived from the urogenital tract does not seem to be androgen-dependent, since it is acquired before the onset of testosterone secretion by foetal testis. By contrast, the enzyme that mediates development of certain secondary sex characteristics, such as pilosebaceous gland activity in sexual areas, is clearly androgen-dependent, since it is absent before puberty and in persons with hypogonadism. These differences in the control of the <em>5a</em>lpha-reductase activity mediating the appearance of either primary or secondary sex characteristics are important and may explain the differences in <em>5a</em>-reductase activity observed in adult skin of both sexes derived from different sexual areas. In addition, the knowledge of androgen relation to the skin is necessary in order to understand the action of the anti-androgens, particularly the compounds which may be used by topical administration.