Endometrial extracellular vesicles of recurrent implantation failure patients inhibit the proliferation, migration, and invasion of HTR8/SVneo cells.

PURPOSE: Endometrial extracellular vesicles are essential in regulating trophoblasts' function. This study aims to investigate whether endometrial extracellular vesicles (EVs) from recurrent implantation failure (RIF) patients inhibit the proliferation, invasion, and migration of HTR8/SVneo cells. METHODS: Eighteen RIF patients and thirteen fertile women were recruited for endometria collection. Endometrial cells isolated from the endometria were cultured and modulated by hormones, and the conditioned medium was used for EV isolation. EVs secreted by the endometrial cells of RIF patients (RIF-EVs) or fertile women (FER-EVs) were determined by Western blotting, nanoparticle tracking analysis, and transmission electron microscopy. Fluorescence-labeled EVs were used to visualize internalization by HTR8/SVneo cells. RIF-EVs and FER-EVs were co-cultured with HTR8/SVneo cells. Cell Counting Kit-8, transwell invasion, and wound closure assays were performed to determine cellular proliferation, invasion, and migration, respectively, in different treatments. RESULTS: RIF-EVs and FER-EVs were bilayer membrane vesicles, ranging from 100 to 150 nm in size, that expressed the classic EV markers Alix and CD9. RIF-EVs and FER-EVs were internalized by HTR8/SVneo cells within 2 h. The proliferation rate in the FER-EV group was significantly higher than that in the RIF-EV group at 20 µg/mL. Moreover, the invasion and migration capacity of trophoblast cells were decreased in the RIF-EV group relative to the FER-EV group at 20 µg/mL. CONCLUSION: Endometrial EVs from RIF patients inhibited the functions of trophoblasts by decreasing their proliferation, migration, and invasive capacity. Such dysregulations induced by RIF-EVs may provide novel insights for better understanding the pathogenesis of implantation failure.

Peri-implantation cytokine profile differs between singleton and twin IVF pregnancies.

PROBLEM: It is unknown whether maternal cytokine production differs between twin and singleton gestations in the implantation phase. A difference in maternal serum cytokine concentrations in twins would imply a dose-response to the invading embryos, as opposed to a general immune reaction. METHOD OF STUDY: A prospective longitudinal cohort of women aged 18-45 at an academic fertility center undergoing in vitro fertilization and embryo transfer (IVF-ET) underwent routine collection of serial serum samples starting 9 days after ET and then approximately every 48 hours thereafter. Cryopreserved aliquots of these samples were assayed for interleukin-10 (IL-10), tumor necrosis factor-alpha (TNF-α), and C-X-C motif chemokine ligand 10 (CXCL10) using the SimplePlex immunoassay platform. Pregnancies were followed until delivery. Serial measures of serum concentrations of IL-10, CXCL10, and TNF-α in singleton or di-di twin pregnancies from 9 to 15 days after IVF-ET were compared. RESULTS: Maternal serum levels of CXCL10 are significantly lower in women with di-di twin pregnancies in early implantation compared to those with singleton gestation (day 9-11, P = .02). Serum levels of TNF-α and IL-10 were comparable at all studied time points (P > .05). CONCLUSION: Maternal serum levels of CXCL10 are significantly lower in the earliest implantation phase in di-di twins compared to singleton conceptions. Given the known anti-angiogenic role of CXCL10, we hypothesize that lower CXCL10 levels in twin implantations allow an environment that is conducive for the greater vascularization required for the establishment of dual placentation in di-di twins.

Bisphenol A-induced mechanistic impairment of decidualization.

Decidualization is a crucial precedent to embryo implantation, as its impairment is a major contributor to female infertility and pregnancy complications. Unraveling the molecular mechanisms involved in the impairment of decidualization has been a subject of interest in the field of reproductive medicine. Evidence from several experimental settings show that exposure to bisphenol A (BPA), an endocrine-disrupting chemical, affects the expression of several molecules that are involved in decidualization. Both low and high doses of BPA impair decidualization through the dysregulation of estrogen (ER) and progesterone (PR) receptors. Exposure to low doses of BPA leads to decreased levels and activities of several antioxidant enzymes, increased activity of endothelial nitric oxide synthase (eNOS), and increased production of nitric oxide (NO) via the upregulation of ER and PR. Consequently, oxidative stress is induced and decidualization becomes impaired. On the other hand, exposure to high doses of BPA downregulates ER and PR and impairs decidualization through two distinct pathways. One is through the upregulation of early growth response-1 (EGR1) via increased phosphorylation of extracellular signal-regulated protein kinases 1 and 2; and the other is through a reduced serum glucocorticoid-induced kinase-1 (SGK1)-mediated downregulation of epithelial sodium channel-α and the induction of oxidative stress. Thus, regardless of the dose, BPA can impair decidualization to trigger infertility and pregnancy complications. This warrants the need to adopt lifestyles that will decrease the tendency of getting exposed to BPA.

Under Arrest: The Embryo in Diapause.

Embryonic diapause is the reversible arrest in development of mammalian embryos at the blastocyst stage. In this issue of Developmental Cell, Hussein et al. (2020) reveal that alternative splicing of Lkb1 is essential for diapause to persist and find the elevation of glycolytic and lipolytic pathways that were previously considered dormant.

Exploring undiagnosed celiac disease in women with recurrent reproductive failure: The gluten-free diet could improve reproductive outcomes.

PROBLEM: Which is the prevalence and seroprevalence of celiac disease (CD) in women with recurrent reproductive failure? METHOD OF STUDY: Retrospective study performed in a single infertility clinic from September 2016 to December 2017. A total of 690 women with unexplained history of recurrent miscarriage and/or recurrent implantation failure were consecutively recruited. IgA anti-transglutaminase 2 (TG2) antibody data were collected, as well as IgG anti-TG2 and IgA/IgG anti-deamidated gluten peptide (DGP) data in most cases, and IgG anti-gliadin antibodies occasionally. In selected women, HLA-DQ genotyping was requested. Biopsy was suggested to all women with positive serological results or belonging to CD risk groups. Reproductive outcomes were recorded from women with high suspicion of CD and a control group comprised of 49 women. RESULTS: Anti-TG2-positive women comprised 1% of the sample. An additional 4% was observed considering less-specific antibodies (31 women). Only 39% of sero-positive women accepted duodenal biopsy. HLA and biopsy data discarded CD in 14 sero-positive cases (37%), only one with anti-TG2 antibodies. CD was suggested in 10 sero-positive and three sero-negative women (1.9%). Compared with controls, the live birthrate of the studied women with probable CD was significantly decreased before gluten removal of the diet (P = .015), but significantly increased after that (P = .020). CONCLUSION: One percent CD prevalence should be expected after anti-TG2 serological screening. However, more sensitive approaches should be explored, especially considering the potential beneficial effect of the gluten-free diet on the reproductive outcomes of women with CD.

Spatiotemporal expression and regulation of FoxO1 in mouse uterus during peri-implantation period.

Recent studies indicate that FoxO1 has roles in female reproductive system, especially in maternal endometrium. Although various cellular aspects and molecular pathways have been identified, the exact molecular characteristics of embryo implantation are still not completely understood. In this study, we aimed to investigate uterine expression and regulation of FoxO1 during peri-implantation period in mice. Experimental mouse models including, normal pregnancy, pseudopregnancy, artificial decidualization, and delayed implantation and activation were performed. Our results showed that FoxO1 expression was spatiotemporal in mouse endometrial tissue throughout peri-implantation period and its expression was significantly upregulated in luminal and glandular epithelium at the time of implantation. Moreover, on day 5 morning (09:00 AM) of pregnancy, expression of FoxO1 was cytoplasmic in endometrial luminal epithelial cells where embryo homing takes place. With progressing time on day 5 evening (19:00 PM) of pregnancy FoxO1 expression was nuclear in luminal epithelium at implantation site. Pseudopregnancy and artificial decidualization models indicated that FoxO1 expression was regulated by pregnancy hormones. Delayed implantation and activation model indicated that FoxO1 expression at the time of implantation is dependent upon activation status of blastocyst due to E2 induction and uterine sensitivity to implantation. In conclusion, our findings highlight a perspective for FoxO1 expression and regulation in mouse uterus during peri-implantation period indicating that its expression is regulated by implanting embryo and pregnancy hormones.

Introduction: Endometrial receptivity: evaluation, induction and inhibition.

Endometrial receptivity is an essential component of the complex process of embryo implantation. Its existence is inferred from the observation that not all embryo transfers result in pregnancy. The endometrium is a unique tissue which undergoes dramatic and rapid changes throughout the menstrual cycle. There appears to be a window of implantation, a time of optimal endometrial receptivity, when embryos are most likely to implant. The assessment of the timing and duration of this window of implantation has been a topic of interest and debate since the 1950s. The existence of the window of implantation led to the development of cycles in which endometrial receptivity is induced with exogenous E2 and P. These cycles are essential to third party parenting and frozen embryo transfers and have therefore become a common part of the practice of assisted reproduction.

Induced endometrial trauma (endometrial scratch) in the mid-luteal menstrual cycle phase preceding first cycle IVF/ICSI versus usual IVF/ICSI therapy: study protocol for a randomised controlled trial.

INTRODUCTION: Endometrial trauma commonly known as endometrial scratch (ES) has been shown to improve pregnancy rates in women with a history of repeated implantation failure undergoing in vitro fertilisation (IVF), with or without intracytoplasmic sperm injection (ICSI). However, the procedure has not yet been fully explored in women having IVF/ICSI for the first time. This study aims to examine the effect of performing an ES in the mid-luteal phase prior to a first-time IVF/ICSI cycle on the chances of achieving a clinical pregnancy and live birth. If ES can influence this success rate, there would be a significant cost saving to the National Health Service through decreasing the number of IVF/ICSI cycles necessary to achieve a pregnancy, increase the practice of single embryo transfer and consequently have a large impact on risks and costs associated with multiple pregnancies. METHODS AND ANALYSIS: This 30-month, UK, multicentre, parallel group, randomised controlled trial includes a 9-month internal pilot and health economic analysis recruiting 1044 women from 16 fertility units. It will follow up participants to identify if IVF/ICSI has been successful and live birth has occurred up to 6 weeks post partum. Primary analysis will be on an intention-to-treat basis. A substudy of endometrial samples obtained during the ES will assess the role of immune factors in embryo implantation. Main trial recruitment commenced on January 2017 and is ongoing.Participants randomised to the intervention group will receive the ES procedure in the mid-luteal phase of the preceding cycle prior to first-time IVF/ICSI treatment versus usual IVF/ICSI treatment in the control group, with 1:1 randomisation. The primary outcome is live birth rate after completed 24 weeks gestation. ETHICS AND DISSEMINATION: South Central-Berkshire Research Ethics Committee approved the protocol. Findings will be submitted to peer-reviewed journals and abstracts to relevant national and international conferences. TRIAL REGISTRATION NUMBER: ISRCTN23800982; Pre-results.

The history of the discovery of embryonic diapause in mammals.

The first incidence of embryonic diapause in mammals was observed in the roe deer, Capreolus capreolus, in 1854 and confirmed in the early 1900s. Since then scientists have been fascinated by this phenomenon that allows a growing embryo to become arrested for up to 11 months and then reactivate and continue development with no ill effects. The study of diapause has required unraveling basic reproductive processes we now take for granted and has spanned some of the major checkpoints of reproductive biology from the identification of the sex hormones to the hypothalamic-pituitary axis to microRNA and exosomes. This review will describe the history of diapause from its origins to the current day, including its discovery and efforts to elucidate its mechanisms. It will also attempt to highlight the people involved who were instrumental in progressing this field over the last 160 years. The most recent confirmation of mammalian diapause was in the panda in 2009 and there are still multiple mammals where it has been predicted but not yet confirmed. Furthermore, there are many questions still unanswered which ensure that embryonic diapause will continue to be a topic of research for many years to come. Note that there have recently been several extensive reviews covering the recent advances in embryonic diapause, so they will be mentioned only briefly here. For further information refer to Renfree and Shaw 2014; Fenelon et al 2014; Renfree and Fenelon 2017, and references therein.

Prevalent genotypes of methylenetetrahydrofolate reductase (MTHFR) in recurrent miscarriage and recurrent implantation failure.

OBJECTIVE: To evaluate the association of two common methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms with recurrent miscarriage (RM) and repeated implantation failure (RIF) METHODS: The study comprised of 521 patients, with a history of RM (n = 370) or RIF (n = 151). One hundred forty-four women with fallopian tube blockages who had successfully conceived after the first in vitro fertilization embryo transfer treatment served as the control group. The MTHFR alleles, genotypes, and haplotypes were assessed in different groups. RESULTS: There was no difference in allele frequency and distribution of MTHFR polymorphisms between case and control patients. The 1298AA genotype was represented in a higher frequency, and 1298AC genotype was significantly lower in subfertile group when compared to the control group. A significant relationship was found between the 1298AC genotype and the RIF subgroup. The haplotype 677CC/1298AA was overrepresented in the RM subgroup (> 2 times) and haplotype 677CC/1298AC was underrepresented in the RIF subgroup (P < 0.05). Nevertheless, these two haplotypes were not connected to fertilization and embryo cleavage rates. CONCLUSION: Our findings indicate that the MTHFR gene polymorphism might play a role in the etiology of patients with RM or RIF. No adverse effects of different MTHFR haplotypes on embryo development were detected. Further studies on the biological role are needed to better understand the susceptibility to pregnancy complications.

Cellular prion protein is involved in decidualization of mouse uterus.

Prion protein (PrP) is encoded by a single copy gene Prnp in many cell and tissue types. PrP is very famous for its infectious conformers (PrPSC) resulting in transmissible spongiform encephalopathies. At present, physiological functions of its cellular isoform (PrPC) remain ambiguous. Although PrPC expression has been found in uterus, whether it functions in maternal-fetal dialogue during early pregnant is unknown. In this study, we examined PrPC mRNA and protein in the uterus of peri-implantation mice, and found that they were expressed with a spatiotemporal dynamic pattern. Interestingly, PrPC was significantly increased in the decidual zones around the implanting embryos at the implantation window stage. To further demonstrate that PrPC is involved in the decidualization of mouse uterus during embryo implantation, we constructed the artificial decidualization models and the delayed implantation models. Once the pseudopregnant mice were artificially induced to decidualization, the PrPC expression then increased significantly in the decidua zone. And also, if the delayed implantation embryos were allowed to implant, PrPC protein was also simultaneously improved in stromal cells surrounding the implanting embryos. Moreover, PrPC expression can be inhibited by progesterone but upregulated by estrogen in mouse uterus. These results suggest that PrPC may play an important role in embryo implantation and decidualization.

Diverse endometrial mRNA signatures during the window of implantation in patients with repeated implantation failure.

High endometrial receptivity in the window of implantation (WOI) is essential for successful implantation. However, a diagnostic tool with high specificity for impaired endometrial receptivity remains to be developed. We collected endometrium specimens during the WOI from patients with RIF and women who conceived after one IVF/ICSI attempt. We conducted mRNA microarray on the samples followed by relevant comparative and functional analysis. Microarray analysis revealed 357 dysregulated mRNAs between the two groups. The majority of these mRNAs were found to encode membrane proteins by Gene Ontology (GO) analysis. The major functional biological pathways associated with the down-regulated mRNAs were cytokine-cytokine receptor interaction, the p53 signalling pathway and the complement and coagulation cascades. Up-regulated mRNAs were found mainly to participate in pathways such as PPAR signalling, hematopoietic cell lineage, phosphatidylinositol signalling system, ECM-receptor interaction and notch signalling. AQP3, DPP4 and TIMP3 whose expression patterns were down-regulated in RIF patients both by microarray and real-time PCR had a high correspondence with previous studies demonstrating that these genes may contribute to the defects in endometrial receptivity in RIF patients. Overall, these RIF-associated mRNAs may help devise new diagnostic tools for endometrial receptivity.

Estrogen receptor α is required for oviductal transport of embryos.

Newly fertilized embryos spend the first few days within the oviduct and are transported to the uterus, where they implant onto the uterine wall. An implantation of the embryo before reaching the uterus could result in ectopic pregnancy and lead to maternal death. Estrogen is necessary for embryo transport in mammals; however, the mechanism involved in estrogen-mediated cellular function within the oviduct remains unclear. In this study, we show in mouse models that ciliary length and beat frequency of the oviductal epithelial cells are regulated through estrogen receptor α (ESR1) but not estrogen receptor ß (ESR2). Gene profiling indicated that transcripts in the WNT/ß-catenin (WNT/CTNNB1) signaling pathway were regulated by estrogen in mouse oviduct, and inhibition of this pathway in a whole oviduct culture system resulted in a decreased embryo transport distance. However, selective ablation of CTNNB1 from the oviductal ciliated cells did not affect embryo transport, possibly because of a compensatory mechanism via intact CTNNB1 in the adjacent secretory cells. In summary, we demonstrated that disruption of estrogen signaling in oviductal epithelial cells alters ciliary function and impairs embryo transport. Therefore, our findings may provide a better understanding of etiology of the ectopic pregnancy that is associated with alteration of estrogen signals.-Li, S., O'Neill, S. R. S., Zhang, Y., Holtzman, M. J., Takemaru, K.-I., Korach, K. S., Winuthayanon, W. Estrogen receptor α is required for oviductal transport of embryos.

HOXA-10 and E-cadherin expression in the endometrium of women with recurrent implantation failure and recurrent miscarriage.

OBJECTIVE: To compare the expression of HOXA-10 and E-cadherin in the endometrium of women with recurrent implantation failure (RIF), women with recurrent miscarriage (RM), and women with proven fertility (normal control; NC). DESIGN: Observational cohort study. SETTING: University assisted reproductive unit. PATIENT(S): Fifty women were recruited: 18 NC, 12 unexplained RIF, and 20 RM. INTERVENTIONS(S): None. MAIN OUTCOME MEASURE(S): Endometrial biopsy was precisely timed 7 days after LH surge. The expression of HOXA-10 and E-cadherin were examined by means of immunohistochemistry. H-Scores of staining intensity in the glandular epithelium and stroma were measured. RESULT(S): HOXA-10 signal was mainly localized in the nuclei of stroma cells and the cytoplasm of glandular epithelium cells. E-Cadherin signal was found only in the cytoplasm of glandular epithelium cells. The HOXA-10 H-scores in the RIF group and the RM group were significantly lower than in the control group in both the glandular epithelium and stroma. The E-cadherin H-scores in the RM group were also significantly lower than in the control group. Interestingly, there was a positive correlation between HOXA-10 and E-cadherin H-scores in all of the women examined. CONCLUSION(S): There is a positive correlation between levels of HOXA-10 and E-cadherin expression in the endometrium, both of which are significantly reduced in women with RIF and RM compared with fertile control women. The findings suggest a potential role of HOXA-10 and E-cadherin in the implantation processes and altered expression in women with reproductive failure.

Regulation of diapause in carnivores.

Embryonic diapause is an evolutionary strategy to ensure that offspring are born when maternal and environmental conditions are optimal for survival. In many species of carnivores, obligate embryonic diapause occurs in every gestation. In mustelids, the regulation of diapause and reactivation is influenced by photoperiod, which then acts to regulate the secretion of pituitary prolactin. Prolactin in turn regulates ovarian steroid function. Reciprocal embryo transplant studies indicate that this state of embryonic arrest is conferred by uterine conditions and is presumed to be due to a lack of specific factors necessary for continued development. Studies of global gene expression in the mink (Neovison vison) revealed reduced expression of a cluster of genes that regulate the abundance of polyamines in the uterus during diapause, including the rate-limiting enzyme in polyamine production, ornithine decarboxylase (ODC). In addition, in this species, in vivo inhibition of the conversion of ornithine to the polyamine, putrescine, induces a reversible arrest in embryonic development and an arrest in both trophoblast and inner cell mass proliferation in vitro. Putrescine, at 0.5, 2 and 1,000 µM concentrations induced reactivation of mink embryos in culture, indicated by an increase in embryo volume, observed within five days. Further, prolactin induces ODC1 expression in the uterus, thereby regulating uterine polyamine levels. These results indicate that pituitary prolactin acts on ovarian and uterine targets to terminate embryonic diapause. In summary, our findings suggest that the polyamines, with synthesis under the control of pituitary prolactin, are the uterine factor whose absence is responsible for embryonic diapause in mustelid carnivores.

Uterine flushing proteome of the tammar wallaby after reactivation from diapause.

The marsupial tammar wallaby has the longest period of embryonic diapause of any mammal, up to 11 months, during which there is no cell division or blastocyst growth. Since the blastocyst in diapause is surrounded by acellular coats, the signals that maintain or terminate diapause involve factors that reside in uterine secretions. The nature of such factors remains to be resolved. In this study, uterine flushings (UFs) were used to assess changes in uterine secretions of tammars using liquid chromatography-mass spectrometry (LC-MS/MS) during diapause (day 0 and 3) and reactivation days (d) 4, 5, 6, 8, 9, 11 and 24 after removal of pouch young (RPY), which initiates embryonic development. This study supports earlier suggestions that the presence of specific factors stimulate reactivation, early embryonic growth and cell proliferation. A mitogen, hepatoma-derived growth factor and soluble epidermal growth factor receptors were observed from d3 until at least d11 RPY when these secreted proteins constituted 21% of the UF proteome. Binding of these factors to specific cellular receptors or growth factors may directly stimulate DNA synthesis and division in endometrial gland cells. Proteins involved in the p53/CDKN1A (p21) cell cycle inhibition pathway were also observed in the diapause samples. Progesterone and most of the oestrogen-regulated proteins were present in the UF after d3, which is concomitant with the start of blastocyst mitoses at d4. We propose that once the p21 inhibition of the cell cycle is lost, growth factors including HDGF and EGFR are responsible for reactivation of the diapausing blastocyst via the uterine secretions.

Decreased expression of aquaporin 2 is associated with impaired endometrial receptivity in controlled ovarian stimulation.

Recently, there has been evidence of decreased implantation rates with in vitro fertilisation and embryo transfer due to controlled ovarian stimulation (COS). The aim of this study was to investigate the effect of COS on embryo implantation and the role of aquaporin 2 (AQP2). We recruited eight patients who underwent COS and 40 matched controls. Endometrial samples were collected on Day 4~8 after injection of human chorionic gonadotrophin in the COS group and in the mid-secretory phase in the control group. Human endometrial morphological changes after COS were examined and expression of AQP2, leukaemia inhibitory factor (LIF) and integrin B3 (ITGB3) were determined by quantitative polymerase chain reaction, western blotting and immunohistochemistry in human endometrium and Ishikawa cells. Attachment rates were obtained using the embryo attachment test. The results showed that endometrial epithelial cells from the COS group were disrupted and lacked pinopodes. Messenger RNA and protein levels of AQP2, LIF and ITGB3 decreased in endometrial samples from the COS group. Knockdown of AQP2 resulted in reduced expression of LIF and ITGB3 and reduced embryo attachment rates. In conclusion, impaired endometrial receptivity in patients who underwent COS is correlated with a decreased expression of AQP2.

Hmgn1 acts downstream of C/EBPß to regulate the decidualization of uterine stromal cells in mice.

Although Hmgn1 is involved in the regulation of gene expression and cellular differentiation, its physiological roles on the differentiation of uterine stromal cells during decidualization still remain unknown. Here we showed that Hmgn1 mRNA was highly expressed in the decidua on days 6-8 of pregnancy. Simultaneously, increased expression of Hmgn1 was also observed in the artificial and in vitro induced decidualization models. Hmgn1 induced the proliferation of uterine stromal cells and expression of Ccna1, Ccnb1, Ccnb2 and Cdk1 in the absence of estrogen and progesterone. Overexpression of Hmgn1 could enhance the expression of Prl8a2 and Prl3c1 which were 2 well-known differentiation markers for decidualization, whereas inhibition of Hmgn1 with specific siRNA could reduce their expression. Further studies found that Hmgn1 could mediate the effects of C/EBPß on the expression of Prl8a2 and Prl3c1 during in vitro decidualization. In the uterine stromal cells, cAMP analog 8-Br-cAMP could stimulate the expression of Hmgn1 via C/EBPß. Moreover, siRNA-mediated down-regulation of Hmgn1 could attenuate the effects of cAMP on the differentiation of uterine stromal cells. During in vitro decidualization, Hmgn1 might act downstream of C/EBPß to regulate the expression of Cox-2, mPGES-1 and Vegf. Progesterone could up-regulate the expression of Hmgn1 in the ovariectomized mouse uterus, uterine epithelial cells and stromal cells. Knockdown of C/EBPß with siRNA alleviated the up-regulation of progesterone on Hmgn1 expression. Collectively, Hmgn1 may play an important role during mouse decidualization.

Reproductive Biology Including Evidence for Superfetation in the European Badger Meles meles (Carnivora: Mustelidae).

The reproductive biology of the European badger (Meles meles) is of wide interest because it is one of the few mammal species that show delayed implantation and one of only five which are suggested to show superfetation as a reproductive strategy. This study aimed to describe the reproductive biology of female Irish badgers with a view to increasing our understanding of the process of delayed implantation and superfetation. We carried out a detailed histological examination of the reproductive tract of 264 female badgers taken from sites across 20 of the 26 counties in the Republic of Ireland. The key results show evidence of multiple blastocysts at different stages of development present simultaneously in the same female, supporting the view that superfetation is relatively common in this population of badgers. In addition we present strong evidence that the breeding rate in Irish badgers is limited by failure to conceive, rather than failure at any other stages of the breeding cycle. We show few effects of age on breeding success, suggesting no breeding suppression by adult females in this population. The study sheds new light on this unusual breeding strategy of delayed implantation and superfetation, and highlights a number of significant differences between the reproductive biology of female Irish badgers and those of Great Britain and Swedish populations.

[Autopsy of children, neonates, fetuses and embryos: Preparation, technique and report writing]. / Obduktion von Kindern, Neonaten, Feten und Embryonen : Vorbereitung, Präparation und Berichterstellung.

Information obtained from the autopsies of children, neonates, fetuses and embryos, may not only be useful to explain the loss experienced by the parents but also to estimate the risk of recurrence. The detection of diseases by an autopsy helps to reduce the risk as well as with the planning of the next pregnancy and the optimal care of mother and fetus. Although incidences are continually dropping, according to the World Health Organization (WHO) statistics each year at least 2.6 million children worldwide suffer intrauterine death after the 28th week of pregnancy. Despite a general decrease in the number of autopsies, the parents agreed to a post-mortem examination in 500 out of 512 cases. The post-mortem examination and interpretation of results of children differ from those obtained from adults. As a supplement to previous publications, this article discusses these differences and may provide standardized instructions on performing autopsies and evaluation of autopsy findings.