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Resume Objectifs Etablir un etat des lieux sur la definition, le diagnostic, les facteurs de risque, les symptomes, et les traitements de l’isthmocele. Methodes Revue de la litterature, reflexions critiques, et conseils pragmatiques. Resultats La definition de l’isthmocele n’est pas consensuelle. Certains proposent une indentation ≥ 2 mm du myometre de la cicatrice de cesarienne, mais cet aspect present chez plus de la moitie des uterus cicatriciels ne prend pas en compte les symptomes de la femme. La methode de diagnostic la plus performante semble etre l’echographie ± hysterosonographie. Les situations a risque d’isthmocele sont l’uterus multicicatriciel, la cesarienne pendant le travail avec une incision trop basse, et l’uterus retroverse. Les symptomes de l’isthmocele sont des saignements gynecologiques anormaux et des douleurs pelviennes, et leur presence signe le « syndrome de la cicatrice de cesarienne ». Les risques d’une grossesse en cas d’isthmocele ont ete peu etudies, mais elle ne semble pas contre-indiquee, meme si l’isthmocele n’est pas traite. Le traitement de l’isthmocele est avant tout chirurgical, conservateur, et semble reserve aux patientes symptomatiques, et aux infertilites secondaires en echec de traitement. Les patientes avec myometre residuel ≥ 2,5 mm peuvent beneficier d’un traitement hysteroscopique de premiere intention, tandis qu’on proposera un abord cœlioscopique ou vaginal dans les autres cas. Conclusions Une definition pragmatique de l’isthmocele maladie en incluant la presence de symptomes est le pre-requis necessaire a la prise en charge des femmes. Le traitement est avant tout chirurgical, conservateur ou non suivant le desir de grossesse ulterieur.

International audience; The interest in and understanding of the human microbiome has grown remarkably over recent years. Advances in molecular techniques have allowed researchers to identify and study the microbiota and also use this information to develop therapeutic solutions for a spectrum of conditions. Alongside the growing interest in the microbiome, societal changes have resulted in many couples looking to start families later in life, therefore increasing the demand for assisted reproductive technologies. Combining these trends, it makes sense that clinicians are eager to understand and exploit the microbiome of their patients, i.e. the reproductive microbiome, in order to help them achieve their goal of becoming parents. This paper aims to provide an overview of the current and future research into the reproductive microbiome in relation to fertility and also share clinical practice recommendations for physicians who are new to this field or unsure about how they can utilise what is known to help their patients.

International audience; Objective: To provide a snapshot of the surgical management of endometriosis in French high-volume activity centers.Methods: Analysis of prospectively collected data between November 2015 and May 2017 in 21 centers with a high volume of endometriosis surgery in France. Each facility could include up to 40 patients undergoing laparoscopy for endometriosis. Data were collected before and two months after surgery.Results: 361 patients were enrolled in the study. Twenty-seven patients (7.48%) were lost to follow-up at the month 2 visit. Endometriosis stage was I-II in 33.70% of patients and III-IV in 66.30%. Uterosacral ligament resection was the most frequently performed procedure (50.97%) followed by rectal surgery (31.58%), ovarian procedures for endometrioma, procedures for ureters (21.33%) and the bladder (11.91%). Antiadhesion agents were employed in 215/361 (59.56%) patients. The median length of hospital stay after surgery was 2 (IQR 1 − 4) days. Post-operative complications were recorded in 9.34% of patients. Rectovaginal fistulae occurred in 8 patients (2.41%), pelvic abscess in 4 (1.20%) and bladder atony in 3 (0.90%). 17 patients (5.14%) required a second surgical procedure after a median time of 31 days (IQR 9 − 81). Two months after surgery, 95.09% of patients reported being satisfied or very satisfied with the surgery.Conclusion: Our study shows that surgical management of endometriosis in centers with a high volume of endometriosis surgery, mainly concerns women presenting with severe disease and deep localizations, with an overall risk of major complications inferior to 10% and a high rate of patient satisfaction.

It is already known that children born after slow frozen embryo replacement have a significantly higher birth weight compared to children born after fresh embryo transfer. Similar data have been reported related to frozen embryo transfer using an open vitrification system. However, few data relative to birth weight using a complete embryo closed vitrification system has been reported. The purpose of this study was to know if frozen embryo in closed vitrification system is associated with a higher birth weight compared to fresh embryo replacement.Design: This was a monocentric retrospective cohort study, 371 children were issued from fresh embryo replacement and 127 from vitrified embryo transfer.Materials and methods: All singletons born after fresh or vitrified embryo transfer between January 2011 and April 2015 were included. Births from the vitrified or fresh transfers of egg or sperm donation, and preimplantation genetic diagnosis were excluded. In addition, pregnancies with more than one gestational sac at the first ultrasound were excluded. An analysis of covariance (ANCOVA) was used for multivariate analysis.Results: Mean birth weight was 205g higher in the frozen embryo compared with fresh embryos transfer groups (3368g vs. 3163g respectively, P, Les enfants nés après un transfert d’embryon congelé par congélation lente ou vitrifié en système ouvert ont un poids de naissance plus élevé que les enfants nés suite à un transfert d’embryon frais. Cependant, il existe peu de données sur le poids de naissance et la vitrification en système fermé. Cette technique de vitrification est la plus utilisée en France. L’objectif de cette étude est de savoir si la vitrification embryonnaire en système fermé était associée à des poids de naissance plus élevés que les transferts d’embryons frais.Méthodes :Il s’agit d’une étude unicentrique de cohorte rétrospective. Tous les singletons nés suite à un transfert d’embryon frais ou vitrifié en dehors d’un don de gamète entre janvier 2011 et avril 2015 ont été inclus. Les embryons ayant bénéficié d’un diagnostic pré-implantatoire ou dont la grossesse était initialement gémellaire ont été exclus. Une analyse de covariance (ANCOVA) a été utilisée pour l’analyse multivariée.Résultats :Nous avons étudié 371 singletons nés après transfert d’embryons frais et 127 singletons nés après transfert d’embryons vitrifiés en système fermé. Le poids de naissance des enfants nés après replacement d’embryons vitirifés était significativement plus élevé de 205 g en comparaison avec les enfants nés après replacements d’embryons frais (3368 g vs 3163 g respectivement, p < 0,001). Cette différence était toujours significative après analyse multivariée incluant les facteurs confondants que sont l’âge maternel, l’IMC maternel, la parité, l’exposition tabagique et le nombre d’embryons transférés (p < 0,001).Conclusion :Les embryons vitrifiés en système fermé sont associés à un poids de naissance plus élevé que les embryons frais. Ces données sont cohérentes avec les données de la congélation lente et la vitrification en système ouvert. Les effets de l’hyperstimulation ovarienne contrôlée, les conditions de culture ex vivo et les effets de la cryopréservation sur le poids de naissance nécessitent d’être étudiés plus largement.

The transfer of a competent embryo into a uterine cavity during the implantion window is a major objective to ensure success in in vitro fertilization (IVF). My thesis work contributes to (i) the identification of new biomarkers for the evaluation of endometrial receptivity (ii) the understanding of embryo competence in IVF.Concerning the first axis, the team has developed the Win-Test®, an endometrial receptivity evaluation test based on the search for a specific transcriptomic signature in endometrial biopsies taken from patients during their theoretical implantation window. The purpose of this test is to determine the appropriate time to replace the embryo. When the “receptive” period is identify, it allows the embryo transfer date to be adapted to the patient’s effective window of implantation, significantly increasing the pregnancy rate after personalized frozen embryo(s) transfer (Haouzi et al, 2020). To date, this test requires two endometrial biopsies and can only be performed on a frozen/thawed embryo transfer. In order to overcome these two constraints, the objective of my thesis was to participate in the development of a non-invasive approach in the evaluation of endometrial receptivity in IVF. Analysis of microRNA expression profiles from endometrial biopsies allowed us to identify certain endometrial microRNAs associated with endometrial receptivity ("receptive" versus "non-receptive") as well as with the success of the attempt ("implantation failure" versus "sucessfull implantation" and "miscarriage" versus "birth") (Drissennek, Baron, et al, 2020). My preliminary results indicate that some of these microRNAs are detected in the serum of patients, thus opening interesting perspectives for the development of a specific non-invasive test of endometrial receptivity.The second axis of my thesis focused on the improvement of the development and the early implant potential of human embryos. In vivo, the embryo evolves in the fallopian tube until the morula stage and arrives in the uterus at the blastocyst stage around D6 post fertilization, in an endometrium supposed to be receptive which will be able to ensure implantation. Physiologically, it has been shown that the oxygen level is 5% in the fallopian tubes and 2% in the uterus. To date, the majority of IVF laboratories in the world use 5% oxygen from D0 to D6 for the in vitro culture of human embryos. Our results showed that biphasic in vitro culture at 5% oxygen from D0 to D3 and then at 2% oxygen from D3 to D5/D6 (thus mimicking physiological oxygen concentrations) significantly improves human embryo development and live birth rates in IVF. Using a transcriptomic approach, our results highlighted the different expression of 707 genes depending on whether the embryos were cultured at 5% oxygen or at 5% and then 2% oxygen, with an overexpression of the majority (93.8%) of dysregulated transcripts in embryos cultured with biphasic strategy (5-2% oxygen). Functional analysis revealed the involvement of these RNAs in key cellular processes of embryonic growth and implant potential; such as proliferation, DNA repair and maintenance of pluripotency. These results open new perspectives for the improvement of embryonic culture conditions, subject to confirmation in a larger scale study., Le transfert d’un embryon compétent dans la cavité utérine pendant la fenêtre d’implantation est un objectif majeur pour garantir le succès en Fécondation in vitro (FIV). Mon travail de thèse contribue (i) à l’identification de nouveaux biomarqueurs d’évaluation de la réceptivité endométriale (ii) à la compréhension de la compétence embryonnaire en FIV.Concernant le premier axe, l’équipe a mis au point le Win-Test®, test d’évaluation de la réceptivité endométriale basé sur la recherche d’une signature transcriptomique spécifique au sein de biopsies endométriales prélevées chez des patientes pendant leur fenêtre théorique d'implantation. Le but de ce test est de déterminer le moment adéquat pour replacer l’embryon. Lorsque la période « réceptive » est identifiée, il permet d’adapter la date du transfert embryonnaire à la fenêtre implantatoire effective de la patiente, augmentant significativement le taux de grossesses après transfert personnalisé d’embryon(s) congelé(s) (Haouzi et al, 2020). A ce jour, ce test nécessite la réalisation de deux biopsies endométriales, et ne peut être réalisé que sur un transfert d’embryon congelé/décongelé. Pour tenter de s’affranchir de ces deux contraintes, l’objectif de ma thèse a été de participer à la mise au point d’une approche non-invasive de l’évaluation de la réceptivité endométriale en FIV. L’analyse des profils d’expression des microARNs issus de biopsies endométriales nous a permis d’identifier certains microARNs endométriaux associés à la réceptivité endométriale (« réceptif » versus « non réceptif ») ainsi qu'au succès de la tentative (« échec d'implantation » versus « implantation réussie » et « fausse couche » versus « naissance ») (Drissennek, Baron, et al, 2020). Mes résultats préliminaires indiquent que certains de ces microARNs sont détectés dans le sérum des patientes, ouvrant ainsi des perspectives intéressantes pour la mise au point d’un test non-invasif spécifique de la réceptivité endométriale.Le second axe de ma thèse a porté sur l’amélioration du développement et du potentiel implantatoire précoce des embryons humains. En effet, l’embryon évolue in vivo dans la trompe de Fallope jusqu’au stade morula et arrive dans l’utérus au stade de blastocyste vers J6 post-fécondation, dans un endomètre supposé réceptif qui pourra assurer l’implantation. Physiologiquement, il a été montré que le taux d’oxygène était de 5% dans les trompes de Fallope, puis de 2% dans l’utérus. A ce jour, la majorité des laboratoires de FIV dans le monde utilise pourtant 5% d’oxygène de J0 à J6 pour la culture in vitro des embryons humains. Nos résultats ont montré qu’une culture in vitro séquentielle à 5% d’oxygène de J0 à J3 puis à 2% d’oxygène de J3 à J5/J6 (mimant ainsi les concentrations physiologiques en oxygène) améliorerait significativement le développement embryonnaire humain et les taux de naissances vivantes en FIV. En utilisant une approche transcriptomique, nos résultats mettent en évidence la différentiation d’expression de 707 gènes selon que les embryons aient été cultivés à 5% d’oxygène ou à 5% puis 2% d’oxygène, avec une surexpression de la majorité (93,8%) des transcrits dérégulés dans les embryons cultivés en stratégie séquentielle (5-2% d’oxygène). L’analyse fonctionnelle a mis en évidence l’implication de ces ARNs dans des processus cellulaires clés de la croissance et du potentiel implantatoire embryonnaire ; tels que la prolifération, la réparation de l’ADN et le maintien de la pluripotence. Ces résultats ouvrent de nouvelles perspectives quant à l’amélioration des conditions de culture embryonnaire sous réserve de la confirmation sur une étude à plus large échelle.

International audience; Vasculogenesis and angiogenesis are key processes of placental development, which occur throughout pregnancy. Placental vasculogenesis occurs during the first trimester of pregnancy culminating in the formation of hemangioblasts from intra-villous stem cells. Placental angiogenesis occurs subsequently, forming new blood vessels from existing ones. Angiogenesis also takes place at the fetomaternal interface, allowing essential spiral arteriole remodeling to establish the fetomaternal circulation. Vasculogenesis and angiogenesis in animal models and in humans have been studied in a wide variety of in vitro, physiological and pathological conditions, with a focus on the pro- and anti-angiogenic factors that control these processes. Recent studies revealed roles for new families of proteins, including direct participants such as the prokineticin family, and regulators of these processes such as the homeobox genes. This review summarizes recent advances in understanding the molecular mechanisms of actions of these families of proteins. Over the past decade, evidence suggests increased production of placental anti-angiogenic factors, as well as angiogenic factors are associated with fetal growth restriction (FGR) and preeclampsia (PE): the most threatening pathologies of human pregnancy with systemic vascular dysfunction. This review also reports novel clinical strategies targeting members of these family of proteins to treat PE and its consequent effects on the maternal vascular system.

International audience; Human ovarian tissue cryopreservation (OTC) is increasingly used worldwide to preserve female fertility in prepubertal girls and women at risk of premature ovarian insufficiency (POI) in the context of urgent gonadotoxic treatments or ovarian surgery. Fertility preservation is challenging because there is no consensus regarding patient management, preservation fertility strategies, or even technical laboratory protocols, which implies that each procedure must be adapted to the characteristics of the patient profile and its own risk-benefit ratio. During OTC, mature/immature oocytes can be aspirated directly from large/small antral follicles within ovarian tissue samples and/or be released into culture media from growing follicles during ovarian tissue dissection in prepubertal girls and women. In this manuscript, we present a protocol that combines ovarian tissue freezing with the cryopreservation of mature/immature oocytes retrieved from ovarian tissue samples, improving the reproductive potential of fertility preservation. Appropriate collection, handling, and storage of ovarian tissue and oocytes before, during, and after the cryopreservation will be described. The subsequent use and safety of cryopreserved/thawed ovarian tissue samples and oocytes will also be discussed, as well as the optimal timing for in vitro maturation of immature oocytes. We recommend the systematic use of this protocol in fertility preservation of prepubertal girls and women as it increases the whole reproductive potential of fertility preservation (i.e., oocyte vitrification in addition of OTC) and also improves the safety and use of fertility preservation (i.e., thawing of oocytes versus ovarian graft), maximizing the chance of successful childbirth for the patients at risk of POI.

BACKGROUNDA dynamic balance between pro- and anti-inflammatory factors contributes to regulating human female reproduction. Chronic low-grade inflammation has been detected in several female reproductive conditions, from anovulation to embryo implantation failure. C-reactive protein (CRP) is a reliable marker of inflammation that is extensively used in clinical practice. Recent studies quantified CRP in the serum of infertile women undergoing ART and suggested its potential for the prediction of ART reproductive outcomes.OBJECTIVE AND RATIONALEThe first objective of this systematic review of the available literature was to evaluate the association between pre-implantation circulating CRP concentration and pregnancy rates in women undergoing ART. The second objective was to describe serum CRP concentration changes after early embryo implantation. The changes in circulating CRP throughout the ART cycle, clinical implications of CRP quantification for the management of women undergoing ART, and future therapeutic options will also be discussed.SEARCH METHODSThe MEDLINE database was systematically searched from inception to March 2019 using the following key words: (C-reactive protein) AND (assisted reproductive techniques OR ovulation induction OR insemination OR in vitro fertilization). Only articles in English were considered. Studies were selected based on title and abstract. The full text of potentially relevant articles was retrieved and assessed for inclusion by two reviewers (S.B. and S.H.). The protocol was registered in the International prospective register of systematic reviews (PROSPERO; registration number: CRD148687).OUTCOMESIn total, 10 studies were included in this systematic review. Most of these studies reported lower circulating CRP values before the window of implantation and higher circulating CRP values during the peri-implantation period in women with successful ART outcome (biochemical or clinical pregnancy) compared to women without a successful outcome. Several lifestyle factors and/or drugs that reduce the concentration of circulating CRP significantly improve ART outcomes. Subgroup analyses according to female BMI and baseline circulating CRP concentration are highly recommended in future analyses.WIDER IMPLICATIONSThese findings highlight a possible detrimental impact of preconception high circulating CRP concentration on ART outcomes. However, the biochemical or clinical pregnancy rate endpoints used in the studies examined here are insufficient (there were no data on live birth outcome), and the impact of major variables that can influence CRP and/or ART, for example maternal age, BMI, number of transferred embryos, and use of anti-inflammatory drugs, were not considered in the analyses. CRP quantification may be a potential marker of ART outcome, but its predictive value still needs to be investigated in large prospective studies. In future, the quantification of circulating CRP before starting ART could help to identify patients with a poor ART prognosis, leading to ART cycle cancellation or to preconception treatment to minimize the medical risks and costs.

International audience; This is a retrospective study to evaluate if the miRNome profile of endometrium samples collected during the implantation window predicts Assisted Reproduction Technology (ART) outcomes. We first investigated the endometrial miRNome profile according to the receptivity status in 20 patients with repeated implantation failures (RIF) (discovery cohort). After customized embryo transfer, the miRNome profiles of receptive patients with a positive or negative β-hCG, and with early miscarriage or live birth were analysed. Some differentially expressed miRNAs were selected for validation by RT-qPCR in endometrial samples from 103 RIF patients (validation cohort). Analysis of the different miRNome profiles identified endometrial receptivity, implantation failure, and early miscarriage-associated miRNA signatures that included 11, 261, and 76 miRNAs, respectively. However, only four miRNAs associated with the endometrial receptivity status (miR-455-3p and miR-4423-3p) and implantation failure (miR-152-3p and miR-155-5p) were significantly validated in endometrial samples. The miRNome profile of endometrial tissues during the implantation window can predict the pregnancy outcome. These data are crucial for opening new perspectives to predict implantation failure and consequently, to increase ART success.

International audience; The genital microbiota actively participates in women's reproductive health. Indeed, a genital dysbiosis (microbial imbalance associated with adverse effects on host health) can lead to vaginal infections (such as mycoses or bacterial vaginosis). Recent data reported that genital dysbiosis (e.g. vaginal or endometrial) was associated with fewer chances of live births in assisted reproductive technologies (ART), via decreased pregnancy rates and an increased risk of miscarriages. The presence or diversity of certain bacterial strains (in particular Gardenellavaginalis, Proteobacteria, Lactobacillusjensenii, Lactobacilluscrispatus or Atopobiumvaginae) within the genital microbiota seem to be associated with the outcomes of ART cycles, suggesting new approaches to improve ART results. In this review, we aim at presenting the state of art on the association between the female genital microbiota and ART success. The diagnostic and therapeutic approaches (i.e. probiotics, antibiotic therapy and transplantation of vaginal microbiota) in the management of patients with altered microbiota will also be discussed. The confirmation of these data in the coming years could significantly improve the management of infertile patients in ART with a more personalized approach partially based on the female genital microbiotic profile.

International audience; Although many studies have demonstrated the superiority of ultra-fast freezing compared with slow freezing, the debate is still ongoing concerning the best type of vitrification method: direct exposure to liquid nitrogen (i.e., open systems), or sterile system without contact with liquid nitrogen (i.e., closed systems). The aims of this study were to share our experience on closed vitrification systems in the framework of our egg donation programme with fully asynchronous cycles, and to identify predictive factors of successful outcome in this context. Logistic regression analysis indicated that the number of vitrified oocytes was the only factor predictive of the oocyte survival rate and of clinical pregnancy. The addition of one vitrified oocyte increased by 15 % the odds of oocyte survival. When the oocyte survival rate was considered as a continuous variable, the following results were obtained: 7 % of clinical pregnancy probability for 50 % survival rate, 15 % for 75 % survival rate, and 32 % for 100 % survival rate. The rates of oocyte survival and fertilization, embryo implantation, and clinical pregnancy were in agreement with the recommended values established by ALPHA Scientists in Reproductive Medicine in 2012. On the basis of these results, and according to the European directives on safety, we validate the routine use of closed oocyte vitrification systems for egg donation programmes. These results must be confirmed in larger samples before extrapolation to all patient types.

Context Prokineticin 1 (PROK1) quantification in global follicular fluid (FF) has been recently reported as a predictive biomarker of in vitro fertilization (IVF) outcome. It is now necessary to evaluate its clinical usefulness in individual follicles. Objectives To evaluate the clinical value of PROK1 secretion in individual FF to predict oocyte competence. To determine the impact of follicular size, oocyte maturity, and gonadotropin treatments on PROK1 secretion. Design and setting Prospective cohort study from May 2015 to May 2017 at the University Hospital of Grenoble. Patients A total of 69 infertile couples underwent IVF. Intervention(s) Collection of 298 individual FF from 44 women undergoing IVF; 52 individual cumulus cell (CC) samples and 15 CC primary cultures from 25 women undergoing IVF-intracytoplasmic sperm injection (ICSI). Main Outcome Measure(s) Oocyte competence was defined as the ability to sustain embryo development to the blastocyst stage. Follicular size was measured by 2D-sonography. PROK1 concentration was quantified by ELISA assay. Results PROK1 concentration was correlated to follicular size (r = 0.85, P = 2.2 × 10−16). Normalized PROK1 concentration in FF was predictive of subsequent oocyte competence (AUROC curve = 0.76 [95% CI, 0.69–0.83]; P = 1.7 × 10−9), irrespectively of day-2 embryo morphokinetic parameters. The expression and secretion of PROK1 were increased in FF and CC of mature oocytes (P < 0.01). Follicle Stimulating Hormone and hCG up-regulated PROK1 secretion in CC primary cultures (P < 0.01; P < 0.05), probably through the cAMP pathway (P < 0.01). Conclusions PROK1 quantification in individual FF could constitute a new predictive biomarker of oocyte competence in addition with embryo morphokinetic parameters. Trial registration number none.

International audience; It is already known that children born after slow frozen embryo replacement have a significantly higher birth weight compared to children born after fresh embryo transfer. Similar data have been reported related to frozen embryo transfer using an open vitrification system. However, few data relative to birth weight using a complete embryo closed vitrification system has been reported. The purpose of this study was to know if frozen embryo in closed vitrification system is associated with a higher birth weight compared to fresh embryo replacement.Design: This was a monocentric retrospective cohort study, 371 children were issued from fresh embryo replacement and 127 from vitrified embryo transfer.Materials and methods: All singletons born after fresh or vitrified embryo transfer between January 2011 and April 2015 were included. Births from the vitrified or fresh transfers of egg or sperm donation, and preimplantation genetic diagnosis were excluded. In addition, pregnancies with more than one gestational sac at the first ultrasound were excluded. An analysis of covariance (ANCOVA) was used for multivariate analysis.Results: Mean birth weight was 205g higher in the frozen embryo compared with fresh embryos transfer groups (3368g vs. 3163g respectively, P

International audience; Repression of retrotransposons is essential for genome integrity during germ cell development and is tightly controlled through epigenetic mechanisms. In primordial germ cells, protein arginine N-methyltransferase (Prmt5) is involved in retrotransposon repression by methylating Piwi proteins , which is part of the piRNA pathway. Here, we show that in mice, genetic inactivation of coprs (which is highly expressed in testis and encodes a histone-binding protein required for the targeting of Prmt5 activity) affects the maturation of spermatogonia to spermatids. Mass spectrom-etry analysis revealed the presence of Miwi in testis protein lysates immunoprecipitated with an anti-Coprs antibody. The observed deregulation of Miwi and pachytene pre-piRNAs levels and the derepression of LINE1 repetitive sequences observed in coprs-/-mice suggest that Coprs is implicated in genome surveillance mechanisms. Spermatogenesis is a multistep process that takes place in the seminiferous tubules whereby mature spermato-zoa are continuously produced during adulthood reproduction lifetime. Specifically, primordial germ cells (PGCs) generate spermatogonia that either self-renew to maintain a pool of stem cells or undergo differentiation. After two meiotic divisions, secondary spermato-cytes are produced. Then, during spermiogenesis, they differentiate into elongating spermatids and finally spermatozoa [1-5]. Importantly, these changes are associated with chromatin compaction, transcription arrest, and specific epigenetic modifications that contribute to the production of mature sperm. Concomi-tantly, to maintain genome integrity, PGCs need to repress repetitive DNA elements because their spread can lead to inherited diseases. These repetitive sequences [for instance, long interspersed nuclear elements 1 (LINEs), short interspersed nuclear element (SINEs), and intracisternal A-particle (IAP) sequences] derive mostly from transposable elements and represent Abbreviations Coprs, coordinator of Prmt5 and differentiation stimulator; IAP, intracisternal A-particle; LINE1, long interspersed nuclear element 1; piRNA, Piwi-interacting RNA; Prmt5, protein arginine N-methyltransferase; SINE, short interspersed nuclear element. 159 FEBS Open Bio 9 (2019) 159-168 ª

International audience; Does autologous endometrial cell co-culture (AECC) improve the number of good-quality blastocysts obtained by IVF/intracytoplasmic sperm injection (ICSI), compared with conventional embryo culture medium in a broad group of patients referred to assisted reproductive technology (ART)?Design: This interventional, randomized, double-blind study took place at Clinique Ovo from March 2013 to October 2015 and included 207 healthy patients undergoing an IVF or ICSI protocol, of which 71 were excluded before randomization. On the previous cycle, all participants underwent an endometrial biopsy at D5 to D7 post-ovulation, following which the endometrial cells were prepared for AECC.Results: The data demonstrated that AECC significantly increased the incidence of good-quality blastocysts compared with culture in conventional media (42.6% vs 28.4%, P < 0.001). No significant differences were found in pregnancy and live birth rates.Conclusion: This study demonstrated the benefits of AECC on blastocyst quality compared with conventional embryo culture medium, in a broader category of patients referred to ART as opposed to other studies that concentrated on specific causes of infertility only. However, limitations of the study design should be taken into consideration; the analysis was performed using embryos rather than patients and a follow-up of children born following the treatments could not be conducted

Birth weight (BW) is higher after frozen embryo transfer (FET) than after fresh embryo replacement. No study has compared the BW of siblings conceived using the same oocyte/embryo cohort. The aim of this study was to determine whether the freezing-thawing procedure is involved in such difference. Multicenter study at Montpellier University Hospital, Clinique Ovo, Canada and Grenoble-Alpes University Hospital. The first cohort (Fresh/FET) included in vitro fertilization (IVF) cycles where the older was born after fresh embryo transfer (n = 158) and the younger after transfer of frozen supernumerary embryos (n = 158). The second cohort (FET/FET) included IVF cycles where older and younger were born after FET of embryos from the same cohort. The mean adjusted BW of the FET group was higher than that of the fresh group (3508.9 ± 452.4 g vs 3237.7 ± 463.3 g; p vs 3337.1 ± 391.9 g; p = 0.3789). Our results strongly suggest that cryopreservation is directly involved in the BW variation. Comparing BW difference between Fresh/FET cohort and FET/FET one, it suggests that parity is not the only responsible, increasing the role of cryopreservation step in BW variation.

International audience; Research question: To analyse why unbalanced viable offspring are derived mainly from the 3:1 segregation mode in t(11;22)(q23;q11.2) reciprocal translocation. Design: Retrospective analysis of 24 pre-implantation genetic testing for chromosomal structural re-arrangements (PGT-SR) cycles was performed on seven male and five female carriers of t(11;22) translocation. Sperm analysis was performed on each male carrier. These patients were directed to the study centre after several years of miscarriages and/or abortions, primary infertility for male carriers or birth of an affected child. Results: Twenty-four PGT-SR cycles were performed to exclude imbalances in both male and female carriers. The unbalanced embryos derived from the adjacent-1 segregation mode were the most represented in both male and female carriers (68.4% and 50%, respectively). These results were positively related with meiotic segregation analysis of reciprocal translocation in spermatozoa. A thorough analysis of the unbalanced embryo karyotypes determined that the expected viable +der22 karyotype resulting from 3:1 malsegregation was less represented at 5.3%. Conclusions: These findings highlight the divergence that may exist between meiotic segregation and post-zygotic selection. Post-zygotic selection would be responsible for the elimination of unbalanced embryos derived from the adjacent-1 segregation mode. The combined action of several factors occurs at the beginning of post-zygotic selection. Genetic counselling must consider the risk of a birth related to the adjacent-1 segregation mode, irrespective of the sex of the translocation carrier. These results will allow deeper understanding of the PGT results of t(11;22) carriers, which often include a high number of aneuploid embryos.

Survival and cell death signals are crucial for mammalian embryo preimplantation development. However, the knowledge on the molecular mechanisms underlying their regulation is still limited. Mouse studies are widely used to understand preimplantation embryo development, but extrapolation of these results to humans is questionable. Therefore, we wanted to analyse the global expression profiles during early mouse and human development with a special focus on genes involved in the regulation of the apoptotic and survival pathways. We used DNA microarray technology to analyse the global gene expression profiles of preimplantation human and mouse embryos (metaphase II oocytes, embryos at the embryonic genome activation stage, and blastocysts). Components of the major apoptotic and survival signalling pathways were expressed during early human and mouse embryonic development; however, most expression profiles were species-specific. Particularly, the expression of genes encoding components and regulators of the apoptotic machinery were extremely stable in mouse embryos at all analysed stages, while it was more stage-specific in human embryos.CASP3,CASP9, andAIFwere the only apoptosis-related genes expressed in both species and at all studied stages. Moreover, numerous transcripts related to the apoptotic and survival pathway were reported for the first time such asCASP6andIL1RAPL1that were specific to MII oocytes;CASP2,ENDOG, andGFERto blastocysts in human. These findings open new perspectives for the characterization and understanding of the survival and apoptotic signalling pathways that control early human and mouse embryonic development.

International audience; Microsatellite instability in colorectal cancer predicts favorable outcomes. However, the mechanistic relationship between microsatellite instability, tumor-infiltrating immune cells, Immunoscore, and their impact on patient survival remains to be elucidated. We found significant differences in mutational patterns, chromosomal instability, and gene expression that correlated with patient microsatellite instability status. A prominent immune gene expression was observed in microsatellite-instable (MSI) tumors, as well as in a subgroup of microsatellite-stable (MSS) tumors. MSI tumors had increased frameshift mutations, showed genetic evidence of immunoediting, had higher densities of Th1, effector-memory T cells, in situ proliferating T cells, and inhibitory PD1-PDL1 cells, had high Immunoscores, and were infiltrated with mutation-specific cytotoxic T cells. Multivariate analysis revealed that Immunoscore was superior to microsatellite instability in predicting patients' disease-specific recurrence and survival. These findings indicate that assessment of the immune status via Immunoscore provides a potent indicator of tumor recurrence beyond microsatellite-instability staging that could be an important guide for immunotherapy strategies.

We present data relating to the interactome of MCM9 from the nuclei of human cells. MCM9 belongs to the AAA+ superfamily, and contains an MCM domain and motifs that may confer DNA helicase activity. MCM9 has been shown to bind MCM8, and has been implicated in DNA replication and homologous recombination. However, the mechanistic basis of MCM9’s role in DNA repair is poorly understood, and proteins with which it interacts were hitherto unknown. We performed tandem affinity purification of MCM9 and its interacting proteins from nuclear extracts of human cells, followed by proteomic analysis, thereby generating a set of mass spectrometry data corresponding to the MCM9 interactome [1]. The proteomic data set comprises 29 mass spectrometry RAW files, deposited to the ProteomeXchange Consortium, and freely available from the PRIDE partner repository with the data set identifier http://www.ebi.ac.uk/pride/archive/projects/PXD000212. A set of 22 interacting proteins identified from the proteomic data was used to create an MCM9-centered interactive network diagram, using the Cytoscape program. These data allow the scientific community to access, mine and explore the human nuclear MCM9 interactome. Keywords: Proteomics Data, Mass Spectrometry, DNA Replication And Repair, Affinity Purification, Protein-Protein Interactions

International audience

Access to in vitro fertilization (IVF) for obese women varies across centers, and the impact of obesity on IVF outcomes is widely discussed. We assessed the impact of obesity and its severity on live birth rate (LBR) after IVF. We included women treated for IVF in our center. Data were prospectively collected in the BabySentryTM software. LBR per cycle and cumulative LBR including all attempts of the couple were calculated, considering transfer of both fresh and frozen embryos. Of 1588 included women (2379 controlled ovarian stimulations), 70.2%, 19.5%, 7.9%, and 2.4% were normal-weight, overweight, class I obesity, and class II/III obesity, respectively. For each cycle, LBR did not differ according to BMI category. Adjusted odds ratios (95% confidence intervals) for obtaining a live birth at the first cycle were 1.11 (0.78&ndash, 1.58) for overweight, 1.17 (0.70&ndash, 1.95) for class I obese, and 1.05 (0.48&ndash, 2.31) for class II/III obese women, as compared with normal-weight women. Similarly, no significant associations were found at cycles 2, 3, and 4. Cumulative LBR increased with the number of cycles, independently of the BMI class (p log-rank = 0.91). After adjustment, obesity status did not impact significantly the miscarriage rate, regardless of the cycle. In conclusion, neither women obesity nor its severity impacted the cumulative LBR after IVF.

International audience; OBJECTIVE: To compare frozen-thawed embryo transfer (FET) outcomes in natural cycles according to ovulation induction: spontaneous versus recombinant human chorionic gonadotrophin (r-hCG) triggering. METHODS: This retrospective study included all patients monitored for natural cycle FET during one year. When serial monitoring were performed until spontaneous LH rise, patients were included in group A (n=38) whereas those receiving r-hCG for ovulation triggering formed group B (n=43). All embryos had been cryopreserved by a vitrification method following a previous IVF cycle. No luteal phase support had been given. We compared outcomes between the 2 groups. RESULTS: After checking groups comparability, we didn't find significant difference for the implantation rate, clinical pregnancy rate and live birth (31% vs 45%, 32% vs 51% et 21% vs 32%, respectively for group A and B). The number of monitoring was significantly lower in group B (1,9\textpm0,8 versus 2,5\textpm1, P=0,006). DISCUSSION: Although no consensus has been yet established, natural cycle seems indicated for normo-ovulating patients but the question of ovulation induction is still debated. In our study, triggering ovulation by r-hCG, respecting strict criteria, seems provide good results while reducing both protocol's constraints and cost.

International audience; The aim of this study was to update our acknowledgment if there is a link between assisted embryo cryopreservation and epigenetics in human? Animal studies have demonstrated epigenetics consequence and especially imprinting disorders due to in vitro culture. In human, it is important to note that after frozen embryo transfer birth weight is significantly increased by 81 to 250g. But these studies cannot identify the reasons of such difference. This review strongly suggests that embryo cryopreservation is responsible for birth weight variations but mechanisms not yet elucidated. Epigenetics is probably one of these but to date, none study is able to prove it. We have to be attentive on a possible link between assisted reproductive technology (ART) and epigenetics reprogrammation.

International audience; OBJECTIVES: The objectives of this study were to assess (1) the expected cost of a live birth (LB) after in vitro fertilization with two different gonadotropin treatments [high purified human menopausal gonadotropin (HP-hMG) and recombinant follicle-stimulating hormone (rFSH)] as the single cost variable, and (2) the cost effectiveness of HP-hMG relative to rFSH in the context of the routine practice of assisted reproductive technology (ART) in France. METHODS: A Markov model was developed to simulate the therapeutic management, the in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) courses, and the effects of complications in hypothetical cohorts of 30,000 patients undergoing IVF/ICSI with fresh embryo transfer (up to four attempts) using data from the MERIT and MEGASET clinical trials or from French routine ART practice. RESULTS: The cost per LB was estimated at \texteuro12,145 and at \texteuro14,247 with HP-hMG and rFSH, respectively, using efficacy data from published clinical trials. The resulting incremental cost-effectiveness ratio (ICER) was -~\texteuro11,616 per LB. HP-hMG was less expensive by around \texteuro15.0 million and more effective by 1289 additional LBs. Using French clinical data, the cost per LB was \texteuro16,415 and \texteuro18,7531 with HP-hMG and rFSH, respectively. The ICER for HP-hMG versus rFSH was estimated at -~\texteuro7,719 per LB with a saving of about \texteuro8.54 million and 1097 additional LBs. Deterministic sensitivity analyses showed that the main ICER drivers were the LB rate, followed by the total gonadotropin doses. The probabilistic sensitivity analysis indicated that HP-hMG was the dominant strategy in 71.2% of cases using the clinical trial data and in 50.2% of cases using the French data. CONCLUSION: This analysis indicates that compared with rFSH, HP-hMG is less costly for IVF/ICSI management from the French healthcare payer's viewpoint. The results of the present Markov model analysis are consistent with previous findings in other European countries.

International audience; DNA mismatch repair (MMR) is an evolutionarily conserved process that corrects DNA polymerase errors during replication to maintain genomic integrity. In E. coli, the DNA helicase UvrD is implicated in MMR, yet an analogous helicase activity has not been identified in eukaryotes. Here, we show that mammalian MCM9, a protein involved in replication and homologous recombination, forms a complex with MMR initiation proteins (MSH2, MSH3, MLH1, PMS1, and the clamp loader RFC) and is essential for MMR. Mcm9−/− cells display microsatellite instability and MMR deficiency. The MCM9 complex has a helicase activity that is required for efficient MMR since wild-type but not helicase-dead MCM9 restores MMR activity in Mcm9−/− cells. Moreover, MCM9 loading onto chromatin is MSH2-dependent, and in turn MCM9 stimulates the recruitment of MLH1 to chromatin. Our results reveal a role for MCM9 and its helicase activity in mammalian MMR.

International audience; The frequency of polycystic ovary syndrome (PCOS) and the consequent fertility disorders cause many difficulties in the management of the assisted reproductive technics. Some studies are focused on different additional treatments, stimulation protocols or techniques that could optimize the in vitro fertilization cycles. The quality of the oocytes and embryos of these patients is also an outstanding issue. They remain difficult to actually evaluate during management, and none of the few published studies on this subject demonstrated any inferiority, compared to control patients. However, many differences have been highlighted, studying intra- and extra-ovarian factors. The advent of new genetic techniques could allow a better understanding of the pathophysiological mechanisms of the syndrome, as well as refining the evaluation of oocytes and embryos, in order to better predict the results of in vitro fertilization attempts. Pregnancy and birth rates, however, appear to be comparable to those of the general population.

In human eggs, aneuploidy increases with age and can result in infertility and genetic diseases. Studies in mouse oocytes suggest that reduced centromere cohesion and spindle assembly checkpoint (SAC) activity could be at the origin of chromosome missegregation. Little is known about these two features in humans. Here, we show that in human eggs, inter-kinetochore distances of bivalent chromosomes strongly increase with age. This results in the formation of univalent chromosomes during metaphase I (MI) and of single chromatids in metaphase II (MII). We also investigated SAC activity by checking the localization of BUB1 and BUBR1. We found that they localize at the kinetochore with a similar temporal timing than in mitotic cells and in a MPS1-dependent manner, suggesting that the SAC signalling pathway is active in human oocytes. Moreover, our data also suggest that this checkpoint is inactivated when centromere cohesion is lost in MI and consequently cannot inhibit premature sister chromatid separation. Finally, we show that the kinetochore localization of BUB1 and BUBR1 decreases with the age of the oocyte donors. This could contribute to oocyte aneuploidy.

We respond to the comments made in the BBC commissioned article by Heneghan and colleagues and the Panorama programme by Deborah Cohen about pre-implantation genetic screening (PGS), which was among the three “add on” treatments highlighted in the programme and the 41 listed in the article.12 Currently an extensive evidence base supports the efficacy of PGS: more than 20 retrospective studies and four randomised controlled trials suggest that, if performed to a high standard, PGS can, and does, improve IVF success for some patient groups.34567 We accept, however, that all studies are open to criticism and thus support further investigations, randomised and retrospective. However, the programme, …

Male infertility is a devastating problem that affects many couples worldwide. However, the molecular mechanisms and causes of idiopathic male infertility remain unclear. Circulating cell-free nucleic acids have an important role in human physiology and emerging evidence suggests that they play a role in male infertility. This review summarizes recent results on cell-free and intracellular nucleic acids in male infertility and discusses their potential use as biomarkers of male infertility in the clinical practice.L’infertilité masculine est un problème qui touche de nombreux couples. Cependant, aujourd’hui les mécanismes moléculaires et les causes de l’infertilité masculine idiopathique ne sont pas élucidés. Les acides nucléiques circulant ont un rôle important dans la physiologie et des évidences suggèrent qu’ils jouent un rôle dans l’infertilité masculine. L’objectif de cette revue est de mettre en avant les nouvelles avancées scientifiques sur les acides nucléiques circulant et non-circulant en lien avec l’infertilité masculine et de fournir une vue d’ensemble de leurs utilisation comme biomarqueurs en pratique clinique.

Several studies suggest a decrease in sperm quality in men in the last decades. Therefore, the aim of this work was to assess the influence of male factors (sperm quality and paternal age) on the outcomes of conventional in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI).This retrospective study included all couples who underwent IVF or ICSI at Montpellier University Hospital, France, between 1 January 2010 and 31 December 2015. Exclusion criteria were cycles using surgically retrieved sperm or frozen sperm, with pre-implantation genetic diagnosis or using frozen oocytes. The primary outcomes were the blastulation rate (number of blastocysts obtained at day 5 or day 6/number of embryos in prolonged culture at day 3) and the clinical pregnancy rate. The secondary outcomes were the fertilization and early miscarriage rates.In total, 859 IVF and 1632 ICSI cycles were included in this study. The fertilization rate after ICSI was affected by oligospermia. Moreover, in ICSI, severe oligospermia (lower than 0.2 million/ml) led to a reduction of the blastulation rate. Reduced rapid progressive motility affected particularly IVF, with a decrease of the fertilization rate and number of embryos at day 2 when progressive motility was lower than 32%. Paternal age also had a negative effect. Although it was difficult to eliminate the bias linked to the woman's age, pregnancy rate was reduced in IVF and ICSI when the father was older than 51 and the mother older than 37 years.These results allow adjusting our strategies of fertilization technique and embryo transfer. In the case of severe oligospermia, transfer should be carried out at the cleaved embryo stage (day 2-3) due to the very low blastulation rate. When the man is older than 51 years, couples should be aware of the reduced success rate, especially if the woman is older than 37 years. Finally, promising research avenues should be explored, such as the quantification of free sperm DNA, to optimize the selection of male gametes.De nombreuses données suggèrent une altération des paramètres spermatiques ces dernières décennies. Le but de ce travail est d’évaluer l’impact de facteurs masculins tels la qualité du sperme et l’âge paternel sur les résultats en fécondation in vitro classique (FIVc) et en fécondation in vitro avec injection intra-cytoplasmique de spermatozoïde (ICSI).L’étude a porté sur l’ensemble des couples ayant fait l’objet d’une tentative de FIVc ou d’ICSI entre le 1En ICSI, le taux de fécondation est affecté par l’oligospermie. Par ailleurs, une oligospermie extrême (inférieure à 0,2 M/ml) entraîne une diminution du taux de blastulation. La mobilité progressive avant préparation a plus d’impact en FIVc, où les taux de fécondation et le nombre d’embryons obtenus à J2 vont être plus bas lorsque la mobilité progressive est inférieure à 32%. Même s’il est difficile d’éliminer le biais lié à l’âge de la partenaire, il semblerait qu’il y ait une diminution du taux de grossesse en FIVc et en ICSI à partir de 51 ans chez l’homme avec une partenaire âgée de plus de 37 ans.Ces résultats permettront essentiellement d’ajuster nos stratégies de choix de technique de mise en fécondation et de transfert. Pour les oligospermies extrêmes, il semble préférable de proposer un transfert précoce au stade embryon clivé (J2 - J3) car le taux de blastulation est très réduit dans ce cas. Lorsque l’homme est âgé, il faudra également informer le couple de la diminution des taux de réussite, d’autant plus si sa partenaire a plus de 37 ans. Enfin, différentes pistes prometteuses de recherche sont encore à explorer, comme le dosage de l’ADN libre spermatique afin d’optimiser la sélection des gamètes masculins et ainsi améliorer les résultats en AMP.

International audience; Background: Proper folliculogenesis is fundamental to obtain a competent oocyte that, once fertilized, can support the acquisition of embryo developmental competence and pregnancy. MicroRNAs (miRNAs) are crucial regulators of folliculogenesis, which are expressed in the cumulus-oocyte complex and in granulosa cells and some can also be found in the bloodstream. These circulating miRNAs are intensively studied and used as diagnostic/prognostic markers of many diseases, including gynecological and pregnancy disorders. In addition, serum contains small amounts of cell-free DNA (cfDNA), presumably resulting from the release of genetic material from apoptotic/necrotic cells. The quantification of nucleic acids in serum samples could be used as a diagnostic tool for female infertility.Methods: An overview of the published literature on miRNAs, and particularly on the use of circulating miRNAs and cfDNA as non-invasive biomarkers of gynecological diseases, was performed (up to January 2014).Results: In the past decade, cell-free nucleic acids have been studied for potential use as biomarkers in many diseases, particularly in gynecological cancers, ovarian and endometrial disorders, as well as in pregnancy-related pathologies and fetal aneuploidy. The data strongly suggest that the concentration of cell-free nucleic acids in serum from IVF patients or in embryo culture medium could be related to the ovarian hormone status and embryo quality, respectively, and be used as a non-invasive biomarker of IVF outcome.Conclusions: The profiling of circulating nucleic acids, such as miRNAs and cfDNA, opens new perspectives for the diagnosis/prognosis of ovarian disorders and for the prediction of IVF outcomes, namely (embryo quality and pregnancy).

ObjectivesThe surgical management of advanced ovarian cancer involves complex surgery. Implementation of a quality management program has a major impact on survival. The goal of this work was to develop a list of quality indicators (QIs) for advanced ovarian cancer surgery that can be used to audit and improve the clinical practice. This task has been carried out under the auspices of the European Society of Gynaecologic Oncology (ESGO).MethodsQuality indicators were based on scientific evidence and/or expert consensus. A 4-step evaluation process included a systematic literature search for the identification of potential QIs and the documentation of scientific evidence, physical meetings of an ad hoc multidisciplinarity International Development Group, an internal validation of the targets and scoring system, and an external review process involving physicians and patients.ResultsTen structural, process, or outcome indicators were selected. Quality indicators 1 to 3 are related to achievement of complete cytoreduction, caseload in the center, training, and experience of the surgeon. Quality indicators 4 to 6 are related to the overall management, including active participation to clinical research, decision-making process within a structured multidisciplinary team, and preoperative workup. Quality indicator 7 addresses the high value of adequate perioperative management. Quality indicators 8 to 10 highlight the need of recording pertinent information relevant to improvement of quality. An ESGO-approved template for the operative report has been designed. Quality indicators were described using a structured format specifying what the indicator is measuring, measurability specifications, and targets. Each QI was associated with a score, and an assessment form was built.ConclusionsThe ESGO quality criteria can be used for self-assessment, for institutional or governmental quality assurance programs, and for the certification of centers. Quality indicators and corresponding targets give practitioners and health administrators a quantitative basis for improving care and organizational processes in the surgical management of advanced ovarian cancer.

Circulating or “extracellular” microRNAs (miRNAs) detected in biological fluids, could be used as potential diagnostic and prognostic biomarkers of several disease, such as cancer, gynecological and pregnancy disorders. However, their contributions in female infertility and in vitro fertilization (IVF) remain unknown. This study investigated the expression profiles of five circulating miRNAs (let-7b, miR-29a, miR-30a, miR-140 and miR-320a) in human follicular fluid from 91 women with normal ovarian reserve and 30 with polycystic ovary syndrome (PCOS) and their ability to predict IVF outcomes. The combination of FF miR-30a, miR-140 and let-7b expression levels discriminated between PCOS and normal ovarian reserve with a specificity of 83.8% and a sensitivity of 70% (area under the ROC curve, AUC = 0.83 [0.73–0.92]; p 2 (p = 0.04). Moreover, FF let-7b predicted the development of expanded blastocysts with 70% sensitivity and 64.3% specificity (AUC = 0.67 [0.54–0.79]; p = 0.02) and FF miR-29a potential to predict clinical pregnancy outcome reached 0.68 [0.55–0.79] with a sensitivity of 83.3% and a specificity of 53.5% (p = 0.01). Therefore, these miRNAs could provide new helpful biomarkers to facilitate personalized medical care during IVF.

International audience

International audience; Screening with breast ultrasound in combination with mammography is needed to investigate a clinical breast mass (Grade B), colored single-pore breast nipple discharge (Grade C), or mastitis (Grade C). The BI-RADS system is recommended for describing and classifying abnormal breast imaging findings. For a breast abscess, a percutaneous biopsy is recommended in the case of a mass or persistent symptoms (Grade C). For mastalgia, when breast imaging is normal, no MRI or breast biopsy is recommended (Grade C). Percutaneous biopsy is recommended for a BI-RADS category 4-5 mass (Grade B). For persistent erythematous nipple or atypical eczema lesions, a nipple biopsy is recommended (Grade C). For distortion and asymmetry, a vacuum core-needle biopsy is recommended due to the risk of underestimation by simple core-needle biopsy (Grade C). For BI-RADS category 4-5 microcalcifications without any ultrasound signal, a minimum 11-G vacuum core-needle biopsy is recommended (Grade B). In the absence of microcalcifications on radiography cores additional samples are recommended (Grade B). For atypical ductal hyperplasia, atypical lobular hyperplasia, lobular carcinoma in situ, flat epithelial atypia, radial scar and mucocele with atypia, surgical excision is commonly recommended (Grade C). Expectant management is feasible after multidisciplinary consensus. For these lesions, when excision margins are not clear, no new excision is recommended except for LCIS characterized as pleomorphic or with necrosis (Grade C). For grade 1 phyllodes tumor, surgical resection with clear margins is recommended. For grade 2 phyllodes tumor, 10 mm margins are recommended (Grade C). For papillary breast lesions without atypia, complete disappearance of the radiological signal is recommended (Grade C). For papillary breast lesions with atypia, complete surgical excision is recommended (Grade C)

Mesenchymal stem cells (MSC) are multipotent cells with great potential in therapy, reflected by more than 500 MSC-based clinical trials registered with the NIH. MSC are derived from multiple tissues but require invasive harvesting and imply donor-to-donor variability. Embryonic stem cell-derived MSC (ESC-MSC) may provide an alternative, but how similar they are to ex vivo MSC is unknown. Here we performed an in depth characterization of human ESC-MSC, comparing them to human bone marrow-derived MSC (BM-MSC) as well as human embryonic stem cells (hESC) by transcriptomics (RNA-seq) and quantitative proteomics (nanoLC-MS/MS using SILAC). Data integration highlighted and validated a central role of vesicle-mediated transport and exosomes in MSC biology and also demonstrated, through enrichment analysis, their versatility and broad application potential. Particular emphasis was placed on comparing profiles between ESC-MSC and BM-MSC and assessing their equivalency. Data presented here shows that differences between ESC-MSC and BM-MSC are similar in magnitude to those reported for MSC of different origin and the former may thus represent an alternative source for therapeutic applications. Finally, we report an unprecedented coverage of MSC CD markers, as well as membrane associated proteins which may benefit immunofluorescence-based applications and contribute to a refined molecular description of MSC.

Identifying genes where a variant allele is preferentially expressed in tumors could lead to a better understanding of cancer biology and optimization of targeted therapy. However, tumor sample heterogeneity complicates standard approaches for detecting preferential allele expression. We therefore developed a novel approach combining genome and transcriptome sequencing data from the same sample that corrects for sample heterogeneity and identifies significant preferentially expressed alleles. We applied this analysis to epithelial ovarian cancer samples consisting of matched primary ovary and peritoneum and lymph node metastasis. We find that preferentially expressed variant alleles include germline and somatic variants, are shared at a relatively high frequency between patients, and are in gene networks known to be involved in cancer processes. Analysis at a patient level identifies patient-specific preferentially expressed alleles in genes that are targets for known drugs. Analysis at a site level identifies patterns of site specific preferential allele expression with similar pathways being impacted in the primary and metastasis sites. We conclude that genes with preferentially expressed variant alleles can act as cancer drivers and that targeting those genes could lead to new therapeutic strategies.

International audience; A 31 year-old woman had a single embryo transfer on day 3 after assisted hatching (AH) and intracytoplasmic sperm injection (ICSI) treatment. Ultrasound examination performed 6 weeks after oocyte retrieval revealed a triplet pregnancy combining monochorionic diamniotic twins and a singleton. If zygote splitting resulting in monochorionic triamniotic triplets following IVF has already been described, this case is about an incredibly rare phenomenon after single embryo transfer. Naturally, a concurrent spontaneous conception cannot be excluded. To our knowledge, this is the first time a dichorionic triplet pregnancy after single embryo transfer is reported.

Identifying genes where a variant allele is preferentially expressed in tumors could lead to a better understanding of cancer biology and optimization of targeted therapy. However, tumor sample heterogeneity complicates standard approaches for detecting preferential allele expression. We therefore developed a novel approach combining genome and transcriptome sequencing data from the same sample that corrects for sample heterogeneity and identifies significant preferentially expressed alleles. We applied this analysis to epithelial ovarian cancer samples consisting of matched primary ovary and peritoneum and lymph node metastasis. We find that preferentially expressed variant alleles include germline and somatic variants, are shared at a relatively high frequency between patients, and are in gene networks known to be involved in cancer processes. Analysis at a patient level identifies patient-specific preferentially expressed alleles in genes that are targets for known drugs. Analysis at a site level identifies patterns of site specific preferential allele expression with similar pathways being impacted in the primary and metastasis sites. We conclude that genes with preferentially expressed variant alleles can act as cancer drivers and that targeting those genes could lead to new therapeutic strategies., Author Summary Identifying genes that contribute to cancer biology is complicated partly because cancers can have dozens of somatic mutations and thousands of germline variants. Somatic mutations are gene variants that arise after conception in an organism while germline variants are gene variants present at conception in an organism. Most methods to identify cancer drivers have focused on determining somatic mutations. In this study we attempt to identify, from a tumor sample, important germline and somatic variants by determining if a variant is expressed (made into RNA) more than expected from the amount of the variant in the genome. The preferred expression of a variant could benefit cancer cells. When applying our analysis to ovarian cancer samples we found that despite the apparent heterogeneity, different patients frequently share the same genes with preferentially expressed variants. These genes in many cases are known to affect cancer processes such as DNA repair, cell adhesion and cell signaling and are targetable with known drugs. We therefore conclude that our analysis can identify germline and somatic gene variants that contribute to cancer biology and can potentially guide individualized therapies.

Summary The ability to reliably express fluorescent reporters or other genes of interest is important for using human pluripotent stem cells (hPSCs) as a platform for investigating cell fates and gene function. We describe a simple expression system, designated GAPTrap (GT), in which reporter genes, including GFP, mCherry, mTagBFP2, luc2, Gluc, and lacZ are inserted into the GAPDH locus in hPSCs. Independent clones harboring variations of the GT vectors expressed remarkably consistent levels of the reporter gene. Differentiation experiments showed that reporter expression was reliably maintained in hematopoietic cells, cardiac mesoderm, definitive endoderm, and ventral midbrain dopaminergic neurons. Similarly, analysis of teratomas derived from GT-lacZ hPSCs showed that β-galactosidase expression was maintained in a spectrum of cell types representing derivatives of the three germ layers. Thus, the GAPTrap vectors represent a robust and straightforward tagging system that enables indelible labeling of PSCs and their differentiated derivatives., Graphical Abstract Image 1, Highlights • GAPTrap vector system targets transgenes to the ubiquitously expressed GAPDH locus • Targeting transgenes to the GAPDH locus yields reliable transgene expression • Transgenes at this locus are robustly expressed in differentiated cells • Generation of GAPTrap targeted human PSC lines is simple and efficient, In this article Stanley, Elefanty, and colleagues describe a simple vector system, GAPTrap, for constitutively expressing transgenes in pluripotent stem cells and their differentiated derivatives. GAPTrap vectors target transgenes to the GAPDH locus, ensuring robust and reliable expression in a wide variety of cell types, and providing an ideal base for cell tagging and gene overexpression experiments.

International audience; In assisted reproduction, about 30% of embryo implantation failures are related to inadequate endometrial receptivity. To identify molecules involved in endometrial receptivity acquisition, we investigated, using a SELDI-TOF approach, the protein expression profile of early-secretory and mid-secretory endometrium samples. Among the proteins upregulated in mid-secretory endometrium, we investigated the function of S100A10 in endometrial receptivity and implantation process. S100A10 was expressed in epithelial and stromal cells of the endometrium of fertile patients during the implantation windows. Conversely, it was downregulated in the mid-secretory endometrium of infertile patients diagnosed as non-receptive. Transcriptome analysis of human endometrial epithelial and stromal cells where S100A10 was silenced by shRNA revealed the deregulation of 37 and 256 genes, respectively, related to components of the extracellular matrix and intercellular connections. Functional annotations of these deregulated genes highlighted alterations of the leukocyte extravasation signaling and angiogenesis pathways that play a crucial role during implantation. S100A10 silencing also affected the migration of primary endometrial epithelial and stromal cells, decidualization and secretory transformation of primary endometrial stromal cells and epithelial cells respectively, and promoted apoptosis in serum-starved endometrial epithelial cells. Our findings identify S100A10 as a player in endometrial receptivity acquisition.

// Ikhlak Ahmed 1 , Thasni Karedath 1 , Simeon S. Andrews 1 , Iman K. Al-Azwani 2 , Yasmin Ali Mohamoud 2 , Denis Querleu 3 , Arash Rafii 4 , Joel A. Malek 1, 2 1 Department of Genetic medicine, Weill Cornell Medicine-Qatar, Education City, Ar-Rayyan, Qatar 2 Genomics Core, Weill Cornell Medicine-Qatar, Education City, Ar-Rayyan, Qatar 3 Department of Gynecologic Oncology, Universite Montepllier 1, Montpellier, France 4 Stem Cell and Microenvironment Laboratory, Weill Cornell Medicine-Qatar, Education City, Ar-Rayyan, Qatar Correspondence to: Joel A. Malek, e-mail: jom2042@qatar-med.cornell.edu Keywords: circular RNA, ovarian cancer, peritoneal metastasis, lymph node metastasis, miRNA Received: December 07, 2015 Accepted: April 02, 2016 Published: April 22, 2016 ABSTRACT Recently, a class of endogenous species of RNA called circular RNA (circRNA) has been shown to regulate gene expression in mammals and their role in cellular function is just beginning to be understood. To investigate the role of circRNAs in ovarian cancer, we performed paired-end RNA sequencing of primary sites, peritoneal and lymph node metastases from three patients with stage IIIC ovarian cancer. We developed an in-house computational pipeline to identify and characterize the circRNA expression from paired-end RNA-Seq libraries. This pipeline revealed thousands of circular isoforms in Epithelial Ovarian Carcinoma (EOC). These circRNAs are enriched for potentially effective miRNA seed matches. A significantly larger number of circRNAs are differentially expressed between tumor sites than mRNAs. Circular and linear expression exhibits an inverse trend for many cancer related pathways and signaling pathways like NFkB, PI3k/AKT and TGF-β typically activated for mRNA in metastases are inhibited for circRNA expression. Further, circRNAs show a more robust expression pattern across patients than mRNA forms indicating their suitability as biomarkers in highly heterogeneous cancer transcriptomes. The consistency of circular RNA expression may offer new candidates for cancer treatment and prognosis.

International audience; Purpose: Few studies have investigated the endometrial receptivity status of oocyte-donation (OD) recipient patients at the specific hormone replacement therapy (HRT)-cycle timing (5 to 6 days after progesterone administration) where embryos at blastocyst stage were mostly replaced. The aim of our study was to analyse, during the implantation window, (i) the global endometrial gene expression profile, and (ii) the endometrial receptivity exploration by the Win-Test® in OD recipient patients under HRT compared to spontaneous cycle in patients awaiting for IVF.Material and methods: This study included OD recipient patients without (n=7) or with (OD RIF, n=20) repeated implantation failures and 12 normal responder patients in spontaneous cycles used as control. Endometrial biopsies were performed during the peri-implantation phase. Samples were analysed using DNA microarrays and the endometrial gene expression profiles of HRT-treated OD recipient patients and of patients in spontaneous cycles were compared. Then, specific biomarkers of endometrial receptivity were assessed in the two groups of HRT-treated OD patients in comparison to control patientsResults: The global gene expression profile of peri-implantation endometrial samples from HRT-treated OD recipients and from patients in spontaneous cycles was different with significant alterations in the oestrogen receptor signalling [GTF2H2B, POLR2B, POLR2E], VEGF family ligand-receptor interactions [VEGFR1, VEGFB] and integrin signalling [ITGAL, PAK7, ILK]. Using specific biomarkers of human endometrial receptivity, we found that endometrium was non-receptive (29 % and 43 % in OD and RIF OD patients, respectively) or partially receptive (71 and 43 % in OD and RIF OD patients, respectively), at Pg+5/+6, in majority of HRT-treated patients. In OD RIF patients, a delay of the implantation window was observed. However, by targeting personalized embryos transfer by identifying for each patient the HRT-cycle day where endometrium is receptive, with respect of the synchronization of embryo-endometrium dialogue, high pregnancy rate per frozen-thawed embryo replacement (50%) was obtained in OD RIF patients.Conclusion: This study underlines the need to take into account the individual patient’s response to HRT cycles and to move to a patient-tailored care management.

International audience; Pluripotency is at the crossroads of stem cell research and biology of reproduction. The mature metaphase II oocyte contains the key factors for pluripotency induction and maintenance as assessed by its capacity to reprogram somatic nuclei. The cumulus cells (CCs) niche that surrounds the oocyte is crucial for its maturation and presumably for the oocyte to acquire its competence to confer pluripotency. In this study, we examined whether cells cultured from the human mature metaphase II oocyte CC niche (hCC) could be used as feeders for the propagation of human induced pluripotent stem cells. The induced pluripotent (iPS) cells cultured on hCC (hCC-iPS) were assessed for their pluripotency potential by their expression of pluripotency-associated genes such as Oct4, Nanog, and TRA1-60 and their competence to differentiate into the three germ layers in vitro (embryoid bodies) as well as in vivo (teratoma formation). We show that not only the hCC-iPS cells maintained their pluripotency potential, but they also exhibited much better self-renewal performance in terms of proliferation rate compared to the same cells cultured on human foreskin fibroblast (hFF) feeders (hFF-iPS). A comparative gene expression profile study of hCC and hFF revealed significant differences (P < 0.05) in expression of cellular matrix components and an upregulation in hCC of genes known to be important players in cell proliferation such as interleukin 6 gene (IL6).

International audience; Mutations of the PARK2 and PINK1 genes, encoding the cytosolic E3 ubiquitin-protein ligase Parkin and the mitochondrial serine/threonine kinase PINK1, respectively, cause autosomal recessive early-onset Parkinson's disease (PD). Parkin and PINK1 cooperate in a biochemical mitochondrial quality control pathway regulating mitochondrial morphology, dynamics and clearance. This study identifies the multifunctional PD-related mitochondrial matrix enzyme 17-β hydroxysteroid dehydrogenase type 10 (HSD17B10) as a new Parkin substrate. Parkin overproduction in cells increased mitochondrial HSD17B10 abundance by a mechanism involving ubiquitin chain extension, whereas PARK2 downregulation or deficiency caused mitochondrial HSD17B10 depletion in cells and mice. HSD17B10 levels were also found to be low in the brains of PD patients with PARK2 mutations. Confocal and Förster resonance energy transfer (FRET) microscopy revealed that HSD17B10 recruited Parkin to the translocase of the outer membrane (TOM), close to PINK1, both in functional mitochondria and after the collapse of mitochondrial membrane potential (ΔΨm). PD-causing PARK2 mutations impaired interaction with HSD17B10 and the HSD17B10-dependent mitochondrial translocation of Parkin. HSD17B10 overproduction promoted mitochondrial elongation and mitigated CCCP-induced mitochondrial degradation independently of enzymatic activity. These effects were abolished by overproduction of the fission-promiting dynamin-related protein 1 (Drp1). By contrast, siRNA-mediated HSD17B10 silencing enhanced mitochondrial fission and mitophagy. These findings suggest that the maintenance of appropriate mitochondrial HSD17B10 levels is one of the mechanisms by which Parkin preserves mitochondrial quality. The loss of this protective mechanism may contribute to mitochondrial dysfunction and neuronal degeneration in autosomal recessive PD.

International audience; OBJECTIVE:To identify risk factors for multiple pregnancies in intrauterine insemination (IUI) cycles with recombinant follicle-stimulating hormone (r-FSH).STUDY DESIGN:A retrospective study including 205 IUI cycles with r-FSH which led to clinical pregnancies was conducted. A total of 145, singleton pregnancies and 60 multiple pregnancies were compared according to clinical characteristics and parameters of ovarian stimulation and IUI procedure. The relationships between size and number of follicles and serum estradiol (E2) levels and the risk of multiple pregnancies were investigated using multiple logistic regression analyses.RESULTS:The means of infertility length, serum E2 levels, the number of follicles 10 mm, 12 mm, and ≥ 16 mm, and the number of intermediate follicles (from 12 to ≤ 15 mm) at the day of ovulation triggering were significantly higher in the multiple pregnancy group as compared to in the singleton pregnancy group (p < 0.05). We first demonstrated that high E2 levels (≥ 1,000 pg/nL) and the number of intermediate follicles represent 2 independent and significant risk factors for multiple gestation in IUI cycles that used ovarian stimulation by r-FSH (p = 0.002 and p = 0.007, respectively).CONCLUSION:This study shows that high E2 levels and the number of intermediate follicles, independently of large follicles, can predict an increased risk of multiple pregnancy in r-FSH IUI cycles.

International audience; Breast inflammation, excluding breast-feeding and pregnancy, is a rare breast pathology. We conducted a PubMed database search of all studies focusing on mastitis or breast inflammation exploration. While the most frequent aetiologies are infectious and inflammatory, inflammatory breast cancer can be diagnosed (LE2). Aetiologic diagnostic is difficult due to the absence of any clinical and imaging specific signs (LE3). The presence of mass, suspect lymph nodes or skin thickening in a woman older than 40 years old should orient toward inflammatory breast cancer (LE3). A suspect lesion must lead to perform a biopsy under sonography (grade A). In the absence of evidence for a malignant pathology after initial evaluation, we recommend starting an antibiotic treatment (grade C) with a clinical follow-up at the end of the treatment (grade B). If the symptoms persist, we recommend a new imaging (± MRI) (grade C) and a biopsy (grade C). Benign inflammatory pathologies may require a biopsy to exclude an inflammatory breast cancer and precise the diagnosis. Their specific management and treatment are presented in detail in the following chapters and may involve steroids.

International audience; Circulating nucleic acids (cell-free DNA and microRNAs) have for particularity to be easily detectable in the biological fluids of the body. Therefore, they constitute biomarkers of interest in female and male infertility care. Indeed, in female, they can be used to detect ovarian reserve disorders (polycystic ovary syndrome and low functional ovarian reserve) as well as to assess follicular microenvironment quality. Moreover, in men, their expression levels can vary in case of spermatogenesis abnormalities. Finally, circulating nucleic acids have also the ability to predict successfully the quality of in vitro embryo development. Their multiple contributions during assisted reproductive technology (ART) make of them biomarkers of interest, for the development of new diagnostic and/or prognostic tests, applied to our specialty. Circulating nucleic acids would so offer the possibility of personalized medical care for infertile couples in ART.

International audience