Insight on the Intracrinology of Menopause: Androgen Production within the Human Vagina

Endocrinology ◽  
2020 ◽  
Vol 162 (2) ◽  
Author(s):  
Ilaria Cellai ◽  
Vincenza Di Stasi ◽  
Paolo Comeglio ◽  
Elisa Maseroli ◽  
Tommaso Todisco ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Steroid Receptors ◽  
Estrogen Receptor Α ◽  
Steroidogenic Enzymes ◽  
Vaginal Tissue ◽  
Liquid Chromatography Tandem Mass ◽  
Human Vagina ◽  
Polymerase Chain ◽  
Signaling Activation ◽  
Benign Gynecological Diseases

Abstract In this study, we investigated steroidogenic gene mRNA expression in human vaginas and verified the ability of human vagina smooth muscle cells (hvSMCs) to synthesize androgens from upstream precursor dehydroepiandrosterone (DHEA). As a readout for androgen receptor (AR) activation, we evaluated the mRNA expression of various androgen-dependent markers. hvSMCs were isolated from vagina tissues of women undergoing surgery for benign gynecological diseases. In these cells, we evaluated mRNA expression of several steroidogenic enzymes and sex steroid receptors using real time reverse transcription-polymerase chain reaction. Androgen production was quantified with liquid chromatography tandem-mass spectrometry (LC-MS/MS). In vaginal tissues, AR mRNA was significantly less expressed than estrogen receptor α, whereas in hvSMCs, its mRNA expression was higher than progestin and both estrogen receptors. In hvSMCs and in vaginal tissue, when compared to ovaries, the mRNA expression of proandrogenic steroidogenic enzymes (HSD3β1/β2, HSD17β3/β5), along with 5α-reductase isoforms and sulfotransferase, resulted as being more abundant. In addition, enzymes involved in androgen inactivation were less expressed than in the ovaries. The LC-MS/MS analysis revealed that, in hvSMCs, short-term DHEA supplementation increased Δ4-androstenedione levels in spent medium, while increasing testosterone and DHT secretion after longer incubation. Finally, androgenic signaling activation was evaluated through AR-dependent marker mRNA expression, after DHEA and T stimulation. This study confirmed that the human vagina is an androgen-target organ with the ability to synthesize androgens, thus providing support for the use of androgens for local symptoms of genitourinary syndrome in menopause.

scholarly journals Promoter Methylation ofSFRP3Is Frequent in Hepatocellular Carcinoma

2014 ◽  
Vol 2014 ◽  
pp. 1-10 ◽  
Author(s):  
Ya-Wen Lin ◽  
Yu-Lueng Shih ◽  
Gi-Shih Lien ◽  
Fat-Moon Suk ◽  
Chung-Bao Hsieh ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Chronic Hepatitis ◽  
Mrna Expression ◽  
Cell Lines ◽  
Promoter Methylation ◽  
Promoter Hypermethylation ◽  
Polymerase Chain ◽  
Related Proteins ◽  
And Control

Oncogenic activation of the Wnt/β-catenin signaling pathway is common in human cancers. The secreted frizzled-related proteins (SFRPs) function as negative regulators of Wnt signaling and have important implications in carcinogenesis. Because there have been no reports about the role ofSFRP3in hepatocellular carcinoma (HCC), we investigated the level of methylation and transcription ofSFRP3. Four HCC cell lines, 60 HCCs, 23 cirrhosis livers, 37 chronic hepatitis livers, and 30 control livers were prescreened forSFRP3promoter methylation by methylation-specific polymerase chain reaction (MS-PCR) and bisulfite sequencing.SFRP3promoter methylation was observed in 100%, 60%, 39.1%, 16.2%, and 0% in HCC cell lines, primary HCCs, cirrhosis livers, chronic hepatitis livers, and control livers, respectively. Demethylation treatment with 5-aza-2′-deoxycytidine in HCC cells restored or increased theSFRP3mRNA expression. We next used quantitative MS-PCR (QMSP) to analyze the methylation level ofSFRP3in 60 HCCs and their corresponding nontumor tissues. Methylation ofSFRP3promoter region in HCCs increased significantly compared with control tissues. There is a positive correlation between promoter hypermethylation andSFRP3mRNA downregulation. Our data suggest that promoter hypermethylation ofSFRP3is a common event in HCCs and plays an important role in regulation ofSFRP3mRNA expression.

scholarly journals The effects of tamoxifen and estradiol on myometrial differentiation and organization during early uterine development in the CD1 mouse

Reproduction ◽  
2009 ◽  
Vol 138 (2) ◽  
pp. 341-350 ◽  
Author(s):  
Mohamed K Mehasseb ◽  
S C Bell ◽  
M A Habiba
Keyword(s):  
Extracellular Matrix ◽  
Image Analysis ◽  
Steroid Receptors ◽  
Smooth Muscle Actin ◽  
Estrogen Receptor Α ◽  
Matrix Proteins ◽  
Muscle Actin ◽  
Uterine Adenomyosis ◽  
Circular Arrangement ◽  
Uterine Development

We used a neonatal mouse model to examine the histogenesis of uterine adenomyosis, and to test whether adenomyosis is due to an abnormality in myometrial differentiation, or in extracellular matrix proteins expression. We also studied the effects of tamoxifen and estradiol on uterine development, myometrial differentiation, and organization. Female CD1 pups were treated with oral tamoxifen (1 mg/kg) (n=27) or estradiol (0.1 mg/kg) (n=24) from age 1 to 5 days. Uteri from control (n=27) and treated mice were obtained on days 2, 5, 10, 15, and 42 of age. We examined the sections histologically, using image analysis and immunohistochemistry for α-smooth muscle actin (α-SMA), desmin, vimentin, laminin, fibronectin, and estrogen receptor-α. Following tamoxifen exposure, all uteri showed adenomyosis by 6 weeks of age (seen as early as day 10). The inner myometrium showed thinning, lack of continuity, disorganization, and bundling. α-SMA expression was normal. Desmin expression normally showed a wave of maturation that was absent in tamoxifen-treated mice. In the estradiol group, adenomyosis was not observed. All uterine layers were normally developed, but hypertrophied. The inner myometrium retained its circular arrangement. There was no difference in the localization of laminin or fibronectin between groups (laminin expression was reduced in the tamoxifen treated uteri). Vimentin could not be detected in all groups. Our results suggest that the development of the inner myometrium is particularly sensitive to estrogen antagonism, and can be affected by steroid receptors modulation. Disruption of the inner myometrium may play a role in the development of uterine adenomyosis.

G-protein Coupled Estrogen Receptor Expression in Growth Hormone Secreting and Non-Functioning Adenomas

2020 ◽  
Author(s):  
Hande Mefkure Ozkaya ◽  
Muge Sayitoglu ◽  
Nil Comunoglu ◽  
Eda Sun ◽  
Fatma Ela Keskin ◽  
...  
Keyword(s):  
Gene Expression ◽  
Estrogen Receptor ◽  
Protein Expression ◽  
G Protein ◽  
Estrogen Receptor Α ◽  
Receptor Expression ◽  
Positive Correlation ◽  
Polymerase Chain ◽  
G Protein Coupled ◽  
Transforming Gene

Abstract Purpose To evaluate the expression of G-protein coupled estrogen receptor (GPER1), aromatase, estrogen receptor α (ERα), estrogen receptor β (ERβ), pituitary tumor transforming gene (PTTG), and fibroblast growth factor 2 (FGF2) in GH-secreting and non-functioning adenomas (NFA). Methods Thirty patients with acromegaly and 27 patients with NFA were included. Gene expression was determined via quantitative reverse transcription polymerase chain reaction (QRT-PCR). Protein expression was determined via immunohistochemistry. Results There was no difference, in terms of gene expression of aromatase, ERα, PTTG, and FGF2 between the two groups (p>0.05 for all). ERβ gene expression was higher and GPER1 gene expression was lower in GH-secreting adenomas than NFAs (p<0.05 for all). Aromatase and ERβ protein expression was higher in GH-secreting adenomas than NFAs (p=0.01). None of the tumors expressed ERα. GPER1 expression was detected in 62.2% of the GH-secreting adenomas and 45% of NFAs. There was no difference in terms of GPER1, PTTG, FGF2 H scores between the two groups (p>0.05 for all). GPER1 gene expression was positively correlated to ERα, ERβ, PTTG, and FGF2 gene expression (p<0.05 for all). There was a positive correlation between aromatase and GPER1 protein expression (r=0.31; p=0.04). Conclusions GPER1 is expressed at both gene and protein level in a substantial portion of GH-secreting adenomas and NFAs. The finding of a positive correlation between GPER1 and ERα, ERβ, PTTG, and FGF2 gene expression and aromatase and GPER1 protein expression suggests GPER1 along with aromatase and classical ERs might mediate the effects of estrogen through upregulation of PTTG and FGF2.

scholarly journals Comparative study of estrogen receptor α, β mRNA expressions of endometriosis and normal endometrium in women and analysis of potential synthetic anti-estrogens in silico

2018 ◽  
Vol 91 (2) ◽  
Author(s):  
Eldafira Eldafira ◽  
Abinawanto Abinawanto ◽  
Luthfiralda Sjahfirdi ◽  
Asmarinah Asmarinah ◽  
Purnomo Soeharso ◽  
...  
Keyword(s):  
Estrogen Receptor ◽  
Mrna Expression ◽  
Estrogen Receptor Α ◽  
Assay Method ◽  
Enzyme Linked Immunosorbent Assay ◽  
Normal Endometrium ◽  
Pcr Method ◽  
Genetic And Environmental Factors

Endometriosis is a multifactorial disease in which genetic and environmental factors interact causing its pathogenesis. The aim of this study was to investigate the expression pattern of estrogen receptor α (ERα) and β (ERβ) in endometriosis patients compared to normal endometrioum (n=18) as a control by using Quantitative Real Time PCR method. Moreover, we also measured serum estradiol levels of endometriosis patients in the proliferation phase of the menstrual cycle using the enzyme-linked immunosorbent assay method. The mRNA expression of ERβ was significantly higher in the endometriosis group compared to control, and the result of t-test showed that were significantly different (P<0.05). Overexpression of ERβ in endometriosis was likely to have other significant important impacts in the pathology of endometriosis that allowed ERβ to stimulate prostaglandin production in endometriosis tissue and cells. Estradiol content did not correlate with the ERα expression, and it is weakly correlated with ERβ mRNA expression. Molecular docking analysis showed that ERα and ERβ have different binding interactions with synthetic antiestrogens, whereas the best inhibitor was Ral2 to ERα and Aco1 to ERβ. Thus, both inhibitors could be used as leads in further investigation of ERα, ERβ inhibitory activities in vitro and in vivo.

scholarly journals Two Approaches for Evaluating the Effects of Galangin on the Activities and mRNA Expression of Seven CYP450

Molecules ◽  
2019 ◽  
Vol 24 (6) ◽  
pp. 1171 ◽  
Author(s):  
Yin-Ling Ma ◽  
Feng Zhao ◽  
Jin-Tuo Yin ◽  
Cai-Juan Liang ◽  
Xiao-Li Niu ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Quantitative Polymerase Chain Reaction ◽  
Treated Group ◽  
Oral Drug ◽  
Pharmacokinetic Parameters ◽  
Sprague Dawley ◽  
Drug Bioavailability ◽  
Marker Compound ◽  
Liquid Chromatography Tandem Mass ◽  
Clinical Drugs

Galangin is a marker compound of honey and Alpinia officinarum Hance that exhibits great potential for anti-microbial, anti-diabetic, anti-obesity, anti-tumour and anti-inflammatory applications. Galangin is frequently consumed in combination with common clinical drugs. Here, we evaluated the effects of galangin on cytochrome P450 (CYP)-mediated metabolism, using two different approaches, to predict drug–drug interactions. Male Sprague Dawley rats were administered galangin daily for 8 weeks. A “cocktail-probes” approach was employed to evaluate the activities of different CYP450 enzymes. Blood samples of seven probe drugs were analysed using liquid chromatography-tandem mass spectrometry in positive and negative electrospray-ionisation modes. Pharmacokinetic parameters were calculated to identify statistical differences. CYP mRNA-expression levels were investigated in real-time quantitative polymerase chain reaction experiments. The galangin-treated group showed significantly decreased AUC0–∞ and Cmax values for CYP1A2, and CYP2B3. The galangin-treated group showed significantly increased AUC0–∞ and Cmax values for CYP2C13 and CYP3A1. No significant influences were observed in the pharmacokinetic profiles of CYP2C11, CYP2D4 and CYP2E1. The mRNA-expression results were consistent with the pharmacokinetic results. Thus, CYP450 enzyme activities may be altered by long-term galangin administration, suggesting galangin to be a promising candidate molecule for enhancing oral drug bioavailability and chemoprevention and reversing multidrug resistance.

Age-related changes in human Leydig cell status

2020 ◽  
Vol 35 (12) ◽  
pp. 2663-2676
Author(s):  
Valentina Mularoni ◽  
Valentina Esposito ◽  
Sara Di Persio ◽  
Elena Vicini ◽  
Gustavo Spadetta ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Sertoli Cell ◽  
Leydig Cell ◽  
Leydig Cells ◽  
Cell Number ◽  
Organ Donors ◽  
Steroidogenic Enzymes ◽  
Age Related ◽  
Age Range

Abstract STUDY QUESTION What are the consequences of ageing on human Leydig cell number and hormonal function? SUMMARY ANSWER Leydig cell number significantly decreases in parallel with INSL3 expression and Sertoli cell number in aged men, yet the in vitro Leydig cell androgenic potential does not appear to be compromised by advancing age. WHAT IS KNOWN ALREADY There is extensive evidence that ageing is accompanied by decline in serum testosterone levels, a general involution of testis morphology and reduced spermatogenic function. A few studies have previously addressed single features of the human aged testis phenotype one at a time, but mostly in tissue from patients with prostate cancer. STUDY DESIGN, SIZE, DURATION This comprehensive study examined testis morphology, Leydig cell and Sertoli cell number, steroidogenic enzyme expression, INSL3 expression and androgen secretion by testicular fragments in vitro. The majority of these endpoints were concomitantly evaluated in the same individuals that all displayed complete spermatogenesis. PARTICIPANTS/MATERIALS, SETTING, METHODS Testis biopsies were obtained from 15 heart beating organ donors (age range: 19–85 years) and 24 patients (age range: 19–45 years) with complete spermatogenesis. Leydig cells and Sertoli cells were counted following identification by immunohistochemical staining of specific cell markers. Gene expression analysis of INSL3 and steroidogenic enzymes was carried out by qRT-PCR. Secretion of 17-OH-progesterone, dehydroepiandrosterone, androstenedione and testosterone by in vitro cultured testis fragments was measured by LC-MS/MS. All endpoints were analysed in relation to age. MAIN RESULTS AND THE ROLE OF CHANCE Increasing age was negatively associated with Leydig cell number (R = −0.49; P &lt; 0.01) and concomitantly with the Sertoli cell population size (R= −0.55; P &lt; 0.001). A positive correlation (R = 0.57; P &lt; 0.001) between Sertoli cell and Leydig cell numbers was detected at all ages, indicating that somatic cell attrition is a relevant cellular manifestation of human testis status during ageing. INSL3 mRNA expression (R= −0.52; P &lt; 0.05) changed in parallel with Leydig cell number and age. Importantly, steroidogenic capacity of Leydig cells in cultured testis tissue fragments from young and old donors did not differ. Consistently, age did not influence the mRNA expression of steroidogenic enzymes. The described changes in Leydig cell phenotype with ageing are strengthened by the fact that the different age-related effects were mostly evaluated in tissue from the same men. LIMITATIONS, REASONS FOR CAUTION In vitro androgen production analysis could not be correlated with in vivo hormone values of the organ donors. In addition, the number of samples was relatively small and there was scarce information about the concomitant presence of potential confounding variables. WIDER IMPLICATIONS OF THE FINDINGS This study provides a novel insight into the effects of ageing on human Leydig cell status. The correlation between Leydig cell number and Sertoli cell number at any age implies a connection between these two cell types, which may be of particular relevance in understanding male reproductive disorders in the elderly. However aged Leydig cells do not lose their in vitro ability to produce androgens. Our data have implications in the understanding of the physiological role and regulation of intratesticular sex steroid levels during the complex process of ageing in humans. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by grants from Prin 2010 and 2017. The authors have no conflicts of interest. TRIAL REGISTRATION NUMBER N/A.

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