The true incidence of recurrent implantation failure.

PURPOSE OF REVIEW: To review the merits and limitations of current definitions of recurrent implantation failure (RIF), how they translate into estimates of incidence and to summarize how emerging concepts of individualizing the recognition of this condition can assist in changing the way RIF is identified, studied and managed. RECENT FINDINGS: The notion of a one size fits all definition of RIF is seen to be of limited clinical value, as the individual risk of repeated IVF failure has many determinants and causes. Novel approaches provide a means of identifying 'actionable' RIF in individual patients. SUMMARY: Uncertainties as to what constitutes, causes and defines RIF have served to limit progress in its management. A new approach promises to permit progress from the current impasse.

Peri-implantation glucocorticoid administration for assisted reproductive technology cycles.

BACKGROUND: The use of peri-implantation glucocorticoids has been advocated to improve embryo implantation during assistive reproductive technology (ART) cycles such as in vitro fertilisation (IVF) or intracytoplasmic sperm injection (ICSI). It has been proposed that glucocorticoids may improve the intrauterine environment by acting as immunomodulators to reduce the uterine natural killer (NK) cell count and activity, normalising the cytokine expression profile in the endometrium and by suppression of endometrial inflammation. OBJECTIVES: To evaluate the effectiveness and safety of glucocorticoids versus no glucocorticoids administered around the time of anticipated implantation in women undergoing IVF or ICSI. SEARCH METHODS: We searched the Cochrane Gynaecology and Fertility (CGF) Group specialised register, CENTRAL (now also containing output from two trial registers and CINAHL), MEDLINE and Embase, on 20 December 2021, together with reference checking, contact with experts in the field and relevant conference proceedings to identify additional studies. This review is an update of the review first published in  2007 and last updated in 2012. SELECTION CRITERIA: Randomised controlled trials (RCTs) comparing the efficacy of supplementary systemic administration of glucocorticoids in the peri-implantation period with a placebo or no glucocorticoids in subfertile women undergoing IVF or ICSI were included. DATA COLLECTION AND ANALYSIS: We used standard methodological procedures recommended by Cochrane. The primary review outcomes were live birth rate and multiple pregnancy. MAIN RESULTS: We included 16 RCTs (2232 couples analysed). We are uncertain whether glucocorticoids improved live birth rates (odds ratio (OR) 1.37, 95% confidence interval (CI) 0.69 to 2.71; 2 RCTs, n = 366; I2 = 7%; very low-certainty evidence). This suggests that if the chance of live birth following no glucocorticoids/placebo is assumed to be 9%, the chance following glucocorticoids would be between 6% and 21%. We are also uncertain whether there was a difference between peri-implantation glucocorticoids on multiple pregnancy rates per couple (OR 0.86, 95% CI 0.33 to 2.20; 4 RCTs, n = 504; I2 = 53%; very low-certainty evidence). The I2 of 53% may represent moderate statistical heterogeneity and results have to be interpreted with caution. With regard to pregnancy rates, we are uncertain whether there was a difference between ongoing pregnancy rates after glucocorticoids versus no glucocorticoids/placebo (OR 1.19, 95% CI 0.80 to 1.76; 3 RCTs, n = 476; I2 = 0%; very low-certainty evidence) and clinical pregnancy rates after glucocorticoids versus no glucocorticoids/placebo (OR 1.17, 95% CI 0.95 to 1.44; 13 RCTs, n = 1967; I2 = 0%; low-certainty evidence). This suggests that if the chance of clinical pregnancy following no glucocorticoids/placebo is assumed to be 25%, the chance following glucocorticoids would be between 24% and 32%. Furthermore, we are also uncertain whether peri-implantation glucocorticoids influenced miscarriage rates per couple (OR 1.09, 95% CI 0.63 to 1.87; 6 RCTs, n = 821; I2 = 0%; very low-certainty evidence), the incidence of ectopic pregnancies per couple (OR 2.28, 95% CI 0.33 to 15.62; 3 RCTs, n = 320; I2 = 0%; very low-certainty evidence) and ovarian hyperstimulation syndrome (OHSS) per couple (OR 1.07, 95% CI 0.60 to 1.90; 3 RCTs, n = 370; I2 = 0%; very low-certainty evidence) compared to no glucocorticoids/placebo. The evidence was very low to low certainty: the main limitations were serious risk of bias due to poor reporting of study methods, and serious imprecision. AUTHORS' CONCLUSIONS: Overall, there was insufficient evidence that administration of peri-implantation glucocorticoids in IVF/ICSI cycles influenced clinical outcomes. These findings were limited to the routine use of glucocorticoids in subfertile women undergoing IVF or ICSI.

Increased obstetric and neonatal risks in artificial cycles for frozen embryo transfers?

RESEARCH QUESTION: What are the obstetric and neonatal risks for women conceiving via frozen-thawed embryo transfer (FET) during a modified natural cycle compared with an artificial cycle method. DESIGN: A follow-up study to the ANTARCTICA randomized controlled trial (RCT) (NTR 1586) conducted in the Netherlands, which showed that modified natural cycle FET (NC-FET) was non-inferior to artificial cycle FET (AC-FET) in terms of live birth rates. The current study collected data on obstetric and neonatal outcomes of 98 women who had a singleton live birth. The main outcome was birthweight; additional outcomes included hypertensive disorder of pregnancy, premature birth, gestational diabetes, obstetric haemorrhage and neonatal outcomes including Apgar scores and admission to the neonatal ward or the neonatal intensive care unit and congenital anomalies. RESULTS: Data from 82 out of 98 women were analysed according to the per protocol principle. There was no significant difference in the birthweights of children born between groups (mean difference -124 g [-363 g to 114 g]; P = 0.30). Women who conceived by modified NC-FET have a decreased risk of hypertensive disorders of pregnancy compared with AC-FET (relative risk 0.27; 95% CI 0.08-0.94; P = 0.031). Other outcomes, such as rates of premature birth, gestational diabetes or obstetric haemorrhage and neonatal outcomes, were not significantly different. CONCLUSIONS: The interpretation is that modified NC-FET is the preferred treatment in women with ovulatory cycles undergoing FET when the increased risk of obstetrical complications and potential neonatal complications in AC-FET are considered.

Influence of endometrial thickness on pregnancy rates in modified natural cycle frozen-thawed embryo transfer.

INTRODUCTION: Pregnancy after frozen-thawed embryo transfer (FET) is a multifactorial process. Although embryo quality is a key factor in determining pregnancy, other factors, including maternal determinants, are also considered to be predictive. Even though an association between endometrial thickness measured by transvaginal ultrasound and pregnancy rates has been reported in patients undergoing various assisted reproductive technology treatments, whether endometrial thickness predicts achieving pregnancy after natural cycle FET (NC-FET) remains unclear. MATERIAL AND METHODS: In this cohort study, 463 patients allocated to the modified NC-FET (mNC-FET) arm of a previously published randomized controlled trial were included. Monitoring in mNC-FET cycles consisted of regular ultrasound scans, measuring both dominant follicle and endometrial thickness. When the dominant follicle reached a size of 16-20 mm, an injection of human chorionic gonadotrophin was administered and embryo thawing and transfer planned. No minimal endometrial thickness was defined below which transfer was to be deferred. The primary endpoint was ongoing pregnancy rate. RESULTS: Overall, the ongoing pregnancy rate per started FET cycle was 12.5%. Multivariate regression analyses showed that embryo quality was the only significant predictor for ongoing pregnancy. Mean endometrial thickness did not differ between patients achieving ongoing pregnancy and those who did not (9.0 vs. 8.8 mm, p = 0.4). Comparable results were obtained with regard to clinical pregnancy, live birth and miscarriage rates. The area under the receiver operator curve was 0.5, indicating little discriminatory value of endometrial thickness. CONCLUSIONS: Given that endometrial thickness was not found to be predictive of pregnancy after mNC-FET, cancellation based on endometrial thickness alone may not be justified.

The effect of elevated progesterone levels before HCG triggering in modified natural cycle frozen-thawed embryo transfer cycles.

Recent studies suggest that elevated late follicular phase progesterone concentrations after ovarian stimulation for IVF may result in embryo-endometrial asynchrony, reducing the chance of successful implantation after fresh embryo transfer. It remains unclear to what extent elevated late follicular phase progesterone levels may occur in unstimulated cycles before frozen-thawed embryo transfer, or what affect they may have on outcomes. In this cohort study, 271 patients randomized to the modified natural cycle arm of a randomized controlled trial comparing two endometrial preparation regimens underwent late follicular phase progesterone and LH testing. A receiver operating characteristic curve was constructed to identify a progesterone cut-off level with the best predictive value for live birth (progesterone level ≥4.6 nmol/l). A total of 24.4% of patients revealed an isolated elevated serum progesterone of 4.6 nmol/l or greater, and 44.3% showed an elevated progesterone level in association with a rise in LH. Neither endocrine disruption affected outcomes, with live birth rates of 12.9% versus 10.6% (OR 0.6, 95% CI 0.19 to 1.9) and 11.9% versus 17.5% (OR 1.6, 95% CI 0.79 to 3.1), respectively. Whether monitoring of progesterone and LH in natural cycle frozen-thawed embryo transfer has added clinical value should studied further.

Amino acid composition of human uterine fluid: association with age, lifestyle and gynaecological pathology.

STUDY QUESTION: Do the amino acid levels of human uterine fluid vary with age, BMI, phase of menstrual cycle, benign pathology or diet? SUMMARY ANSWER: The levels of 18 amino acids in human uterine fluid were shown to be affected only by maternal diet. WHAT IS KNOWN ALREADY: Murine, bovine and ovine uterine amino acid content has been reported, but no reliable data on the human exist. Murine studies have demonstrated that the intrauterine periconceptional nutritional environment is affected by maternal diet. STUDY DESIGN, SIZE, DURATION: Uterine secretions were aspirated from 56 women aged 18-45 years. The women were recruited preoperatively from gynaecological theatre operating schedules or hysterosalpingo-contrast-sonography (HyCoSy) lists. A proportion of these women had proven fertility; however, the majority were being investigated for subfertility. The BMI, gynaecological history and dietary pattern of these women were also assessed. PARTICIPANTS/MATERIALS, SETTING, METHODS: Reverse phase high performance liquid chromatography was used to analyse the concentrations of 18 amino acids within the uterine fluid and blood serum. The results were analysed against the women's stage of cycle, age, BMI and diet. MAIN RESULTS AND THE ROLE OF CHANCE: The profile of 18 amino acids in uterine fluid was described. In total, human uterine fluid was observed to contain an amino acid concentration of 3.54 mM (interquartile range: 2.27-6.24 mM). The relative concentrations of 18 amino acids were not significantly altered by age, BMI, cycle phase or the presence of specific benign gynaecological pathologies. However, a diet identified by a validated scoring system as being less healthy was associated with higher concentrations of asparagine (P = 0.018), histidine (P = 0.011), serine (P = 0.033), glutamine (P = 0.049), valine (P = 0.025), phenylalanine (P = 0.019), isoleucine (P = 0.025) and leucine (P = 0.043) in the uterine fluid compared with a healthier diet, defined as one with a higher intake of fresh vegetables, fruit, whole-grain products and fish and a low intake of red and processed meat and high fat dairy products. There were no significant correlations between serum amino acid concentrations and those in the uterine fluid. LIMITATIONS, REASONS FOR CAUTION: Our results enabled us to detect the effect of diet on the concentrations of amino acids in human uterine fluid; however, the study may not have had sufficient numbers to detect mild effects of BMI or age. WIDER IMPLICATIONS OF THE FINDINGS: These findings increase our understanding of the nutritional environment encountered by the preimplantation embryo, and indicate how periconceptional diet may alter this. Given the importance of early embryo environment for programming of development and future health, this information may aid in the development of nutritional interventions aimed at optimizing the preimplantation phase of human embryo development in vivo. STUDY FUNDING/COMPETING INTERESTS: This work was funded by the NIHR, the Medical Research Council (G0701153) and the University of Southampton and was supported by the NIHR BRC in Nutrition and Southampton University NHS Foundation Trust. The authors declare no conflicts of interest.

Lower values of VEGF in endometrial secretion are a possible cause of subfertility in non-atopic asthmatic patients.

OBJECTIVE: Using endometrial secretion analysis, we assessed whether altered inflammatory cytokine levels can be detected in the uterine environment in asthma patients, thereby providing a possible cause of reduced fertility in asthmatics. METHODS: Forty-four unexplained infertile women (aged 28-44) underwent asthma and allergy testing, questionnaires, endometrial secretion and blood samples in the mid-secretory phase of the menstrual cycle (day 19-23) during assisted reproduction. Differences in cytokines and growth factors were analyzed. RESULTS: Mean log-VEGF in uteri was lower in asthma patients compared with controls (2.29 versus 2.70, p = 0.028). This was mainly due to lower values of VEGF among women with non-atopic asthma compared with women with atopic asthma (1.86 versus 2.72, p = 0.009) and with healthy controls (1.86 versus 2.70, p = 0.01). Asthma treatment status had no effect on VEGF levels in uteri. Serum high sensitivity CRP was negatively correlated with VEGF in endometrial secretions. No other significant correlations were observed between peripheral blood values and markers found in utero. CONCLUSION: Asthma is associated with lower values of VEGF in uterine endometrial secretions, which might affect the receptiveness of the endometrium and thereby increase time to pregnancy. The effect appears to be associated with non-atopic asthma with general increased systemic inflammation.

The human endometrium as a sensor of embryo quality.

Human reproduction is characterized by a high degree of embryo wastage, which is largely ascribed to a high prevalence of embryo aneuploidy. It is proposed that maternal strategies have evolved that prevent inappropriate investment in invasive, but poorly viable embryos. Key to this is the emerging concept of the endometrium as biosensor, first identified in human in vitro embryo/decidualized stromal cell coculture systems and recently confirmed in an in vivo mouse model. In this review, the growing supporting experimental evidence for the biosensor component of decidualized endometrium is outlined, and recent insights into the nature of the embryo-derived signal detected by the endometrium and the biological processes by which this signal is thought to be converted into a go or no-go endometrial response are described. Finally, the clinical implications of this new paradigm of the choosy uterus are addressed.

The science of implantation emerges blinking into the light.

Although embryo implantation is essential for human survival, it remains an enigmatic biological phenomenon. Following fertilization, the resulting blastocyst must signal its presence to the mother, attach to the luminal epithelium of the endometrium and embed into the decidualising stroma. Failure to do so results in infertility, which affects around 9% of women. Subsequent placental development requires remodelling of maternal blood vessels by trophoblast cells from the placenta, that invade deep into the decidua. Failure in these very early stages can compromise fetal development, resulting in diseases of pregnancy such as intrauterine growth restriction or pre-eclampsia which can also impact on health in adulthood. Abnormal implantation therefore constitutes a significant disease burden in humans. Although we have known for many years that successful implantation requires an embryo that is competent to implant and an endometrium that is receptive, the molecular basis of these processes remains poorly understood. Our inability to identify implantation-competent embryos or to diagnose/treat the non-receptive endometrium therefore limits our ability to intervene through assisted reproduction techniques. This Implantation Symposium aims to review recent exciting developments in our understanding of the biology of early implantation and to highlight the rapid progress being made to translate these into improved diagnosis and treatment.

In-vitro model systems for the study of human embryo-endometrium interactions.

Implantation requires highly orchestrated interactions between the developing embryo and maternal endometrium. The association between abnormal implantation and reproductive failure is evident, both in normal pregnancy and in assisted reproduction patients. Failure of implantation is the pregnancy rate-limiting step in assisted reproduction, but, as yet, empirical interventions have largely failed to address this problem. Better understanding of the mechanisms underlying human embryo-endometrium signalling is a prerequisite for the further improvement of assisted reproduction outcomes and the development of effective interventions to prevent early pregnancy loss. Studying human embryo implantation is challenging since in-vivo experiments are impractical and unethical, and studies in animal models do not always translate well to humans. However, in recent years in-vitro models have been shown to provide a promising way forward. This review discusses the principal models used to study early human embryo development and initial stages of implantation in vitro. While each model has limitations, exploiting these models will improve understanding of the molecular mechanisms and embryo-endometrium cross-talk at the early implantation site. They provide valuable tools to study early embryo development and pathophysiology of reproductive disorders and have revealed novel disease mechanisms such as the role of epigenetic modifications in recurrent miscarriage.

Reprint of: In-vitro model systems for the study of human embryo-endometrium interactions.

Implantation requires highly orchestrated interactions between the developing embryo and maternal endometrium. The association between abnormal implantation and reproductive failure is evident, both in normal pregnancy and in assisted reproduction patients. Failure of implantation is the pregnancy rate-limiting step in assisted reproduction, but, as yet, empirical interventions have largely failed to address this problem. Better understanding of the mechanisms underlying human embryo-endometrium signalling is a prerequisite for the further improvement of assisted reproduction outcomes and the development of effective interventions to prevent early pregnancy loss. Studying human embryo implantation is challenging since in-vivo experiments are impractical and unethical, and studies in animal models do not always translate well to humans. However, in recent years in-vitro models have been shown to provide a promising way forward. This review discusses the principal models used to study early human embryo development and initial stages of implantation in vitro. While each model has limitations, exploiting these models will improve understanding of the molecular mechanisms and embryo-endometrium cross-talk at the early implantation site. They provide valuable tools to study early embryo development and pathophysiology of reproductive disorders and have revealed novel disease mechanisms such as the role of epigenetic modifications in recurrent miscarriage.

Embryo implantation: biology, evaluation, and enhancement.

PURPOSE OF REVIEW: Implantation is an essential step in the development of a pregnancy, but often fails in humans. In assisted reproductive technologies, implantation failure continues to impair treatment outcomes, with distressing results for patients and physicians. RECENT FINDINGS: Morphokinetics, comprehensive chromosome screening, and the analysis of embryo-derived products detectable in spent culture media offer new means of assessing embryo viability. However, all await validation in randomized controlled trials. Genomic, transcriptomic, and secretomic technologies are similarly being exploited to define specific biomarkers of endometrial receptivity with the aim of identifying novel therapeutic interventions. However, to date no single, clinically relevant molecular marker capable of indicating endometrial receptivity has been reported. Recent work continues to describe the key signalling pathways which result in acceptance or rejection of the implanting embryo. In-vitro studies have revealed that the decidualized endometrium plays an important role in natural embryo selection, which could change our understanding of the aetiology and treatment of reproductive failure. SUMMARY: Recent developments in analytical techniques have initiated a search for biomarkers of embryo quality and endometrial receptivity, and in-vitro studies have revealed novel roles for the decidualized endometrium as a biosensor of embryo quality.

Peri-implantation glucocorticoid administration for assisted reproductive technology cycles.

BACKGROUND: In order to improve embryo implantation for in vitro fertilisation (IVF) or intracytoplasmic sperm injection (ICSI) cycles the use of glucocorticoids has been advocated. It has been proposed that glucocorticoids may improve the intrauterine environment by acting as immunomodulators to reduce the uterine natural killer (NK) cell count and normalise the cytokine expression profile in the endometrium and by suppression of endometrial inflammation. OBJECTIVES: To investigate whether the administration of glucocorticoids around the time of implantation improved clinical outcomes in subfertile women undergoing IVF or ICSI when compared to no glucocorticoid administration. SEARCH METHODS: The Cochrane Menstrual Disorders and Subfertility Group Trials Register (September 2011), the Cochrane Central Register of Controlled Trials (CENTRAL) (September 2011), MEDLINE (1966 to September 2011), EMBASE (1976 to September 2011), CINAHL (1982 to September 2011) and Science Direct (1966 to September 2011) were searched. Reference lists of relevant articles and relevant conference proceedings were handsearched. SELECTION CRITERIA: All randomised controlled trials (RCTs) addressing the research question were included. DATA COLLECTION AND ANALYSIS: Two review authors independently assessed eligibility and quality of trials and extracted relevant data. MAIN RESULTS: Fourteen studies (involving 1879 couples) were included. Three studies reported live birth rate and these did not identify a significant difference after pooling the (preliminary) results (OR 1.21, 95% CI 0.67 to 2.19). With regard to pregnancy rates, there was also no evidence that glucocorticoids improved clinical outcome (13 RCTs; OR 1.16, 95% CI 0.94 to 1.44). However, a subgroup analysis of 650 women undergoing IVF (6 RCTs) revealed a significantly higher pregnancy rate for women using glucocorticoids (OR 1.50, 95% CI 1.05 to 2.13). There were no significant differences in adverse events, but these were poorly and inconsistently reported. AUTHORS' CONCLUSIONS: Overall, there was no clear evidence that administration of peri-implantation glucocorticoids in ART cycles significantly improved the clinical outcome. The use of glucocorticoids in a subgroup of women undergoing IVF (rather than ICSI) was associated with an improvement in pregnancy rates of borderline statistical significance and should be interpreted with care. These findings were limited to the routine use of glucocorticoids and cannot be extrapolated to women with autoantibodies, unexplained infertility or recurrent implantation failure. Further well designed randomised studies are required to elucidate the possible role of this therapy in well defined patient groups.

Cryo-thawed embryo transfer: natural versus artificial cycle. A non-inferiority trial. (ANTARCTICA trial).

BACKGROUND: Frozen thawed embryo transfer (FET) is a cost-effective adjunct to IVF or IVF-ICSI treatment. In order to optimize treatment outcome, FET should be carried out during a period of optimal endometrial receptivity. To optimize implantation several methods for endometrium preparation have been proposed. In natural cycle FET (NC-FET), the endometrium develops under endogenous hormonal stimulation. The development of the dominant follicle and endometrium is monitored by ultrasound and FET is timed after triggering ovulation induction or determination of the spontaneous LH surge. In an artificial cycle FET (AC-FET) estrogens and progesterone are administered to prepare the endometrium for implantation. While the currently available data show no significant difference in pregnancy rates between these methods, well designed randomized controlled trials are lacking. Moreover there is little literature on difference in cancellation rates, cost-efficiency and adverse events. METHODS AND DESIGN: In this randomized, multi-centre, non-inferiority trial we aim to test the hypothesis that there is no significant difference in live birth rates between patients undergoing NC-FET versus AC-FET. The primary outcome will be live birth rate per embryo transfer procedure. Secondary outcomes will be ongoing and clinical pregnancy rate, cancellation rate, (serious) adverse events and cost-efficiency. Based on a live birth rate of 20% and a minimal clinical important difference of 7.5% (one-sided alpha 2.5%, beta 20%) a total of 1150 patients will be needed. Analyzes will be performed using both per protocol as well as intention to treat analyses. DISCUSSION: This prospective, randomized, non-inferiority trial aims to address the hypothesis that there is no significant difference in live birth rates between patients undergoing NC-FET versus patients undergoing AC-FET. Moreover it addresses cost-efficiency as well as the perceived burden of both treatments. TRIAL REGISTER: Netherlands trial register (NTR): 1586.

Recurrent pre-clinical pregnancy loss is unlikely to be a 'cause' of unexplained infertility.

BACKGROUND: A proportion of women with 'unexplained' infertility may present with subfertility because their pregnancies fail before they are clinically recognized. In order to test whether pre-clinical early pregnancy losses (EPL) occur more frequently in women with unexplained infertility, serial urinary hCG concentrations were measured to compare EPL per cycle rates following spontaneous conception in patients with unexplained infertility versus healthy volunteers. METHODS: Sixty patients under 39 years of age with unexplained infertility and 60 healthy controls, who were trying to conceive spontaneously, participated in this study. All participants were asked to collect daily urine samples from cycle day 14 until menstruation for three consecutive cycles or until a positive pregnancy test was obtained. Urinary hCG and creatinine levels were measured by immunoassay. Implantation was detected when urinary hCG levels rose above reference levels constructed from samples obtained from 12 women not attempting to conceive. EPL rates were determined by a linear mixed model using logarithmically transformed hCG/creatinine data. RESULTS: In the 133 cycles of 60 women with unexplained infertility, just one implantation was detected, which became an ongoing pregnancy. In contrast, in 103 such cycles in 46 control patients, 30 implantations were detected (24 clinical pregnancies, 6 cases of EPL). The odds ratio for EPL/cycle in the unexplained versus control group was 0 (95% confidence interval: 0-0.795, P = 0.026). CONCLUSIONS: Our data do not support the hypothesis that recurrent EPL may present as unexplained infertility. Post-implantation failure is therefore unlikely to contribute significantly to the presentation of subfertility.

The science behind 25 years of ovarian stimulation for in vitro fertilization.

To allow selection of embryos for transfer after in vitro fertilization, ovarian stimulation is usually carried out with exogenous gonadotropins. To compensate for changes induced by stimulation, GnRH analog cotreatment, oral contraceptive pretreatment, late follicular phase human chorionic gonadotropin, and luteal phase progesterone supplementation are usually added. These approaches render ovarian stimulation complex and costly. The stimulation of multiple follicular development disrupts the physiology of follicular development, with consequences for the oocyte, embryo, and endometrium. In recent years, recombinant gonadotropin preparations have become available, and novel stimulation protocols with less detrimental effects have been developed. In this article, the scientific background to current approaches to ovarian stimulation for in vitro fertilization is reviewed. After a brief discussion of the relevant aspect of ovarian physiology, the development, application, and consequences of ovarian stimulation strategies are reviewed in detail.

The molecular basis of recurrent pregnancy loss: impaired natural embryo selection.

Recurrent pregnancy loss (RPL) is a common and distressing disorder. Chromosomal errors in the embryo are the single most common cause, whereas uterine factors are invariably invoked to explain non-chromosomal miscarriages. These uterine factors are, however, poorly defined. The ability of a conceptus to implant in the endometrium is normally restricted to a few days in the menstrual cycle. A limited 'window of implantation' ensures coordinated embryonic and endometrial development, thereby minimizing the risk of late implantation of compromised embryos. In this paper, we review emerging evidence, indicating that RPL is associated with impaired differentiation of endometrial stromal cells into specialized decidual cells. From a functional perspective, this differentiation process, termed decidualization, is not only critical for placental development but also signals the end of the implantation window and bestows on the endometrium the ability to recognize, respond to and eliminate implanting compromised embryos. Thus, we propose that spontaneous decidualization of the human endometrium, which inevitably causes menstrual shedding in the absence of a viable conceptus, serves as functional 'window for natural embryo selection'. Conversely, impaired decidualization predisposes to late implantation, negates embryo quality control and causes early placental failure, regardless of the embryonic karyotype. This pathological pathway also explains the common observation that many RPL patients seem exceptionally fertile, often conceiving within one or two cycles. Thus, as the clinical correlate of inappropriate uterine receptivity, 'superfertility' should be considered as a genuine reproductive disorder that requires targeted intervention.

The fate of the mosaic embryo: chromosomal constitution and development of Day 4, 5 and 8 human embryos.

BACKGROUND: Post-zygotic chromosome segregation errors are very common in human embryos after in vitro fertilization, resulting in mosaic embryos. However, the significance of mosaicism for the developmental potential of early embryos is unknown. We assessed chromosomal constitution and development of embryos from compaction to the peri-implantation stage. METHODS: From 112 cryopreserved Day 4 human embryos donated for research, 21 were immediately fixed and all cells were analysed by fluorescent in situ hybridization (FISH) for chromosomes 1, 7, 13, 15, 16, 18, 21, 22, X and Y. The remaining 91 embryos were thawed, with 54 embryos undergoing biopsy of one or two cells which were fixed and analysed by FISH. Biopsied embryos were kept in standard culture conditions for 24 h. Embryos arrested before cavitation (n = 24) were fixed whereas developing Day 5 blastocysts (n = 24) were co-cultured for a further 72 h on an endometrial monolayer followed by fixation. Cell numbers were counted and all nuclei were analysed by FISH. Data from a previous FISH analysis on cryopreserved good-quality Day 5 blastocysts (n = 36) were also included in the present study. RESULTS: FISH analysis was successful for 18 Day 4 fixed embryos and, according to our definition, 83% were mosaic and 11% showed a chaotic chromosomal constitution. FISH analysis of two blastomeres from Day 4 developing embryos showed that 54% were mosaic, 40% were normal and 6% were abnormal. Analysis of Day 4, 5 and 8 whole embryos showed a decrease in incidence of mosaicism over time, from 83% on Day 4 to 42% on Day 8. A significant positive correlation was observed between the total cell number and the percentage of normal cells in developing Day 5 and Day 8 embryos but not in developing Day 4 or embryos arrested before cavitation. CONCLUSIONS: These data suggest that both the developmental arrest of a significant proportion of mosaic embryos on Day 4, and the cell death or reduced proliferation of aneuploid cells within an embryo may be responsible for the observed decrease of aneuploid blastomeres from compaction to the peri-implantation stage.

The impact of ovarian stimulation for IVF on the developing embryo.

The use of assisted reproductive technologies (ART) has been increasing over the past three decades, and, in developed countries, ART account for 1-3% of annual births. In an attempt to compensate for inefficiencies in IVF procedures, patients undergo ovarian stimulation using high doses of exogenous gonadotrophins to allow retrieval of multiple oocytes in a single cycle. Although ovarian stimulation has an important role in ART, it may also have detrimental effects on oogenesis, embryo quality, endometrial receptivity and perinatal outcomes. In this review, we consider the evidence for these effects and address possible underlying mechanisms. We conclude that such mechanisms are still poorly understood, and further knowledge is needed in order to increase the safety of ovarian stimulation and to reduce potential effects on embryo development and implantation, which will ultimately be translated into increased pregnancy rates and healthy offspring.

Is bacterial vaginosis associated with a pro-inflammatory cytokine profile in endometrial secretions of women undergoing IVF?

The objective of this prospective cohort study was to elucidate whether bacterial vaginosis (BV) is associated with a pro-inflammatory endometrial secretion cytokine profile and whether there is a relationship between BV and the concentrations of a number of key regulatory cytokines, chemokines and growth factors. A total of 198 women undergoing IVF treatment were included. Prior to embryo transfer, participants underwent screening for BV according to Nugent criteria by a Gram-stained cervical smear. The concentrations of 17 soluble mediators of human implantation were measured by multiplex immunoassay in endometrial secretions aspirated prior to embryo transfer. Seventeen (8.6%) women had BV (Nugent score >6). Multivariable logistic regression showed a significant positive association between interleukin-beta and the presence of BV (P=0.011; Nugent score >6 versus 6) and a significant negative association between eotaxin and BV (P=0.003). No significant differences were found in the ratios of distinct pro- and anti-inflammatory cytokines in endometrial secretions from women with or without BV. In conclusion, BV is associated with higher concentrations of interleukin-beta in endometrial secretions compared with women without BV. However, no distinct difference in pro- and anti-inflammatory profiles is present. An effect on endometrial receptivity is unlikely.