Which clinical and embryological factors should be considered to apply double embryo transfer (DET) instead of elective single embryo transfer (eSET)? No clinical or embryological factor per se justifies a recommendation of DET instead of eSET in IVF/ICSI. DET is correlated with a higher rate of multiple pregnancy, leading to a subsequent increase in complications for both mother and babies. These complications include preterm birth, low birthweight, and other perinatal adverse outcomes. To mitigate the risks associated with multiple pregnancy, eSET is recommended by international and national professional organizations as the preferred approach in ART. The guideline was developed according to the structured methodology for development and update of ESHRE guidelines. Literature searches were performed in PUBMED/MEDLINE and Cochrane databases, and relevant papers published up to May 2023, written in English, were included. Live birth rate, cumulative live birth rate, and multiple pregnancy rate were considered as critical outcomes. Based on the collected evidence, recommendations were discussed until a consensus was reached within the Guideline Development Group (GDG). A stakeholder review was organized after the guideline draft was finalized. The final version was approved by the GDG and the ESHRE Executive Committee. The guideline provides 35 recommendations on the medical and non-medical risks associated with multiple pregnancies and on the clinical and embryological factors to be considered when deciding on the number of embryos to transfer. These recommendations include 25 evidence-based recommendations, of which 24 were formulated as strong recommendations and one as conditional, and 10 good practice points. Of the evidence-based recommendations, seven (28%) were supported by moderate-quality evidence. The remaining recommendations were supported by low (three recommendations; 12%), or very low-quality evidence (15 recommendations; 60%). Owing to the lack of evidence-based research, the guideline also clearly mentions recommendations for future studies. The guideline assessed different factors one by one based on existing evidence. However, in real life, clinicians’ decisions are based on several prognostic factors related to each patient’s case. Furthermore, the evidence from randomized controlled trials is too scarce to formulate high-quality evidence-based recommendations. The guideline provides health professionals with clear advice on best practice in the decision-making process during IVF/ICSI, based on the best evidence currently available, and recommendations on relevant information that should be communicated to patients. In addition, a list of research recommendations is provided to stimulate further studies in the field. The guideline was developed and funded by ESHRE, covering expenses associated with the guideline meetings, the literature searches, and the dissemination of the guideline. The guideline group members did not receive payment. DPB declared receiving honoraria for lectures from Merck, Ferring, and Gedeon Richter. She is a member of ESHRE EXCO, and the Mediterranean Society for reproductive medicine and the president of the Croatian Society for Gynaecological Endocrinology and Reproductive Medicine. CDG is the past Chair of the ESHRE EIM Consortium and a paid deputy member of the Editorial board of Human Reproduction. IR declared receiving reimbursement from ESHRE and EDCD for attending meetings. She holds an unpaid leadership role in OBBCSSR, ECDC Sohonet, and AER. KAR-W declared receiving grants for clinical researchers and funding provision to the institution from the Swedish Cancer Society (200170F), the Senior Clinical Investigator Award, Radiumhemmets Forskningsfonder (Dnr: 201313), Stockholm County Council FoU (FoUI-953912) and Karolinska Institutet (Dnr 2020-01963), NovoNordisk, Merck and Ferring Pharmaceuticals. She received consulting fees from the Swedish Ministry of Health and Welfare. She received honoraria from Roche, Pfizer, and Organon for chairmanship and lectures. She received support from Organon for attending meetings. She participated in advisory boards for Merck, Nordic countries, and Ferring. She declared receiving time-lapse equipment and grants with payment to institution for pre-clinical research from Merck pharmaceuticals and from Ferring. SS-R received research funding from Roche Diagnostics, Organon/MSD, Theramex, and Gedeo-Richter. He received consulting fees from Organon/MSD, Ferring Pharmaceuticals, and Merck Serono. He declared receiving honoraria for lectures from Ferring Pharmaceuticals, Besins, Organon/MSD, Theramex, and Gedeon Richter. He received support for attending Gedeon Richter meetings and participated in the Data Safety Monitoring Board of the T-TRANSPORT trial. He is the Deputy of ESHRE SQART special interest group. He holds stock options in IVI Lisboa and received equipment and other services from Roche Diagnostics and Ferring Pharmaceuticals. KT declared receiving payment for honoraria for giving lectures from Merck Serono and Organon. She is member of the safety advisory board of EDQM. She holds a leadership role in the ICCBBA board of directors. ZV received reimbursement from ESHRE for attending meetings. She also received research grants from ESHRE and Juhani Aaltonen Foundation. She is the coordinator of EHSRE SQART special interest group. The other authors have no conflicts of interest to declare. This guideline represents the views of ESHRE, which were achieved after careful consideration of the scientific evidence available at the time of preparation. In the absence of scientific evidence on certain aspects, a consensus between the relevant ESHRE stakeholders has been obtained. Adherence to these clinical practice guidelines does not guarantee a successful or specific outcome, nor does it establish a standard of care. Clinical practice guidelines do not replace the need for application of clinical judgement to each individual presentation, nor variations based on locality and facility type. ESHRE makes no warranty, express or implied, regarding the clinical practice guidelines and specifically excludes any warranties of merchantability and fitness for a particular use or purpose (full disclaimer available at https://www.eshre.eu/Guidelines-and-Legal).

Among the possible risk factors for male reproduction, exposure to phthalates and alkylphenols is widely documented. This study evaluated the possible association between chemical exposure and the quality of the seminal fluid of 105 subjects in a fertility clinic. The urinary levels of seven phthalate metabolites (monoethylphthalate, MEP; monobenzylphthalate, MBzP; mono n-butylphthalate, MnBP; mono-(2-ethylhexyl) phthalate, MEHP; mono(2-ethyl-5-hydroxyhexyl) phthalate, MEHHP; mono-n-octylphthalate, MnOP; mono-isononylphthalate, MiNP) and bisphenol A (BPA), were analysed by high performance liquid chromatography/tandem mass spectrometry HPLC/MS/MS. The regression analysis showed that the semen volume was positively associated with MnBP, MnOP and BPA levels while was negatively associated with MiNP levels. The sperm concentration had a significant inverse relationship with MEP levels. A negative association was found between the use of plastic containers for food storage (p = 0.037) and semen volume (3.06 vs. 2.30 mL as average values, never vs daily). A significant positive correlation emerged (p < 0.005) between the consumption of canned food and the levels of BPA (2.81 vs. 0.14 µg/g creat as average values, daily vs. never) and between the use of perfumes and levels of MEP (389.86 vs. 48.68 µg/g creat, as average values, daily vs. never). No further statistically significant associations were found, even considering the working activity. Some evidence emerged about the possible link between exposure and seminal fluid quality: further case/control or prospective studies will allow us to confirm this causality hypothesis.

MiR-206, a member of the so-called myomiR family, is largely acknowledged as a specific, positive regulator of skeletal muscle differentiation. A growing body of evidence also suggests a tumor suppressor function for miR-206, as it is frequently downregulated in various types of cancers. In this study, we show that miR-206 directly targets cyclin D1 and contributes to the regulation of CCND1 gene expression in both myogenic and non-muscle, transformed cells. We demonstrate that miR-206, either exogenous or endogenous, reduces cyclin D1 levels and proliferation rate in C2C12 cells without promoting differentiation, and that miR-206 knockdown in terminally differentiated C2C12 cells leads to cyclin D1 accumulation in myotubes, indicating that miR-206 might be involved in the maintenance of the post-mitotic state. Targeting of cyclin D1 might also account, at least in part, for the tumor-suppressor activity suggested for miR-206 in previous studies. Accordingly, the analysis of neoplastic and matched normal lung tissues reveals that miR-206 downregulation in lung tumors correlates, in most cases, with higher cyclin D1 levels. Moreover, gain-of-function experiments with cancer-derived cell lines and with in vitro transformed cells indicate that miR-206-mediated cyclin D1 repression is directly coupled to growth inhibition. Altogether, our data highlight a novel activity for miR-206 in skeletal muscle differentiation and identify cyclin D1 as a major target that further strengthens the tumor suppressor function proposed for miR-206.

Growing evidence supports a role of vitamin D (VD) in reproductive health. Vitamin D receptor (VDR) is expressed in the ovary, endometrium, and myometrium. The biological actions of VD in fertility and reproductive tissues have been investigated but mainly using animal models. Conversely, the molecular data addressing the mechanisms underlying VD action in the physiologic endometrium and in endometrial pathologies are still scant. Levels of VDR expression according to the menstrual cycle are yet to be definitively clarified, possibly being lower in the proliferative compared to the secretory phase and in mid-secretory compared to early secretory phase. Endometrial tissue also expresses the enzymes involved in the metabolism of VD. The potential anti-proliferative and anti-inflammatory effects of VD for the treatment of endometriosis have been investigated in recent years. Treatment of ectopic endometrial cells with 1,25(OH)₂D₃ could significantly reduce cytokine-mediated inflammatory responses. An alteration of VD metabolism in terms of increased 24-hydroxylase mRNA and protein expression has been demonstrated in endometrial cancer, albeit not consistently. The effect of the active form of the vitamin as an anti-proliferative, pro-apoptotic, anti-inflammatory, and differentiation-inducing agent has been demonstrated in various endometrial cancer cell lines.