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Conventional IVF (c-IVF) is one of the most practiced assisted reproductive technology (ART) approaches used worldwide. However, in the last years, the number of c-IVF procedures has dropped dramatically in favor of intracytoplasmic sperm injection (ICSI) in cases of non-male-related infertility. In this review, we have outlined advantages and disadvantages associated with c-IVF, highlighting the essential steps governing its success, its limitations, the methodology differences among laboratories and the technical progress. In addition, we have debated recent insights into fundamental questions, including indications regarding maternal age, decreased ovarian reserve, endometriosis, autoimmunity, single oocyte retrieval-cases as well as preimplantation genetic testing cycles. The “overuse” of ICSI procedures in several clinical situations of ART has been critically discussed. These insights will provide a framework for a better understanding of opportunities associated with human c-IVF and for best practice guidelines applicability in the reproductive medicine field.

STUDY QUESTION Are uterine fluid-derived extracellular vesicles (UF-EVs) a ‘liquid biopsy’ reservoir of biomarkers for real-time monitoring of endometrial status? SUMMARY ANSWER The transcriptomic cargo of UF-EVs reflects the RNA profile of the endometrial tissue as well as changes between the non-receptive and the receptive phase, possibly supporting its use for a novel endometrial receptivity test. WHAT IS KNOWN ALREADY EVs have been previously isolated from uterine fluid, where they likely contribute to the embryo-endometrium crosstalk during implantation. Based on a meta-analysis of studies on endometrial tissue implantation-associated genes and the human exosomes database, 28 of the 57 transcripts considered as receptivity markers refer to proteins present in human exosomes. However, the specific transcriptomic content of receptive phase UF-EVs has yet to be defined. STUDY DESIGN, SIZE, DURATION Two experimental series were set up. First, we simultaneously sequenced RNA species derived from paired UF-EVs and endometrial tissue samples collected from physiologically cycling women. Second, we analyzed RNA species of UF-EVs collected during the non-receptive (LH + 2) and receptive (LH + 7) phase of proven fertile women and from the receptive (LH + 7) phase of a population of women undergoing ART and transfer of euploid blastocysts. PARTICIPANTS/MATERIALS, SETTING, METHODS For paired UF—endometrial tissue sampling, endometrial tissue biopsies were obtained with the use of a Pipelle immediately after UF collection performed by lavage of the endometrial cavity. Overall, n = 87 UF samples were collected and fresh-processed for EV isolation and total RNA extraction, while western blotting was used to confirm the expression of EV protein markers of the isolated vesicles. Physical characterization of UF-EVs was performed by Nanoparticle Tracking Analysis. To define the transcriptomic cargo of UF-EV samples, RNA-seq libraries were successfully prepared from n = 83 UF-EVs samples and analyzed by RNA-seq analysis. Differential gene expression (DGE) analysis was used to compare RNA-seq results between different groups of samples. Functional enrichment analysis was performed by gene set enrichment analysis with g:Profiler. Pre-ranked gene set enrichment analysis (GSEA) with WebGestalt was used to compare RNA-seq results with the gene-set evaluated in a commercially available endometrial receptivity array. MAIN RESULTS AND THE ROLE OF CHANCE A highly significant correlation was found between transcriptional profiles of endometrial biopsies and pairwise UF-EV samples (Pearson’s r = 0.70 P LARGE SCALE DATA Transcriptomic data were deposited in NCBI Gene Expression Omnibus (GEO) and can be retrieved using GEO series accession number: GSE158958. LIMITATIONS, REASONS FOR CAUTION Separation of RNA species associated with EV membranes might have been incomplete, and membrane-bound RNA species—rather than the internal RNA content of EVs—might have contributed to our RNA-seq results. Also, we cannot definitely distinguish the relative contribution of exosomes, microvesicles and apoptotic bodies to our findings. When considering patients undergoing ART, we did not collect UFs in the same cycle of the euploid embryo transfer but in the one immediately preceding. We considered this approach as the most appropriate in relation to the novel, explorative nature of our study. Based on our results, a validation of UF-EV RNA-seq analyses in the same cycle in which embryo transfer is performed could be hypothesized. WIDER IMPLICATIONS OF THE FINDINGS On the largest sample size of human EVs ever analyzed with RNA-seq, this study establishes a gene signature to use for less-invasive endometrial receptivity tests. This report is indeed the first to show that the transcriptome of UF-EVs correlates with the endometrial tissue transcriptome, that RNA signatures in UF-EVs change with endometrial status, and that UF-EVs could serve as a reservoir for potential less-invasive collection of receptivity markers. This article thus represents a step forward in the design of less-invasive approaches for real-time monitoring of endometrial status, necessary for advancing the field of reproductive medicine. STUDY FUNDING/COMPETING INTEREST(S) The study was funded by a competitive grant from European Society of Human Reproduction and Embryology (ESHRE Research Grant 2016-1). The authors have no financial or non-financial competing interests to disclose. TRIAL REGISTRATION NUMBER NA.

This meta-analysis aimed to offer a general picture of the available data on the effects of early-life factors on the risk of developing endometriosis in adult life. An advanced, systematic search of the online medical databases PubMed, EMBASE and CINAHL was limited to full-length manuscripts published in English in peer-reviewed journals up to February 2019. Log of relative risk (RR) was employed to calculate the pooled effect sizes using both fixed and random effects modelling and I-squared tests to assess heterogeneity. Funnel plots were used to investigation publication bias. The meta-analysis was registered in PROSPERO (ID CRD42019138668). Six studies that included a total of 2360 women affected by endometriosis were analysed. The pooled results showed that the risk of developing endometriosis in adult life was significantly increased by being born prematurely (logRR 0.21, 95% CI -0.03 to 0.40), having a low birthweight (logRR 0.35, 95% CI -0.15 to 0.54), being formula-fed (logRR 0.65, 95% CI -0.35 to 0.95) and having been exposed to diethylstilbestrol (DES) in utero (logRR 0.65, 95% CI 0.26 to 1.04. Among intrauterine and early neonatal exposures, prematurity, birthweight, formula feeding and DES were risk factors for the development of endometriosis in adult life.

Extracellular vesicles (EVs) derived from different parts of the male reproductive tract can be internalized by human spermatozoa affecting their maturation and regulating their functions. Here we demonstrate that EVs derived from the female tract can be uptaken by sperm and affect their competence. Primary endometrial cells release EVs with a diameter between 50 and 350 nm and bear the standard vesicle and exosome marker proteins CD63, CD9, TSG101 and ALIX. The uptake of dye-labelled endometrial cell-derived EVs by spermatozoa, quantified as fluorescence intensity, was significantly higher when EVs were derived from cells in the proliferative phase. Vital, motile fluorescent sperm could be appreciated after a 48-hour co-incubation with endometrial cells previously labelled with the Vybrant™ DiO dye. EV internalization by sperm was blocked at 4 °C and by incubation with filipin, suggesting an energy-dependent process probably attributable to the lipid-raft domain mediated-endocytosis. Sperm ability to undergo capacitation and acrosome reaction was stimulated by endometrial cell-derived EVs as manifested by the increased protein tyrosine phosphorylation and evident reactivity when stimulated with a calcium ionophore. Based on these findings, EVs exchange may be suggested as an emerging way through which female reproductive tract cells can interact with the passing spermatozoa.

Study question Could EVs secreted by aneuploid embryos a) serve for development of RNA biomarkers for PGT-A and b) elicit a relevant transcriptomic response in decidualised ESCs? Summary answer Aneuploid embryo EVs a) contain PPM1J, LINC00561, ANKRD34C and TMED10 in differential abundance from euploid EVs and b) induce up-regulation of MUC1 in decidualised ESCs. What is known already Embryo aneuploidy accounts for approximately 50% of all recurrent implantation failures in women >35 years old. PGT-A identifies euploid embryos to increase implantation probability but the technology is controversial as it requires an invasive embryo biopsy with an elusive long-term biosafety. The development of non-invasive methods to screen out aneuploid embryos is paramount. It is also critical to decode the embryo-endometrial dialog underlying implantation failure. We have previously reported that IVF embryos secrete EVs that can be internalised by ESCs, conceptualising that successful implantation to the endometrium is facilitated by EVs, which may additionally serve as biomarkers of ploidy status. Study design, size, duration Embryos destined for biopsy on days 5-7 for PGT-A were grown under standard conditions. Spent media (30μl) were collected from euploid (n = 175) and aneuploid embryos (n = 145) at both cleavage (days 1-3) and blastocyst (days 3-5) stage. Media samples from n = 35 cleavage embryos were pooled in order to obtain five euploid and four aneuploidy pools. Blastocyst media were pooled to create one euploid and one aneuploid pool. ESCs were obtained from five women undergoing diagnostic laparoscopy. Participants/materials, setting, methods The study was realised at a research hospital. EVs were isolated from euploid and aneuploid Day3 pools with differential ultracentrifugation and EV-RNA sequencing was performed following the SMARTer Stranded Total RNA-Seq approach. ESCs were decidualised (E2:10nM, P4:1uM, cAMP:0.5 mM twice every 48 hours) and treated for 24 hours with 50 ng/ml euploid or aneuploid EVs extracted from blastocyst media. RNA sequencing was performed on ESCs following the Truseq RNAseq protocol. Main results and the role of chance Aneuploid cleavage stage embryos (n = 4) secreted EVs that were less abundant in RNA fragments originating from the genes PPM1J (log2fc=-5.13, p = 0.011), LINC00561 (log2fc=-7.87, p = 0.010) and ANKRD34C (log2fc=-7.30, p = 0.017) and more abundant in TMED10 (log2fc=1.63 p = 0.025) compared to EVs (n = 5) from euploid embryos. Decidualisation per se induced downregulation of MUC1 (log2FC=-0.54, p = 0.0028) in ESCs as prerequisite for the establishment of receptive endometrium. The expression of MUC1 transcript in decidualised ESCs was significantly increased following treatment with aneuploid compared to euploid embryo-secreted EVs (log2FC=0.85, p = 0.0201). Limitations, reasons for caution The findings of the study may require validation utilising a second cohort of EVs samples. Wider implications of the findings This discovery that the RNA cargo of EVs secreted from aneuploid cleavage stage embryos is diverse from that of euploid embryos potentiates the development of non-invasive methodology for PGT-A. The upregulation of MUC1 in decidualised ESCs following aneuploid embryo EV treatment proposes a new mechanism underlying implantation failure. Trial registration number NA

Background Data about endometrioma ablation using energies with little thermal spread reported good results in terms of ovarian reserve and postoperative pregnancy rates. The aim of the present study was to assess the impact of 'one step' CO2 fiber laser vaporization for endometrioma on subsequent controlled ovarian stimulation. Methods This prospective observational cohort study included a consecutive series of infertile patients who have undergone CO2 fiber laser vaporization for endometrioma treatment. The primary endpoint was to assess the number of follicles per ovary growing during controlled ovarian stimulation. The secondary endpoints included the number of oocytes retrieved, the total number of embryos obtained and the cumulative clinical pregnancy rate per patient treated. Results Twenty-six patients underwent assisted reproductive technology after surgery for endometriosis-related infertility. In unilateral operated ovaries at the end of controlled ovarian stimulation no significant differences emerged from comparison of total recruited follicles in the operated ovary and in the contralateral ovary (p=0.55). If considering only bilateral endometriomas, the number of recruited follicles at the end of controlled ovarian stimulation was similar in both operated ovaries (p=0.79). The number of cumulative clinical pregnancies was 15 (57.7%; 95% CI: 38.5-76.9 %). When comparing women aged ≤ 35 years to those aged > 35 years, controlled ovarian stimulation outcomes were significantly higher in the younger patients. Age at the time of assisted reproductive technology was the only independent predictor for follicular growth during ovarian hyperstimulation (CI: -1.27 to -0.116, p=0.027). Conclusions CO2 laser-treated endometrioma is associated with favorable reproductive assisted reproductive technology outcomes.

Improving in vitro fertilization success is an unmet need. Observational studies have suggested that women with deficient or insufficient vitamin D have lower chances of in vitro fertilization success, but whether supplementation improves clinical pregnancy rate remains unclear.This study aimed to determine whether oral vitamin DThe "supplementation of vitamin D and reproductive outcome" trial is a 2-center randomized superiority double-blind placebo-controlled trial. Subjects were recruited between October 2016 and January 2019. Participants were women aged 18 to 39 years with low vitamin D (peripheral 25-hydroxyvitamin D of30 ng/mL), serum calcium of ≥10.6 mg/dL, body mass index of 18 to 25 kg/mOverall, 630 women were randomized 2 to 12 weeks before the initiation of the in vitro fertilization cycle to receive either a single dose of 600,000 IU of vitamin DIn women with normal weight with preserved ovarian reserve and low vitamin D levels undergoing in vitro fertilization cycles, a single oral dose of 600,000 IU of vitamin D

STUDY QUESTION Does oral Vitamin D supplementation alter the hormonal milieu of follicular fluid (FF) and the transcriptomic profile of luteinised granulosa cells (GCs) in women with Vitamin D deficiency undergoing IVF? SUMMARY ANSWER A transcriptomic signature relevant to oral Vitamin D supplementation in luteinised GCs was demonstrated, although Vitamin D supplementation did not alter hormone levels in FF. WHAT IS KNOWN ALREADY Vitamin D deficiency is linked to lower live birth rates among women undergoing IVF. It is unclear whether Vitamin D elicits a targeted action in reproductive physiology or is a surrogate marker of overall well-being. Several in-vitro studies, but none in vivo, have examined the impact of Vitamin D on the periovulatory follicle, focusing on GCs as a proxy marker of oocyte competence. STUDY DESIGN, SIZE, DURATION We present a report of secondary outcomes from the SUNDRO clinical trial, which was launched in 2016 to determine whether Vitamin D supplementation can improve the IVF outcomes of women who are deficient in Vitamin D ( PARTICIPANTS/MATERIALS, SETTING, METHODS Two academic infertility units were involved in the recruitment of the participants, who received a single dose of oral 25-hydroxyvitamin D (600 000 IU) or placebo, 2–12 weeks before oocyte retrieval. Women in both groups were deficient in Vitamin D, aged 18–39 years with a normal BMI (18–25 kg/m2) and MAIN RESULTS AND THE ROLE OF CHANCE At oocyte retrieval, FF concentration of 25-hydroxyvitamin D was 2.8-fold higher (P LIMITATIONS, REASONS FOR CAUTION Interpretation of the data is influenced by our intervention strategy (2–12 weeks prior to retrieval). As folliculogenesis may last 5–6 months, our protocol can only examine with confidence the impact of Vitamin D on the final stages of follicular growth. Furthermore, we examined the hormonal profile of the dominant follicle only, while the GC data reflect the transcriptome of all (pooled) follicles large enough to be used for IVF. Luteinised GCs from controlled ovarian stimulation were used in this study, which may be functionally distinct from the GCs of developing follicles. Moreover, the sample size for RNA-sequencing analysis was low (n = 3 per group), regardless of validation by RT-PCR that was performed on a larger cohort, introducing complexity to the IPA analysis, which required an input of data with P-adjusted WIDER IMPLICATIONS OF THE FINDINGS This is the first in-vivo study to show that Vitamin D supplementation alters gene expression in luteinised GCs. In contrast to some in-vitro evidence, no effect of the intervention on expression of genes encoding steroidogenic enzymes was observed. Unlike other studies, our results suggest that supplementation with Vitamin D is unlikely to directly influence hormone availability in FF. Our findings instead reinforce the hypothesis that Vitamin D could be considered one of the gatekeepers in protecting against an exaggerated response to ovarian stimulation. STUDY FUNDING/COMPETING INTEREST(S) The study has been funded by the Italian Ministry of Health (RF-2013-02358757) following peer review in the competitive ‘Bando di Ricerca Finalizzata e Giovani Ricercatori 2013’ for the clinical trial SUNDRO (EudraCT registration number 2015-004233-27). There are no competing interests. TRIAL REGISTRATION NUMBER EudraCT registration number 2015-004233-27.

Purpose of review Extracellular vesicles have emerged as a promising field of research for their potential to serve as biomarkers. In the pathophysiology of reproduction, they have attracted significant attention because of their diverse roles in gametogenesis and embryo-endometrial cross-talk. Advances in extracellular vesicle translational potential are herein reviewed with a particular focus in oocyte competence, semen quality diagnostics, embryo selection and detection of endometrial receptivity. Recent findings Specific miRNAs present in follicular fluid-derived extracellular vesicles have been associated with follicle development and oocyte maturation. Some proteins known to regulate sperm function and capacitation such as glycodelin, and CRISP1 have been found as overrepresented in semen exosomes isolated from severe asthenozoospermic compared to normozoospermic men. In vitro developed human embryos can secrete extracellular vesicles whose propitiousness for preimplantation genetic testing is being increasingly investigated. Endometrial cell-derived extracellular vesicles recovered from uterine flushings might represent a reservoir of molecular markers potentially exploited for monitoring the endometrial status. Summary Accumulated knowledge on extracellular vesicles deriving from endometrium, follicular fluid, embryos or male reproductive system may be translated to clinical practice to inform diagnostics in assisted reproduction technology (ART). Validation studies and technology developments are required to implement the profiling of extracellular vesicles as diagnostic tests in ART.

Uterine extracellular matrix (ECM) remodeling occurs throughout pregnancy and at parturition. Imbalanced availability of key mediators in ECM degradation, namely, matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs), is implicated in the pathogenesis of preterm labor (PTL).Examine the expression of MMPs and their inhibitors TIMPs in (a) the mouse uterus throughout normal gestation, at labor, and during inflammation-induced PTL and (b) the human term and preterm myometrium.The expression of Mmp-2/9/3/10 and Timp-1/2 was determined in the uterus of C57BL/6 mice (n = 6/group) during pregnancy (on days (d) 5, 8, 12, 15, 17, and 18), at normal labor, and during lipopolysaccharide-induced PTL (n = 6/group). The expression of MMP-10 and TIMP-1 was determined in human term and preterm myometrium before the onset of labor (TNL, n = 7; PTNL, n = 7) and during active labor (TL, n = 8; PTL, n = 8). Gene expression and tissue localization were assessed by quantitative polymerase chain reaction and immunohistochemistry, respectively.Mmp-10 was higher during murine labor (53-fold vs early pregnancy) in contrast to Mmp-2/3/9 and Timp-1, the expression of which reached a nadir at labor ( P.001 vs d5 [ Mmp-2/ 9] or P.05 vs d8 [ Mmp-3 and Timp-1]). The Mmp-3/10 and Timp-1 were localized to the uterine epithelium and stroma/myometrium. In the human myometrium, TIMP-1 messenger RNA was higher and MMP-10 was lower in TL versus TNL ( P.05), PTL ( P.001), and PTNL ( P.001). MMP-10 and TIMP-1 were localized to the myometrial smooth muscle cells, interstitial fibroblasts, and inflammatory cells.These data implicate MMP-3, TIMP-1, and MMP-10 in the uterine ECM remodeling during physiological and pathological parturition.

Background Improving in vitro fertilization success is an unmet need. Observational studies have suggested that women with deficient or insufficient vitamin D have lower chances of in vitro fertilization success, but whether supplementation improves clinical pregnancy rate remains unclear. Objective This study aimed to determine whether oral vitamin D3 supplementation improves clinical pregnancy in women undergoing an in vitro fertilization cycle. Study Design The “supplementation of vitamin D and reproductive outcome” trial is a 2-center randomized superiority double-blind placebo-controlled trial. Subjects were recruited between October 2016 and January 2019. Participants were women aged 18 to 39 years with low vitamin D (peripheral 25-hydroxyvitamin D of 0.5 ng/mL and starting their first, second, or third treatment cycle of conventional in vitro fertilization or intracytoplasmic sperm injection. The primary outcome was the cumulative clinical pregnancy rate per cycle. Pregnancies obtained with both fresh or frozen embryo transfers were included. Clinical pregnancy was defined as an intrauterine gestational sac with a viable fetus. The primary analysis was performed according to the intention-to-treat principle and could also include natural conceptions. Secondary outcomes included total dose of gonadotropins used, embryologic variables (number of oocytes retrieved, number of suitable oocytes retrieved, fertilization rate, and rate of top-quality embryos), and clinical outcomes (miscarriage rate and live birth rate). Results Overall, 630 women were randomized 2 to 12 weeks before the initiation of the in vitro fertilization cycle to receive either a single dose of 600,000 IU of vitamin D3 (n=308) or placebo (n=322). Interestingly, 113 (37%) and 130 (40%) women achieved a clinical pregnancy in the treatment and placebo groups, respectively (P=.37). The risk ratio of clinical pregnancy in women receiving vitamin D3 was 0.91 (95% confidence interval, 0.75–1.11). Compared with the placebo, vitamin D3 supplementation did not improve the rate of clinical pregnancy. Exploratory subgroup analyses for body mass index, age, indication to in vitro fertilization, ovarian reserve, interval between drug administration and initiation of the cycle, and basal levels of 25-hydroxyvitamin D failed to highlight any clinical situation that could benefit from the supplementation. Conclusion In women with normal weight with preserved ovarian reserve and low vitamin D levels undergoing in vitro fertilization cycles, a single oral dose of 600,000 IU of vitamin D3 did not improve the rate of clinical pregnancy. Although the findings do not support the use of vitamin D3 supplementation to improve in vitro fertilization success rates, further studies are required to rule out milder but potentially interesting benefits and explore the effectiveness of alternative modalities of supplementation.

The incidence of endometriosis in middle-aged women is not minimal compared to that in the reproductive age group. The treatment of affected women after childbearing age to the natural transition toward menopause has received considerably poor attention. Disease management is problematic for these women due to increased contraindications regarding hormonal treatment and the possibility for malignant transformation, considering the increased cancer risk in patients with a long-standing history of the disease. This state-of-the-art review aims for the first time to assess the benefits of the available therapies to help guide treatment decisions for the care of endometriosis in women approaching menopause. Progestins are proven effective in reducing pain and should be preferred in these women. According to the international guidelines that lack precise recommendations, hysterectomy with bilateral salpingo-oophorectomy should be the definitive therapy in women who have completed their reproductive arc, if medical therapy has failed. Strict surveillance or surgery with removal of affected gonads should be considered in cases of long-standing or recurrent endometriomas, especially in the presence of modifications of ultrasonographic cyst patterns. Although rare, malignant transformation of various tissues in endometriosis patients has been described, and management is herein discussed.

In vitro fertilization can be an effective tool to manage the endometriosis-associated infertility, which accounts for 10% of the strategy indications. Nevertheless, a negative effect of endometriosis on IVF outcomes has been suggested. The aim of this study was to evaluate the potential effect of endometriosis in the development of embryos at cleavege stage in assisted reproduction treatment cycles. A total of 429 cycles from women previously operated for moderate/severe endometriosis were compared with 851 cycles from non-affected women. Patients were matched by age, number of oocyte retrieved and study period. A total of 3818 embryos in cleavage stage have been analyzed retrospectively. Overall, no difference was found between women with and without endometriosis regarding the number of cleavage stage embryos obtained as well as the percentage of good/fair quality embryos. Excluding cycles in which no transfers were performed or where embryos were frozen in day three, no difference was observed for blastulation rate or the percentage of good/fair blastocysts obtained. Despite similar fertilization rate and number/quality of embryos, a reduction in ongoing pregnancy rate was observed in patients affected, possibly due to an altered endometrial receptivity or to the limited value of the conventional morphological evaluation of the embryo.

Endometrial cells perceive and respond to their microenvironment forming the basis of endometrial homeostasis. Errors in endometrial cell signaling are responsible for a wide spectrum of endometrial pathologies ranging from infertility to cancer. Intensive research over the years has been decoding the sophisticated molecular means by which endometrial cells communicate to each other and with the embryo. The objective of this review is to provide the scientific community with the first overview of key endometrial cell signaling pathways operating throughout the menstrual cycle. On this basis, a comprehensive and critical assessment of the literature was performed to provide the tools for the authorship of this narrative review summarizing the pivotal components and signaling cascades operating during seven endometrial cell fate “routes”: proliferation, decidualization, implantation, migration, breakdown, regeneration, and angiogenesis. Albeit schematically presented as separate transit routes in a subway network and narrated in a distinct fashion, the majority of the time these routes overlap or occur simultaneously within endometrial cells. This review facilitates identification of novel trajectories of research in endometrial cellular communication and signaling. The meticulous study of endometrial signaling pathways potentiates both the discovery of novel therapeutic targets to tackle disease and vanguard fertility approaches.

PURPOSE: To investigate the effect of sperm concentration, motility and advanced paternal age on reproductive outcomes. METHODS: A retrospective analysis of 1266 intracytoplasmic sperm injection (ICSI) cycles between 2013 and 2017. The cohort was divided into four groups according to semen concentration based on the WHO criteria (2010): group A (conc.

Endometriosis is an inflammatory disease affecting 7-10% of women throughout their reproductive life. Few data are available regarding the age at menopause of women affected by this disease. Most of the available evidence, although scanty, would indicate that both ovarian surgery for endometriomas and endometriosis per se appear to influence age at menopause. Other factors might interfere with the age at menopause of women affected by endometriosis. The purpose of this review was to examine the age at menopause in women with a history of endometriosis and to identify factors that might affect the timing of menopause.

Mouse models are used to study mechanisms that link intrauterine infection and preterm birth (PTB). To mimic intrauterine infection, lipopolysaccharide (LPS) is commonly injected into the uterus via minilaparotomy, which is invasive, and can cause PTB in control animals. We hypothesized that less-invasive ultrasound-guided intrauterine LPS injection or intravaginal LPS administration could induce PTB by stimulating an inflammatory response of the uteroplacental tissues, while minimizing PTB in control animals. On day 17 of gestation mice received LPS intravaginally (10 to 240 μg; n = 3 to 8) or into the uterus (20 μg) under ultrasound guidance (n = 7) or via laparotomy (n = 7). Control animals received phosphate-buffered saline (PBS; n = 5 to 7). Intrauterine administration of LPS, both under ultrasound guidance and via laparotomy, induced delivery earlier than in PBS control groups (P

BACKGROUND Understanding the physiology of pregnancy enables effective management of pregnancy complications that could otherwise be life threatening for both mother and fetus. A functional uterus (i) retains the fetus in utero during pregnancy without initiating stretch-induced contractions and (ii) is able to dilate the cervix and contract the myometrium at term to deliver the fetus. The onset of labour is associated with successful cervical remodelling and contraction of myometrium, arising from concomitant activation of uterine immune and endocrine systems. A large body of evidence suggests that actions of local steroid hormones may drive changes occurring in the uterine microenvironment at term. Although there have been a number of studies considering the potential role(s) played by progesterone and estrogen at the time of parturition, the bio-availability and effects of androgens during pregnancy have received less scrutiny. The aim of this review is to highlight potential roles of androgens in the biology of pregnancy and parturition.METHODS A review of published literature was performed to address (i) androgen concentrations, including biosynthesis and clearance, in maternal and fetal compartments throughout gestation, (ii) associations of androgen concentrations with adverse pregnancy outcomes, (iii) the role of androgens in the physiology of cervical remodelling and finally (iv) the role of androgens in the physiology of myometrial function including any impact on contractility. RESULTS Some, but not all, androgens increase throughout gestation in maternal circulation. The effects of this increase are not fully understood; however, evidence suggests that increased androgens might regulate key processes during pregnancy and parturition. For example, androgens are believed to be critical for cervical remodelling at term, in particular cervical ripening, via regulation of cervical collagen fibril organization. Additionally, a number of studies highlight potential roles for androgens in myometrial relaxation via non-genomic, AR-independent pathways critical for the pregnancy reaching term. Understanding of the molecular events leading to myometrial relaxation is an important step towards development of novel targeted tocolytic drugs.CONCLUSIONS The increase in androgen levels throughout gestation is likely to be important for establishment and maintenance of pregnancy and initiation of parturition. Further investigation of the underlying mechanisms of androgen action on cervical remodelling and myometrial contractility is needed. The insights gained may facilitate the development of new therapeutic approaches to manage pregnancy complications such as preterm birth.

Glucocorticoids play a primary role in the maturation of fetal organs and may contribute to the onset of labour. Glucocorticoid activity depends on the 11β-hydroxysteroid dehydrogenase family (11β-HSDs), catalysing the interconversion between “active” cortisol into inactive cortisone. No definitive study exists on 11β-HSD expression profile in human decidua and myometrium during pregnancy. We investigated the implications of 11β-HSD1 in the regulation of uterine activity in pregnancy, examining its role on contraction of a myocyte cell line and murine 11β-hsd1 levels in utero. Murine 11β-hsd1 mRNA and protein levels in utero progressively increased until the last day of gestation and significantly decreased at the onset of labour (P n = 3 to 5 in the various gestational days analysed). Experiments on human myometrial samples confirm the significant fall in 11β-hsd1 mRNA levels at labour, compared to end pregnancy samples ( n = 5 to 8). In vitro experiments showed that human myometrial contraction is inhibited by using a non-selective inhibitor of 11β-HSD1. The present study shows the temporal localisation of 11β-HSD1 in uterus, highlighting its importance in the timing of gestation and suggesting its contribution in the myometrium contraction.

Premature prelabor rupture of fetal membranes accounts for 30% of all premature births and is associated with detrimental long-term infant outcomes. Premature cervical remodeling, facilitated by matrix metalloproteinases, may trigger rupture at the zone of the fetal membranes overlying the cervix. The similarities and differences underlying cervical remodeling in premature prelabor rupture of fetal membranes and spontaneous preterm labor with intact membranes are unexplored.We aimed to perform the first transcriptomic assessment of the preterm human cervix to identify differences between premature prelabor rupture of fetal membranes and preterm labor with intact membranes and to compare the enzymatic activities of matrix metalloproteinases-2 and -9 between premature prelabor rupture of fetal membranes and preterm labor with intact membranes.Cervical biopsies were collected following preterm labor with intact membranes (n = 6) and premature prelabor rupture of fetal membranes (n = 5). Biopsies were also collected from reference groups at term labor (n = 12) or term not labor (n = 5). The Illumina HT-12 version 4.0 BeadChips microarray was utilized, and a novel network graph approach determined the specificity of changes between premature prelabor rupture of fetal membranes and preterm labor with intact membranes. Quantitative reverse transcription-polymerase chain reaction and Western blotting confirmed the microarray findings. Immunofluorescence was used for localization studies and gelatin zymography to assess matrix metalloproteinase activity.PML-RARA-regulated adapter molecule 1, FYVE-RhoGEF and PH domain-containing protein 3 and carcinoembryonic antigen-ralated cell adhesion molecule 3 were significantly higher, whereas N-myc downstream regulated gene 2 was lower in the premature prelabor rupture of fetal membranes cervix when compared with the cervix in preterm labor with intact membranes, term labor, and term not labor. PRAM1 and CEACAM3 were localized to immune cells at the cervical stroma and NDRG2 and FGD3 were localized to cervical myofibroblasts. The activity of matrix metalloproteinase-9 was higher (1.22 ± 4.403-fold, P.05) in the cervix in premature prelabor rupture of fetal membranes compared with preterm labor with intact membranes.We identified 4 novel proteins with a potential role in the regulation of cervical remodeling leading to premature prelabor rupture of fetal membranes. Our findings contribute to the studies dissecting the mechanisms underlying premature prelabor rupture of fetal membranes and inspire further investigations toward the development of premature prelabor rupture of fetal membranes therapeutics.

CONTEXT: Uterine quiescence must be maintained until pregnancy reaches term. Premature activation of myometrial contractility leads to preterm labor and delivery.OBJECTIVE: To scrutinize the potential of androgens to relax the myometrium and the mechanism of their action.SAMPLES: A pregnancy-derived myometrial smooth muscle cell line (PHM1-41) and myometrial strips prepared from tissues obtained from pregnant women (lean, n = 9; obese, n = 6) undergoing elective cesarean section at term and from nonpregnant C57BL/6 mice (n=5) were each utilized.DESIGN: The contraction of collagen-embedded PHM1-41s and the stretch-induced contraction of human and murine myometrial strips were assessed after incubation with Testosterone (T), dihydrotestosterone (DHT), and T conjugated to BSA. Intracellular calcium ([Ca(2+)]) and phosphorylated myosin light chain concentrations were quantified in PHM1-41s using a Fluo-4 Ca(2+) assay and in-cell Westerns, respectively.SETTING: University research institute.RESULTS: DHT and T, but not T conjugated to BSA, impaired the contractile function of PHM1-41s and of human and murine myometrial strips. The response was rapid (observed within minutes), was sustainable for up to 48 hours, and was not abolished on knockdown of the androgen receptor. DHT (100 μm) reduced the amplitude of lean strip contraction to 2 ± 2% of the pretreatment value and T (100 μm) to 3.3 ± 1%. These values for obese strips were 15 ± 6.7% and 11± 6.7%, respectively. At the same doses, in murine strips, DHT reduced the amplitude to 4.8 ± 3% and T to 4.9 ± 3%. DHT (50 μm) pretreatment reduced the oxytocin-stimulated increase in [Ca(2+)] (P < .0001; n = 6) and phosphorylated myosin light chain (P < .05; n = 5) in PHM1-41s.CONCLUSION: Lipid-soluble androgens could be developed as tocolytic agents for the treatment of preterm labor.