High-quality human preimplantation embryos actively influence endometrial stromal cell migration.

PURPOSE: The purpose of this paper is to study whether human preimplantation embryos regulate endometrial stromal cell (hESC) migration. METHODS: Primary hESCs were isolated from fertile patients undergoing hysterectomy for benign conditions (uterine scar niche n = 3, dysmenorrhea n = 2; no hormonal treatment). Migration and proliferation assays were performed by culturing decidualized or non-decidualized hESCs in the presence of embryo conditioned medium (ECM) from high-quality embryos (fragmentation ≤ 20%) or from low-quality embryos (fragmentation > 20%) or in non-conditioned medium from the same dishes (control). ECM samples from 425 individually cultured human embryos were used in this study. RESULTS: ECM from high-quality embryos, i.e., with a low percentage of fragmentation, actively stimulated decidualized hESC migration (p < 0.001). This effect was consistent throughout embryonic development from cleavage stage embryos with 2-7 cells (high quality vs. control; p = 0.036), 8-18 cells (high quality vs. control; p < 0.001) to morulae (high quality vs. control; p = 0.003). Additionally, linear regression analysis showed that hESC migration was influenced by embryo quality (fragmentation, ß - 0.299; p = 0.025) and not developmental stage (cell number, ß 0.177; p = 0.176) or maternal age (ß - 0.036; p = 0.78). Opposite to decidualized hESCs, the migration response of non-decidualized hESCs was inhibited by ECM from high-quality embryos (p = 0.019). ECM from low-quality embryos, i.e., with a high percentage of fragmentation, did not cause an altered migration response in decidualized hESCs (p = 0.860) or non-decidualized hESCs (p = 0.986). Furthermore, ECM of both high- and low-quality human embryos did not influence the number of proliferating cells (p = 0.375) and the cell cycle time (p = 0.297) of non-decidualized or decidualized hESCs. CONCLUSION: This study reveals a mechanism by which high-quality human preimplantation embryos actively interact with the endometrium to increase their chances of successful implantation.

GATA4 represses an ileal program of gene expression in the proximal small intestine by inhibiting the acetylation of histone H3, lysine 27.

GATA4 is expressed in the proximal 85% of small intestine where it promotes a proximal intestinal ('jejunal') identity while repressing a distal intestinal ('ileal') identity, but its molecular mechanisms are unclear. Here, we tested the hypothesis that GATA4 promotes a jejunal versus ileal identity in mouse intestine by directly activating and repressing specific subsets of absorptive enterocyte genes by modulating the acetylation of histone H3, lysine 27 (H3K27), a mark of active chromatin, at sites of GATA4 occupancy. Global analysis of mouse jejunal epithelium showed a statistically significant association of GATA4 occupancy with GATA4-regulated genes. Occupancy was equally distributed between down- and up-regulated targets, and occupancy sites showed a dichotomy of unique motif over-representation at down- versus up-regulated genes. H3K27ac enrichment at GATA4-binding loci that mapped to down-regulated genes (activation targets) was elevated, changed little upon conditional Gata4 deletion, and was similar to control ileum, whereas H3K27ac enrichment at GATA4-binding loci that mapped to up-regulated genes (repression targets) was depleted, increased upon conditional Gata4 deletion, and approached H3K27ac enrichment in wild-type control ileum. These data support the hypothesis that GATA4 both activates and represses intestinal genes, and show that GATA4 represses an ileal program of gene expression in the proximal small intestine by inhibiting the acetylation of H3K27.

The addition of a low-quality embryo as part of a fresh day 3 double embryo transfer does not improve ongoing pregnancy rates.

STUDY QUESTION: Does the addition of a low-quality embryo in fresh Day 3 double embryo transfer (DET) affect the ongoing pregnancy rate (OPR) and multiple gestation rate in patients with only one or no high-quality embryos available? SUMMARY ANSWER: In patients with only one- or no high-quality embryo available, the addition of a low-quality embryo in fresh Day 3 DET does not improve the OPR but increases multiple gestation rates in fresh DET. WHAT IS KNOWN ALREADY: Pregnancy rates after DET are considered to be higher compared to single embryo transfer (SET) when analyzed per first embryo transfer only. However, these conclusions are based on RCTs in which mostly patients with two or more high-quality embryos were included, and can therefore not be applied to patients with only one or no high-quality embryo available. This is particularly relevant since it has been suggested that low-quality embryos could impair the implantation of simultaneously transferred embryos by paracrine signaling. Hence, we investigated in patients with only one or no high-quality embryo available whether the addition of a low-quality embryo in DET affects the OPR, multiple gestation rate and miscarriage rate. STUDY DESIGN SIZE DURATION: This was a retrospective cohort study of 5050 patients receiving 7252 fresh embryo transfers on Day 3 after fertilization in IVF/ICSI cycles from 2012 to 2015 in two academic hospitals. PARTICIPANTS/MATERIALS SETTING METHODS: We included all women that received fresh SET or DET with any combination of high-quality embryos (7, 8 or 9 blastomeres, with equal to or <20% fragmentation) or low-quality embryos (all other embryos). Outcomes were OPR (primary outcome, defined as a positive fetal heartbeat by transvaginal ultrasound at least 10 weeks after oocyte retrieval), miscarriage rate and multiple gestation rate. We used a generalized estimating equations model adjusting for maternal age, number of oocytes retrieved, center of treatment and the interaction between maternal age and number of oocytes retrieved. Other baseline characteristics, including infertility diagnosis, fertilization method and the number of consecutive fresh embryo transfers per patient, did not contribute significantly to the GEE model and were therefore excluded, and not adjusted for. MAIN RESULTS AND THE ROLE OF CHANCE: Compared to SET with one high-quality embryo, DET with two high-quality embryos resulted in a higher OPR (adjusted odds ratio (OR) 1.38, 95% CI 1.14-1.67), while DET with one high- and one low-quality embryo resulted in a lower OPR (adjusted OR 0.65, 95% CI 0.49-0.90). However, SET in patients with only one high-quality embryo available resulted in a lower OPR compared to SET in patients with two or more high-quality embryos available (adjusted OR 0.52, 95% CI 0.39-0.70). After adjusting for this confounding factor, we found that both DET with two high-quality embryos (adjusted OR 0.99, 95% CI 0.74-1.31) and DET with one high- and one low-quality embryo (adjusted OR 0.78, 95% CI 0.47-1.27) resulted in a not significantly different OPR compared to SET with one high-quality embryo. If only low-quality embryos were available, DET did not increase the OPR as compared to SET with one low-quality embryo (adjusted OR 0.84, 95% CI 0.55-1.28). Multiple gestation rates were higher in all DET groups compared to SET (DET with ≥1 high-quality embryo(s) compared to SET with one high-quality embryo; DET with two low-quality embryos compared to SET with one low-quality embryo; all comparisons P < 0.001). Miscarriage rates were not different in all DET groups compared to SET (DET with ≥1 high-quality embryo(s) compared to SET with one high-quality embryo; DET with two low-quality embryos compared to SET with one low-quality embryo; all comparisons P > 0.05). LIMITATIONS REASONS FOR CAUTION: Limitations to this study include the retrospective design and possible bias between study groups related to embryo transfer policies between 2012 and 2015. Consequently, we may have underestimated pregnancy chances in all DET groups. Furthermore, the OPR was calculated as a percentage of the number of fresh embryo transfers in each study group, and not the total number of started IVF/ICSI cycles. Therefore, the reported pregnancy outcomes may not truly reflect the pregnancy chances of couples at the start of treatment. A possible confounding effect of maternal age in our study is acknowledged but we could not compare clinical outcomes in different age groups separately owing to small sample sizes. Analysis of pregnancy outcomes in lower prognosis patients (higher maternal age, fewer oocytes retrieved) separately is an avenue for future research. WIDER IMPLICATIONS OF THE FINDINGS: The decision to perform DET rather than SET in order to increase the OPR per fresh embryo transfer seems not to be justified for those patients with only one or no high-quality embryo(s) available. However, owing to the limitations of this study, prospective RCTs are needed that specifically investigate pregnancy outcomes in patients with only one or no high-quality embryo(s) available in SET and DET. STUDY FUNDING/COMPETING INTERESTS: This study was funded by a grant from the joint Amsterdam Reproduction & Development Institute of the Academic Medical Center and VU University Medical Center (www.amsterdam-reproduction-and-development.org). The authors have no conflicts of interest to declare.