D-chiro-inositol in clinical practice: A perspective from The Experts Group on Inositol in Basic and Clinical Research (EGOI).

BACKGROUND: D-chiro-inositol is a natural molecule that, in association with its well-studied isomer myo-inositol, may play a role in treating various metabolic and gynecological disorders. OBJECTIVES: This perspective seeks to explore the mechanisms and functions of D-chiro-inositol, laying the foundations to discuss its use in clinical practice, across dysmetabolism, obesity, and hormonal dysregulation. METHODS: A narrative review of all the relevant papers known to the authors was conducted. OUTCOME: D-chiro-inositol acts through a variety of mechanisms, acting as an insulin sensitizer, inhibiting the transcription of aromatase, in addition to modulating white adipose tissue/brown adipose tissue trans differentiation. These different modes of action have potential applications in a variety of therapeutic fields including: PCOS, dysmetabolism, obesity, hypoestrogenic/hyperandrogenic disorders, and bone health. CONCLUSIONS: D-chiro-inositol mode of action has been studied in detail in recent years, resulting in a clear differentiation between D-chiro-inositol and its isomer myo-inositol. The insulin sensitizing activities of D-chiro-inositol are well understood; however, its potential applications in other fields, in particular obesity and hyperestrogenic/hypoandrogenic disorders in men and women, represent promising avenues of research that require further clinical study.

Glass wool column filtration for stallion sperm selection: A comparative analysis with the single-layer colloid centrifugation.

Glass wool column filtration (GWCF) selects human, bull, boar, dog and buffalo spermatozoa, but reports in the horse are scarce. Single-layer colloid centrifugation with Androcoll-E™ is currently the standard procedure to select good-quality equine sperm. This study was designed to assess GWCF (50 and 75 mg columns; GWCF-50 and GWCF-75, respectively) efficacy to select good-quality sperm from fresh and frozen-thawed equine semen, and to compare its performance with Androcoll-E™ colloid centrifugation. Percentage total motile (TM), progressively motile (PM), morphologically normal (MN), osmotically competent (HOS+) and acrosome-intact/osmotically competent (AI/HOS+) sperm were determined. In studies done with fresh semen samples (n = 17), suspensions subjected to GWCF-50 showed an improvement (p < .05) in PM and HOS+ sperm after selection. With GWCF-75, an increase (p < .05) in PM, MN and HOS+ sperm was observed. Results with GWCF were comparable or better than with Androcoll-E™ selection. Sperm recovery was similar between procedures for all semen parameters. Total sperm count recovery was lower after GWCF-75 (GWCF-50 = 60.0; GWCF-75 = 51.0; Androcoll-E™ = 76.0 million sperm; median; p = .013), but results on total progressive sperm count were similar (GWCF-50 = 23.0; GWCF-75 = 27.0; Androcoll-E™ = 24.0 million sperm; median; p = .3850). Using frozen-thawed semen samples (n = 16), an improvement (p < .05) in TM, PM, NM, HOS+ and AI/HOS+ sperm was observed in GWCF-75 filtrates. Results were comparable to Androcoll-E™ centrifugation, except HOS+ that increased (p < .05) only after GWCF-75. Recovery was comparable for all parameters in frozen samples. GWCF is a simple and low-cost procedure that selects equine sperm with a quality comparable to colloid centrifugation with Androcoll-E™.

Association between meteorological variables and semen quality: a retrospective study.

Spermatogenesis is a temperature-dependent process, and high summer temperatures have been linked to lower sperm concentration and count. However, reports describing the association between other meteorological variables and semen quality are scarce. This study evaluated the association between semen quality and temperature, humidity, pressure, apparent temperature (AT), temperature-humidity index (THI), simplified wet-bulb global temperature (sWBGT), and sunshine duration. Semen samples were obtained at the Laboratorio de Andrología y Reproducción (LAR, Argentina), from men undergoing routine andrology examination (n=11657) and computer-assisted sperm analysis (n=4705) following WHO 2010 criteria. Meteorological variables readings were obtained from the Sistema Meteorológico Nacional. Sperm quality parameters were negatively affected in summer when compared to winter. Additionally, there was a significant decrease in sperm kinematics between winter and spring. Branch and bound variable selection followed by multiple regression analysis revealed a significant association between semen quality and meteorological variables. Specifically, changes in sunshine duration and humidity reinforced the prognosis of semen quality. Highest/lowest sunshine duration and humidity quantiles resulted in decreased sperm concentration, count, motility, vitality and membrane competence, nuclear maturity, and sperm kinematics associated to highest sunshine duration and lowest humidity. Findings from this report highlight the relevance of environmental studies for predicting alterations in male reproductive health associated to variations in meteorological variables, especially considering the current climate changes around the planet due to global warming and its consequences for human health.

Inositols: From Established Knowledge to Novel Approaches.

Myo-inositol (myo-Ins) and D-chiro-inositol (D-chiro-Ins) are natural compounds involved in many biological pathways. Since the discovery of their involvement in endocrine signal transduction, myo-Ins and D-chiro-Ins supplementation has contributed to clinical approaches in ameliorating many gynecological and endocrinological diseases. Currently both myo-Ins and D-chiro-Ins are well-tolerated, effective alternative candidates to the classical insulin sensitizers, and are useful treatments in preventing and treating metabolic and reproductive disorders such as polycystic ovary syndrome (PCOS), gestational diabetes mellitus (GDM), and male fertility disturbances, like sperm abnormalities. Moreover, besides metabolic activity, myo-Ins and D-chiro-Ins deeply influence steroidogenesis, regulating the pools of androgens and estrogens, likely in opposite ways. Given the complexity of inositol-related mechanisms of action, many of their beneficial effects are still under scrutiny. Therefore, continuing research aims to discover new emerging roles and mechanisms that can allow clinicians to tailor inositol therapy and to use it in other medical areas, hitherto unexplored. The present paper outlines the established evidence on inositols and updates on recent research, namely concerning D-chiro-Ins involvement into steroidogenesis. In particular, D-chiro-Ins mediates insulin-induced testosterone biosynthesis from ovarian thecal cells and directly affects synthesis of estrogens by modulating the expression of the aromatase enzyme. Ovaries, as well as other organs and tissues, are characterized by a specific ratio of myo-Ins to D-chiro-Ins, which ensures their healthy state and proper functionality. Altered inositol ratios may account for pathological conditions, causing an imbalance in sex hormones. Such situations usually occur in association with medical conditions, such as PCOS, or as a consequence of some pharmacological treatments. Based on the physiological role of inositols and the pathological implications of altered myo-Ins to D-chiro-Ins ratios, inositol therapy may be designed with two different aims: (1) restoring the inositol physiological ratio; (2) altering the ratio in a controlled way to achieve specific effects.

Deficiency of fibroblast growth factor 2 (FGF-2) leads to abnormal spermatogenesis and altered sperm physiology.

In previous studies, we described the presence of fibroblast growth factor 2 (FGF-2) and its receptors (FGFRs) in human testis and sperm, which are involved in spermatogenesis and in motility regulation. The aim of the present study was to analyze the role of FGF-2 in the maintenance of sperm physiology using FGF-2 knockout (KO) mice. Our results showed that in wild-type (WT) animals, FGF-2 is expressed in germ cells of the seminiferous epithelium, in epithelial cells of the epididymis, and in the flagellum and acrosomal region of epididymal sperm. In the FGF-2 KO mice, we found alterations in spermatogenesis kinetics, higher numbers of spermatids per testis, and enhanced daily sperm production compared with the WT males. No difference in the percentage of sperm motility was detected, but a significant increase in sperm concentration and in sperm head abnormalities was observed in FGF-2 KO animals. Sperm from KO mice depicted reduced phosphorylation on tyrosine residues (a phenomenon that was associated with sperm capacitation) and increased acrosomal loss after incubation under capacitating conditions. However, the FGF-2 KO males displayed no apparent fertility defects, since their mating with WT females showed no differences in the time to delivery, litter size, and pup weight in comparison with WT males. Overall, our findings suggest that FGF-2 exerts a role in mammalian spermatogenesis and that the lack of FGF-2 leads to dysregulated sperm production and altered sperm morphology and function. FGF-2-deficient mice constitute a model for the study of the complex mechanisms underlying mammalian spermatogenesis.

The O-glycosylated ectodomain of FXYD5 impairs adhesion by disrupting cell-cell trans-dimerization of Na,K-ATPase ß1 subunits.

FXYD5 (also known as dysadherin), a regulatory subunit of the Na,K-ATPase, impairs intercellular adhesion by a poorly understood mechanism. Here, we determined whether FXYD5 disrupts the trans-dimerization of Na,K-ATPase molecules located in neighboring cells. Mutagenesis of the Na,K-ATPase ß1 subunit identified four conserved residues, including Y199, that are crucial for the intercellular Na,K-ATPase trans-dimerization and adhesion. Modulation of expression of FXYD5 or of the ß1 subunit with intact or mutated ß1-ß1 binding sites demonstrated that the anti-adhesive effect of FXYD5 depends on the presence of Y199 in the ß1 subunit. Immunodetection of the plasma membrane FXYD5 was prevented by the presence of O-glycans. Partial FXYD5 deglycosylation enabled antibody binding and showed that the protein level and the degree of O-glycosylation were greater in cancer than in normal cells. FXYD5-induced impairment of adhesion was abolished by both genetic and pharmacological inhibition of FXYD5 O-glycosylation. Therefore, the extracellular O-glycosylated domain of FXYD5 impairs adhesion by interfering with intercellular ß1-ß1 interactions, suggesting that the ratio between FXYD5 and α1-ß1 heterodimer determines whether the Na,K-ATPase acts as a positive or negative regulator of intercellular adhesion.

Involvement of fibroblast growth factor 2 (FGF2) and its receptors in the regulation of mouse sperm physiology.

Fibroblast growth factor 2 (FGF2) and its receptors (FGFRs) have been described in several tissues, where they regulate cellular proliferation, differentiation, motility and apoptosis. Although FGF2/FGFRs expression in the male reproductive tract has been reported, there is scarce evidence on their presence in the female reproductive tract and their involvement in the modulation of sperm function. Therefore, the objective of this study was to determine the expression of FGF2 in the female reproductive tract and to assess the role of the FGF2/FGFRs system in the regulation of sperm physiology using the murine model. FGF2 was detected in uterus and oviduct protein extracts, and it was immunolocalized in epithelial cells of the uterus, isthmus and ampulla, as well as in the cumulus oophorus-oocyte complex. The receptors FGFR1, FGFR2, FGFR3 and FGFR4 were immunodetected in the flagellum and acrosomal region of sperm recovered from the cauda epididymis. Analysis of testis sections showed the expression of FGFRs in germ cells at different stages of the spermatogenesis, suggesting the testicular origin of the sperm FGFRs. Sperm incubation with recombinant FGF2 (rFGF2) led to increased sperm motility and velocity and to enhanced intracellular Ca2+ levels and acrosomal loss compared to the control. In conclusion, this study shows that FGF2 is expressed in tissues of the female reproductive tract. Also, the fact that functional FGFRs are present in mouse sperm and that rFGF2 affects sperm motility and acrosomal exocytosis, suggests the involvement of this system in the in vivo regulation of sperm function.

Identification of a Novel Human E-Cadherin Splice Variant and Assessment of Its Effects Upon EMT-Related Events.

Epithelial Cadherin (E-cadherin) is involved in calcium-dependent cell-cell adhesion and signal transduction. The E-cadherin decrease/loss is a hallmark of Epithelial to Mesenchymal Transition (EMT), a key event in tumor progression. The underlying molecular mechanisms that trigger E-cadherin loss and consequent EMT have not been completely elucidated. This study reports the identification of a novel human E-cadherin variant mRNA produced by alternative splicing. A bioinformatics evaluation of the novel mRNA sequence and biochemical verifications suggest its regulation by Nonsense-Mediated mRNA Decay (NMD). The novel E-cadherin variant was detected in 29/42 (69%) human tumor cell lines, expressed at variable levels (E-cadherin variant expression relative to the wild type mRNA = 0.05-11.6%). Stable transfection of the novel E-cadherin variant in MCF-7 cells (MCF7Ecadvar) resulted in downregulation of wild type E-cadherin expression (transcript/protein) and EMT-related changes, among them acquisition of a fibroblastic-like cell phenotype, increased expression of Twist, Snail, Zeb1, and Slug transcriptional repressors and decreased expression of ESRP1 and ESRP2 RNA binding proteins. Moreover, loss of cytokeratins and gain of vimentin, N-cadherin and Dysadherin/FXYD5 proteins was observed. Dramatic changes in cell behavior were found in MCF7Ecadvar, as judged by the decreased cell-cell adhesion (Hanging-drop assay), increased cell motility (Wound Healing) and increased cell migration (Transwell) and invasion (Transwell w/Matrigel). Some changes were found in MCF-7 cells incubated with culture medium supplemented with conditioned medium from HEK-293 cells transfected with the E-cadherin variant mRNA. Further characterization of the novel E-cadherin variant will help understanding the molecular basis of tumor progression and improve cancer diagnosis. J. Cell. Physiol. 232: 1368-1386, 2017. © 2016 Wiley Periodicals, Inc.

Epithelial and neural cadherin expression in the mammalian reproductive tract and gametes and their participation in fertilization-related events.

Mammalian fertilization involves a series of well-orchestrated cell-cell interaction steps between gametes, as well as among spermatozoa and somatic cells of both the male and female reproductive tracts. Cadherins are Ca(2+)-dependent glycoproteins that have been involved in cellular adhesion and signaling in somatic cells. Taking into account that Ca(2+) ions are required during fertilization, the involvement of these proteins in adhesion events during this process can be anticipated. This report presents an overview on two members of classical cadherins, Epithelial (E-) and Neural (N-) cadherin in reproductive biology. Its provides evidence of studies done by several research groups about the expression of E- and N-cadherin during spermatogenesis, oogenesis and folliculogenesis, and their involvement in gamete transport in the reproductive tracts. Moreover, it describes current knowledge of E- and N-cadherin presence in cells of the cumulus-oocyte complex and spermatozoa from several mammalian species, and shows gathered evidence on their participation in different steps of the fertilization process. A brief summary on general information of both proteins is also presented.

Secretory Leukocyte Protease Inhibitor (SLPI) expression downregulates E-cadherin, induces ß-catenin re-localisation and triggers apoptosis-related events in breast cancer cells.

BACKGROUND INFORMATION: Epithelial cadherin (E-cadherin) is involved in cell-cell adhesion through its extracellular domain, whereas the intracellular domain interacts with adaptor proteins, i.e. ß-catenin, links E-cadherin to the actin cytoskeleton and participates in signal transduction events. E-cadherin protects mammary epithelial cells from apoptosis and its loss during tumour progression has been documented. Secretory Leukocyte Protease Inhibitor (SLPI) has anti- and pro-tumourigenic activities but its role in breast cancer has not been fully elucidated. Notwithstanding its relevance, how SLPI affects E-cadherin in breast cancer is still unknown. This study evaluated the effect of SLPI upon E-cadherin/ß-catenin expression and apoptosis-related markers in murine (F3II) and human (MCF-7) breast tumour cells either treated with exogenous recombinant human SLPI (rhSLPI) or stably transfected with a plasmid encoding its sequence. RESULTS: Addition of rhSLPI to F3II cells caused a decrease (P < 0.05) in E-cadherin transcript and protein levels. Similar results were observed in SLPI-stable F3II transfectants (2C1), and treatment of 2C1 cells with a siRNA toward SLPI restored E-cadherin to control levels. SLPI-expressing cells showed disruption of E-cadherin/ß-catenin complex and increased (P < 0.05) percentage of cells depicting nuclear ß-catenin localisation. Associated to these changes, 2C1 cells showed increased Bax/Bcl-2 ratio and p21 protein levels, decreased c-Myc protein levels and decreased Cyclin D1 and Claudin-1 transcript levels. No differences in N- and P-cadherin were observed between SLPI-transfected cells and controls. Addition of rhSLPI to MCF-7 cells or stable transfection with SLPI caused a decrease (P < 0.05) in E-cadherin expression (transcript/protein) and its redistribution to the cytoplasm, as well as ß-catenin re-localisation to the cell nucleus. CONCLUSIONS: Expression of SLPI was associated to a decrease in E-cadherin expression and re-localisation of E-cadherin to the cell cytoplasm and ß-catenin to the cell cytoplasm and nucleus, and had pro-apoptotic and cell cycle-arrest effects.

Proteomics, oxidative stress and male infertility.

Oxidative stress has been established as one of the main causes of male infertility and has been implicated in many diseases associated with infertile men. It results from high concentrations of free radicals and suppressed antioxidant potential, which may alter protein expression in seminal plasma and/or spermatozoa. In recent years, proteomic analyses have been performed to characterize the protein profiles of seminal ejaculate from men with different clinical conditions, such as high oxidative stress. The aim of the present review is to summarize current findings on proteomic studies performed in men with high oxidative stress compared with those with physiological concentrations of free radicals, to better understand the aetiology of oxidative stress-induced male infertility. Each of these studies has suggested candidate biomarkers of oxidative stress, among them are DJ-1, PIP, lactotransferrin and peroxiredoxin. Changes in protein concentrations in seminal plasma samples with oxidative stress conditions were related to stress responses and to regulatory pathways, while alterations in sperm proteins were mostly associated to metabolic responses (carbohydrate metabolism) and stress responses. Future studies should include assessment of post-translational modifications in the spermatozoa as well as in seminal plasma proteomes of men diagnosed with idiopathic infertility. Oxidative stress, which occurs due to a state of imbalance between free radicals and antioxidants, has been implicated in most cases of male infertility. Cells that are in a state of oxidative stress are more likely to have altered protein expression. The aim of this review is to better understand the causes of oxidative stress-induced male infertility. To achieve this, we assessed proteomic studies performed on the seminal plasma and spermatozoa of men with high levels of oxidative stress due to various clinical conditions and compared them with men who had physiological concentrations of free radicals. A variety of sperm and seminal plasma proteins were found to be expressed either in abundance (over-expressed) or in a lesser amount (underexpressed), while other proteins were found to be unique either to men with oxidative stress or to men with a balanced ratio of antioxidants/free radicals. Each study included in this review suggested several proteins that could possibly act as biomarkers of oxidative stress-induced male infertility, such as protein DJ-1, PIP, lactotransferrin and peroxiredoxin. Pathway analysis performed in these studies revealed that the changes in seminal plasma proteins in men with oxidative stress could be attributed to stress responses and regulatory pathways, while changes in sperm proteins were linked to stress responses and metabolic responses. Subsequent studies could look into post-translational modifications in the protein profile of men with idiopathic infertility. We hope that the information in this review will contribute to a better understanding of the main causes of idiopathic male infertility.

Impact of heat waves on semen quality: A retrospective study in Argentina between 2005 and 2023.

Heat waves, defined as periods with daily temperatures surpassing the historical average for a specific region, have become more frequent worldwide in recent years. Previous studies have reported a negative association between temperature and semen quality, but the focus has mainly been on Asian and European populations. The study included 54,926 men (18-60 years) undergoing routine semen analysis between 2005 and 2023 at CEUSA-LAEH andrology unit, in Buenos Aires, Argentina. Hourly temperature readings were provided by the Servicio Meteorológico Nacional. R programming (R Studio v2022.07.2) was used to define heat waves, calculate key characteristics, visualize results, and perform statistical tests at the IBYME laboratory. During the period studied, a total of 124 days had heat waves (defined after at least 3 consecutive days with 32.3 °C and 22 °C). Men exposed to heat waves during spermatogenesis exhibited lower sperm number (concentration and count; P < 0.0001) and decreased normal morphology (percentage of normal sperm and normal motile count; P < 0.05) compared to those not exposed. These differences were most pronounced between semen samples from years with several heat waves (2013, 2023) and none (2005, 2007, 2016), displaying 4-5 times higher fold changes (P < 0.05). Further analysis employing multiple regression revealed a significantly negative association between semen quality and heat wave length, suggesting that a prolonged exposure may be more detrimental than an acute exposure. Subsequent analysis focusing on prolonged exposure (≥6-days heat wave) during spermatogenesis revealed a negative (P < 0.05) association between early exposure (spermatocytogenesis: 64-90 days prior semen collection) and semen quality. This study underscores the negative association between early exposure to heat waves during sperm development and semen quality, raising concerns about its possible association with the worldwide declining male fertility. A comprehensive collaborative approach is crucial, involving global governmental policies, sustainable practices, and coordinated efforts across scientific, healthcare, and policy domains.

A 2-step remote TUNEL approach for sperm DNA fragmentation assessment. Analysis in donors and patients.

OBJECTIVES: Infertility is a disease of the male or female reproductive systems. Male reproductive workup is based on routine semen analysis, although of limited value. The 2021 WHO Manual incorporated Sperm DNA Fragmentation (SDF) assessment, and highlighted the need for individual laboratories to define suitable thresholds. This study aimed to present an alternative to address this issue, determine an SDF cut-off value with fertile donors, and characterize SDF in a patient cohort and their relationship with semen parameters. STUDY DESIGN: A service unit was established to remotely perform TUNEL assay in a 2 step-process. Semen samples were received at andrology laboratories, subjected to routine semen analysis (WHO, 2010), partially processed and transported to the service unit for SDF evaluation. Using this setting, studies were done in fertile donors (n = 15) to define the cut-off value, and in men undergoing infertility workup (n = 318). RESULTS: A cut-off value of 9.17 % was determined with the fertile donor cohort. With this cut-off, a 64.46 % abnormal SDF incidence was determined in the patient cohort. SDF negatively correlated with sperm number, vitality and motility, and positively with abnormal morphology and male age (P < 0.05). TUNEL-positive cases depicted lower sperm quality and higher male age (P < 0.05). A similar abnormal SDF incidence was determined among patients with semen abnormalities. Asthenozoospermic and ≥40 years patient samples depicted higher (P < 0.05) SDF than those of the general population. SDF incidence was also high in normozoospermic patients. CONCLUSIONS: Using a 2-step remote approach with a standardized procedure and an SDF cut-off value established with fertile donors, high SDF incidence in semen samples depicting normal and abnormal quality were identified in men consulting for infertility, highlighting the relevance of its evaluation as part of the male fertility workup.

Frequency, morbidity and equity - the case for increased research on male fertility.

Currently, most men with infertility cannot be given an aetiology, which reflects a lack of knowledge around gamete production and how it is affected by genetics and the environment. A failure to recognize the burden of male infertility and its potential as a biomarker for systemic illness exists. The absence of such knowledge results in patients generally being treated as a uniform group, for whom the strategy is to bypass the causality using medically assisted reproduction (MAR) techniques. In doing so, opportunities to prevent co-morbidity are missed and the burden of MAR is shifted to the woman. To advance understanding of men's reproductive health, longitudinal and multi-national centres for data and sample collection are essential. Such programmes must enable an integrated view of the consequences of genetics, epigenetics and environmental factors on fertility and offspring health. Definition and possible amelioration of the consequences of MAR for conceived children are needed. Inherent in this statement is the necessity to promote fertility restoration and/or use the least invasive MAR strategy available. To achieve this aim, protocols must be rigorously tested and the move towards personalized medicine encouraged. Equally, education of the public, governments and clinicians on the frequency and consequences of infertility is needed. Health options, including male contraceptives, must be expanded, and the opportunities encompassed in such investment understood. The pressing questions related to male reproductive health, spanning the spectrum of andrology are identified in the Expert Recommendation.

Current global status of male reproductive health.

BACKGROUND: The widespread interest in male reproductive health (MRH), fueled by emerging evidence, such as the global decline in sperm counts, has intensified concerns about the status of MRH. Consequently, there is a pressing requirement for a strategic, systematic approach to identifying critical questions, collecting pertinent information, and utilizing these data to develop evidence-based strategies. The methods for addressing these questions and the pathways toward their answers will inevitably vary based on the variations in cultural, geopolitical, and health-related contexts. To address these issues, a conjoint ESHRE and Male Reproductive Health Initiative (MRHI) Campus workshop was convened. OBJECTIVE AND RATIONALE: The three objectives were: first, to assess the current state of MRH around the world; second, to identify some of the key gaps in knowledge; and, third, to examine how MRH stakeholders can collaboratively generate intelligent and effective paths forward. SEARCH METHODS: Each expert reviewed and summarized the current literature that was subsequently used to provide a comprehensive overview of challenges related to MRH. OUTCOMES: This narrative report is an overview of the data, opinions, and arguments presented during the workshop. A number of outcomes are presented and can be summarized by the following overarching themes: MRH is a serious global issue and there is a plethora of gaps in our understanding; there is a need for widespread international collaborative networks to undertake multidisciplinary research into fundamental issues, such as lifestyle/environmental exposure studies, and high-quality clinical trials; and there is an urgent requirement for effective strategies to educate young people and the general public to safeguard and improve MRH across diverse population demographics and resources. LIMITATIONS REASONS FOR CAUTION: This was a workshop where worldwide leading experts from a wide range of disciplines presented and discussed the evidence regarding challenges related to MRH. While each expert summarized the current literature and placed it in context, the data in a number of areas are limited and/or sparse. Equally, important areas for consideration may have been missed. Moreover, there are clear gaps in our knowledge base, which makes some conclusions necessarily speculative and warranting of further study. WIDER IMPLICATIONS: Poor MRH is a global issue that suffers from low awareness among the public, patients, and heathcare professionals. Addressing this will require a coordinated multidisciplinary approach. Addressing the significant number of knowledge gaps will require policy makers prioritizing MRH and its funding. STUDY FUNDING/COMPETING INTERESTS: The authors would like to extend their gratitude to ESHRE for providing financial support for the Budapest Campus Workshop, as well as to Microptic S.L. (Barcelona) for kindly sponsoring the workshop. P.B. is the Director of the not-for-profit organization Global Action on Men's Health and receives fees and expenses for his work, (which includes the preparation of this manuscript). Conflicts of interest: C.J.D.J., C.L.R.B., R.A.A., P.B., M.P.C., M.L.E., N.G., N.J., C.K., AAP, M.K.O., S.R.-H., M.H.V.-L.: ESHRE Campus Workshop 2022 (Travel support-personal). C.J.D.J.: Cambridge University Press (book royalties-personal). ESHRE Annual Meeting 2022 and Yale University Panel Meeting 2023 (Travel support-personal). C.L.R.B.: Ferring and IBSA (Lecture), RBMO editor (Honorarium to support travel, etc.), ExSeed and ExScentia (University of Dundee), Bill & Melinda Gates Foundation (for research on contraception). M.P.C.: Previously received funding from pharmaceutical companies for health economic research. The funding was not in relation to this work and had no bearing on the contents of this work. No funding from other sources has been provided in relation to this work (funding was provided to his company Global Market Access Solutions). M.L.E.: Advisor to Ro, Doveras, Next, Hannah, Sandstone. C.K.: European Academy of Andrology (Past president UNPAID), S.K.: CEO of His Turn, a male fertility Diagnostic and Therapeutic company (No payments or profits to date). R.I.M.: www.healthymale.org.au (Australian Government funded not for profit in men's health sector (Employed as Medical Director 0.2 FET), Monash IVF Pty Ltd (Equity holder)). N.J.: Merck (consulting fees), Gedeon Richter (honoraria). S.R.-H.: ESHRE (Travel reimbursements). C.N.: LLC (Nursing educator); COMMIT (Core Outcomes Measures for Infertility Trials) Advisor, meeting attendee, and co-author; COMMA (Core Outcomes in Menopause) Meeting attendee, and co-author; International Federation of Gynecology and Obstetrics (FIGO) Delegate Letters and Sciences; ReproNovo, Advisory board; American Board of Urology Examiner; American Urological Association Journal subsection editor, committee member, guidelines co-author Ferring Scientific trial NexHand Chief Technology Officer, stock ownership Posterity Health Board member, stock ownership. A.P.: Economic and Social Research Council (A collaborator on research grant number ES/W001381/1). Member of an advisory committee for Merck Serono (November 2022), Member of an advisory board for Exceed Health, Speaker fees for educational events organized by Mealis Group; Chairman of the Cryos External Scientific Advisory Committee: All fees associated with this are paid to his former employer The University of Sheffield. Trustee of the Progress Educational Trust (Unpaid). M.K.O.: National Health and Medical Research Council and Australian Research Council (Funding for research of the topic of male fertility), Bill and Melinda Gates Foundation (Funding aimed at the development of male gamete-based contraception), Medical Research Future Fund (Funding aimed at defining the long-term consequences of male infertility). M.H.V.-L.: Department of Sexual and Reproductive Health and Research (SRH)/Human Reproduction Programme (HRP) Research Project Panel RP2/WHO Review Member; MRHI (Core Group Member), COMMIT (member), EGOI (Member); Human Reproduction (Associate Editor), Fertility and Sterility (Editor), AndroLATAM (Founder and Coordinator).

Male Attitudes towards Infertility: Results from a Global Questionnaire.

PURPOSE: In general, men are less likely to seek health care than women. Infertility is a global disease that afflicts approximately 15% of reproductive age couples and the male contributes to 40% of the diagnosable cause. Remarkably, no large or multi-national population data exist regarding men's perceptions about their infertility. The purpose of this study was to advance our knowledge about the infertile male's social experience regarding: (1) how they feel about their infertility, (2) what motivated them to seek health care, (3) how likely are they to talk with others about their infertility, (4) their awareness of male infertility support groups, and (5) what their primary source for information is regarding male infertility? Based on the results from this study, these simple questions now have clearer definition. MATERIALS AND METHODS: An Institutional Review Board-approved, male-directed, anonymous questionnaire translated into 20 languages was made globally available through the Fertility Europe website (https://fertilityeurope.eu). Males (n=1,171) age 20-49 years were invited to complete the online survey after informed consent. RESULTS: Most respondents were European (86%). Of European men, <15.8% were self-motivated to seek medical help. Further, their physician was not the primary source of information regarding their infertility. While most men (59%) viewed their infertility positively, a large majority were not very likely (73%) to talk about it. Most respondents indicated a lack of awareness or absence of male infertility support groups. CONCLUSIONS: These are the first multi-national population data revealing men's feelings about their infertility, what motivates them to seek help and their awareness of resources for peer support and information. These findings also serve to highlight significant gaps that exist in the provision of male reproductive health care and in supportive resources for men suffering from infertility. We offer recommendations on how to address the problem(s).

Corrigendum: FXYD5/Dysadherin, a biomarker of endometrial cancer myometrial invasion and aggressiveness: its relationship with TGF-ß1 and NF-κB pathways.

[This corrects the article DOI: 10.3389/fonc.2019.01306.].

ETV5 transcription factor is overexpressed in ovarian cancer and regulates cell adhesion in ovarian cancer cells.

Epithelial ovarian cancer is the most lethal gynecological malignancy and the fifth leading cause of cancer deaths in women in the Western world. ETS transcription factors are known to act as positive or negative regulators of the expression of genes that are involved in various biological processes, including those that control cellular proliferation, differentiation, apoptosis, tissue remodeling, angiogenesis and transformation. ETV5 belongs to the PEA3 subfamily. PEA3 subfamily members are able to activate the transcription of proteases, matrix metalloproteinases and tissue inhibitor of metalloproteases, which is central to both tumor invasion and angiogenesis. Here, we examined the role of the ETV5 transcription factor in epithelial ovarian cancer and we found ETV5 was upregulated in ovarian tumor samples compared to ovarian tissue controls. The in vitro inhibition of ETV5 decreased cell proliferation in serum-deprived conditions, induced EMT and cell migration and decreased cell adhesion to extracellular matrix components. ETV5 inhibition also decreased cell-cell adhesion and induced apoptosis in anchorage-independent conditions. Accordingly, upregulation of ETV5 induced the expression of cell adhesion molecules and enhanced cell survival in a spheroid model. Our findings suggest that the overexpression of ETV5 detected in ovarian cancer cells may contribute to ovarian tumor progression through the ability of ETV5 to enhance proliferation of ovarian cancer cells. In addition, upregulation of ETV5 would play a role in ovarian cancer cell dissemination and metastasis into the peritoneal cavity by protecting ovarian cancer cells from apoptosis and by increasing the adhesion of ovarian cancer cells to the peritoneal wall through the regulation of cell adhesion molecules.

D-chiro-inositol, an aromatase down-modulator, increases androgens and reduces estrogens in male volunteers: a pilot study.

BACKGROUND: Androgen deficiency affects men in the adulthood, causing several harmful effects at the reproductive and behavioural levels. Since aromatase is an enzyme that catalyses the conversion of androgens to estrogens, and it is responsible for an adequate balance of both sex hormones in males and females, the administration of molecules acting as down modulators may contribute to restore an abnormal enzymatic activity. A prospective pilot study was carried out to investigate the effect of D-chiro-inositol, a putative aromatase down-modulator, on serum levels of testosterone, estradiol, estrone, dehydroepiandrosterone and epiandrosterone from a group of adult male volunteers. Glucose, insulin, follicle-stimulating hormone, luteinizing hormone, inhibin B, D-chiro-inositol and myo-inositol serum levels were also measured. RESULTS: Male volunteers were selected according to age and body mass index. Subjects with altered glycemia and/or hormonal status, due to advanced age or abnormal weight, were enrolled in the study. Each of the 10 volunteers enrolled took oral D-chiro-inositol (1 g/day) for 1 month. Serum assays of selected markers were performed at baseline (control) and after treatment. D-chiro-inositol administration was associated to reduced serum levels of estrone (- 85.0%) and estradiol (- 14.4%), and increased serum levels of testosterone (+ 23.4%) and dehydroepiandrosterone (+ 13.8%). In addition, epiandrosterone levels were higher (+39%) after treatment. On the other hand, follicle-stimulating hormone, luteinizing hormone and inhibin B did not change. A trend toward a decrease of glycemia, insulinemia and Homeostatic Model Assessment index was observed after D-chiro-inositol treatment, although differences did not reach statistical significance. D-chiro-inositol treatment did not cause any noticeable adverse effect. CONCLUSIONS: Increased androgens and decreased estrogens seem to confirm that D-chiro-inositol acts as an aromatase down-modulator, but with a still unknown mechanism of action. This pilot study opens up new perspectives of research and therapeutic applications for D-chiro-inositol at different dosages and length of treatment. Authorization number 005/2020 released by the Local Ethics Committee of Alma Res Fertility Center, Rome. TRIAL REGISTRATION NUMBER: NCT04615767 (registry: ClinicalTrials.gov) Date of registration: November 3, 2020.

RéSUMé: CONTEXTE: L'insuffisance en androgènes affecte les hommes à l'âge adulte, causant plusieurs effets nocifs aux niveaux reproductif et comportemental. Puisque l'aromatase est. une enzyme qui catalyse la conversion des androgènes en œstrogènes, et qu'elle est. responsable d'un équilibre adéquat des hormones sexuelles chez les hommes et les femmes, l'administration de molécules agissant comme freinateurs peut contribuer à restaurer une activité enzymatique anormale. Une étude prospective pilote a été menée pour étudier l'effet du D-chiro-inositol, un potentiel freinateur de l'aromatase, sur les taux sériques de testostérone, estradiol, estrone, déhydroépiandrostérone et d'épiandrostérone d'un groupe d'hommes adultes volontaires. Le glucose, l'insuline, l'hormone folliculostimulante, l'hormone lutéinisante, l'inhibine B, le D-chiro-inositol et les taux sériques de myo-inositol ont également été mesurés. RéSULTATS: Les hommes volontaires ont été sélectionnés selon l'âge et l'indice de masse corporelle. Les hommes qui présentaient une glycémie et/ou un statut hormonal altérés en raison d'un âge avancé ou d'un poids anormal, ont été inclus dans l'étude. Chacun des 10 volontaires enrôlés a pris du D-chiro-inositol (1 g/jour) par voie orale pendant un mois. Des dosages sériques des marqueurs sélectionnés ont été réalisés avant (témoin) et après le traitement. L'administration de D-chiro-inositol a été associée à une réduction des taux sériques de l'estrone (− 85.0%) et de l'estradiol (− 14,4%), et a une augmentation des taux sériques de testostérone (+ 23,4%) et de déhydroépiandrostérone (+ 13,8%). En outre, les taux d'épiandrostérone étaient plus élevés (39%) après le traitement. D'autre part, les taux d'hormone folliculostimulante, d'hormone lutéinisante et d'inhibine B n'ont pas été modifiés. Une tendance à la diminution de la glycémie, de l'insulinémie et de l'indice d'évaluation du modèle homéostatique a été observée après traitement par D-chiro-inositol, bien que les différences n'aient pas atteint une signification statistique. Le traitement par D-chiro-inositol n'a causé aucun effet indésirable notable. CONCLUSIONS: L'augmentation des androgènes et la diminution des œstrogènes semblent confirmer que le D-chiro-inositol agit comme un freinateur de l'aromatase, mais avec un mécanisme d'action encore inconnu. Cette étude pilote ouvre de nouvelles perspectives de recherche et d'applications thérapeutiques pour le D-chiro-inositol à différents dosages et durées de traitement.

Epithelial and Neural Cadherin in Mammalian Fertilization: Studies in the Mouse Model.

Successful mammalian fertilization requires a well-orchestrated sequence of molecular events leading to gamete fusion. Since this interaction involves Ca2+-dependent adhesion events, the participation of the Ca+2-dependent cell-cell adhesion proteins Epithelial (E-cad) and Neural (N-cad) cadherin is envisaged. We have previously reported the expression of E-cad and N-cad in human gametes and showed evidence of their involvement in sperm-oocyte adhesion events leading to fertilization. To overcome ethical limitations associated with the use of human gametes in fertilization-related studies, the mouse has been selected worldwide as the experimental model for over 4 decades. Herein, we report a detailed study aimed at characterizing the expression of E-cad and N-cad in murine gametes and their involvement in murine fertilization using specific antibodies and blocking peptides towards both adhesion proteins. E-cad and N-cad protein forms, as well as other members of the adhesion complex, specifically ß-catenin and actin, were identified in spermatozoa, cumulus cells and oocytes protein extracts by means of Western immunoblotting. In addition, subcellular localization of these proteins was determined in whole cells using optical fluorescent microscopy. Gamete pre-incubation with anti-E-cad (ECCD-1) or N-cad (H-63) antibodies resulted in decreased (p < 0.05) In Vitro Fertilization (IVF) rates, when using both cumulus-oocytes complexes and cumulus-free oocytes. Moreover, IVF assays done with denuded oocytes and either antibodies or blocking peptides against E-cad and N-cad led to lower (p < 0.05) fertilization rates. When assessing each step, penetration of the cumulus mass was lower (p < 0.05) when spermatozoa were pre-incubated with ECCD-1 or blocking peptides towards E-cad or towards both E- and N-cad. Moreover, sperm-oolemma binding was impaired (p < 0.0005) after sperm pre-incubation with E-cad antibody or blocking peptide towards E-cad, N-cad or both proteins. Finally, sperm-oocyte fusion was lower (p < 0.05) after sperm pre-incubation with either antibody or blocking peptide against E-cad or N-cad. Our studies demonstrate the expression of members of the adherent complex in the murine model, and the use of antibodies and specific peptides revealed E-cad and N-cad participation in mammalian fertilization.