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Study Question: Is the microRNA (miRNA) expression pattern of cumulus oophorus cells (COCs) in women undergoing medically assisted reproduction (MAR) procedures differentially modulated according to patient age and gonadotropin treatment strategy?Summary Answer: Maternal age is an independent factor impacting miRNA expression in COCs while gonadotropin treatment may affect follicular miRNA expression and IVF efficacy.What Is Known Already: Epigenetic mechanisms in female infertility are complex and poorly studied. DNA methylation, histone modifications, miRNAs and nucleosome positioning influence cellular machinery through positive and negative feedback mechanisms either alone or interactively. miRNAs are important regulators during oogenesis, spermatogenesis and early embryogenesis, and are reported to play a role in regulating crosstalk between the oocyte and COCs. Although miRNome analysis has been performed in female human reproductive tissues (endometrium, myometrium, cervix and ovaries), epigenetic modifications in women with infertility have not been explored in detail. In addition, the impact of gonadotropin treatments during MAR on miRNA expression in COCs has not been fully investigated.Study Design, Size, Duration: This study was carried out in 53 COC samples obtained from mature metaphase II (MII) oocytes in 53 women undergoing MAR treatment. A total of 38 samples for assay development were pooled by maternal age and gonadotropin treatment into four predetermined subgroups: ≥36 years and recombinant human FSH (r-hFSH), n = 10; ≥36 years and r-hFSH+ recombinant human-luteinizing hormone (r-hLH), n = 10; ≤35 years and r-hFSH, n = 9; ≤35 years and r-hFSH+r-hLH, n = 9. miRNome profiles were determined and compared between subgroups. Expression of defined miRNAs was validated in the remaining fifteen samples, representative of each subgroup, by quantitative polymerase chain reaction (PCR).Participants/materials, Setting, Methods: COCs were processed for miRNA-enriched total RNA extraction and pooled in homogeneous subgroups to obtain a sufficient amount and quality of starting material to perform the analysis. Each pooled sample underwent miRNA profiling using PCR assay system to examine expression of 752 human miRNAs without pre-amplification. Data were analyzed using the delta-delta Ct method for relative quantitation and prediction of target genes (with at least four algorithms predicting the same miRNA-gene interaction pair (HIT)>4). The miRSystem database provided functional annotation enrichment (raw P-value <0.05) of co-expressed miRNAs.Main Results and the Role Of Chance: We found distinctive miRNA expression profiles in each subgroup correlating with age and MAR stimulation. In addition, a number of selective and co-expressed miRNAs were revealed by comparative analysis. A cluster of 37 miRNAs were commonly but differentially expressed in all four pools. Significant differences were observed in expression regulation of 37 miRNAs between age groups (≤35 or ≥36) in women receiving r-hFSH+r-hLH compared to those receiving r-hFSH alone. Higher concentrations and increased numbers of miRNAs were recorded in younger than in older patients, regardless of treatment. Functional and expression studies performed to retrieve common miRNome profiles revealed an enrichment of biological functions in oocyte growth and maturation, embryo development, steroidogenesis, ovarian hyperstimulation, apoptosis and cell survival, glucagon and lipid metabolism, and cell trafficking. The highest scored pathways of target genes of the 37 common miRNAs were associated with mitogen-activated protein kinase (MAPK) signaling pathways, G alpha signaling, transcription regulation, tight junctions, RNA polymerase I and III, and mitochondrial transcription. We identified a potential age- and MAR stimulation-dependent signature in the miRNA landscape of COCs.Limitations, Reasons For Caution: We cannot rule out the possibility that other unknown individual genetic or clinical factors may have interfered with the reported results. Since miRNA profiling was conducted with a predefined array of target probes, other miRNA molecules, potentially modulated by age and hormonal stimulation, may have been missed in this study.Wider Implications Of the Findings: miRNA expression in COCs is modulated by gonadotropin treatment and correlates strongly with age. A better understanding of the expression patterns and functions of miRNAs may lead to the development of novel therapeutics to treat ovarian dysfunction and improve fertility in older women.Study Funding/competing Interest: This study was funded by Merck KGaA, Darmstadt, Germany. All authors declared no competing interest, except SL and TD who are fully employed by Merck KGaA.Trial Registration Number: N/A. [ABSTRACT FROM AUTHOR]

Study Question: Can the Endometriosis Fertility Index (EFI) be estimated accurately before surgery?Summary Answer: The EFI can be estimated accurately based on mere clinical/ultrasound information, with some improvement after adding data from diagnostic laparoscopy.What Is Known Already: The EFI is a validated clinical instrument predicting the probability of pregnancy after endometriosis surgery without the use of ART. Being an end-of-surgery-score, it implies the decision for operative laparoscopy to be made in advance-hence, its role in the pre-surgical decision-making process remains to be established.Study Design, Size, Duration: Single-cohort prospective observational study in 82 patients undergoing complete endometriosis excision (between June and December 2016). Two methods were used to estimate the final EFI: type A based on non-surgical clinical/ultrasound findings only, and type B based on the combination of non-surgical clinical/ultrasound findings and diagnostic laparoscopy data. To calculate EFI type A, an algorithm was created to translate non-surgical clinical/imaging information into rASRM (revised American Society of Reproductive Medicine)-and EFI points. EFI type A and type B estimates were assessed for their clinical and numerical agreement with the final EFI score. Agreement was defined as clinical if EFI scores were within the same range (0-4, 5-6, 7-10), and numerical if their difference was ≤1.Participants/materials, Setting, Methods: All 82 patients underwent complete laparoscopic CO2-laser excision of any rASRM stage of endometriosis in the Leuven University Fertility Centre (LUFC) of University Hospitals Leuven, a tertiary referral centre for both endometriosis and infertility. An anonymized clinical research file was created. For each patient, three different data sets were created, in order to allow the estimation of the (surgical part) EFI and of the rASRM scores, defined as follows: 'Estimated type A' contained only non-surgical clinical/imaging data, 'Estimated type B' included type A information plus the information of the diagnostic laparoscopy and 'Final EFI' included information of type A, type B and all intra-operative information required to calculate the final EFI. To calculate EFI type A without surgical information, a set of rules was used to translate pre-surgical clinical/imaging information into (rASRM and EFI points). Scoring was done by one person (C.T.), with a time interval of 4 weeks between sessions for each EFI type. Next to the EFI, also rASRM score and stage were calculated.Main Results and the Role Of Chance: Agreement rate between estimated EFI type A and final EFI was high for both the clinical (0.915; 95% CI 0.832-0.965) and numerical definition (0.878; 95% CI 0.787-0.940). Agreement rates between estimated EFI type B and final EFI were even higher (clinical (0.988; 95% CI 0.934-1.000), numerical (0.963; 95% CI 0.897-0.992)).Limitations, Reasons For Caution: Type A estimation is dependent on high-level gynaecological ultrasound expertise, which may not be available in all clinics. A small number of patients had no prior clinical, ultrasound (hard markers) or surgical confirmation of the diagnosis of endometriosis. When applying the estimated EFI type A in clinical practice, a priori assumptions of the presence or absence of endometriosis will need to be made in adjunct to the estimation of the estimated type A EFI when counselling patients on the potential benefit of an (at least diagnostic) laparoscopy. The level of agreement for type A or B should also be taken into account when counselling patients on the type of efforts undertaken to attempt to diagnose or rule out endometriosis.Wider Implications Of the Findings: As this study reports, the EFI can be estimated accurately based on clinical/ultrasound data only without the need for any surgical data. This means that the EFI could be used as an instrument to guide joint physician-patient decision-making between surgery, ART or other fertility management options for the individualized treatment of women with endometriosis-related infertility.Study Funding/competing Interest(s): During this study period, C.T. was supported by FWO (Research Fund Flanders, Grant number 1700816N) and UZ Leuven KOF (University Hospitals Leuven, Klinisch Onderzoeksfonds).The LUFC received unrestricted research grants from Ferring Pharmaceuticals and Merck SA. Gedeon Richter and MSD sponsored travel to and attendance at scientific meetings. C.M. received consultancy fees from Lumenis (paid to KU Leuven, no private revenue). T.D. has been vice-president and head of global medical affairs infertility for the multinational pharmaceutical company Merck (Darmstadt, Germany) since 1 October 2015. He continues his academic appointment on a part-time basis as Professor of Reproductive Medicine at the University of Leuven (KU Leuven). T.D. has been vice-president and head of global medical affairs infertility for the multinational pharmaceutical company Merck (Darmstadt, Germany) since October 2015. He is also a Guest Professor in Reproductive Medicine and Biology at the Department of Development and Regeneration, Group Biomedical Sciences, KU Leuven (University of Leuven), Belgium, and an Adjunct Professor at the Department of Obstetrics and Gynecology in the University of Yale, New Haven, USA. This work was initiated before he joined Merck KGaA in October 2015, and completed during the subsequent years.Trial Registration Number: study registration number at UZ Leuven Clinical Trial Centre: S59221. [ABSTRACT FROM AUTHOR]