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Salicylate preservatives are widely used in consumer products and pharmaceuticals. This study investigates their potential endocrine-disrupting effects on neurosteroidogenesis, focusing on 5α-reductase type 1 (SRD5A1). We evaluated the effects of 13 salicylates on human SRD5A1 using SF126 glioblastoma cell microsomes and rat brain microsomes, examining dihydrotestosterone production in SF126 cells. Results revealed a hierarchy of inhibitory potency against human SRD5A1, with methyl salicylate (IC 50 , 71.93 μM) to menthyl salicylate (2.41 μM), indicating increasing potency. Kinetic analysis indicates their mixed/noncompetitive inhibitions. In SF126 cells, all salicylates at 100 μM significantly reduced dihydrotestosterone production. Rat SRD5A1 showed reduced sensitivity, with menthyl salicylate as the most potent inhibitor (IC 50 , 17.12 μM). Docking analysis suggests salicylates bind to the reduced nicotinamide adenine dinucleotide phosphate site of both human and rat SRD5A1. Bivariate correlation analysis highlights the influence of LogP, molecular weight, carbon number in the alcohol moiety, and p K a on inhibitory potency. 3D-QSAR revealed the importance of hydrophobic aromatic regions in SRD5A1 binding. This study delineates the inhibitory effects of salicylates and binding mechanisms on human and rat SRD5A1, providing insights into their impact on neurosteroid production.

SRD5A3-CDG is a congenital disorder of glycosylation (CDG) resulting from pathogenic variants in SRD5A3 and follows an autosomal recessive inheritance pattern. The enzyme encoded by SRD5A3, polyprenal reductase, plays a crucial role in synthesizing lipid precursors essential for N-linked glycosylation. Despite insights from functional studies into its enzymatic function, there remains a gap in understanding global changes in patient cells. We sought to identify N-glycoproteomic and proteomic signatures specific to SRD5A3-CDG, potentially aiding in biomarker discovery and advancing our understanding of disease mechanisms. Using tandem mass tag (TMT)-based relative quantitation, we analyzed fibroblasts derived from five patients along with control fibroblasts. N-glycoproteomics analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS) identified 3,047 glycopeptides with 544 unique N-glycosylation sites from 276 glycoproteins. Of these, 418 glycopeptides showed statistically significant changes with 379 glycopeptides decreased (P < 0.05) in SRD5A3-CDG patient-derived samples. These included high mannose, complex and hybrid glycan-bearing glycopeptides. High mannose glycopeptides from protocadherin Fat 4 and integrin alpha-11 and complex glycopeptides from CD55 were among the most significantly decreased glycopeptides. Proteomics analysis led to the identification of 5,933 proteins, of which 873 proteins showed statistically significant changes. Decreased proteins included cell surface glycoproteins, various mitochondrial protein populations and proteins involved in the N-glycosylation pathway. Lysosomal proteins such as N-acetylglucosamine-6-sulfatase and procathepsin-L also showed reduced levels of phosphorylated mannose-containing glycopeptides. Our findings point to disruptions in glycosylation pathways as well as energy metabolism and lysosomal functions in SRD5A3-CDG, providing clues to improved understanding and management of patients with this disorder., (© The Author(s) 2024. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Androgenetic alopecia (AGA) is the leading cause of hair loss in adults. Its pathogenesis remains unclear, but studies have shown that the androgen-mediated 5α-reductase-AR receptor pathway and the Wnt/β-catenin signaling pathway play significant roles. Camellia oleifera is an oil plant, and its fruits have been documented in folklore as having a hair cleansing effect and preventing hair loss. In this study, we used UPLC-Q-TOF-MS/MS to identify the structure of the substances contained in the polyphenols of Camellia oleifera seed shell. These polyphenols are mainly used for shampooing and anti-hair loss purposes. Next, we used molecular docking technology to dock 41 polyphenols and steroidal 5 alpha reductase 2 (SRD5A2). We found that the docking scores and docking sites of 1,3,6-tri-O-galloylglucose (TGG) and finasteride were similar. We constructed a mouse model of DHT-induced AGA to evaluate the effects of Camellia oleifera seed shell polyphenols (CSSP) and TGG in vivo. Treatment with CSSP and TGG alleviated alopecia symptoms and reduced DHT levels. Additionally, CSSP and TGG were able to reduce androgen levels by inhibiting the SRD5A2-AR receptor signaling pathway. Furthermore, by regulating the secretion of growth factors and activating the Wnt/β-catenin signaling pathway, CSSP and TGG were able to extend the duration of hair growth. In conclusion, our study showed that CSSP and TGG can improve AGA in C57BL/6 J mice and reduce the effect of androgen on hair follicle through the two signaling pathways mentioned above. This provides new insights into the material basis and mechanism of the treatment of AGA by CSSP., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Introduction: This study combined the bioinformatics and in vitro experiment-related technologies to analyze the impact of steroid 5 alpha-reductase 3 (SRD5A3) on the prognosis and immune microenvironment of Liver Hepatocellular Carcinoma (LIHC)., Method: Gene expression and clinical data were obtained from public databases. The prognosis was evaluated using survival, multifactor Cox, enrichment, and mutation analyses. This was then verified through in vitro experiments., Results: The expression level of SRD5A3 in LIHC tissues was significantly higher than that in the adjacent tissues. Kaplan-Meier survival analysis showed that high SRD5A3 expression was associated with poor overall survival (OS) and short progression-free survival in patients with LIHC. Multivariate Cox regression analysis revealed that positive SRD5A3 expression was an independent risk factor for OS in patients with LIHC. Expression of SRD5A3 was negatively correlated with immune cell infiltration of CD4+ T, CD8+ T, and B cells. GO and KEGG enrichment analyses showed that SRD5A3 was significantly enriched in signaling- and tumor metastasis-related pathways. Nomogram and calibration curve showed that the predicted performance of the model was consistent with the actual results. In vitro results confirmed that SRD5A3 knockdown inhibited the migration, invasion, and proliferation of LIHC cells., Conclusions: SRD5A3 is actively expressed in LIHC, and positive expression of SRD5A3 is an independent risk factor for different prognoses in patients with LIHC. SRD5A3 can promote the proliferation, migration, and invasion of liver cancer cells and is related to short immune infiltration in patients with LIHC.

Background: Endometrial cancer (EC) is a prevalent gynecological malignancy globally, with a rising incidence trend. While classic androgens have been implicated with EC risk, the role of their 11-oxygenated metabolites is poorly understood. Here, we studied 11-oxyandrogen formation from steroid precursors in EC for the first time., Methods: We performed in vitro studies on a panel of four EC cell lines of varying differentiation degree and molecular subtype and a control cell line of normal endometrium to assess 11-oxyandrogen formation from steroid precursors. We also characterized the transcriptomic effects of dihydrotestosterone (DHT) and 11-keto-DHT on Ishikawa and RL95-2. Key molecular players in 11-oxyandrogen metabolism and action were explored in endometrial tumors using public transcriptomic datasets., Results: We discovered that within endometrial tumors, the formation of 11-oxyandrogens does not occur from classic androgen precursors. However, we observed distinct regulatory mechanisms at a pre-receptor level in normal endometrium compared to cancerous tissue, and between low- and high-grade tumors. Specifically, in vitro models of low-grade EC formed higher levels of bioactive 11-keto-testosterone from 11-oxyandrogen precursors compared to models of noncancerous endometrium and high-grade, TP53-mutated EC. Moreover, the potent androgen, DHT and its 11-keto homologue induced mild transcriptomic effects on androgen receptor (AR)-expressing EC model, Ishikawa. Finally, using public transcriptomic datasets, we found HSD11B2 and SRD5A2 , coding for key enzymes in steroid metabolism, to be associated with better disease-specific survival, whereas higher intra-tumoral AR expression correlated with lower recurrence in TP53-wt tumors., Conclusions: The intra-tumoral metabolism of 11-oxyandrogen precursors is characteristic for low-grade EC of non-TP53-alt molecular subtypes. Our findings support further exploration of circulating 11-oxyandrogens as prognostic biomarkers in EC., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author TLR declared that she was an Associate Editor of Frontiers in Pharmacology, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Gjorgoska, Šturm and Lanišnik Rižner.)

Dolichol is a lipid critical for N-glycosylation as a carrier for activated sugars and nascent oligosaccharides. It is commonly thought to be directly produced from polyprenol by the enzyme SRD5A3. Instead, we found that dolichol synthesis requires a three-step detour involving additional metabolites, where SRD5A3 catalyzes only the second reaction. The first and third steps are performed by DHRSX, whose gene resides on the pseudoautosomal regions of the X and Y chromosomes. Accordingly, we report a pseudoautosomal-recessive disease presenting as a congenital disorder of glycosylation in patients with missense variants in DHRSX (DHRSX-CDG). Of note, DHRSX has a unique dual substrate and cofactor specificity, allowing it to act as a NAD + -dependent dehydrogenase and as a NADPH-dependent reductase in two non-consecutive steps. Thus, our work reveals unexpected complexity in the terminal steps of dolichol biosynthesis. Furthermore, we provide insights into the mechanism by which dolichol metabolism defects contribute to disease., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Rationale: In inflammatory diseases such as rheumatoid arthritis (RA), steroid metabolism is a central component mediating the actions of immuno-modulatory glucocorticoids and sex steroids. However, the regulation and function of cellular steroid metabolism within key leukocyte populations such as macrophages remain poorly defined. In this study, the inflammatory regulation of global steroid metabolism was assessed in RA macrophages., Methods: Bulk RNA-seq data from RA synovial macrophages was used to assess transcripts encoding key enzymes in steroid metabolism and signalling. Changes in metabolism were assessed in synovial fluids, correlated to measures of disease activity and functionally validated in primary macrophage cultures., Results: RNA-seq revealed a unique pattern of differentially expressed genes, including changes in genes encoding the enzymes 11β-HSD1, SRD5A1, AKR1C2 and AKR1C3. These correlated with disease activity, favouring increased glucocorticoid and androgen levels. Synovial fluid 11β-HSD1 activity correlated with local inflammatory mediators (TNFα, IL-6, IL-17), whilst 11β-HSD1, SRD5A1 and AKR1C3 activity correlated with systemic measures of disease and patient pain (ESR, DAS28 ESR, global disease activity). Changes in enzyme activity were evident in inflammatory activated macrophages in vitro and revealed a novel androgen activating role for 11β-HSD1. Together, increased glucocorticoids and androgens were able to suppress inflammation in macrophages and fibroblast-like-synoviocytes., Conclusions: This study underscores the significant increase in androgen and glucocorticoid activation within inflammatory polarized macrophages of the synovium, contributing to local suppression of inflammation. The diminished profile of inactive steroid precursors in postmenopausal women may contribute to disturbances in this process, leading to increased disease incidence and severity., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Steroid 5α reductase 2 (SRD5A2) converts testosterone to dihydrotestosterone and is crucial for prostatic development. 5α reductase inhibitors (5ARI) reduce prostate size in benign prostate hyperplasia (BPH) and ameliorate lower urinary tract symptoms secondary to BPH. However, the mechanisms of 5ARI functioning are still not fully understood. Here, we used a Srd5a2 -/- mouse model and employed single-cell RNA sequencing to explore the impact of SRD5A2 absence on prostate cellular heterogeneity. Significant alterations in luminal epithelial cell (LE) populations were observed, alongside an increased proportion and proliferative phenotype of estrogen receptor 1 (ESR1) + LE2 cells, following an SRD5A2-independent ESR1 differentiation trajectory. LE2 cells exhibited enhanced estrogen response gene signatures, suggesting an alternative pathway for prostate growth when SRD5A2 is absent. Human prostate biopsy analysis revealed an inverse correlation between the expressions of SRD5A2 and LE2 markers (ESR1/PKCα), and an inverse correlation between SRD5A2 and the clinical efficiency of 5ARI. These findings provide insights into 5ARI resistance mechanisms and potential alternative therapies for BPH-related lower urinary tract symptoms. © 2024 The Pathological Society of Great Britain and Ireland., (© 2024 The Pathological Society of Great Britain and Ireland.)

Epitestosterone is a stereoisomer of the active androgen testosterone and its circulating concentrations are similar to those of testosterone in women and children. However, its biological function and pathways of metabolism remain unknown. The structural similarity to testosterone suggests a potential function in the modulation of androgen receptor signalling. It is well established that the conversion of testosterone to 5α-dihydrotestosterone enhances local androgen receptor signalling. In this study, we show that epitestosterone is metabolized to 5α-dihydroepitestosterone by both human steroid 5α-reductase isoforms, SRD5A1 and SRD5A2. Using two different variations of a reporter assay for transactivation of the human androgen receptor, we show that epitestosterone is a partial AR agonist and that the 5α-reduction of epitestosterone increases its androgenic activity. In line with this, we show that 5α-reduction of epitestosterone reduces its ability to antagonize 5α-dihydrotestosterone-induced androgen receptor transactivation. In conclusion, we provide evidence that steroid 5α-reductases regulate the modulatory effect of epitestosterone on androgen receptor signalling., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Hypospadias, an oft-occurring penis anomaly, ranks among neonatal's foremost birth defects. The SRD5A2 can affect male reproductive system development and is abnormally expressed in its epithelial cells. This study exploration aimed at understanding the role of SRD5A2 in the development of hypospadias from a molecular perspective. SRD5A2 levels in hypospadias primary cells were analyzed by Western blot, while targeted interaction with miR-1199-5p was ascertained by dual-luciferase gene reporter assay. In vitro biological experiments were used to confirm the biological function of SRD5A2 in hypospadias. SRD5A2 expression was significantly upregulated, and miR-1199-5p expression was significantly downregulated in hypospadias primary cells. Intervention of SRD5A2 expression can affect cell proliferation, migration, invasion, EMT, and the expression of cell cycle-related proteins. Additionally, we found that SRD5A2 is regulated by upstream miR-1199-5p and can enhance the effect of SRD5A2 on hypospadias cells. Conclusions Silencing SRD5A2 promotes cell proliferation, invasion, and migration blocks the cell cycle at the G1 phase, and simultaneously promotes EMT, cell cycle, and cell proliferation-related protein expression. The biological function of SRD5A2 in hypospadias cells is regulated by miR-1199-5p. SRD5A2 may be an effective therapeutic target for hypospadias.

Selective Androgen Receptor Modulators (SARMs), particularly (17α,20E)-17,20-[(1-methoxyethylidene)bis(oxy)]-3-oxo-19-norpregna-4,20-diene-21-carboxylic-acid-methyl-ester (YK11), are increasingly popular among athletes seeking enhanced performance. Serving as an Androgen Receptor (AR) agonist, YK11 stimulates muscle growth while inhibiting myostatin. Our study delved into the impact of YK11 on the rat hippocampus, analyzing potential alterations in neurochemical mechanisms and investigating its synergistic effects with exercise (EXE), based on the strong relationship between SARM users and regular exercise. Utilizing Physiologically Based Pharmacokinetic (PBPK) modeling, we demonstrated YK11 remarkable brain permeability, with molecular docking analysis revealing YK11 inhibitory effects on 5-alpha-reductase type II (5αR2), suggesting high cell bioavailability. Throughout a 5-week experiment, we divided the animals into the following groups: Control, YK11 (0.35 g/kg), EXE (swimming exercise), and EXE + YK11. Our findings showed that YK11 displayed a high binding affinity with AR in the hippocampus, influencing neurochemical function and modulating aversive memory consolidation, including the downregulation of the BDNF/TrkB/CREB signaling, irrespective of EXE combination. In the hippocampus, YK11 increased pro-inflammatory IL-1β and IL-6 cytokines, while reducing anti-inflammatory IL-10 levels. However, the EXE + YK11 group counteracted IL-6 effects and elevated IL-10. Analysis of apoptotic proteins revealed heightened p38 MAPK activity in response to YK11-induced inflammation, initiating the apoptotic cascade involving Bax/Bcl-2/cleaved caspase-3. Notably, the EXE + YK11 group mitigated alterations in Bcl-2 and cleaved caspase-3 proteins. In conclusion, our findings suggest that YK11, at anabolic doses, significantly alters hippocampal neurochemistry, leading to impairments in memory consolidation. This underscore concerns about the misuse risks of SARMs among athletes and challenges common perceptions of their minimal side effects., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Per- and polyfluoroalkyl substances (PFAS) are widely used synthetic chemicals that persist in the environment and bioaccumulate in animals and humans. There is growing evidence that PFAS exposure adversely impacts neurodevelopment and neurological health. Steroid 5α-reductase 1 (SRD5A1) plays a key role in neurosteroidogenesis by catalyzing the conversion of testosterone or pregnenolone to neuroactive steroids, which influence neural development, cognition, mood, and behavior. This study investigated the inhibitory strength and binding interactions of 18 PFAS on human and rat SRD5A1 activity using enzyme assays, molecular docking, and structure-activity relationship analysis. Results revealed that C9-C14 PFAS carboxylic acid at 100 μM significantly inhibited human SRD5A1, with IC 50 values ranged from 10.99 μM (C11) to 105.01 μM (C14), and only one PFAS sulfonic acid (C8S) significantly inhibited human SRD5A1 activity, with IC 50 value of 8.15 μM. For rat SRD5A1, C9-C14 PFAS inhibited rat SRD5A1, showing the similar trend, depending on carbon number of the carbon chain. PFAS inhibit human and rat SRD5A1 in a carbon chain length-dependent manner, with optimal inhibition around C11. Kinetic studies indicated PFAS acted through mixed inhibition. Molecular docking revealed PFAS bind to the domain between NADPH and testosterone binding site of both SRD5A1 enzymes. Inhibitory potency correlated with physicochemical properties like carbon number of the carbon chain. These findings suggest PFAS may disrupt neurosteroid synthesis and provide insight into structure-based inhibition of SRD5A1., Competing Interests: Declaration of competing interest The authors have no conflict of interest to be disclosed., (Copyright © 2024. Published by Elsevier B.V.)

18-β-Glycyrrhetinic acid, a major component of licorice, stimulated the proliferation of both dermal papilla cells and outer root sheath cells isolated from human hair follicles. Thus, suggesting that this compound promotes hair growth. Furthermore, this compound inhibited the activity of testosterone 5α-reductase, an enzyme responsible for converting androgen to dihydroandrogen, with an IC 50 of 137.1 µM. 18-β-Glycyrrhetinic acid also suppressed the expression of transforming growth factor-β1 (TGF-β1), which shifts the hair cycle from the anagen phase to the telogen phase. It suggested that this compound may prolong the anagen phase. Based on these findings, this compound could be a potentially effective treatment for androgenetic alopecia.

The cloning and characterization of the complete coding sequence of the Clarias magur SRD5A1 (CmSRD5A1) gene, which encodes an enzyme responsible for regulating steroid levels by converting testosterone into 5α-dihydrotestosterone (DHT), have been successfully achieved. DHT plays a vital role in enabling the complete expression of testosterone's actions in neuroendocrine tissues. The ORF of the full-length cDNA sequence of SRD5A1 was 795 bp, translating into 265 amino acids, with a total length of 836 bp including UTRs. Like other vertebrates, the signal peptide analysis revealed that SRD5A1 is a non-secretory protein, and hydropathy profiles indicated that it is hydrophobic in nature. The 3D structure of CmSRD5A1 sequence generated above was predicted using highly accurate AlphaFold 2 in Google Colab online platform. CmSRD5A1 contains seven transmembrane helices connected by six loops, with the N-termini located on the periplasmic side and C-termini on the cytosolic side. Structural superimposition with known bacterial and human SRD5As showed very high structural similarity. The electrostatic potential calculation and surface analysis of CmSRD5A1 revealed the presence of a large cavity with two openings one highly electropositive towards the cytosolic side and another relatively neutral towards the transmembrane region. The structural comparison revealed that the electropositive side of the cavity should bind to NADPH and the steroid hormone in the hydrophobic environment. Polar residues binding to NADPH are highly conserved and the same as known strictures. The conserved residues involved in hydrogen bonding with the ketone group at C-3 in the steroids hence fevering Δ4 double-bond reduction are identified as E66 and Y101. Our findings showed that SRD5A1 expression was lower during the spawning phase than the preparatory phase in female fish, while the administration of Ovatide (a GnRH analogue) resulted in up-regulation of expression after 6 h of injection in the ovary. In males, the lowest expression was observed during the preparatory phase and peaked at 16 h post- Ovatide injection in the testis. The expression of SRD5A1 in the brain of female fish was slightly higher during the Ovatide stimulation phase than the spawning phase. This study represents the first report on the cloning and characterization of the full-length cDNA of SRD5A1 in Indian catfish., (© 2023. The Author(s).)

Cinnamon and its bioactive compounds inhibit prostate cancer cell proliferation in vitro. The aim of the current study was to assess the chemopreventive efficacy of cinnamon (CN) and its bioactive compounds in vivo using N-methyl-N-nitrosourea (MNU) and testosterone (T) to induce prostate carcinogenesis in male Wistar/National Institute of Nutrition rats. Cancer-induced (CI) rats (n = 10) developed prostatic hyperplasia and prostatic intraepithelial neoplasia. These histopathologic changes were diminished in CI rats fed for 4 months with diets supplemented with either CN (n = 20) or its bioactive compounds (cinnamaldehyde, n = 10 and procyanidin B2, n = 10). Androgen receptor (AR) expression was lower in the prostates of CI rats than in control, but the AR target gene, probasin, was robustly upregulated. Treatment of CI rats with CN or its bioactive compounds upregulated AR expression but inhibited the expression of the 5-alpha reductase genes (Srd5a1 and Srd5a2) and did not further increase probasin expression, suggesting blunted transcriptional activity of AR due to the limited availability of dihydrotestosterone. MNU+T induced an altered oxidant status in rat prostate, which was reflected by an increase in lipid peroxidation and DNA oxidation. These changes were completely or partially corrected by treatment with CN or the bioactive compounds. CN and its active components increased the activity of the apoptotic enzymes caspase-8 and caspase-3 in the prostates of CI rats. In conclusion, our data demonstrate that CN and its bioactive compounds have inhibitory effects on premalignant prostate lesions induced by MNU + T and, therefore, may be considered for the chemoprevention of prostate cancer., Prevention Relevance: The research work presented in this article demonstrates the chemopreventive efficacy of CN and its bioactive compounds in a rat model of premalignant prostate cancer., (©2022 American Association for Cancer Research.)

The two human steroid 5α-reductase (5αR) enzymes catalyze the conversion 3-keto-Δ 4 -steroids to their 5α-reduced congeners. In the genital skin and prostate, the type 2 isoenzyme converts testosterone (T) to the more potent androgen 5α-dihydrotestosterone (DHT), and intracellular DHT is essential for the morphogenesis of the undifferentiated external genitalia to the male phenotype. Both isoenzymes also metabolize other 19- and 21-carbon 3-keto-Δ 4 -steroids, both endogenous compounds and some steroid-based drugs. Rigorous biochemical studies have been limited due to the extremely hydrophobic nature of these proteins. We have described the heterologous expression of these enzymes in bacteria, their purification with affinity chromatography, and the reconstitution of activity in liposomes. This article details these procedures, as well as reconstitution in phospholipid nanodiscs and enzyme assay., (Copyright © 2023. Published by Elsevier Inc.)

Decidualization is the hormone-dependent process of endometrial remodeling that is essential for fertility and reproductive health. It is characterized by dynamic changes in the endometrial stromal compartment including differentiation of fibroblasts, immune cell trafficking and vascular remodeling. Deficits in decidualization are implicated in disorders of pregnancy such as implantation failure, intra-uterine growth restriction, and pre-eclampsia. Androgens are key regulators of decidualization that promote optimal differentiation of stromal fibroblasts and activation of downstream signaling pathways required for endometrial remodeling. We have shown that androgen biosynthesis, via 5α-reductase-dependent production of dihydrotestosterone, is required for optimal decidualization of human stromal fibroblasts in vitro , but whether this is required for decidualization in vivo has not been tested. In the current study we used steroid 5α-reductase type 1 (SRD5A1) deficient mice ( Srd5a1-/- mice) and a validated model of induced decidualization to investigate the role of SRD5A1 and intracrine androgen signaling in endometrial decidualization. We measured decidualization response (weight/proportion), transcriptomic changes, and morphological and functional parameters of vascular development. These investigations revealed a striking effect of 5α-reductase deficiency on the decidualization response. Furthermore, vessel permeability and transcriptional regulation of angiogenesis signaling pathways, particularly those that involved vascular endothelial growth factor (VEGF), were disrupted in the absence of 5α-reductase. In Srd5a1-/- mice, injection of dihydrotestosterone co-incident with decidualization restored decidualization responses, vessel permeability, and expression of angiogenesis genes to wild type levels. Androgen availability declines with age which may contribute to age-related risk of pregnancy disorders. These findings show that intracrine androgen signaling is required for optimal decidualization in vivo and confirm a major role for androgens in the development of the vasculature during decidualization through regulation of the VEGF pathway. These findings highlight new opportunities for improving age-related deficits in fertility and pregnancy health by targeting androgen-dependent signaling in the endometrium., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Shaw, Kirkwood, Rebourcet, Cousins, Ainslie, Livingstone, Smith, Saunders and Gibson.)

Garrod's concept of 'chemical individuality' has contributed to comprehension of the molecular origins of human diseases. Untargeted high-throughput metabolomic technologies provide an in-depth snapshot of human metabolism at scale. We studied the genetic architecture of the human plasma metabolome using 913 metabolites assayed in 19,994 individuals and identified 2,599 variant-metabolite associations (P < 1.25 × 10 -11 ) within 330 genomic regions, with rare variants (minor allele frequency ≤ 1%) explaining 9.4% of associations. Jointly modeling metabolites in each region, we identified 423 regional, co-regulated, variant-metabolite clusters called genetically influenced metabotypes. We assigned causal genes for 62.4% of these genetically influenced metabotypes, providing new insights into fundamental metabolite physiology and clinical relevance, including metabolite-guided discovery of potential adverse drug effects (DPYD and SRD5A2). We show strong enrichment of inborn errors of metabolism-causing genes, with examples of metabolite associations and clinical phenotypes of non-pathogenic variant carriers matching characteristics of the inborn errors of metabolism. Systematic, phenotypic follow-up of metabolite-specific genetic scores revealed multiple potential etiological relationships., (© 2022. The Author(s).)

Background: Benign prostatic hyperplasia (BPH) is a urinary system disease with high prevalence among the middle and elder men. In BPH, proliferation of prostate cells and the imbanlance between androgen and estrogen are both important inducers. Previous studies have demonstrated that compounds from Ligustri Lucidi Fructus (LLF) and Ecliptae Herba (EH) are of phytoestrogenic or phytoandrogenic activities. The combination of LLF with EH at the ratio of 1:1 on crude drugs quantity is called Erzhi formula (EZF), which is used for in vivo research of our study., Purpose: This study aimed to investigate potential mechanisms of EZF and its active pharmaceutical ingredients on BPH in vitro and in vivo., Methods: Therapeutic effects of EZF was evaluated in E 2 /testosterone (1:100) induced BPH rats model. The pathological changes of prostate, concentrations of testosterone, DHT, E 2 , PSA in rats' plasma and prostate were detected. The expressions of PCNA, AR, ERα, ERβ, SRD5A1, SRD5A2 were measured in BPH rat prostates and E 2 -stimulated human benign prostatic epithelial cells (BPH-1)., Results: EZF treatment significantly attenuated rat prostate enlargement, alleviated BPH pathological features, and decreased the expression of PCNA. The up-regulation of AR, ERα, SRD5A1/2 expressions, and down-regulation of ERβ expression at prostate of rat BPH model were significantly blocked by EZF administration. The expression levels of testosterone, DHT, E 2 , PSA were strongly inhibited by EZF treatment. At the cellular level, ligustrosidic acid and echinocystic acid inhibited E 2 -induced BPH-1 cell proliferation and PCNA expressions, which were consistent with the results in vivo. And these two ingredients also down-regulated the expressions of AR, ERα, SRD5A1/2 and up-regulated the expression of ERβ in BPH-1 cells., Conclusion: EZF, ligustrosidic acid from LLF and echinocystic acid from EH showed inhibitive effects on BPH via down-regulating prostatic AR, ERα, SRD5A1/2 expressions and up-regulating ERβ expression., (Copyright © 2022. Published by Elsevier GmbH.)

Prostate cancer (PCa) is the most frequent malignancy in male urogenital system around worldwide. We performed molecular subtyping and prognostic assessment based on consensus genes in patients with PCa. Five cohorts containing 1,046 PCa patients with RNA expression profiles and recorded clinical follow-up information were included. Univariate, multivariate Cox regression analysis and least absolute shrinkage and selection operator (LASSO) Cox regression were used to select prognostic genes and establish the signature. Immunohistochemistry staining, cell proliferation, migration and invasion assays were used to assess the biological functions of key genes. Thirty-nine intersecting consensus prognostic genes from five independent cohorts were identified. Subsequently, an eleven-consensus-gene classifier was established. In addition, multivariate Cox regression analyses showed that the classifier served as an independent indicator of recurrence-free survival in three of the five cohorts. Combined receiver operating characteristic (ROC) analysis achieved synthesized effects by combining the classifier with clinicopathological features in four of five cohorts. SRD5A2 inhibits cell proliferation, while ITGA11 promotes cell migration and invasion, possibly through the PI3K/AKT signaling pathway. To conclude, we established and validated an eleven-consensus-gene classifier, which may add prognostic value to the currently available staging system., (© 2022. The Author(s).)

Oily skin from overactive sebaceous glands affects self-confidence and personality. There is report of an association between steroid 5-alpha reductase gene ( SRD5A ) expression and facial sebum production. There is no study of the effect of Asparagus racemosus Willd. root extract on the regulation of SRD5A mRNA expression and anti-sebum efficacy. This study extracted A. racemosus using the supercritical carbon dioxide fluid technique with ethanol and investigated its biological compounds and activities. The A. racemosus root extract had a high content of polyphenolic compounds, including quercetin, naringenin, and p -coumaric acid, and DPPH scavenging activity comparable to that of the standard L-ascorbic acid. A. racemosus root extract showed not only a significant reduction in SRD5A1 and SRD5A2 mRNA expression by about 45.45% and 90.86%, respectively, but also a reduction in the in vivo anti-sebum efficacy in male volunteers, with significantly superior percentage changes in facial sebum production and a reduction in the percentages of pore area after 15 and 30 days of treatment. It can be concluded that A. racemosus root extract with a high content of polyphenol compounds, great antioxidant effects, promising downregulation of SRD5A1 and SRD5A2 , and predominant facial sebum reduction and pore-minimizing efficacy could be a candidate for an anti-sebum and pore-minimizing active ingredient to serve in functional cosmetic applications.

Hepatocellular carcinoma (HCC) is the most common malignant liver tumor with high mortality and poor prognosis worldwide. This study aimed to identify hub genes and investigate the underlying molecular mechanisms in HCC progression by integrated bioinformatics analysis and experimental validation. Based on the Gene Expression Omnibus (GEO) databases and The Cancer Genome Atlas (TCGA), 12 critical differential co-expression genes were identified between tumor and normal tissues. Via survival analysis, we found higher expression of LCAT, ACSM3, IGF1, SRD5A2, THRSP and ACADS was associated with better prognoses in HCC patients. Among which, THRSP was selected for the next investigations. We found that THRSP mRNA expression was negatively correlated with its methylation and closely associated with clinical characteristics in HCC patients. Moreover, THRSP expression had a negative correlation with the infiltration levels of several immune cells (e.g., B cells and CD4+ T cells). qRT-PCR verified that THRSP was lower expressed in HCC tissues and cell lines compared with control. Silencing of THRSP promoted the migration, invasion, proliferation, and inhibited cell apoptosis of HCCLM and Huh7 cell lines. Decreased expression of THRSP promoted HCC progression by NF-κB, ERK1/2, and p38 MAPK signaling pathways. In conclusion, THRSP might serve as a novel biomarker and therapeutic target of HCC.

Background: Women facing increased energetic demands in childhood commonly have altered adult ovarian activity and shorter reproductive lifespan, possibly comprising a strategy to optimize reproductive success. Here, we sought to understand the mechanisms of early-life programming of reproductive function, by integrating analysis of reproductive tissues in an appropriate mouse model with methylation analysis of proxy tissue DNA in a well-characterized population of Bangladeshi migrants in the UK. Bangladeshi women whose childhood was in Bangladesh were found to have later pubertal onset and lower age-matched ovarian reserve than Bangladeshi women who grew-up in England. Subsequently, we aimed to explore the potential relevance to the altered reproductive phenotype of one of the genes that emerged from the screens., Results: Of the genes associated with differential methylation in the Bangladeshi women whose childhood was in Bangladesh as compared to Bangladeshi women who grew up in the UK, 13 correlated with altered expression of the orthologous gene in the mouse model ovaries. These mice had delayed pubertal onset and a smaller ovarian reserve compared to controls. The most relevant of these genes for reproductive function appeared to be SRD5A1, which encodes the steroidogenic enzyme 5α reductase-1. SRD5A1 was more methylated at the same transcriptional enhancer in mice ovaries as in the women's buccal DNA, and its expression was lower in the hypothalamus of the mice as well, suggesting a possible role in the central control of reproduction. The expression of Kiss1 and Gnrh was also lower in these mice compared to controls, and inhibition of 5α reductase-1 reduced Kiss1 and Gnrh mRNA levels and blocked GnRH release in GnRH neuronal cell cultures. Crucially, we show that inhibition of this enzyme in female mice in vivo delayed pubertal onset., Conclusions: SRD5A1/5α reductase-1 responds epigenetically to the environment and its downregulation appears to alter the reproductive phenotype. These findings help to explain diversity in reproductive characteristics and how they are shaped by early-life environment and reveal novel pathways that might be targeted to mitigate health issues caused by life-history trade-offs., (© 2021. The Author(s).)

Alterations in glucocorticoid metabolism may contribute to the development of obesity and insulin resistance (IR). Obesity in turn affects the androgen balance. The peripheral metabolism of steroids is equally an important determinant of their bioavailability and activity. The aim of this study was to evaluate steroid metabolism in obese children and to define which enzyme alterations are associated with IR. Clinical characteristics and anthropometric measurements were determined in 122 obese children and adolescents (72 girls, 50 boys) aged 8 - 18 years. 26 of them (21.3%) were diagnosed with IR (13 boys, 13 girls). Routine laboratory tests were performed and 24h urinary steroid excretion profiles were analyzed by gas chromatography/mass spectrometry. Positive relationship between 5α-reductase (SRD5A) activity and IR was found. According to the androsterone to etiocholanolone (An/Et) ratio the activity of SRD5A was significantly increased in obese children with IR, but the difference remained insignificant once the 5α-dihydrotestosterone to testosterone (5αDHT/T) ratio was considered. Furthermore, this relationship persisted in boys but was not observed in girls. The activity of 20α-hydroxysteroid dehydrogenase (20αHSD) and 20β-hydroxysteroid dehydrogenase (20βHSD) was reduced only in obese girls with IR. Conclude, in the context of obese children and adolescents with IR, we surmise that increased SRD5A represents a compensatory mechanism to reduce local glucocorticoid availability. This phenomenon is probably different in the liver (restriction) and in the adipose tissue (expected increase in activity). We show significant changes in 20αHSD and 20βHSD activity in obese girls with IR, but it is difficult to clearly determine whether the activity of these enzymes is an indicator of the function in their ovaries or adrenal glands., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Sumińska, Podgórski, Fichna and Fichna.)

Benign prostatic hyperplasia (BPH) is a frequently encountered disease in older men that affects sexual function and is capable of causing lower urinary tract dysfunction. Unfortunately, current treatment options for BPH primarily seek to address the lower urinary tract dysfunction aspect of the disease and do not improve sexual function. Yohimbine has been effectively used for decades to treat erectile dysfunction. Therefore, the objective of this study was to evaluate the inhibitory effect of yohimbine on BPH and explore the associated underlying mechanisms. Thirty-six rats were randomly divided into the control, BPH, finasteride (1 mg/kg), and yohimbine (2, 4, and 8 mg/kg) groups. Except for the rats in the control group, those in the other groups were subcutaneously injected testosterone propionate (5 mg/kg/day) daily for a period of 4 weeks to establish BPH models. They were also administration the corresponding drug daily for a period of 6 weeks. After the treatments, in addition to determining prostate wet weight and index, the histopathological status of the prostate was observed, and the levels of testosterone, dihydrotestosterone, prostatic acid phosphatase, the prostate-specific antigen, proliferating cell nuclear antigen, and steroid 5α-reductase were determined. Specifically, the administration of 2, 4, and 8 mg/kg yohimbine inhibited prostatic index increase by 46.7, 55.1, and 69.3%, respectively, in BHP rats. Further, yohimbine significantly reduced the levels of dihydrotestosterone, prostatic acid phosphatase, prostate-specific antigen, proliferating cell nuclear antigen, and steroid 5α-reductase, suggesting that it exerts beneficial effects against BPH by modulating the steroid 5α-reductase pathway., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Network-based methods for the analysis of drug-target interactions have gained attention and rely on the paradigm that a single drug can act on multiple targets rather than a single target. In this study, we have presented a novel approach to analyze the interactions between the chemicals in the medicinal plants and multiple targets based on the complex multipartite network of the medicinal plants, multi-chemicals, and multiple targets. The multipartite network was constructed via the conjunction of two relationships: chemicals in plants and the biological actions of those chemicals on the targets. In doing so, we introduced an index of the efficacy of chemicals in a plant on a protein target of interest, called target potency score (TPS). We showed that the analysis can identify specific chemical profiles from each group of plants, which can then be employed for discovering new alternative therapeutic agents. Furthermore, specific clusters of plants and chemicals acting on specific targets were retrieved using TPS that suggested potential drug candidates with high probability of clinical success. We expect that this approach may open a way to predict the biological functions of multi-chemicals and multi-plants on the targets of interest and enable repositioning of the plants and chemicals.

Although 11-ketotestosterone (11KT) and testosterone (T) are major androgens in both teleosts and humans, their 5α-reduced derivatives produced by steroid 5α-reductase (SRD5A/srd5a), i.e., 11-ketodihydrotestosterone (11KDHT) and 5α-dihydrotestosterone (DHT), remains poorly characterized, especially in teleosts. In this study, we compared the presence and production of DHT and 11KDHT in Japanese eels and humans. Plasma 11KT concentrations were similar in both male and female eels, whereas T levels were much higher in females. In accordance with the levels of their precursors, 11KDHT levels did not show sexual dimorphism, whereas DHT levels were much higher in females. It is noteworthy that plasma DHT levels in female eels were higher than those in men. In addition, plasma 11KDHT was undetectable in both sexes in humans, despite the presence of 11KT. Three srd5a genes ( srd5a1 , srd5a2a and srd5a2b ) were cloned from eel gonads. All three srd5a genes were expressed in the ovary, whereas only both srd5a2 genes were expressed in the testis. Human SRD5A1 was expressed in testis, ovary and adrenal, whereas SRD5A2 was expressed only in testis. Human SRD5A1, SRD5A2 and both eel srd5a2 isoforms catalyzed the conversion of T and 11KT into DHT and 11KDHT, respectively, whereas only eel srd5a1 converted T into DHT. DHT and 11KDHT activated eel androgen receptor (ar)α-mediated transactivation as similar fashion to T and 11KT. In contrast, human AR and eel arβ were activated by DHT and11KDHT more strongly than T and 11KT. These results indicate that in teleosts, DHT and 11KDHT may be important 5α-reduced androgens produced in the gonads. In contrast, DHT is the only major 5α-reduced androgens in healthy humans., Competing Interests: Author YM was employed by the company ASKA Pharmaceutical Medical Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Yazawa, Inaba, Imamichi, Sekiguchi, Uwada, Islam, Orisaka, Mikami, Ida, Sato, Miyashiro, Takahashi, Khan, Suzuki, Umezawa and Kitano.)

The 5-reductases (5α-reductase types 1, 2 and 3 [5αR1-3], 5β-reductase [5βR]) are steroid hormone metabolising enzymes that hold fundamental roles in human physiology and pathology. They possess broad substrate specificity converting many steroid hormones to their 5α- and 5β-reduced metabolites, as well as catalysing crucial steps in bile acid synthesis. 5αRs are fundamentally important in urogenital development by converting testosterone to the more potent androgen 5α-dihydrotestosterone (5αDHT); inactivating mutations in 5αR2 lead to disorders of sexual development. Due to the ability of the 5αRs to generate 5αDHT, they are an established drug target, and 5αR inhibitors are widely used for the treatment of androgen-dependent benign or malignant prostatic diseases. There is an emerging body of evidence to suggest that the 5-reductases can impact upon aspects of health and disease (other than urogenital development); alterations in their expression and activity have been associated with metabolic disease, polycystic ovarian syndrome, inflammation and bone metabolism. This review will outline the evidence base for the extra-urogenital role of 5-reductases from in vitro cell systems, pre-clinical models and human studies, and highlight the potential adverse effects of 5αR inhibition in human health and disease., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Steroid 5α-reductase type I (SRD5A1) is a validated oncogene in many sex hormone-related cancers, but its role in multiple myeloma (MM) remains unknown. Based on gene expression profiling (GEP) of sequential MM samples during the disease course, we found that the aberrant expression of SRD5A1 was correlated with progression and poor prognosis in MM patients. In this study, the oncogenic roles of SRD5A1 were validated in human MM cell lines (ARP1 and H929) and the xenograft MM model as well as the 5TMM mouse model. MTT and flow cytometry were used to assess MM cell proliferation, cell cycle, and apoptosis post inducible knockdown SRD5A1 by lentivirus-mediated short-hairpin RNA (shRNA). Transcriptomic sequencing, immunofluorescence, and western blot were used to investigate the effects of SRD5A1 suppression on cell apoptosis and autophagy. Mechanistically, SRD5A1 downregulation simultaneously regulated both the Bcl-2 family protein-mediated apoptosis and the autophagic process via PI3K/Akt/mTOR signaling pathway in MM cells. Meanwhile, the autophagy inhibitor (3-methyladenine) and SRD5A1 inhibitor (Dutasteride) were utilized to evaluate their anti-myeloma effect. Thus, our results demonstrated that SRD5A1 downregulation simultaneously regulated both the apoptosis and the autophagic process in MM cells. The dual autophagy-apoptosis regulatory SRD5A1 may serve as a biomarker and potential target for MM progression and prognosis.

Steroid 5-α reductase (5AR) is responsible for the reduction of steroids to 5-α reduced metabolites, such as the reduction of testosterone to 5-α dihydrotestosterone (DHT). A new adverse outcome pathway (AOP) for 5AR inhibition to reduce female reproduction in fish (AOP 289) is under development to clarify the antiestrogenic effects of 5AR inhibitors in female fish. A sensitive method for the DHT analysis using chemical derivatization and liquid chromatography-tandem mass spectrometry was developed. A cell-based 5AR inhibition assay that utilizes human cell lines, a transient overexpression system, and fish cell lines was developed. The measured IC 50 values of two well-known 5AR inhibitors, finasteride and dutasteride, were comparable in the different systems. However, the IC 50 of dutasteride in the fish cell lines was lower than that in the human cell lines. Finasteride showed a higher IC 50 against the RTG-2 cell line. These results demonstrated that 5ARs inhibition could differ in terms of structural characteristics among species. The assay has high sensitivity and reproducibility and is suitable for the application in 5AR inhibition screening for various endocrine disruption chemicals (EDCs). Future studies will continue to evaluate the quantitative inhibition of 5AR by EDCs to compare the endocrine-disrupting pathway in different species.

Steroid hormones are essential in stress response, immune system regulation, and reproduction in mammals. Steroids with 3-oxo-Δ 4 structure, such as testosterone or progesterone, are catalyzed by steroid 5α-reductases (SRD5As) to generate their corresponding 3-oxo-5α steroids, which are essential for multiple physiological and pathological processes. SRD5A2 is already a target of clinically relevant drugs. However, the detailed mechanism of SRD5A-mediated reduction remains elusive. Here we report the crystal structure of PbSRD5A from Proteobacteria bacterium, a homolog of both SRD5A1 and SRD5A2, in complex with the cofactor NADPH at 2.0 Å resolution. PbSRD5A exists as a monomer comprised of seven transmembrane segments (TMs). The TM1-4 enclose a hydrophobic substrate binding cavity, whereas TM5-7 coordinate cofactor NADPH through extensive hydrogen bonds network. Homology-based structural models of HsSRD5A1 and -2, together with biochemical characterization, define the substrate binding pocket of SRD5As, explain the properties of disease-related mutants and provide an important framework for further understanding of the mechanism of NADPH mediated steroids 3-oxo-Δ 4 reduction. Based on these analyses, the design of therapeutic molecules targeting SRD5As with improved specificity and therapeutic efficacy would be possible.

Aims and Objective: In this study, a novel quantitative structure activity relationship (QSAR) model has been developed for inhibitors of human 5-alpha reductase type II, which are used to treat benign prostate hypertrophy (BPH)., Methods: The dataset consisted of 113 compounds-mainly nonsteroidal-with known inhibitory concentration. Then 3D structures of compounds were optimized and molecular structure descriptors were calculated. The stepwise multiple linear regression was used to select descriptors encoding the inhibitory activity of the compounds. Multiple linear regression (MLR) was used to build up the linear QSAR model., Results: The results obtained revealed that the descriptors which best describe the activity were atom type electropological state, carbon type, radial distribution function (RDF), barysz matrix and molecular linear free energy relation. The suggested model could achieve satisfied square correlation coefficient of R 2 = 0.72, higher than of many previous studies, indicating its superiority. Rigid validation criteria were met using external data with Q 2 ˃ 0.5 and R 2 = 0.75, reflecting the predictive power of the model., Conclusion: The QSAR model was applied for screening botanical components of herbal preparations used to treat BPH, and could predict the activity of some, among others, making reasonable attribution to the proposed effect of these preparations. Gamma tocopherol was found to be an active inhibitor, in consistence with many previous studies, anticipating the power of this model in the prediction of new candidate molecules and suggesting further investigations., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Background: Circadian rhythm disruption impacts a wide range of physiological processes, including fertility. However, the effect of circadian disruption on male spermatogenesis and fertility, and treatments for these effects have been largely unexplored at the molecular level., Methods: In this study, we examined the effects of genipin on improving the reproductive health problems caused by circadian disruption. Three groups of animals were fed under different conditions: control group (normal T cycle with saline), group of shortened T cycles (Light/Dark = 4 hours/4 hours) with saline, and a group of shortened T cycles with genipin by oral gavage. The male fertility was evaluated by fertility study and pups parameters analysis after successful sexual behavior and mating with female mice. We sacrificed the treated animals after 5 or 10 weeks and collected the testis, sperm and serum for histological analysis, sperm motility assay, and serum hormone detection, respectively. Furthermore, the effect of genipin was assessed by detection of progesterone secretion and steroidogenic key proteins expression, including StAR and CYP11A1, in mouse Leydig tumor MLTC-1 cells., Results: Male mice exposed to shortened light-dark cycles, much shorter than 24 hours, had reduced fertility with decreased sperm concentrations and sperm motility. Male mice under circadian disruption have reduced testis size and abnormal morphology, leading to lower fertility rates, reduced litter size and pup body weight. Treatment with exogenous genipin, a natural plant-derived compound, alleviated circadian disruption-induced damage to fertility and spermatogenesis and normalized testosterone, dihydrotestosterone (DHT), and androstenedione (ASD) levels in the male mice. The levels of key proteins involved in steroidogenesis, StAR and CYP11A1, were reduced in mouse testes after the circadian disruption, but genipin treatment restored the reduction. The mRNA expression of SRD5A1, which encodes an androgen synthesis enzyme, was also upregulated by genipin treatment. Furthermore, genipin treatment showed a positive effect on steroidogenesis in MLTC-1 cells, resulting in an increase in hormone secretion and the upregulation of StAR and CYP11A1., Conclusions: Our results showed an association between circadian disruption and reproductive health problems in male mice and indicated that treatments with genipin have positive effects on the reproductive health of male mice with circadian rhythm disorders.

Steroid 5 alpha-reductase 3 (SRD5A3) is an important molecule in glycosylation metabolism and steroid hormone formation. It is differentially expressed in human fetal liver, endometrial cancer and prostate cancer; however, its prognostic value and biological function in hepatocellular carcinoma (HCC) remain unclear. Here, bioinformatics analysis was employed to explore the expression and prognostic significance of SRD5A3 in various cancers including HCC. Additionally, clinical specimens of HCC were applied to analyze the expression of SRD5A3. SRD5A3-underexpressed HCC cell lines were established to test the effect of SRD5A3 on cell proliferation in in vitro and in vivo . We found that the elevated expression of SRD5A3 was common in many cancers with poor prognosis. Moreover, public datasets and our specimens revealed that SRD5A3 was also upregulated in HCC tissues and associated with clinical stage and patient's gender. Kaplan-Meier survival analysis showed that higher SRD5A3 level predicted poor overall survival, progression-free survival, relapse-free survival and disease specific survival in HCC patients. Further experiments showed that the lack of SRD5A3 inhibited the growth of HCC. Collectively, these findings indicate that SRD5A3 functions as an oncogene and might serve as a potential biomarker for prognosis and a therapeutic target for HCC.

Testosterone (T) affects β-cell function in men and women. T is a prohormone that undergoes intracrine conversion in target tissues to the potent androgen dihydrotestosterone (DHT) via the enzyme 5α-reductase (5α-R) or to the active estrogen 17β-estradiol (E2) via the aromatase enzyme. Using male and female human pancreas sections, we show that the 5α-R type 1 isoform (SRD5A1) and aromatase are expressed in male and female β-cells. We show that cultured male and female human islets exposed to T produce DHT and downstream metabolites. In these islets, exposure to the 5α-R inhibitors finasteride and dutasteride inhibited T conversion into DHT. We did not detect T conversion into E2 from female islets. However, we detected T conversion into E2 in islets from two out of four male donors. In these donors, exposure to the aromatase inhibitor anastrozole inhibited E2 production. Notably, in cultured male and female islets, T enhanced glucose-stimulated insulin secretion (GSIS). In these islets, exposure to 5α-R inhibitors or the aromatase inhibitor both inhibited T enhancement of GSIS. In conclusion, male and female human islets convert T into DHT and E2 via the intracrine activities of SRD5A1 and aromatase. This process is necessary for T enhancement of GSIS., (© 2020 by the American Diabetes Association.)

Human steroid 5α-reductase 2 (SRD5A2) is an integral membrane enzyme in steroid metabolism and catalyzes the reduction of testosterone to dihydrotestosterone. Mutations in the SRD5A2 gene have been linked to 5α-reductase deficiency and prostate cancer. Finasteride and dutasteride, as SRD5A2 inhibitors, are widely used antiandrogen drugs for benign prostate hyperplasia. The molecular mechanisms underlying enzyme catalysis and inhibition for SRD5A2 and other eukaryotic integral membrane steroid reductases remain elusive due to a lack of structural information. Here, we report a crystal structure of human SRD5A2 at 2.8 Å, revealing a unique 7-TM structural topology and an intermediate adduct of finasteride and NADPH as NADP-dihydrofinasteride in a largely enclosed binding cavity inside the transmembrane domain. Structural analysis together with computational and mutagenesis studies reveal the molecular mechanisms of the catalyzed reaction and of finasteride inhibition involving residues E57 and Y91. Molecular dynamics simulation results indicate high conformational dynamics of the cytosolic region that regulate NADPH/NADP + exchange. Mapping disease-causing mutations of SRD5A2 to our structure suggests molecular mechanisms for their pathological effects. Our results offer critical structural insights into the function of integral membrane steroid reductases and may facilitate drug development.

Background and Objective: Our patient cohort revealed that obesity is strongly associated with steroid-5α reductase type 2 (SRD5A2) promoter methylation and reduced protein expression. The underlying mechanism of prostatic growth in this population is poorly understood. Here we addressed the question of how obesity, inflammation, and steroid hormones affect the development of benign prostatic hyperplasia (BPH)., Material and Methods: We used preadipocytes, macrophages, primary human prostatic stromal cells, prostate tissues from high-fat diet-induced obese mice, and 35 prostate specimens that were collected from patients who underwent transurethral resection of the prostate (TURP). RNA was isolated and quantified with RT-PCR. Genome DNA was extracted and SRD5A2 promoter methylation was determined. Sex hormones were determined by high-performance liquid chromatography-tandem mass spectrometry. Protein was extracted and determined by ELISA test., Results: In prostatic tissues with obesity, the levels of inflammatory mediators were elevated. SRD5A2 promoter methylation was promoted, but SRD5A2 expression was inhibited. Inflammatory mediators and saturated fatty acid synergistically regulated aromatase activity. Obesity promoted an androgenic to estrogenic switch in the prostate., Conclusions: Our findings suggest that obesity-associated inflammation induces androgenic to estrogenic switch in the prostate gland, which may serve as an effective strategy for alternative therapies for management of lower urinary tract symptoms associated with BPH in select individuals.

Testosterone and its 5α-reduced form, 5α-dihydrotestosterone, were previously thought to represent the only active androgens in humans. However, recent studies have shown that the potent androgen, 11-ketotestosterone, derived from the adrenal androgen precursor, 11β-hydroxyandrostenedione, may in fact serve as the primary androgen in healthy women. Yet, despite recent renewed interest in these steroids, their downstream metabolism has remained undetermined. We therefore set out to investigate the metabolism of 11-ketotestosterone by characterising the 5α- or 5β-reduction commitment step. We show that inactivation of 11-ketotestosterone is predominantly driven by AKR1D1, which efficiently catalyses the 5β-reduction of 11-ketotestosterone, committing it to a metabolic pathway that terminates in 11-ketoetiocholanolone. We demonstrate that 5α-reduction of 11-ketotestosterone is catalysed by SRD5A2, but not SRD5A1, and terminates in 11-ketoandrosterone, but is only responsible for a minority of 11-ketotestosterone inactivation. However, as 11-ketoetiocholanolone is also generated by the metabolism of the glucocorticoid cortisone, 11-ketoandrosterone should be considered a more specific urinary marker of 11-ketotestosterone production., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The potent androgen 5α-dihydrotestosterone irreversibly derives from testosterone via the activity of steroid 5α-reductases (5αRs). The major 5αR isoforms in most species, 5αR1 and 5αR2, have not been purified to homogeneity. We report here the heterologous expression of polyhistidine-tagged, codon-optimized human 5αR1 and 5αR2 cDNAs in Escherichia coli. A combination of the nonionic detergents Triton X-100 and Nonidet P-40 enabled solubilization of these extremely hydrophobic integral membrane proteins and facilitated purification with affinity and cation-exchange chromatography methods. For functional reconstitution, we incorporated the purified isoenzymes into Triton X-100-saturated dioleoylphosphatidylcholine liposomes and removed excess detergent with polystyrene beads. Kinetic studies indicated that the 2 isozymes differ in biochemical properties, with 5αR2 having a lower apparent Km for testosterone, androstenedione, progesterone, and 17-hydroxyprogesterone than 5αR1; however, 5αR1 had a greater capacity for steroid conversion, as reflected by a higher Vmax than 5αR2. Both enzymes preferred progesterone as substrate over other steroids, and the catalytic efficiency of purified reconstituted 5αR2 exhibited a sharp pH optimum at pH 5. Intriguingly, we found that the prostate-cancer drug-metabolite 3-keto-∆ 4-abiraterone is metabolized by 5αR1 but not 5αR2, which may serve as a structural basis for isoform selectivity and inhibitor design. The functional characterization results with the purified reconstituted isoenzymes paralleled trends obtained with HEK-293 cell lines stably expressing native 5αR1 and 5αR2. Access to purified human 5αR1 and 5αR2 will advance studies of these important enzymes and might help to clarify their contributions to steroid anabolism and catabolism., (© Endocrine Society 2020. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Human steroid 5α-reductase 2 (SRD5A2) plays a determinative role in the masculinization of external genitalia. To date, approximately 114 different mutations of the SRD5A2 gene have been reported; however, little information is available about their impact on catalytic function or their three-dimensional (3D) structures. We determined the effect of point mutations on the testosterone-depend kinetic constants (K m,app and V max,app ) and structural characteristics of SRD5A2 from Mexican patients with 46,XY-steroid 5α-reductase 2 deficiency. PCR-SSCP assays identified ten distinct gene variants and sequencing analysis identified missense mutations [p.V3I, p.S14R, p.A52T, p.F118L, p.R145W, p.R171S, p.L226P, p.F229S, p.S245Y, and p.A248V]. Mutations were re-created by site-directed mutagenesis and expressed in HEK293 cells. Functional studies demonstrated that 8 variants led to partial (K m,app = 0.16-2.6 μM; V max,app = 224-2640 pmol/mg P/min) or complete losses of activity compared to the wild-type enzyme (K m,app = 0.7 μM; V max,app = 4044 pmol/mg P/min). All the mutations were assessed using multiple software tools and the results predicted that all of the mutations were associated with disease or damage. Mapping mutations on the model of a 3D structure of SRD5A2 demonstrated alterations in contact sites with their proximal amino acids. Our data show that mutations affect the catalytic efficiency (V max /K m ) or result in residual enzymatic activity, which could be due to erroneous interactions between amino acid residues, the substrate testosterone, or NADPH., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Neurosteroids are potent allosteric modulators of GABA A receptors (GABA A Rs). Although the effects of exogenous neurosteroids on GABA A R function are well documented, less is known about effects of neurosteroids produced by local endogenous biosynthesis. The neurosteroidogenic enzymes 5α-reductase and 3α-hydroxysteroid dehydrogenase are expressed in two nuclei of somatosensory thalamus, the thalamic reticular nucleus (nRT) and ventrobasal nucleus (VB). Here, the effects of acute blockade of neurosteroidogenesis by the 5α-reductase inhibitor finasteride on phasic and tonic GABA A R-mediated currents were examined in nRT and VB of mice. In nRT, finasteride altered the decay and amplitude, but not the frequency, of phasic currents, with no effect on tonic inhibition. In VB neurons, by contrast, finasteride reduced both the size and frequency of phasic currents, and also reduced the degree of tonic inhibition. These studies thus provide novel evidence for endogenous modulation of GABA A R function by 5α-reduced neurosteroids in the mature thalamus., (© 2019 Wiley Periodicals, Inc.)

The aim of the present study was to examine the effect of prolonged use of finasteride on serum levels of dihydrotestosterone (DHT), estradiol (E2), progesterone, testosterone and androstenedione in women during the menstrual period. Further, to screen and compare the 5α-reductase activities through the expression of SRD5A1, SRD5A2 and AR gene and to determine the level of VEGF, VKOR and SAA gene expression and DNA damage. A total of 30 Saudi women aged between 25 and 35 years were enrolled in the study. The selected women were divided into two groups. The first group (n = 15) received 5 mg finasteride/day for prolonged period of one year and second group (n = 15) was taken as a healthy control. ELISA technique was used for measuring the serum levels of the targeted hormones, and Comet assay was used for checking the DNA integrity. Our findings revealed significant decrement of DHT, E2, progesterone and androstenedione levels and elevated levels of testosterone in group treated with daily oral doses of 5 mg finasteride/day compared with the control subjects. mRNA expression suggested that finasteride has concrete effects on the gene expression of the selected genes from the treated group in comparison with the control group. In addition, finasteride induced DNA damage, and heavy menstrual bleeding was noted in women treated with finasteride. In conclusion, the present findings revealed that finasteride has adverse health effects in women associated with gonadal sex steroids alterations, DNA damage and heavy menstrual bleeding with no consensus in the treatment of androgenetic alopecia in women., (© 2020 The Author(s).)

In Mongolia, yak (Bos grunniens) are able to live in alpine areas and their products greatly influence the lives of the local people. Increased vigour in hybridized yak and cattle can offer benefits for livestock farmers. However, male hybrids show reproductive defects resulting from spermatogenesis arrest, affecting the conservation and maintenance of dominant traits in the next generation. The underlying mechanisms involved in hybrid cattle-yak infertility have recently been investigated; however, the genetic cause is still unclear. Androgens and androgen receptor (AR) signalling are required for spermatogenesis. We, therefore, evaluated the expression of AR, 3β-hydroxysteroid dehydrogenase (3βHSD) and 5α-reductase 2 (SRD5A2) in Leydig cells to investigate their function in cattle-yak spermatogenesis. Testicular tissues from yaks (1-3 years old) and hybrids (F1-F3, 2 years old) were collected and subjected to immunohistochemistry and image analyses to investigate the expression of each parameter in the Leydig cells. After maturation at 2 years, the expression levels of AR increased and the levels of 3βHSD decreased, but the SRD5A2 levels remained constant in yak. However, the cattle-yak hybrid F2 showed immature testicular development and significantly different expression levels of AR and 3βHSD compared with mature yak. These results suggest that the decreased expression of AR and increased expression of 3βHSD in the Leydig cells of cattle-yak hybrid testes may represent one of the causes of infertility. Our study might help in solving the problem of infertility in crossbreeding., (© 2019 Blackwell Verlag GmbH.)

Life-course experiences have been postulated to program hypothalamus-pituitary-adrenal (HPA) axis activity, suggesting that HPA axis activity is, at least partially, stable over time. Yet, there is paucity of data on the long-term stability of cortisol production and metabolism. We performed a prospective follow-up study in twins recruited from a nationwide register to estimate the stability of cortisol production and metabolism over time, and the contribution of genetic and environmental factors to this stability. In total, 218 healthy mono- and dizygotic twins were included. At the ages of 9, 12 and 17 years, morning urine samples were collected for assessment (by gas chromatography-tandem mass spectrometry) of cortisol metabolites, enabling the calculation of cortisol metabolite excretion rate and cortisol metabolism activity. Our results showed a low stability for both cortisol metabolite excretion rate (with correlations <.20) and cortisol metabolism activity indices (with correlations of .25 to .46 between 9 and 12 years, -.02 to .15 between 12 and 17 years and .09 to .28 between 9 and 17 years). Because of the low stability over time, genetic and environmental contributions to this stability were difficult to assess, although it seemed to be mostly determined by genetic factors. The low stability in both cortisol production and metabolism between ages 9 and 17 years reflects the dynamic nature of the HPA axis.

Background: Nandrolone decanoate (ND) is an anabolic-androgenic steroid, and its indiscriminate use leads to subclinical alterations in the hypothalamic-pituitary-gonadal axis and androgen-dependent organs., Objectives: To evaluate the effects of ND, either alone or in combination with resistance exercise (RE), on the levels of sex hormones, converting enzymes, and steroid receptors and the morphology of the ventral prostate (VP) in adult and aged rats., Methods: Forty Sprague-Dawley adult and aged rats were divided into four groups each, sedentary and trained with and without ND. The groups received treatments over 8 weeks. Adult animals were sacrificed immediately following treatment completion, while the aged groups were left untreated until 300 days of age., Results: Adult and aged animals showed reductions in testosterone levels following the different treatments, and 17β-estradiol levels were decreased in the ND-treated groups. The level of 5α-reductase type 2 (5αR2) and aromatase was increased significantly in the prostates of adult animals that performed RE. However, aromatase levels were decreased in the prostates of aged animals that performed RE and were treated with ND, while 5αR2 levels were reduced in aged animals that performed RE without ND treatment. When sex receptors levels were examined, the aged and trained animals presented low androgen receptor (AR) levels. Estrogen receptors (ERs) levels were increased in the prostates of adult animals that received ND. ERβ levels were reduced after treatments in aged animals. The heights of the prostatic epithelium were reduced in all adult treated animals, coinciding with increases in PCNA and PAR4 levels., Discussion: ND and RE alter the levels of hormone, converting enzymes, and sex steroid receptors and the morphology of the VP. These effects were observed in both adult and aged rats., Conclusion: ND, either with or without RE, during post-puberty stage is able to interfere with the morphophysiology of the prostate., (© 2019 American Society of Andrology and European Academy of Andrology.)

Androgens exert important effects both in androgen-responsive tissues and in the intestinal tract. To determine the impact of the gut microbiota (GM) on intestinal androgen metabolism, we measured unconjugated (free) and glucuronidated androgen levels in intestinal contents from the small intestine, with a low bacterial density, and from cecum and colon, with a high bacterial density. Using a specific, sensitive gas chromatography-tandem mass spectrometry method, we detected high levels of glucuronidated testosterone (T) and dihydrotestosterone (DHT) in small intestinal content of mice of both sexes, whereas in the distal intestine we observed remarkably high levels of free DHT, exceeding serum levels by >20-fold. Similarly, in young adult men high levels of unconjugated DHT, >70-fold higher than in serum, were detected in feces. In contrast to mice with a normal GM composition, germ-free mice had high levels of glucuronidated T and DHT, but very low free DHT levels, in the distal intestine. These findings demonstrate that the GM is involved in intestinal metabolism and deglucuronidation of DHT and T, resulting in extremely high free levels of the most potent androgen, DHT, in the colonic content of young and healthy mice and men.

Context: Lespedeza cuneata G. Don (Fabaceae), has been used as a traditional treatment of various diseases. There is a report L. cuneata effects on hormone replacement therapy for endocrine-related disease. However, studies related to benign prostatic hyperplasia (BPH) have not been investigated., Objective: The effects of L. cuneata aqueous extract (LCW) on testosterone-induced prostatic hyperplasia (TPH) were examined., Materials and Methods: Male Wistar rats (10 weeks, 330-350 g) were randomly divided to 6 groups (n = 6): Control group; TPH group (3 mg/kg, s.c, daily); TPH + LCW (25, 50, 100 mg/kg); TPH + Finasteride 10 mg/kg for 6 weeks. At the end of treatment, histological change of prostate, serum dihydrotestosterone (DHT) level, mRNA expression of 5α-reductase, inflammatory factors, proliferating cell nuclear antigen (PCNA) and fibroblast growth factor-2 (FGF-2) in prostate were examined. Then, LCW was treated with BPH-1, a human BPH cell line, at 25, 50, 100 μg/mL for 24 h and examine mRNA level of androgen receptor (AR) and prostate-specific antigen (PSA). In addition, the content of vicenin-2 was analyzed., Results: LCW treatment of TPH inhibited serum DHT levels by 54.5, 51.2 and 54.1% and mRNA expression of 5α-reductase were inhibited 54.3, 61.3 and 73.6%, respectively. In addition, mRNA expression of inflammatory factors, PCNA and FGF-2 were decreased in the prostate of rats. Also, LCW attenuated mRNA level of AR and PSA in BPH-1 cell. The content of vicenin-2 in the LCW was analyzed to 0.89 mg/g., Discussion and Conclusions: Based on the results, LCW is a potential pharmacological candidate for the treatment of prostatic hyperplasia.

The dorsal motor nucleus of the vagus (DMV) contains the preganglionic motor neurons important in the regulation of glucose homeostasis and gastrointestinal function. Despite the role of sex in the regulation of these processes, few studies examine the role of sex and/or ovarian cycle in the regulation of synaptic neurotransmission to the DMV. Since GABAergic neurotransmission is critical to normal DMV function, the present study used in vitro whole cell patch-clamping to investigate whether sex differences exist in GABAergic neurotransmission to DMV neurons. It additionally investigated whether the ovarian cycle plays a role in those sex differences. The frequency of phasic GABA A receptor-mediated inhibitory postsynaptic currents in DMV neurons from females was lower compared with males, and this effect was TTX sensitive and abolished by ovariectomy (OVX). Amplitudes of GABAergic currents (both phasic and tonic) were not different. However, females demonstrated significantly more variability in the amplitude of both phasic and tonic GABA A receptor currents. This difference was eliminated by OVX in females, suggesting that these differences were related to reproductive hormone levels. This was confirmed for GABAergic tonic currents by comparing females in two ovarian stages, estrus versus diestrus. Female mice in diestrus had larger tonic current amplitudes compared with those in estrus, and this increase was abolished after administration of a 5α-reductase inhibitor but not modulation of estrogen. Taken together, these findings demonstrate that DMV neurons undergo GABA A receptor activity plasticity as a function of sex and/or sex steroids. NEW & NOTEWORTHY Results show that GABAergic signaling in dorsal vagal motor neurons (DMV) demonstrates sex differences and fluctuates across the ovarian cycle in females. These findings are the first to demonstrate that female GABA A receptor activity in this brain region is modulated by 5α-reductase-dependent hormones. Since DMV activity is critical to both glucose and gastrointestinal homeostasis, these results suggest that sex hormones, including those synthesized by 5α-reductase, contribute to visceral, autonomic function related to these physiological processes.

Introduction: The extraction of salient information from the environment is modulated by the activation of dopamine receptors. Using rodent models, we previously reported that gating deficits caused by dopamine receptor activation - as measured by the prepulse inhibition (PPI) of startle - are effectively opposed by inhibitors of the steroidogenic enzyme 5α-reductase (5αR). The specific 5αR isoenzyme and steroids implicated in these effects, however, remain unknown., Methods: The effects of the selective D 1 dopamine receptor agonist SKF-82958 (SKF, 0.3 mg/kg, IP) and D 2 receptor agonist quinpirole (QUIN, 0.5 mg/kg, IP) were tested in the startle reflex and PPI of knockout (KO) mice for either 5αR type 1 (5αR1) or type 2 (5αR2). Furthermore, we established whether these effects may be modified by the 5α-reduced steroids dihydroprogesterone (DHP), allopregnanolone (AP), dihydrotestosterone (DHT), 5α-androstane-3α,17β-diol (3α-diol), or androsterone. To test the mechanisms whereby 5αR products may alter the PPI-disrupting properties of D 1 agonists, we studied the involvement of GABA-A and PXR, two receptors targeted by neuroactive steroids. Specifically, we tested the effects of SKF in combination with the GABA-A antagonist bicuculline, as well as in KO mice for the GABA-A δ subunit and PXR., Results: 5αR1, but not 5αR2, knockout (KO) mice were insensitive to the PPI-disrupting effects of SKF. This sensitivity was reinstated by AP (3 mg/kg, IP), but not other 5α-reduced steroids. The PPI deficits induced by SKF were not modified by bicuculline, δ-subunit KO mice and PXR KO mice., Conclusions: These results collectively suggest that 5αR1 enables the negative effects of D 1 dopamine receptor activation on information processing via production of AP. The contribution of AP to the PPI-disrupting mechanisms of D 1 receptor agonists, however, do not appear to be mediated by either GABA-A or PXR receptors., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Flurbiprofen is one of the nonsteroidal anti-inflammatory drugs. Whether flurbiprofen affects androgen biosynthesis in Leydig cells is still unknown. Immature Leydig cells (ILCs) isolated from 35-day-old male Sprague-Dawley rats were cultured with 0-100 μM flurbiprofen for 24 h and medium androgen levels and Leydig cell mRNA levels were measured. Immature Leydig cells were also incubated with 100 μM flurbiprofen for 3 h in combination with luteinizing hormone (LH), 8bromo-cAMP, 22R-OH-cholesterol, pregnenolone, progesterone, androstenedione, testosterone, and dihydrotestosterone, respectively, and medium androgen levels were measured. The ROS generation and apoptosis rate were also investigated. The direct effects of flurbiprofen on androgen biosynthetic and metabolizing enzyme activities were measured. Flurbiprofen significantly inhibited basal, LH, and 8bromo-cAMP stimulated androgen production at 10 and 100 μM. Further study demonstrated that flurbiprofen competitively inhibited rat and human testis 3β-hydroxysteroid dehydrogenase (HSD3B) activity with the half maximal inhibitory concentration (IC 50 ) values of 0.95 μM for rat enzyme and 6.31 μM for human enzyme. In addition, flurbiprofen down-regulated the expression of Srd5a1 and Akr1c14 at 1, 10, and 100 μM. Flurbiprofen also down-regulated Lhcgr expression at 100 μM. Flurbiprofen at 10 and 100 μM increased ROS production and apoptosis rate of rat Leydig cells. In conclusion, flurbiprofen directly inhibits HSD3B activity and the expression levels of Srd5a1 and Akr1c14 in rat Leydig cells, thus leading to the reduction of androgen secretion.