Article

Time of Implantation of the Conceptus and Loss of Pregnancy

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Abstract

Implantation of the conceptus is a key step in pregnancy, but little is known about the time of implantation or the relation between the time of implantation and the outcome of pregnancy. We collected daily urine samples for up to six months from 221 women attempting to conceive after ceasing to use contraception. Ovulation was identified on the basis of the ratio of urinary estrogen metabolites to progesterone metabolites, which changes rapidly with luteinization of the ovarian follicle. The time of implantation was defined by the appearance of chorionic gonadotropin in maternal urine. There were 199 conceptions, for 95 percent of which (189) we had sufficient data for analysis. Of these 189 pregnancies, 141 (75 percent) lasted at least six weeks past the last menstrual period, and the remaining 48 pregnancies (25 percent) ended in early loss. Among the pregnancies that lasted six weeks or more, the first appearance of chorionic gonadotropin occurred 6 to 12 days after ovulation; 118 women (84 percent) had implantation on day 8, 9, or 10. The risk of early pregnancy loss increased with later implantation (P<0.001). Among the 102 conceptuses that implanted by the ninth day, 13 percent ended in early loss. This proportion rose to 26 percent with implantation on day 10, to 52 percent on day 11, and to 82 percent after day 11. In most successful human pregnancies, the conceptus implants 8 to 10 days after ovulation. The risk of early pregnancy loss increases with later implantation.

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... Embryo implantation involves the close interaction between a 'competent' blastocyst and a receptive endometrium, occurring during the 'window of implantation' (WOI). In most women, the WOI is open during the mid-luteal phase, 8-10 days after ovulation, driven by the sequential actions of estrogen and P 4 (Wilcox et al., 1999). ...
... To pinpoint the WOI in an NC, Wilcox et al. (1999) collected daily urine samples for up to 6 months from 221 women with no history of infertility, attempting to conceive. Timing of ovulation was identified by the ratio of urinary E 2 metabolite (estrone 3-glucuronide) to P 4 metabolite (pregnanediol 3-glucuronide) (Baird et al., 1991). ...
... The authors discussed several reasons for the high early loss rates of late-implanting conceptuses, such as decreased receptivity during the luteal phase, a less hCG responsive corpus luteum or intrinsic embryonic factors resulting in a lower production of hCG. In most successful ongoing pregnancies, the embryo implanted 8-10 days after ovulation, coinciding with days 22-24 of the cycle (Wilcox et al., 1999). A large sample size with a 6-month follow-up for each patient is the strength of this milestone study; however, determining the day of ovulation using urinary E 2 and P 4 metabolites without transvaginal ultrasound is a limitation. ...
Article
Background: Efficient and safe embryo vitrification techniques have contributed to a marked worldwide increase in the use of elective frozen embryo transfer (FET). Pinpointing the day of ovulation, more commonly by documentation of the LH surge and less commonly by ultrasonography, is crucial for timing of FET in a true natural cycle (t-NC) to maximize the reproductive outcome. Objective and rationale: The definition of the onset of the LH surge should be standardized in t-NC FET cycles; however, a clear definition is lacking in the available literature. The first search question concerns the definition of the onset of the LH surge in a natural cycle. The second search question relates to the duration between the onset of the LH surge and ovulation. Search methods: We searched PubMed, Web of Science and Cochrane Library databases for two search questions from inception until 31 August 2021. 'Luteinizing hormone'[MeSH] OR 'LH' AND 'surge' terms were used to identify eligible articles to answer the first question, whereas 'Luteinizing hormone'[MeSH] OR 'LH' AND 'surge' OR 'rise' AND 'ovulation'[MeSH] OR 'follicular rupture' OR 'follicular collapse' were the terms used regarding the second question. The included publications were all written in the English language, conducted in women of reproductive age with regular ovulatory cycles and in whom serial serum or urine LH measurement was performed. For the quality and risk of bias assessment of the included studies, the Strengthening the Reporting of Observational Studies in Epidemiology and modified Newcastle Ottawa Scale were used. Outcomes: A total of 10 and 8 studies were included for search Questions 1 and 2, respectively. Over the years, through different studies and set-ups, testing in either serum or urine, different definitions for the onset of the LH surge have been developed without a consensus. An increase in LH level varying from 1.8- to 6-fold above the baseline LH level was used in seven studies and an increase of at least two or three standard deviations above the mean of the preceding LH measurements was used in two studies. An LH level exceeding the 30% of the amplitude (peak-baseline LH level) of the LH surge was defined as the onset day by one study. A marked inter-personal variation in the time interval between the onset of the LH surge and ovulation was seen, ranging from 22 to 56 h. When meta-analysis was performed, the mean duration in hours between the onset of the LH surge and ovulation was 33.91 (95% CI = 30.79-37.03: six studies, 187 cycles). Wider implications: The definition of the onset of the LH surge should be precisely defined in future well-designed studies employing state-of-art laboratory and ultrasonographic equipment. The window of implantation in a natural cycle is still a black box, and future research is warranted to delineate the optimal interval to time the embryo transfer in t-NC FET cycles. Randomized controlled trials employing different precise endocrine and/or ultrasonographic criteria for timing of FET in a t-NC are urgently required.
... With the development of egg freezing techniques and egg donation programs, oocyte maturation and induction of endometrial receptivity are often dissociated and assessed separately. Studies in the 1990's suggested that a critical window in the menstrual cycle determines whether implantation will or will not occur (6,11,12). One study showed that detection of urinary human chorionic gonadotropin (hCG) occurred from day 8-10 postovulation in 84% of the successful pregnancies (11). ...
... Studies in the 1990's suggested that a critical window in the menstrual cycle determines whether implantation will or will not occur (6,11,12). One study showed that detection of urinary human chorionic gonadotropin (hCG) occurred from day 8-10 postovulation in 84% of the successful pregnancies (11). The result suggests that the optimal implantation rates occur with embryo-endometrial asynchrony of ± 1.5 days or less. ...
Article
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Suboptimal endometrial receptivity and altered embryo-endometrial crosstalk account for approximately two-thirds of human implantation failures. Current tests of the window of implantation, such as endometrial thickness measurements and the endometrial receptivity assay, do not consistently improve clinical outcomes as measured by live birth rates. Understanding the mechanisms regulating the endometrial receptivity during the window of implantation is a critical step toward developing clinically meaningful tests. In this narrative review, the available literature is evaluated regarding mechanisms that regulate the endometrial receptivity during the window of implantation and the current tests developed. Overall, both animal and human studies point to five possible and interrelated mechanisms regulating the endometrial window of implantation: suitable synchrony between endometrial cells, adequate synchrony between the endometrium and the embryo, standard progesterone signaling and endometrial responses to progesterone, silent genetic variations, and typical morphological characteristics of the endometrial glands. The biological basis of current clinical markers or tests of window of implantation is poor. Future studies to elucidate the mechanisms shaping the window of implantation and to investigate the potential markers based on these mechanisms are required. In addition, molecular testing of the endometrium at single-cell resolution should be an initial step toward developing clinically meaningful tests for the optimal window of implantation. As understanding of the optimal window of implantation continues to evolve, one can envision the future development of non-invasive, mechanism-based testing of the window of implantation.
... As mentioned previously, laboratory findings of pregnancy include detection of human chorionic gonadotropin (hCG) in blood or urine. hCG is secreted into the maternal circulation after implantation, which may occur as early as 6 days after ovulation but typically occurs eight to ten days after ovulation [4][5][6][7]. This is the earliest that hCG can be detected with a standard serum hCG test. ...
... This is the earliest that hCG can be detected with a standard serum hCG test. However, the ovulation-toimplantation interval has been observed to vary by up to six days in naturally conceived pregnancies [4]. The hCG concentration doubles every 29 to 53 hours during the first 30 days after implantation of a viable, intrauterine pregnancy [1]. ...
Article
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Pregnancy should be suspected whenever a woman in her childbearing years misses a menstrual period. Clinical suspicion is increased if she also reports any sexual activity while not using contraception or is inconsistent in her use of contraception. Laboratory findings that aid in the diagnosis of pregnancy include the detection of human chorionic gonadotropin (hCG) in blood or urine. Hydatidiform mole (HM) is part of a group of diseases classified under gestational trophoblastic disease (GTD), which originate in the placenta and have the potential to locally invade the uterus and metastasize. Although molar pregnancies are designated as benign, they have the potential to develop into a malignancy. In this case study, we present a 48-year-old peri-menopausal female patient, with a 1+ year history of irregular menses, who presented to the clinic with signs and symptoms of pregnancy, unprotected sexual activity, and a positive at-home pregnancy test. Upon further workup of the patient, it was diagnosed that the patient had a hydatidiform molar pregnancy. It is interesting to note that benign gestational trophoblastic diseases generally occur in younger women, of "reproductive age" (generally in their twenties to early thirties), and is extremely rare in peri- and post-menopausal women.
... A short delay of implantation (deferred implantation) caused by targeted gene deletion, such as Pla2g4a, Lpar3, Klf5 or Msx1 missing in mice, created a negative ripple effects on the late events of pregnancy leading to poor pregnancy outcome [7][8][9][10]. In human, late implantation may cause miscarriage [11]. At present, several different protocols have been used to induce delayed implantation in wide type mice and rats. ...
Preprint
Implantation timing is key for a successful pregnancy. Short delay in embryo implantation caused by targeted gene ablation produced a cascading problem in the later stages of the pregnancy. Although several delayed implantation models have been established in wild mice, almost none of them is suitable for investigating the delay on the late events of pregnancy. Here, we report a new delayed implantation model established by the intraperitoneally administration of letrozole at 5 mg/kg body weight on the day 3 of pregnancy. In these mice, initiation of implantation was induced at will by the injection of estradiol (E2). When the estradiol (3 ng) was injected on day 4 of pregnancy (i.e., without delay), the embryo implantation restarted, and the pregnancy continued normally. However, high dose of estrogen (25 ng) caused compromised implantation. We also found that only 67% of the female mice could be pregnant normally and finally gave birth when the injection of estradiol (3 ng) was on day 5 of pregnancy (i.e., one day delay). Most of the failed pregnancies had impaired decidualization, decreased plasma progesterone levels and compromised angiogenesis. Progesterone supplementation could rescue decidualization failure in the mice. Collectively, we established a new model of delayed implantation by letrozole, which can be easily used to study the effect and mechanisms of delay of embryo implantation on the progression of late pregnancy events.
... 32 The idea of an optimal WOI is supported by a subsequent study that demonstrated that delayed implantation on the edge of the WOI can lead to an increased risk of early pregnancy loss due to abnormal placentation. 33,34 In our study, the rates of abnormal chromosomal detection in the POC were similar between the pET and npET groups. If the endometrial receptivity test could reduce the miscarriage rate rather than chromosomal abnormality, the chromosomal abnormality rate would have been higher in the pET group TA B L E 1 RIF patients' profiles and reproductive outcomes before propensity score matching of the personalized embryo transfer (pET) and non-personalized embryo transfer (npET) groups Abbreviations: POC, products of conception; RIF, recurrent implantation failure; SD, standard deviation. ...
Article
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Purpose: To assess the clinical efficacy of personalized embryo transfer (pET) guided by a new endometrial receptivity test, ERPeakSM, in patients with recurrent implantation failure (RIF). Methods: Recurrent implantation failure patients of all ages at two private Japanese clinics from April 2019 to June 2020 were retrospectively analyzed. The intervention group (n = 244) received pET in accordance with endometrial receptivity testing results and was compared to control group (n = 306) receiving standardized timing, non-personalized embryo transfer (npET). In propensity score matching analysis, the clinical pregnancy rate (CPR) and live birth rate (LBR) were compared between groups, and a subanalysis of advanced maternal age (AMA) (≥38 years old) versus non-AMA (<38 years old) patients was also conducted. Results: The CPR and LBR of the pET group were significantly higher than those of the npET group (37.7% vs. 20.0%, adjusted OR: 2.64; 95%CI, 1.70-4.11, p < 0.001 and 29.9% vs. 9.7%, adjusted OR: 4.13; 95%CI, 2.40-7.13, p < 0.001, respectively). Furthermore, in the subanalyses, the CPR and LBR of the pET group were significantly higher than those of the npET group in both the AMA non-AMA patients. Conclusions: The new ERPeakSM endometrial receptivity test is a useful alternative diagnostic tool for poor-prognosis patients, regardless of age.
... When the uterus is well prepared, a short period, known as window of implantation, occurs for embryos to implant. Implantation out of window of implantation would lead to spontaneous miscarriages (3). Through the processes of apposition, attachment (adhesion) and penetration, the blastocyst implant to the receptive endometrium. ...
Article
Full-text available
Extracellular vesicles (EVs) are membrane-coating nanoparticles derived from cells. The effect of cell-to-cell communication mediated by EVs has been investigated in different fields of physio-logical as well as pathological process in recent years. Reproduction, regarded as a definitive characteristic of organisms, has been a focus in both animal and medical sciences. It is well agreed that implantation is a critical event during early pregnancy in viviparous animals, and a proper implantation is essential for the establishment and maintenance of normal pregnancy. However, successful implantation requires the synchronized development of both the uterus and the embryo, therefore, in which well communication and opportune regulation are necessary. This review focuses on the progression of studies that reveal the role of EVs in early pregnancy, especially during implantation. Based on current evidence, EVs are produced and exist in the environment for implantation. It has been proved that EVs of different origins such as endometrium and embryo, have positive influences on embryo implantation. With their cargos of proteins and nucleic acids (especially microRNAs), EVs exert their effects including information transportation, immune stimulation and regulation of gene expression.
... In the midst of these complex changes exists a "window of implantation," a roughly 3-6-day period in which the architecture and hormonal make-up of the endometrium is optimal for implantation to occur [2,6]. While the sequential actions of estrogen and progesterone are clearly central in the timing of this critical period, the exact molecular mechanisms governing the transition from non-receptive to receptive endometrium remain poorly understood. ...
Article
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Purpose of Review The purpose of this review is to update readers on our current understanding of endometrial receptivity and to review the literature regarding endometrial assessment and pathologic states which may influence receptivity. Recent Findings Topics discussed include molecular mechanisms implicated in endometrial receptivity on the basis of which three commercially available yet controversial tests have been developed (Endometrial Function Test, Endometrial Receptivity Assay, and ReceptivaDx), as well as current evidence surrounding how common gynecologic conditions such as polyps, fibroids, adenomyosis, and chronic endometritis may influence endometrial receptivity and the success of assisted reproductive technologies (ART). Summary While the sequential action of estrogen and progesterone is central to the development of endometrial receptivity, the exact molecular mechanisms governing the transition from non-receptive to receptive endometrium remain poorly understood. Commercially available tests attempt to harness molecular markers to assess and potentially improve endometrial receptivity; however, current evidence is insufficient to support their routine clinical application. Polyps, fibroids, adenomyosis, and chronic endometritis may negatively impact implantation either directly by impeding embryo transport and attachment or indirectly by influencing endometrial receptivity. Given the likely heterogenous genetic construct and molecular expression of any pathologic condition, further research is needed to develop molecular testing to guide approach to clinical treatment.
... embryo, meanwhile, delayed implantation will bring early pregnancy loss (17). Several studies have been conducted to explore the question that "What is the optimal duration of progesterone administration before transferring a vitrifiedwarmed blastocyst", but no consensus has been reached (12,13). ...
Article
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Background: Progesterone administration before transfer in hormone replacement treatment (HRT) is crucial to pregnancy outcomes in frozen-thawed blastocyst transfer (FET), but the optimal progesterone duration is inconsistent. The objective of this study was to investigate live birth rate (LBR) of different progesterone duration before blastocyst transfer in HRT-FET cycles. Method: In this retrospective cohort study, patients underwent first HRT-FET (including suppression HRT) from January 2016 to December 2019 were included. Logit-transformed propensity score matching (PSM) was performed to assess covariates. The primary outcome was live birth rate after 28 weeks' gestation. Basing on different duration of progesterone before transfer, patients were classified into P6-protocol (blastocyst transfer performed on the sixth day), or P7-protocol (blastocyst transfer performed on the seventh day). Subgroup analyses were conducted as follows: age stratification (-35, 35-38, 38-), development days of blastocyst (D5 or D6), blastocyst quality (high-quality or poor-quality), and endometrial preparation protocols (HRT or suppression HRT). Result: After case matching with propensity score methods, a total of 1,400 patients were included finally: 700 with P6-protocol and 700 with P7-protocol. Significantly higher live birth rate (38.43% versus 31.57%, respectively, P = 0.01) and clinical pregnant rate (50.43% versus 44.14%, respectively, P = 0.02) were observed in P6-protocol than those of P7-protocol. First-trimester abortion rates (18.13% versus 20.71%, P = 0.40) and ectopic pregnancy rates (2.27% versus 1.94%, P = 0.77) were similar between P6- and P7-groups. Preterm birth rate, low birth weight rate, newborn sex proportion, neonatal malformation rate were comparable between groups. Significantly higher LBRs were observed in patients with: age under 35, D5 blastocyst transfer, high-quality blastocyst transfer, and undergoing HRT cycles combined P6-protocol. Conclusion: Frozen-thawed blastocyst transfer on the sixth day of progesterone administration in first HRT cycle is related to higher live birth rate compared with transfer on the seventh day, especially among patients aged under 35, D5 blastocyst and/or high-quality blastocyst transfer.
... The duration of the pre-implantation phase of the embryo varies from species to species. In mice, implantation occurs 4 days after intercourse: in humans it is on average 9 days, while in cows, implantation takes place 30 days after fertilization [26]. According to the different types of blastocyst-uterine cell interactions, there are three main types of implantation: central, eccentric and interstitial [27]. ...
Article
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Endometrial receptivity plays a crucial role in fertilization as well as pregnancy outcome in patients faced with fertility challenges. The optimization of endometrial receptivity may help with normal implantation of the embryo, and endometrial receptivity may be affected by numerous factors. Recently, the role of lipids in pregnancy has been increasingly recognized. Fatty acids and their metabolites may be involved in all stages of pregnancy and play a role in supporting cell proliferation and development, participating in cell signaling and regulating cell function. Polyunsaturated fatty acids, in particular, are essential fatty acids for the human body that can affect the receptivity of the endometrium through in a variety of methods, such as producing prostaglandins, estrogen and progesterone, among others. Additionally, polyunsaturated fatty acids are also involved in immunity and the regulation of endometrial decidualization. Fatty acids are essential for fetal placental growth and development. The interrelationship of polyunsaturated fatty acids with these substances and how they may affect endometrial receptivity will be reviewed in this article.
... Because the successful blastocyst implantation is closely related to the state of the endometrium and its acceptance time window, the establishment and maintenance of a normal pregnancy require sufficient crosstalk between the endometrium and the embryo and the embryo best microenvironment in uterus. [13][14][15] In addition, abnormal hormone and inflammatory environments may disturb the dialog of immune-endocrine interaction between the decidua and trophoblast during embryo implantation, leading to pregnancy complications. [16][17][18][19] In this review, we would like to explain the risk of intrauterine exposure to the SARS-CoV-2 virus in female patients with COVID-19 and whether it could interrupt the development of pregnancy. ...
Article
Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) is a new type of coronavirus that has caused fatal infectious diseases and global spread. This novel coronavirus attacks target cells through the interaction of spike protein and angiotensin‐converting enzyme II (ACE2), leading to different clinical symptoms. However, for a successful pregnancy, a well‐established in‐uterine environment includes a specific immune environment, and multi‐interactions between specific cell types are prerequisites. The immune‐related changes in patients infected with novel coronavirus could interfere with the immune microenvironment in the uterus, leading to fetal loss. We first reviewed the intrauterine environment in the normal development process and the possible pregnancy outcome in the infection state. Then, we summarized the immune response induced by SARS‐CoV‐2 in patients and analyzed the changes in ACE2 expression in the female reproductive system. Finally, the present observational evidence of infection in pregnant women was also reviewed. This article is protected by copyright. All rights reserved
... We previously provided evidence that embryo implantation is aberrantly delayed in suboptimal uterine conditions, including AS [22,28,44]. The unsynchronized interactions between the blastocyst and unhealthy uterus, such as in AS, often lead to IUGR and/or pregnancy loss in both mice and humans [22,28,45]. Patients with AS have a relatively higher risk of developing IUGR during pregnancy [46,47]. ...
Article
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Asherman’s syndrome (AS) is caused by intrauterine adhesions and inactive endometrium from repeated curettage of the uterine endometrium. AS is a major cause of recurrent implantation failure and miscarriage and is very difficult to treat because of the poor recovery of endometrial basal cells. Platelet-rich plasma (PRP) has abundant growth factors that may induce angiogenesis and cell proliferation. Here, we demonstrate that human PRP (hPRP) significantly enhances angiogenesis to restore embryo implantation, leading to successful pregnancy in mice with AS. In mice with AS, hPRP treatment considerably reduced the expression of fibrosis markers and alleviated oligo/amenorrhea phenotypes. Mice with AS did not produce any pups, but the hPRP therapy restored their infertility. AS-induced abnormalities, such as aberrantly delayed embryo implantation and intrauterine growth retardation, were considerably eliminated by hPRP. Furthermore, hPRP significantly promoted not only the elevation of various angiogenic factors, but also the migration of endometrial stromal cells. It also increased the phosphorylation of STAT3, a critical mediator of wound healing, and the expression of tissue remodeling genes in a fibrotic uterus. PRP could be a promising therapeutic strategy to promote angiogenesis and reduce fibrosis in impaired uterine environments, leading to successful embryo implantation for better clinical outcomes in patients with AS.
... Successful embryo implantation requires two conditions to be met; the presence of a highquality embryo and a receptive endometrium. Endometrial receptivity is a limited time period during the menstrual cycle when the endometrium enables embryo implantation and is also called the window of implantation (WOI) [1]. It has been estimated that one-third of embryo implantation failures occur due to an unreceptive endometrium [2]. ...
Article
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Obesity and being overweight are growing worldwide health problems that also affect women of reproductive age. They impair women’s fertility and are associated with lower IVF success rates. The mechanism by which increased body weight disrupts fertility has not yet been established. One possibility is that it affects the process of embryo implantation on the endometrial level. The purpose of our study was to determine the differences in enriched biological pathways in the endometrium of overweight and obese women undergoing IVF procedures. For this purpose, 14 patients (5 pregnant, 9 non-pregnant) were included in the study. Endometrial samples were obtained during the window of implantation and RNA sequencing was performed. There were no differences in general patient’s and IVF cycle characteristics between pregnant and non-pregnant women. In the endometrial samples of women who did not conceive, pathways related to the immune response, inflammation, and reactive oxygen species production were over-expressed. Our findings show that the reason for implantation failure in overweight and obese women could lie in the excessive immune and inflammatory response at the endometrial level.
... Studies have previously discovered that failure in pregnancy arises mostly during the period of embryo implantation [5]. The complex but vital step of embryo implantation in establishing pregnancy can only occur in a limited window once the unreceptive uterine lining converts to a hospitable environment that is able to receive the embryo [6]. ...
Article
Full-text available
Extracellular vesicles (EVs) are small, nanometre sized, membrane-enclosed structures released by cells and are thought to be crucial in cellular communication. The cargo of these vesicles includes lipids, proteins, RNAs and DNA, and control various biological processes in their target tissues depending on the parental and receiver cell’s origin and phenotype. Recently data has accumulated in the role of EVs in embryo implantation and pregnancy, with EVs identified in the uterine cavity of women, sheep, cows, horses, and mice, in which they aid blastocyst and endometrial preparation for implantation. Herein is a critical review to decipher the role of extracellular vesicles in endometrial receptivity and their potential in reproductive therapies and diagnosis. The current knowledge of the function of embryo and endometrial derived EVs and their cargoes, with regards to their effect on implantation and receptivity are summarized and evaluated. The findings of the below review highlight that the combined knowledge on EVs deriving from the endometrium and embryo have the potential to be translated to various clinical applications including treatment, a diagnostic biomarker for diseases and a drug delivery tool to ultimately improve pregnancy rates.
... During this period, the endometrial environment is most suitable for embryo implantation. An embryo can only be successfully implanted during this state of endometrial receptivity (8). Many studies have shown that abnormal endometrial receptivity is associated with decidualization in response to hormone dysfunction (9,10). ...
Article
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Background Recurrent implantation failure (RIF) is a disease associated with endometrial receptivity dysfunction. Retinoic acid receptor alpha (RARα) is an important protein in many biological processes, such as differentiation and development. However, the exact underlying mechanism whereby RARα affects RIF remains unknown. This study investigated RARα expression and its contribution in the mid-luteal phase endometria of patients with RIF. Methods The expression levels of RARα and CCAAT/enhancer-binding protein (C/EBP) β in the endometria of the RIF and normal group were investigated using western blotting and immunohistochemistry. In in vitro experiments, immortal telomerase-transformed human endometrial stromal cells (T-HESCs) were incubated with medroxyprogesterone-17-acetate (MPA) and cyclic adenosine monophosphate (cAMP) for 4 days to induce decidualization. The expression levels of the decidualization markers prolactin (PRL) and insulin-like growth factor-binding protein-1 (IGFBP-1) were determined using quantitative polymerase chain reaction. RARα was knocked down using a small interfering RNA, and C/EBPβ was overexpressed from an adenoviral vector. The transcriptional regulation of CEBPB by RARα was determined by chromatin immunoprecipitation (ChIP) assay and luciferase assays. Results We found that the expression levels of RARα decreased in the mid-luteal endometria of RIF patients. After 4 days of decidualization induction in vitro , RARα knockdown impaired the decidualization of T-HESCs and downregulated the expression of C/EBPβ. The restoration of C/EBPβ expression rescued the RARα knockdown-induced suppression of T-HESC decidualization. In ChIP analysis of lysates from decidualized T-HESCs, the CEBPB promoter region was enriched in chromatin fragments pulled down using an anti-RARα antibody. However, the relationship between CEBPB transcription and RARα expression levels was only observed when the decidualization of T-HESCs was induced by the addition of cAMP and MPA. To identify the binding site of RARα/retinoid X receptor α, we performed luciferase assays. Mutation of the predicted binding site in CEBPB (-2,009/-1,781) decreased the transcriptional activity of the reporter. To confirm this mechanism, the expression levels of C/EBPβ in the mid-luteal endometria of RIF patients were determined and found to decrease with decreased RARα expression levels. Conclusion A deficiency of RARα expression in the mid-luteal endometrium inhibits decidualization due to the downregulation of CEBPB transcription. This is a potential mechanism contributing to RIF.
... Endometrial receptivity refers to the endometrium's ability to accept embryos, indicating that it is in a state that allows embryonic localization, adhesion, and invasion [16][17][18]. This period is also known as the window of implantation (WOI) [19,20]. In 2011, Spanish scholars developed a genetic diagnostic tool for assessing endometrial receptivity based on gene expression microarray technology, called endometrial receptivity array [21]. ...
Article
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Background Today, approximately 10% of participants in assisted reproductive technology (ART) are defined as having recurrent implantation failure (RIF). Recent studies show that endometrial receptivity array can improve pregnancy and implantation rates by nearly 20% in women with RIF. However, these studies are limited, with little published data in the Chinese population. Recently, we have established a transcriptome-based endometrial receptivity assessment (Tb-ERA) method of predicting the endometrial window of implantation (WOI) using transcriptome-profiling data of different phases of the menstrual cycle from healthy fertile Chinese women by RNA-Seq. It is meaningful to conduct a randomized controlled trial (RCT) to assess the clinical efficiency of Tb-ERA in Chinese patients with RIF. Methods In this RCT, a total of 200 RIF patients will be recruited and randomized into 2 groups. Patients in the Tb-ERA group will undergo a Tb-ERA test, after which embryo transfer time will be adjusted according to Tb-ERA results and embryo transfer will be performed again in the next cycle. Patients in the control group will not receive any interventions until the next transfer cycle. We will perform statistical analysis on both groups at the primary endpoint (clinical-pregnancy rate) and at secondary endpoints (rate of WOI displacement, embryo implantation, biochemical pregnancy, early abortion, and ectopic pregnancy). Implications: This study aims to evaluate the effectiveness of our Tb-ERA test in Chinese RIF patients and to determine that whether Tb-ERA could improve the clinical-pregnancy rate in these RIF patients. Trial registration NCT04497558, registered August 4, 2020.
... [10][11][12] It is known that the human endometrium becomes receptive only during the implantation window, 10,13-16 a certain period that results from the synchronized interaction of a variety of molecules (ovarian hormones, growth factors, transcription factors, cytokines, adhesion molecules), with an important role in establishing uterine receptivity. [16][17][18][19][20][21][22] Thus, molecular changes in the eutopic endometrium of these patients could impair their endometrial receptivity, contributing to the infertility observed in women with the disease. ...
Article
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Objective Abnormalities in the eutopic endometrium of women with endometriosis may be related to disease-associated infertility. Although previous RNA-sequencing analysis did not show differential expression in endometrial transcripts of endometriosis patients, other molecular alterations could impact protein synthesis and endometrial receptivity. Our aim was to screen for functional mutations in the transcripts of eutopic endometria of infertile women with endometriosis and controls during the implantation window. Methods Data from RNA-Sequencing of endometrial biopsies collected during the implantation window from 17 patients (6 infertile women with endometriosis, 6 infertile controls, 5 fertile controls) were analyzed for variant discovery and identification of functional mutations. A targeted study of the alterations found was performed to understand the data into disease's context. Results None of the variants identified was common to other samples within the same group, and no mutation was repeated among patients with endometriosis, infertile and fertile controls. In the endometriosis group, nine predicted deleterious mutations were identified, but only one was previously associated to a clinical condition with no endometrial impact. When crossing the mutated genes with the descriptors endometriosis and/or endometrium, the gene CMKLR1 was associated either with inflammatory response in endometriosis or with endometrial processes for pregnancy establishment. Conclusion Despite no pattern of mutation having been found, we ponder the small sample size and the analysis on RNA-sequencing data. Considering the purpose of the study of screening and the importance of the CMKLR1 gene on endometrial modulation, it could be a candidate gene for powered further studies evaluating mutations in eutopic endometria from endometriosis patients.
... Although sparse evidence shows that a large delay between transfer and implantation is possible (e.g., one case study shows a 5-week delay Grinsted and Avery, 1996), large-scale studies have not found such outliers. Naturally conceived human embryos implant 7-11 days after ovulation, although a range of 6-18 days was observed (Wilcox et al., 1999). Importantly, late implantation is associated with a higher rate of pregnancy loss. ...
Article
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The energetically costly mammalian investment in gestation and lactation requires plentiful nutritional sources and thus links the environmental conditions to reproductive success. Flexibility in adjusting developmental timing enhances chances of survival in adverse conditions. Over 130 mammalian species can reversibly pause early embryonic development by switching to a near dormant state that can be sustained for months, a phenomenon called embryonic diapause. Lineage-specific cells are retained during diapause, and they proliferate and differentiate upon activation. Studying diapause thus reveals principles of pluripotency and dormancy and is not only relevant for development, but also for regeneration and cancer. In this review, we focus on the molecular regulation of diapause in early mammalian embryos and relate it to maintenance of potency in stem cells in vitro. Diapause is established and maintained by active rewiring of the embryonic metabolome, epigenome, and gene expression in communication with maternal tissues. Herein, we particularly discuss factors required at distinct stages of diapause to induce, maintain, and terminate dormancy.
... The endometrium is receptive during the window of implantation (WOI) regulated by an incompletely understood endocrine, paracrine, and autocrine factors (136). During the natural cycle, the WOI is limited to days 8-10 after ovulation during which the blastocyst may implant (137). However, there seems to be marked interpersonal differences in the timing of the WOI that cannot be elucidated by ultrasonographic, hormonal or histologic assessments (138). ...
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Despite the worldwide increase in frozen embryo transfer, the search for the best protocol to prime endometrium continues. Well-designed trials comparing various frozen embryo transfer protocols in terms of live birth rates, maternal, obstetric and neonatal outcome are urgently required. Currently, low-quality evidence indicates that, natural cycle, either true natural cycle or modified natural cycle, is superior to hormone replacement treatment protocol. Regarding warmed blastocyst transfer and frozen embryo transfer timing, the evidence suggests the 6th day of progesterone start, LH surge+6 day and hCG+7 day in hormone replacement treatment, true natural cycle and modified natural cycle protocols, respectively. Time corrections, due to inter-personal differences in the window of implantation or day of vitrification (day 5 or 6), should be explored further. Recently available evidence clearly indicates that, in hormone replacement treatment and natural cycles, there might be marked inter-personal variation in serum progesterone levels with an impact on reproductive outcomes, despite the use of the same dose and route of progesterone administration. The place of progesterone rescue protocols in patients with low serum progesterone levels one day prior to warmed blastocyst transfer in hormone replacement treatment and natural cycles is likely to be intensively explored in near future.
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Introduction . It is now generally acknowledged that cytomegalovirus infection (CMVI) is one of the main causes of miscarriage. However, the mechanism of this effect has not been sufficiently studied. At the same time, the influence of acids of the ɷ-3 family and α-tocopherol (αTP) on the placentation process through a pro-angiogenic action is shown. Aim. To study the concentration of αTP and ɷ-3 family acids in the peripheral blood and establish their role in miscarriage in CMV-seropositive women with CMVI reactivation. Materials and methods . A case-control study included 64 women in the first trimester of pregnancy (7-10 weeks), of which 36 were CMV-seropositive with CMVI reactivation (main group) and 28 were CMV-seronegative (control group). CMVI was diagnosed by the determination of class M and G antibodies by ELISA, as well as CMV DNA detected by PCR. The concentration of ɷ-3 acids of the family (eicosapentaenoic – EPA, docosahexaenoic – DHA) in blood serum was studied by capillary gas-liquid chromatography (J.P.Carreau, J.P.Dubacq). The αTP concentration was determined by the fluorometric method (L.G.Hansen, W.I.Warwich). Results . In women of the main group, a significant (p<0.001) decrease in the concentration of αTP to 1.32±0.025 μg/mL was observed in the peripheral blood compared to the same indicator in the control group (1.49±0.029 μg/mL). At the same time, the levels of EPA and DHA were also statistically significant (p<0.001) lower than the same indicator in the control group and amounted to 1.09±0.012 and 6.09±0.015%, respectively (in the control, 1.29±0.071 and 8.80±0.071%, respectively). Conclusion. The obtained results of the study allow us to establish the important role of disorders in the content of α-TF, EPA and DHA in the pathogenesis of miscarriage during reactivation of CMVI in the early periods of gestation, which can serve as a basis for expanding diagnostic and therapeutic measures in this pathology of pregnant women.
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This review discusses why the embryo requires vitamin E (VitE) and shows that its lack causes metabolic dysregulation and impacts morphological changes at very early stages in development, which occur prior to when a woman knows she is pregnant. VitE halts the chain reactions of lipid peroxidation (LPO). Metabolomic analyses indicate that thiols become depleted in E- embryos because LPO generates products that require compensation using limited amino acids and methyl donors that are also developmentally relevant. Thus, VitE protects metabolic networks and the integrated gene expression networks that control development. VitE is critical especially for neurodevelopment, which is dependent on trafficking by the α-tocopherol transfer protein (TTPa). VitE-deficient (E-) zebrafish embryos initially appear normal, but by 12 and 24 hours post-fertilization (hpf) E- embryos are developmentally abnormal with expression of pax2a and sox10 mis-localized in the midbrain-hindbrain boundary, neural crest cells and throughout the spinal neurons. These patterning defects indicate cells that are especially in need of VitE-protection. They precede obvious morphological abnormalities (cranial-facial malformation, pericardial edema, yolksac edema, skewed body-axis) and impaired behavioral responses to locomotor activity tests. The TTPA gene (ttpa) is expressed at the leading edges of the brain ventricle border. Ttpa knockdown using morpholinos is 100% lethal by 24 hpf, while E- embryo brains are often over- or under-inflated at 24 hpf. Further, E- embryos prior to 24 hpf have increased expression of genes involved in glycolysis and the pentose phosphate pathway, and decreased expression of genes involved in anabolic pathways and transcription. Combined data from both gene expression and the metabolome in E- embryos at 24 hpf suggest that the activity mechanistic Target of Rapamycin (mTOR) signaling pathway is decreased, which may impact both metabolism and neurodevelopment. Further evaluation of VitE deficiency in neurogenesis and its subsequent impact on learning and behavior is needed.
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Purpose To analyse the effect of ulipristal acetate (UPA) as emergency contraception (EC) on the gene expression of human endometrial cell line (HEC-1A) and endometrium from fertile women treated with UPA after ovulation. Materials and methods HEC-1A cells were treated with UPA, and endometrial tissue from four healthy women was collected in cycles before, during and 2 months after post-ovulation pill intake. Ovulation and luteal phase were monitored, and endometrial biopsies were obtained at day LH + 7 in each cycle. In all cases, we analysed the expression profile of 192 genes associated to endometrial receptivity. Results We observed a significant change in total transcriptomic activity of UPA-treated HEC-1A cells compared to controls. In vivo, we also observed a trend to down-regulation of genes in the UPA-treated cycle that was partially restored in the post-treatment cycle. Altogether, our results supported a partially reversible effect of UPA in gene expression associated with uterine receptivity. Conclusions When UPA was administered after ovulation, it seems to induce a down-regulation of the main genes involved in conditioning the endometrium for implantation. This effect is partially restored two months after pill intake. The action of UPA on the endometrium for users of EC should be further investigated.
Article
Research Question Is there any difference in the live birth rate between the natural cycle (NC) and hormone replacement therapy (HRT) endometrial preparation protocols for women with regular menstrual cycles undergoing their first single, frozen‒thawed euploid blastocyst transfer? Design This was a retrospective cohort study that enrolled 722 women who underwent frozen‒thawed euploid blastocyst transfer at the First Affiliated Hospital of Zhengzhou University assisted reproductive technology (ART) centre, from January 2013 to December 2019. Univariate and multivariate logistic regression models were used to analyse the relationship between the endometrial preparation protocols and live birth rates. Stratified analyses and sensitivity analyses were performed to ensure the reliability and stability of the results. Results A total of 722 single frozen‒thawed euploid blastocyst transfer cycles were included. Overall, the live birth rates were 50.00% (110/220) in the NC group and 47.61% (239/502) in the HRT group. Multiple logistic regression analyses showed that there was no significant association (adjusted odds ratio, 0.82; 95% confidence interval, 0.56‒1.20; P = 0.313) between NC or HRT protocols and the live birth rate. Interaction analysis showed that there was no significant difference in live birth rates between the two groups for any subgroup after adjusting for confounding factors. Conclusions For single, frozen‒thawed euploid blastocyst transfer, NC and HRT endometrial preparation protocols result in similar live birth rates among women with regular menstrual cycles. Further studies on the effects of endometrial preparation protocols on pregnancy outcomes are required.
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Purpose Does the OPtimization of Thyroid function, Thrombophilia, Immunity, and Uterine Milieu (OPTIMUM) treatment strategy, developed for treating repeated implantation failure (RIF), contribute to improving pregnancy outcomes in patients with a history of recurrent pregnancy loss (RPL)? Methods Between 2018 and 2019, women with RPL after two or more clinical pregnancy losses underwent RPL testing. We treated chronic endometritis with antibiotics, high Th1/Th2 cell ratios with vitamin D and/or tacrolimus, overt/subclinical hypothyroidism with levothyroxine, and thrombophilia with low-dose aspirin. Of 168 consecutive women aged ≤43 years, 115 underwent RPL testing. We compared 100 pregnancies (90 women) and 46 pregnancies (41 women) with and without the OPTIMUM treatment strategy, respectively. Results RPL testing identified intrauterine abnormalities in 66 (57.4%), elevated Th1/Th2 cell ratios in 50 (43.5%), thyroid dysfunction in 33 (28.7%), and thrombophilia in 33 (28.7%). The live birth rate in the OPTIMUM group was significantly higher than that in the control group among women aged <40 years (78.1% and 42.3%, respectively; p = 0.002), but no significant difference was observed in women aged ≥40 years (55.6% and 30.0%, respectively; p = 0.09). Conclusions The OPTIMUM treatment strategy improved pregnancy outcomes in patients with not only RIF but also RPL.
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PurposeTo evaluate optimal warming time, the early warming or the routine warming time, for transferring vitrified-warmed and cultured overnight cleavage stage of the slow-growing embryos on day 3 in frozen embryo transfer (FET) cycle.Methods This was a retrospective cohort study from January 2017 to July 2018. A total of 705 FET patients aged < 40 years were included and 1486 embryos were formed, of which 1366 embryos were eventually transferred.ResultsFor slow-growing embryos, the clinical pregnancy rate of early warming group [152/468 (32.5%)] was significantly higher than that of routine warming group (55/235 (23.4%)) [OR 1.39 (CI 1.06–1.81), p = 0.01], while there was no statistically significant difference in pregnancy loss in early warming group [39/170 (22.9%)] versus in routine warming group [16/62 (25.8%)] [OR 0.89 (CI 0.53–1.47), p = 0.65].Conclusion For slow-growing embryos, higher pregnancy outcomes were shown in early warming strategy as compared to the routine warming, which suggested that the improvement of endometrium–embryo synchronism may correct the time difference brought by the slow-growing embryos.
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Many factors impede embryonic implantation, and excluding obvious known factors such as chronic endometritis, the immune status of the endometrium may be related to pregnancy. Although an abundantly large number of immune cells infiltrate the endometrium during the secretory phase, whether these immune cells can be used as a predictor of prognosis in ART has not yet been clarified. In the present study we therefore retrospectively analyzed 97 CD138-negative women with a previous fresh-embryo-transfer failure. We assessed the expression of CD56+ uNK cells, CD16 + NK cells, CD57 + NK cells, CD68+ pan-macrophages, CD163 + M2 macrophages, CD4 + T cells, CD8 + T cells, FOXP3+ regulatory T cells, and CD19 + B cells in the endometrium by IHC to evaluate mid-luteal endometrial immune cells as prognostic indicators of pregnancy outcome in the next frozen-embryo-transfer cycle. CD19-positive cells and the intraglandular CD163-positivity rate increased significantly in the clinically non-pregnant group (0.47% vs. 0.20%, P = 0.021; 61% vs. 30%, P = 0.017). The ratios of CD4/CD8 were also higher in the non-pregnant group (1.96 vs. 1.45, P = 0.005).The area under the ROC curve of CD19 cell number alone, the intraglandular CD163-positivity alone, and CD19 number combined with the intraglandular CD163-positivity were 0.692 (95% CI, 0.55–0.834), 0.661 (95% CI, 0.514–0.809), and 0.748 (95% CI, 0.614–0.882), respectively. The optimal cut-off value of CD19 was 0.464%, and the clinical pregnancy rate and live-birth rate diminished significantly when the CD19 level was above this cut-off value. Our study suggests that CD19-positive cells and intraglandular CD163-positivity can be used as prognostic indicators of pregnancy outcome in CD138-negative patients who experienced first-fresh-embryo transfer failure.
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Repeated implantation failure (RIF) is a major problem that limits the pregnancy rate associated with assisted reproductive technology. However, the pathogenesis of RIF is still unknown. Recently, the expression levels of circular RNAs (circRNAs) were profiled in the endometrial tissues of patients with RIF. However, the exact role of circRNAs in RIF remains unclear. In our study, we found that circFAM120A levels were significantly down‐regulated in the endometrium at the window of implantation in RIF patients compared with non‐RIF controls. The suppression of circFAM120A expression inhibited decidualization in human endometrial stromal cells (hESCs). Furthermore, RNA‐seq analysis after circFAM120A knockdown revealed ABHD5 as a potential downstream target gene of circFAM120A. As expected, down‐regulating ABHD5 in hESCs also inhibited decidualization. Using the starBase and TargetScan databases, we predicted that miR‐29 may interact with ABHD5, based on nucleotide sequence matching. Luciferase reporter assay showed that miR‐29 bound to the 3′ UTR of ABHD5 at the predicted complementary sites. Moreover, miR‐29 mimics efficiently reduced ABHD5 expression levels and suppressed the decidualization process, whereas a miR‐29 inhibitor partly rescued ABHD5 mRNA expression level and decidualization reduced by the knockdown of circFAM120A. Therefore, circFAM120A modulated decidualization in RIF through the miR‐29/ABHD5 axis.
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With continuous improvement of sow litter size, breeders are gradually paying more attention to the quality of litter traits that directly impact the production efficiency of pig companies, such as the rate of piglets born alive (RBA) and the rate of healthy births (RHB). The objectives of this study are to dissect the genetic basis of litter traits in pig and to identify valuable genes and genetic markers, especially pleiotropic, for pig breeding. Herein, 1140 Duroc pigs and 2046 reproduction records, 5 litter traits, including the number of healthy births (NHB), number of deformed fetuses (NDF), number of stillborn (NSB), RBA, and RHB, were used in this study. Subsequently, a genome-wide association study (GWAS) was performed for the five litter traits in the first two parities from two Duroc populations. A total of 76 significantly related SNPs and 10 potential candidate genes (CAV1, DAB2, FGF12, FHOD3, DYNC2H1, GRHL1, TCTN3, PYROXD2, MMP8, MMP13, and PGR) were detected, including 13 pleiotropic SNPs that affected more than one litter trait. Finally, the functional enrichment analysis of functional genes that were closest to these significant SNPs indicated that most of the significant pathways were associated with hormone secretion and embryo and organ development. This study advances our understanding of the genetic mechanisms of litter traits, especially the survival rate of piglets born, and provides an opportunity to increase the quality of litter using marker-assisted selection or genomic selection in pigs.
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Background A few endocrine disrupting chemicals (EDCs) have been associated with pregnancy loss often as reported by women, though there has been no study of EDC mixtures and pregnancy loss in keeping with the nature of human exposure. Objectives To investigate preconception exposure to a mixture of EDCs to identify important drivers and inform multi-pollutant models of EDCs in relation to incident human gonadrophin chorionic (hCG) pregnancy loss. Methods A cohort of 501 couples were recruited from the general population and prospectively followed until a hCG-confirmed pregnancy or 12 months of trying to become pregnant. Pregnant (n = 344; 69%) women were followed daily through seven weeks post-conception then monthly until delivery. Loss was defined as conversion to negative pregnancy test or a clinical diagnosis. Preconception exposure assessment of EDCs included sixty-three serum chemicals and three blood metals. EDCs were measured using isotope dilution gas chromatography-high resolution mass spectrometry or high-performance liquid chromatography-tandem mass spectrometry, and inductively coupled plasma-mass spectrometry, respectively. Using elastic net variable selection to identify important factors from the exposure mixture, EDC levels and covariates were then included in Cox proportional hazard models to estimate hazard ratios (HRs) and 95% confidence intervals (CIs) of time-to-pregnancy loss in multi-pollutant models. Results Incidence of hCG pregnancy loss was 28%. Nine EDCs of the sixty-six chemical mixture were associated with pregnancy loss; HRs were elevated for polychlorinated biphenyl 194, 2-(N-methyl-perfluorooctane sulfonamido) acetate, polybrominated diphenyl ether 28, and cadmium, even in sensitivity models adjusting for male partners’ EDC concentrations. In final multivariable multi-pollutant Cox proportional hazard models, female partners’polybrominated diphenyl ether 28 (aHR = 1.16, 95% CI: 1.02, 1.31) and cadmium (aHR = 1.23, 95% CI: 1.07, 1.40) remained associated with hCG pregnancy loss. Female partners’ preconception serum polychlorinated biphenyl 194 and 2-(N-methyl-perfluorooctane sulfonamido) acetate concentrations were consistently inversely associated with loss [(aHR = 0.72, 95% CI: 0.56, 0.92) and (aHR = 0.79, 95% CI: 0.65, 0.95), respectively]. Conclusion Assessing exposure to a mixture of 66 persistent EDCs, females’ preconception concentrations of polybrominated diphenyl ether 28 and cadmium were positively associated with incident hCG pregnancy loss in a cohort of couples from the general population trying for pregnancy.
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The goal of this study was to identify genes that were differentially methylated and differentially expressed and their related signaling pathways in ovarian endometriosis tissue. First, the DNA methylation and gene expression profiles in the endometrial tissue of patients with ovarian endometriosis were studied using Illumina 450K methylation microarray analysis and the GSE141549 gene expression dataset. Second, differentially methylated and differentially expressed genes, herein referred to as differentially methylated/expressed genes, were identified and protein-protein interaction networks and functional analysis of these genes were determined. Third, qPCR and immunohistochemistry of patient samples was used to confirm the differential expression of a subset of differentially methylated/expressed genes. Finally, the GSE7305 dataset was used confirm the expression profile of differentially methylated/expressed genes and to determine the potential usefulness of these genes for diagnosis of endometriosis. A total of 37 hypermethylated low-expression genes and 66 hypomethylated high-expression genes were identified in ovarian endometriosis patients. Protein-protein interaction and functional analysis highlighted 8 hypermethylated low-expression genes (KRT19, KRT8, ESR1, PRL, SFN, IL20RA, IL2RB, and PAX8) and 4 hypomethylated high-expression genes (CYP11A1, NR5A1, ME1, and GSTM1). Significantly, both of these gene sets had a diagnostic value for patients with ovarian endometriosis. Signaling pathways that were identified included JAK-STAT (involving IL20RA and IL2RB), prolactin (involving PRL and ESR1), Staphylococcus aureus infection (involving KRT19), viral protein interaction with cytokine and cytokine receptor (involving IL20RA and IL2RB), cytokine-cytokine receptor interaction (involving IL20RA and IL2RB), and drug metabolism-cytochrome P450 (involving GSTM1). The differentially methylated/expressed genes and enriched signaling pathways identified in this study are likely to be associated with the process of ovarian endometriosis.
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Since the advent of ART, technology has continuously evolved to improve embryology and pregnancy outcomes. However, not all technologies that are integrated into practice have convincing evidence of clinical effectiveness, and they often increase the financial burden of fertility care. We discuss here a selection of commonly utilized IVF “add-ons” and discuss the existing evidence for their utility. The procedures included in this review are time-lapse imaging of embryos, assisted hatching, EmbryoGlue, sperm DNA testing, egg activation with calcium ionophore, endometrial receptivity array, and physiological intracytoplasmic sperm injection (PICSI). While there is rather limited supporting evidence for nearly all IVF add-ons that we reviewed, there is strong demand from patients, physicians, and the biotechnology industry to continue further research and development in this arena. We propose that all add-on procedures should provide true efficacy for the patient, and reproductive endocrinologists should inform patients of the costs and benefits of utilizing various technologies before they undergo treatment. In the future, add-ons that show clear evidence of efficacy and justifiable cost should be incorporated into routine practice, while others that do not meet these criteria should be phased out entirely.
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STUDY QUESTION Is fecundability associated with miscarriage history and future miscarriage risk? SUMMARY ANSWER Prior miscarriage was associated with lower fecundability, and participants with a history of subfertility (time-to-pregnancy (TTP) ≥12 months) were at a higher risk of subsequent miscarriage. WHAT IS KNOWN ALREADY Although miscarriage and low fecundability share common risk factors, prior studies have reported both lower and higher fecundability after miscarriage. STUDY DESIGN, SIZE, DURATION In this study, we examined two related associations: one, between miscarriage history and subsequent fecundability and, two, between fecundability and miscarriage risk in the subsequent pregnancy. The study is based on the Norwegian Mother, Father and Child Cohort Study (MoBa). In addition, the outcome of the pregnancy after the MoBa index pregnancy was obtained by linking information from three national health registries: the Medical Birth Registry of Norway, the Norwegian Patient Registry and the general practice database. PARTICIPANTS/MATERIALS, SETTING, METHODS We examined the association between number of prior miscarriages and fecundability in 48 537 naturally conceived, planned pregnancies in participants with at least one prior pregnancy. We estimated fecundability ratios (FRs) and 95% CIs using proportional probability regression. We further estimated the relative risk (RR) of miscarriage in the subsequent pregnancy as a function of TTP in the MoBa index pregnancy for 7889 pregnancies using log-binomial regression. Multivariable analyses adjusted for maternal age, pre-pregnancy maternal BMI, smoking status, cycle regularity, income level and highest completed or ongoing education. MAIN RESULTS AND THE ROLE OF CHANCE Fecundability decreased as the number of prior miscarriages increased. The adjusted FRs among women with one, two and three or more prior miscarriages were 0.83 (95% CI: 0.80–0.85), 0.79 (95% CI: 0.74–0.83) and 0.74 (95% CI: 0.67–0.82), respectively, compared with women with no prior miscarriages. Compared to women with a TTP of <3 months, the adjusted RR of miscarriage in the subsequent pregnancy was 1.16 (0.99–1.35) with TTP of 3–6 months, 1.18 (0.93–1.49) with TTP of 7–11 months and 1.43 (1.13–1.81) with TTP of 12 or more months. LIMITATIONS, REASONS FOR CAUTION Information on TTP and prior miscarriages was obtained retrospectively, and TTP was self-reported. MoBa is a pregnancy cohort, and findings may not be generalizable to all women. We were unable to examine the effect of changing partners between pregnancies, as well as other paternal factors such as seminal parameters. We also did not know what proportion of our participants had changed partners between their prior pregnancies and the index pregnancy. Furthermore, it is likely that many early miscarriages are not recognized. WIDER IMPLICATIONS OF THE FINDINGS The association between miscarriage and fecundability may reflect a contribution of occult pregnancy losses to TTP, as well as shared underlying causes for reduced fecundability and miscarriage. STUDY FUNDING/COMPETING INTEREST(S) The study was funded by the Research Council of Norway through its Medical Student Research Program funding scheme (project number 271555/F20), its Centres of Excellence funding scheme (project number 262700) and through the project ‘Women's fertility – an essential component of health and well-being’ (project number 320656). M.C.M. has received funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (grant agreement number 947684). A.J.W. is supported by the Intramural Program of the National Institute of Environmental Health Sciences at the National Institutes of Health, USA. The authors report no competing interests. TRIAL REGISTRATION NUMBER N/A.
Article
Objective This study aimed to investigate the duration of progesterone (P) therapy on clinical pregnancy rates as measured by the window of implantation (WOI) in the first cycle of frozen embryo transplantation. Methods The study compared the pregnancy rates between 345 cleavage stage transfers and 348 blastocyte transfers of frozen embryos with modified natural cycles in patients from July 1, 2020, to November 30, 2020. Four different P durations were analyzed in the cleavage stage embryo transfer group, i.e., two, three, four, and five days. Five different P durations were analyzed in the blastocyst transfer group, i.e., three, four, five, six, and seven days. Results The baseline demographics and clinical characteristics of the cleavage stage embryos and blastocyst transfer groups were not comparable. The clinical pregnancy rates following the cleavage stage embryo transfer after two, three, four, and five-day P administration were 45.71%, 44.60%, 38.40%, and 30.43%, respectively (the difference among the subgroups was not significant). Following the blastocyst transfer, the clinical pregnancy rates after three, four, five, six, and seven-day P administration were 50.65%, 63.51%, 60.00%, 54.55%, and 61.54%, respectively (the difference among the subgroups was not significant). There was no difference in the clinical pregnancy rates between the cleavage stage embryos and blastocyte transfers after two and three-day P administration. In contrast, these two transfer groups showed significantly different clinical pregnancy rates following four and five-day P exposure (P < 0.05). Conclusion For cleavage-stage embryo transfer, the most effective WOI was found between days two and five of P administration. The effective WOI for blastocyst transfer was observed between days three and seven of P administrations.
Chapter
Routine preoperative pregnancy testing has remained a controversial issue. Even though the prevalence of pregnancy is expected to be low in patients undergoing elective surgery, the discovery of the fewest number of cases is extremely significant. Multiple ethical, legal, and financial variables need to be considered. The method of testing and the resulting sensitivity and specificity of the test should also be a consideration. Although governing bodies have released guidelines providing some direction on this controversial topic, it is ultimately up to each individual institution to choose and adhere to their policy. Nevertheless, when considering the recent body of evidence published over the last couple of years as reviewed in this chapter, one can conclude that pregnancy testing remains a cost-effective method and should be offered to all verbally consenting females of childbearing potential. This recommendation does not, however, substitute for an appropriate pregnancy history and physical examination.
Chapter
Although the majority of pregnancies are uneventful, sometimes complications do happen. Pregnancy complications are the conditions or pathological processes associated with pregnancy. They can occur during or after pregnancy and range from minor discomforts to serious diseases that require medical interventions. They can involve the mother's health, the baby's health, or both. Complication of pregnancy can cause maternal morbidity and mortality. The most common causes of maternal mortality are maternal bleeding, maternal sepsis, hypertensive disease, obstructed labour, and pregnancy with the consequence of abortion, which includes miscarriage, ectopic pregnancy, and medical abortion. The primary means of preventing maternal deaths is to provide rapid access to emergency obstetric care, including treatment of haemorrhage, infection, hypertension, and obstructed labour. Proper antenatal care can reduce the maternal mortality rate by reducing the number of pregnancies among women of reproductive age. Thus, adequate monitoring and appropriate intervention strategies should be provided for better maternal and fetal outcome.
Article
BACKGROUND The positive effects of growth hormone (GH) on IVF are often attributed to improvements in oocyte and embryo quality. While emerging evidence emphasizes GH-induced improvements in the endometrium, these results are controversial. OBJECTIVE AND RATIONALE This meta-analysis aimed to evaluate whether GH administration improved endometrial function and reproductive outcomes during IVF cycles and to thus guide clinical practice. SEARCH METHODS A literature search in the Cochrane Central Register of Controlled Trials, PubMed and Embase was performed through to 30 November 2021, without language restrictions. Randomized controlled trials (RCTs) evaluating the effects of GH on IVF outcomes were included. Risk of bias and quality of evidence (QoE) were assessed according to the Cochrane Collaboration’s tool and the Grading of Recommendations Assessment, Development and Evaluation system. Odds ratios (ORs) and mean differences (MDs) with 95% confidence intervals (CIs) were assessed by random-effects models. OUTCOMES A total of 25 trials with 2424 women were included. Seventeen RCTs with poor responders (n = 1723) showed that GH administration significantly increased endometrial thickness (EMT) (MD = 0.38, 95% CI: 0.18–0.59; moderate QoE), which contributed to an improved live birth rate (OR = 1.67, 95% CI: 1.13–2.49; very low QoE) and clinical pregnancy rate (CPR) (OR = 1.97, 95% CI: 1.43–2.72; low QoE). Subgroup analyses showed a dose- and time-dependent relationship between GH cotreatment and IVF outcomes; the optimal recommendation for improving CPR was consistent with that for EMT, rather than for oocytes and embryos. Hence, GH might improve fertility via effects on the endometrium. Administration of GH daily from the follicular phase of previous cycle until the hCG trigger with < 5 IU/day led to a thicker endometrium and a greater chance of becoming pregnant, while 5–10 IU/day or administration from the luteal phase of the previous cycle until the hCG trigger resulted in higher oocyte and embryo quality. Poor responders might benefit from cotreatment with the GnRH agonist long protocol more than other stimulation protocols. Pooled data from four trials (n = 354) on women with a thin endometrium indicated that improved endometrial function might be critical for improving reproductive outcomes during GH treatment, as no improvements in embryo quality were found. GH administration not only increased EMT (MD = 1.48, 95% CI: 1.21–1.75; moderate QoE) but also promoted endometrial morphology (OR = 2.67, 95% CI: 1.36–5.23; low QoE) and perfusion (OR = 5.84, 95% CI: 1.30–26.17; low QoE), thereby improving the CPR (OR = 2.71, 95% CI: 1.69–4.34; P < 0.0001; low QoE). There was insufficient evidence to reach a conclusion regarding the effects of GH in normal responders (n = 80). Due to obvious improvements in the CPR, women with a thin endometrium might be the most appropriate population to benefit from GH administration. WIDER IMPLICATIONS Improving endometrial function might be another vital mechanism by which GH improves IVF outcomes. Optimal treatment should be offered to the target population according to their personal conditions and needs. The QoE was moderate to very low, due to limited sample sizes and methodological problems; thus, the results should be interpreted with caution. More rigorous RCTs with large sample sizes are needed to confirm the effects and determine optimal GH protocols.
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The peri-implantation period from blastula to gastrula is one of the crucial stages of human embryo and stem cell development. During development, human embryos undergo many crucial events, such as embryonic lineage differentiation and development, structural self-assembly, pluripotency state transition, cell communication between lineages, and crosstalk between the embryo and uterus. Abnormalities in these developmental events will result in implantation failure or pregnancy loss. However, because of ethical and technical limits, the developmental dynamics of human peri-implantation embryos and the underlying mechanisms of abnormal development remain in a “black box”. In this review, we summarize recent progress made towards our understanding of human peri-implantation embryogenesis based on extended in vitro cultured embryos and stem cell-based embryoids. These findings lay an important foundation for understanding early life, promoting research into human stem cells and their application, and preventing and treating infertility. We also propose key scientific issues regarding peri-implantation embryogenesis and provide an outlook on future study directions. Finally, we sum up China’s contribution to the field and future opportunities.
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This article discusses the issue of food hazards and child health during the first thousand days of life. The aim is to raise the attention of decision makers, healthcare officers and professionals, including pediatricians, pediatric surgeons, obstetricians, nurses, midwives, dieticians and lactation consultants, on the importance of protecting infants and their families during a most critical period for the mother-child binomial. The conclusions emphasize the importance of encouraging the adoption of integrated strategies, useful at establishing adequate preventive efforts and a game-changing perspective shift in order to develop and adopt efficient monitoring strategies and procedures, able to minimize the risks due to hazards in food throughout the first thousand days of life, as a first line of prevention in children's health.
Article
The biological processes that control endometrial receptivity and embryo implantation are critical for the successful outcome of pregnancy. The endometrium is the complex inner lining of the uterine wall that is under the cyclical control of estrogen and progesterone and is a site of intimate contact between mother and blastocyst. The bone morphogenetic signaling (BMP) pathway is a highly conserved signaling pathway that controls key cellular processes throughout pregnancy and exerts intracellular effects via the SMAD1/5 transcription factors. To delineate the endometrial compartment-specific roles of BMP signaling, we generated mice with epithelial-specific conditional deletion of SMAD1/5 using Lactoferrin-icre (Smad1flox/flox;Smad5flox/flox;Lactoferrin-cre, “Smad1/5 cKO”). Histological analysis of the reproductive tracts showed that Smad1/5 cKO mice were developmentally normal and displayed no defects in glandular morphology. In fertility analyses, single SMAD1 or SMAD5 deletion had no effect on fertility, however, double conditional deletion of SMAD1 and SMAD5 resulted in severe subfertility. Timed mating analyses revealed endometrial receptivity defects in the Smad1/5 cKO mice beginning at 3.5 days post coitum (dpc) that perturbed embryo implantation at 4.5 dpc, as demonstrated by the detection of unattached blastocysts in the uterus, decreased COX2 expression, and FOXO1 cytoplasmic mis-localization. We also found that defects that arose during peri-implantation adversely affected embryonic and decidual development at 5.5 and 6.5 dpc. Thus, uterine epithelial BMP/SMAD1/5 signaling is essential during early pregnancy and SMAD1/5 epithelial-specific deletion has detrimental effects on stromal cell decidualization and pregnancy development.
Chapter
This chapter focuses on various modern birth control methods, including combined oral contraceptives, progestogen-only pills, progestogen-only injectables, progestogen-only implants, intrauterine devices, barrier contraceptives, and emergency contraceptive pills. Each contraceptive method is covered in detail, including mechanism of action, effectiveness, health benefits, advantages, disadvantages, risks, and side-effects.
Preprint
Female bonobos exhibit prolonged receptivity. One suggested function of the prolonged receptivity is it lowers male mating competition. However, it is questionable whether easier access to receptive females can reduce male-male competition, given the exclusive nature of male reproductive success. We tested whether males could determine a fertile phase of females. We found that ovulation probability predicted male mating effort. High-ranking males copulated with the female with higher fertility, and male-male agonistic interactions increased when there were fertile females in the party. When there were multiple females with maximal swelling, males concentrated their mating effort on a female with an older infant whose maximal swelling started earlier, and they continued mating efforts until detumescence (rainmaking). These findings suggest that male bonobos distinguish between fertile and non- fertile phases of females and that having more receptive females in the party does not reduce male-male competition for fertile females. Teaser Males use the rainmaker’s rule to meet the periovulatory phase of the female bonobo for better reproductive success.
Article
STUDY QUESTION What are the key considerations for developing an enhanced transcriptomic method for secretory endometrial tissue dating? SUMMARY ANSWER Multiple gene expression signature combinations can serve as biomarkers for endometrial dating, but their predictive performance is variable and depends on the number and identity of the genes included in the prediction model, the dataset characteristics and the technology employed for measuring gene expression. WHAT IS KNOWN ALREADY Among the new generation of transcriptomic endometrial dating (TED) tools developed in the last decade, there exists variation in the technology used for measuring gene expression, the gene makeup and the prediction model design. A detailed study, comparing prediction performance across signatures for understanding signature behaviour and discrepancies in gene content between them, is lacking. STUDY DESIGN, SIZE, DURATION A multicentre prospective study was performed between July 2018 and October 2020 at five different centres from the same group of clinics (Spain). This study recruited 281 patients and finally included in the gene expression analysis 225 Caucasian patients who underwent IVF treatment. After preprocessing and batch effect filtering, gene expression measurements from 217 patients were combined with artificial intelligence algorithms (support vector machine, random forest and k-nearest neighbours) allowing evaluation of different prediction models. In addition, secretory-phase endometrial transcriptomes from gene expression omnibus (GEO) datasets were analysed for 137 women, to study the endometrial dating capacity of genes independently and grouped by signatures. This provided data on the consistency of prediction across different gene expression technologies and datasets. PARTICIPANTS/MATERIALS, SETTING, METHODS Endometrial biopsies were analysed using a targeted TruSeq (Illumina) custom RNA expression panel called the endometrial dating panel (ED panel). This panel included 301 genes previously considered relevant for endometrial dating as well as new genes selected for their anticipated value in detecting the secretory phase. Final samples (n = 217) were divided into a training set for signature discovery and an independent testing set for evaluation of predictive performance of the new signature. In addition, secretory-phase endometrial transcriptomes from GEO were analysed for 137 women to study endometrial dating capacity of genes independently and grouped by signatures. Predictive performance among these signatures was compared according to signature gene set size. MAIN RESULTS AND THE ROLE OF CHANCE Testing of the ED panel allowed development of a model based on a new signature of 73 genes, which we termed ‘TED’ and delivers an enhanced tool for the consistent dating of the secretory phase progression, especially during the mid-secretory endometrium (3–8 days after progesterone (P) administration (P + 3–P + 8) in a hormone replacement therapy cycle). This new model showed the best predictive capacity in an independent test set for staging the endometrial tissue in the secretory phase, especially in the expected window of implantation (average of 114.5 ± 7.2 h of progesterone administered; range in our patient population of 82–172 h). Published sets of genes, in current use for endometrial dating and the new TED genes, were evaluated in parallel in whole-transcriptome datasets and in the ED panel dataset. TED signature performance was consistently excellent for all datasets assessed, frequently outperforming previously published sets of genes with a smaller number of genes for dating the endometrium in the secretory phase. Thus, this optimized set exhibited prediction consistency across datasets. LARGE SCALE DATA The data used in this study is partially available at GEO database. GEO identifiers GSE4888, GSE29981, GSE58144, GSE98386. LIMITATIONS, REASONS FOR CAUTION Although dating the endometrial biopsy is crucial for investigating endometrial progression and the receptivity process, further studies are needed to confirm whether or not endometrial dating methods in general are clinically useful and to guide the specific use of TED in the clinical setting. WIDER IMPLICATIONS OF THE FINDINGS Multiple gene signature combinations provide adequate endometrial dating, but their predictive performance depends on the identity of the genes included, the gene expression platform, the algorithms used and dataset characteristics. TED is a next-generation endometrial assessment tool based on gene expression for accurate endometrial progression dating especially during the mid-secretory. STUDY FUNDING/COMPETING INTEREST(S) Research funded by IVI Foundation (1810-FIVI-066-PD). P.D.-G. visiting scientist fellowship at Oxford University (BEFPI/2010/032) and Josefa Maria Sanchez-Reyes’ predoctoral fellowship (ACIF/2018/072) were supported by a program from the Generalitat Valenciana funded by the Spanish government. A.D.-P. is supported by the FPU/15/01398 predoctoral fellowship from the Ministry of Science, Innovation and Universities (Spanish Government). D.W. received support from the NIHR Oxford Biomedical Research Centre. The authors do not have any competing interests to declare.
Article
Implantation timing is critical for a successful pregnancy. A short delay in embryo implantation caused by targeted gene ablation produced a cascading problem in the later stages of the pregnancy. Although several delayed implantation models have been established in wild mice, almost none of them is suitable for investigating the early delay's effects on the late events of pregnancy. Here, we report a new delayed implantation model established by the intraperitoneal administration of letrozole at 5 mg/kg body weight on day 3 of pregnancy. In these mice, initiation of implantation was induced at will by the injection of estradiol (E2). When the estradiol (3 ng) was injected on day 4 of pregnancy (i.e., without delay), the embryo implantation restarted, and the pregnancy continued normally. However, 25 ng estrogen caused compromised implantation. We also found that 67% of the female mice could be pregnant normally and finally gave birth when the estradiol injection (3 ng) was on day 5 of pregnancy (i.e., 1-day delay). Most failed pregnancies had impaired decidualization, decreased serum progesterone levels, and compromised angiogenesis. Progesterone supplementation could rescue decidualization failure in the mice. Collectively, we established a new model of delayed implantation by letrozole, which can be easily applied to study the effect and mechanisms of delay of embryo implantation on the progression of late pregnancy events.
Article
Full-text available
Background: Thin endometrium is a primary cause of defective endometrial receptivity, resulting in infertility or recurrent miscarriage. Much effort has been devoted toward regenerating thin endometrium by stem cell-based therapies. The human placenta-derived mesenchymal stem cells (HP-MSCs) are emerging alternative sources of MSCs with various advantages. To maximize their retention inside the uterus, we loaded HP-MSCs with cross-linked hyaluronic acid hydrogel (HA hydrogel) to investigate their therapeutic efficacy and possible underlying mechanisms. Methods: Ethanol was injected into the mice uterus to establish the endometrium-injured model. The retention time of HP-MSCs and HA hydrogel was detected by in vivo imaging, while the distribution of HP-MSCs was detected by immunofluorescence staining. Functional restoration of the uterus was assessed by testing embryo implantation rates. The endometrial morphological alteration was observed by H&E staining, Masson staining, and immunohistochemistry. In vitro studies were further conducted using EdU, transwell, tube formation, and western blot assays. Results: Instilled HP-MSCs with HA hydrogel (HP-MSCs-HA) exhibited a prolonged retention time in mouse uteri than normal HP-MSCs. In vivo studies showed that the HP-MSCs-HA could significantly increase the gland number and endometrial thickness (P < 0.001, P < 0.05), decrease fibrous area (P < 0.0001), and promote the proliferation and angiogenesis of endometrial cells (as indicated by Ki67 and VEGF, P < 0.05, P < 0.05, respectively) in mice injured endometrium. HP-MSCs-HA could also significantly improve the embryo implantation rate (P < 0.01) compared with the ethanol group. Further mechanistic study showed the paracrine effects of HP-MSCs. They could not only promote the proliferation and migration of human endometrial stromal cells via the JNK/Erk1/2-Stat3-VEGF pathway but also facilitate the proliferation of glandular cells via Jak2-Stat5 and c-Fos-VEGF pathway. In turn, the increased VEGF in the endometrium promoted the angiogenesis of endothelial cells. Conclusion: Our study suggested the potential therapeutic effects and the underlying mechanisms of HP-MSCs-HA on treating thin endometrium. HA hydrogel could be a preferable delivery method for HP-MSCs, and the strategy represents a promising therapeutic approach against endometrial injury in clinical settings.
Article
Progesterone (P4) is crucial for the achievement and maintenance of a pregnancy and with rising numbers of frozen embryo transfers (FETs) performed worldwide, the search for the 'optimal' P4 levels in HRT FET cycles became a focus of research. Certainly, measurement of systemic P4 levels is an easy applicable tool and P4 levels, considered as being too low, could be addressed by changing and/or increasing exogenously administered P4. However, the question must be raised whether the sole measurement of systemic P4 levels is reflective for the endometrial status and the endometrial receptivity in HRT FET cycles, since systemic P4 levels do not reflect the dynamic of the endometrial changes, deemed necessary to prepare the endometrium for implantation. Moreover, different types of P4 administration routes will exhibit distinct different patterns of P4 release, affecting the process of secretory transformation and last but not least, embryonic factors are almost fully neglected in this concept. This opinion article aims to raise critical points towards the 'sole' focus on systemic P4 levels in HRT FET cycles and raises the question whether 'serum P4 measurements are truly representative for the identification of an adequate luteal phase in HRT FETs'?.
Article
Objective To determine whether endometrial receptivity analysis (ERA) improves live births in patients with and without a history of unsuccessful frozen embryo transfers (FETs). Design Retrospective cohort study. Setting Large reproductive center. Patient(s) Patients with and without ERA before euploid single FET were included in the analysis. Intervention(s) Subjects in the exposed group underwent ERA and ERA-timed FETs. Subjects in the unexposed group followed a standard protocol FET without ERA. Outcomes were compared between nonreceptive and receptive subjects undergoing an ERA-timed FET and between ERA-timed vs. standard protocol FETs. Main Outcome Measure(s) The primary outcome was a live birth; secondary outcomes were biochemical and clinical pregnancy rates. Result(s) A total of 307 ERA-timed FETs and 2,284 standard protocol FETs were analyzed. One hundred twenty-five patients (40.7%) were ERA receptive, and 182 (59.3%) were ERA nonreceptive. After adjusting for the number of the previously failed FETs, there was no difference in the proportion of receptive and nonreceptive ERA results. There were no statistically significant differences in live births in patients with ERA-receptive vs. ERA-nonreceptive results (48.8% and 41.7%, respectively; adjusted odds ratio 1.17; 95% CI, 0.97–1.40). There were no statistically significant differences in live births in patients with or without ERA testing results before FET (44.6% and 51.3%, respectively; adjusted odds ratio 0.87; 95% CI, 0.73–1.04). Conclusion(s) Patients with an increasing number of previous failed euploid FET cycles are not at an increased risk of a displaced window of implantation. Patients categorized as receptive vs. nonreceptive and those without ERA testing results have comparable FET success rates.
Article
Endometriosis is a common estrogen-dependent chronic inflammatory disease that leads to infertility in women of reproductive age. Perhaps infertility reflects the reduced expression of integrin αvβ3 and HOXA10 in endometriosis. Previous studies have shown that administration of letrozole, a non-steroidal aromatase inhibitor for cancer treatment, increased the clinical pregnancy rate in women with endometriosis, but the mechanisms remain to be determined. In this communication, a rat model of endometriosis was established. Animals were treated with letrozole at 2ug/kg of body weight, intragastric administration for 15 consecutive days. Letrozole increased the expression of αvβ3 and HOXA10 in the endometriosis model and endometrial receptivity. Abbreviations: WOI: window of implantation; RGD: Arg-Gly-Asp; HOX: homeobox; E2: estradiol; SPF: specific pathogen-free.
Article
Human embryo implantation is an intricate spatiotemporal process that involves the intimate association between the embryo and the endometrium of the mother. During implantation, the endometrium undergoes a dynamic cascade of gene activation and repression, largely driven by autocrine, paracrine, and endocrine action. Steroid hormones, such as estrogen and progesterone, act on a variety of targets including cellular adhesion molecules (CAMs), cytokines, and growth factors to facilitate the implantation process. Given the synchrony required to achieve implantation, it is unsurprising that embryo implantation represents a substantial problem for infertility patients. This is due to a complex interplay taking place at the level of the endometrium. This review discusses the intricacies of embryo implantation including the window of implantation, the cyclical phases of the endometrium, the implantation process itself, and features of endometrial receptivity. Additionally, we will discuss new research regarding inflammatory reproductive biology, epigenetics and microRNA, and the role of the vaginal and endometrial microbiome in implantation. A better understanding of embryo implantation and the interactions occurring at the level of the blastocyst and the endometrium will improve patient care for infertile patients who experience this frustrating challenge.
Article
Full-text available
STUDY QUESTION Are uterine fluid-derived extracellular vesicles (UF-EVs) a ‘liquid biopsy’ reservoir of biomarkers for real-time monitoring of endometrial status? SUMMARY ANSWER The transcriptomic cargo of UF-EVs reflects the RNA profile of the endometrial tissue as well as changes between the non-receptive and the receptive phase, possibly supporting its use for a novel endometrial receptivity test. WHAT IS KNOWN ALREADY EVs have been previously isolated from uterine fluid, where they likely contribute to the embryo-endometrium crosstalk during implantation. Based on a meta-analysis of studies on endometrial tissue implantation-associated genes and the human exosomes database, 28 of the 57 transcripts considered as receptivity markers refer to proteins present in human exosomes. However, the specific transcriptomic content of receptive phase UF-EVs has yet to be defined. STUDY DESIGN, SIZE, DURATION Two experimental series were set up. First, we simultaneously sequenced RNA species derived from paired UF-EVs and endometrial tissue samples collected from physiologically cycling women. Second, we analyzed RNA species of UF-EVs collected during the non-receptive (LH + 2) and receptive (LH + 7) phase of proven fertile women and from the receptive (LH + 7) phase of a population of women undergoing ART and transfer of euploid blastocysts. PARTICIPANTS/MATERIALS, SETTING, METHODS For paired UF—endometrial tissue sampling, endometrial tissue biopsies were obtained with the use of a Pipelle immediately after UF collection performed by lavage of the endometrial cavity. Overall, n = 87 UF samples were collected and fresh-processed for EV isolation and total RNA extraction, while western blotting was used to confirm the expression of EV protein markers of the isolated vesicles. Physical characterization of UF-EVs was performed by Nanoparticle Tracking Analysis. To define the transcriptomic cargo of UF-EV samples, RNA-seq libraries were successfully prepared from n = 83 UF-EVs samples and analyzed by RNA-seq analysis. Differential gene expression (DGE) analysis was used to compare RNA-seq results between different groups of samples. Functional enrichment analysis was performed by gene set enrichment analysis with g:Profiler. Pre-ranked gene set enrichment analysis (GSEA) with WebGestalt was used to compare RNA-seq results with the gene-set evaluated in a commercially available endometrial receptivity array. MAIN RESULTS AND THE ROLE OF CHANCE A highly significant correlation was found between transcriptional profiles of endometrial biopsies and pairwise UF-EV samples (Pearson’s r = 0.70 P < 0.0001; Spearman’s ρ = 0.65 P < 0.0001). In UF-EVs from fertile controls, 942 gene transcripts were more abundant and 1305 transcripts less abundant in the LH + 7 receptive versus the LH + 2 non-receptive phase. GSEA performed to evaluate concordance in transcriptional profile between the n = 238 genes included in the commercially available endometrial receptivity array and the LH + 7 versus LH + 2 UF-EV comparison demonstrated an extremely significant and consistent enrichment, with a normalized enrichment score (NES)=9.38 (P < 0.001) for transcripts up-regulated in LH + 7 in the commercial array and enriched in LH + 7 UF-EVs, and a NES = −5.40 (P < 0.001) for transcripts down-regulated in LH + 7 in the commercial array and depleted in LH + 7 UF-EVs. When analyzing LH + 7 UF-EVs of patients with successful versus failed implantation after transfer of one euploid blastocyst in the following cycle, we found 97 genes whose transcript levels were increased and 64 genes whose transcript levels were decreased in the group of women who achieved a pregnancy. GSEA performed to evaluate concordance in transcriptional profile between the commercially available endometrial receptivity array genes and the comparison of LH + 7 UF-EVs of women with successful versus failed implantation, demonstrated a significant enrichment with a NES = 2.14 (P = 0.001) for transcripts up-regulated in the commercial array in the receptive phase and enriched in UF-EVs of women who conceived, and a not significant NES = −1.18 (P = 0.3) for transcripts down-regulated in the commercial array and depleted in UF-EVs. In terms of physical features, UF-EVs showed a homogeneity among the different groups analyzed except for a slight but significant difference in EV size, being smaller in women with a successful implantation compared to patients who failed to conceive after euploid blastocyst transfer (mean diameter ± SD 205.5± 22.97 nm vs 221.5 ± 20.57 nm, respectively, P = 0.014). LARGE SCALE DATA Transcriptomic data were deposited in NCBI Gene Expression Omnibus (GEO) and can be retrieved using GEO series accession number: GSE158958. LIMITATIONS, REASONS FOR CAUTION Separation of RNA species associated with EV membranes might have been incomplete, and membrane-bound RNA species—rather than the internal RNA content of EVs—might have contributed to our RNA-seq results. Also, we cannot definitely distinguish the relative contribution of exosomes, microvesicles and apoptotic bodies to our findings. When considering patients undergoing ART, we did not collect UFs in the same cycle of the euploid embryo transfer but in the one immediately preceding. We considered this approach as the most appropriate in relation to the novel, explorative nature of our study. Based on our results, a validation of UF-EV RNA-seq analyses in the same cycle in which embryo transfer is performed could be hypothesized. WIDER IMPLICATIONS OF THE FINDINGS On the largest sample size of human EVs ever analyzed with RNA-seq, this study establishes a gene signature to use for less-invasive endometrial receptivity tests. This report is indeed the first to show that the transcriptome of UF-EVs correlates with the endometrial tissue transcriptome, that RNA signatures in UF-EVs change with endometrial status, and that UF-EVs could serve as a reservoir for potential less-invasive collection of receptivity markers. This article thus represents a step forward in the design of less-invasive approaches for real-time monitoring of endometrial status, necessary for advancing the field of reproductive medicine. STUDY FUNDING/COMPETING INTEREST(S) The study was funded by a competitive grant from European Society of Human Reproduction and Embryology (ESHRE Research Grant 2016-1). The authors have no financial or non-financial competing interests to disclose. TRIAL REGISTRATION NUMBER NA.
Article
Full-text available
Genes for chorionic gonadotrophin (CG) are transcribed by the 16-cell embryo stage in humans, but there is no clear evidence of CG secretion as a bioactive dimer before attachment and trophoblast outgrowth stages of implantation. The studies summarized question the timing of CG expression and secretion, the possible roles of CG for intraembryonic differentiation and at the implantation site, and the recognition of this primate embryo-derived signal in support of the corpus luteum. The data suggest that the implantation window in primates may be broader than in non-primate species, where a closer synchrony between embryonic, tubal and uterine events appears to be necessary for embryonic survival. Some preliminary data concerning an association between peripheral thrombocytopenia, ovarian inhibin secretion and peri-implantation stages of embryo development indicate that an unknown embryonic signal may be secreted before bioactive CG can be detected.
Article
We have developed a method of estimating day of ovulation using urinary ovarian hormone data. The method identifies a day of luteal transition that occurs at the shift from production of follicular oestrogen to luteal progesterone. The algorithm for identifying this shift was evaluated and judged better than specified alternatives in that it resulted in (1) a high concordance between the day of luteal transition and peaks in urinary luteinizing hormone (LH) for cycles with well-defined peaks, (2) a low variance in the length of the luteal phase of the menstrual cycle, which presumably reflects a low measurement error in estimating day of ovulation, and (3) a high proportion of cycles for which an approximate day of ovulation could be determined. To validate the new algorithm, it was applied to an independent data set. The algorithm identified a day of luteal transition in 88 percent of these cycles, and the identified day occurred within two days of the urinary LH peak for all of the cycles with clear LH peaks. Determination of the day of luteal transition to estimate ovulation requires only first-morning urine specimens, requires no correction for day-to-day variations in urine concentration, and can be applied to a mid-ycle window of data.
Article
Human in vitro fertilization is characterized by a low of ficiency of implantation. Possible mechanisms for pregnancy loss are discussed. Embryo viability or quality, abnormal implantation, and delayed or absent corpus luteum rescue may all play a role in pregnancy wastage. Defining the possible mechanism for these losses may allow hormonal treatment to correct specific abnormalities.
Article
Direct radioimmunoassay are described for the measurement of each of three specific estrogen glucosiduronates: estrone glucosiduronate, 17 beta-estradiol-17-glucosiduronate and estriol-16 alpha-glucosiduronate in urine. Each assay utilizes a specific antiserum prepared by complexing the carboxylic acid group of the appropriate glucosiduronate to the epsilon-amino group of lysine in bovine serum albumin or bovine thyroglobulin. The antisera showed little or no cross reactivity toward other estrogens that might be present in significant amounts in urine. These antisera were used for the direct assay of the conjugates in urine from normal men and nonpregnant women without prior extraction or chromatography. The values were similar to those obtained after extraction, chromatographic purification on DEAE-Sephadex and subsequent immunoassay; The following mean values +/- SE (microgram/g creatinine) were obtained: estrone glucosiduronate, male 10.1 +/- 0.6, follicular phase female 17.3+/- 1.6, luteal phase female 31.8 +/- 2.5; 17 beta-estradiol-17-glucosiduronate, male 1.7 +/- 0.3, follicular phase female 2.4 +/- 0.1, luteal phase female 4.2 +/- 0.4; estriol-16 alpha-glucosiduronate, male 1.8 +/- 0.2, follicular phase female 4.7 +/- 0.9, luteal phase female 10.0 +/- 1.6.
Article
An antibody was prepared against pregnanediol-3α-glucuronide-BSA. The hapten (5β-pregnane-20α-ol-3α-yl-glucuronide) used for the preparation of the immunogen and as ‘cold’ standard for RIA was synthesized by an unambiguous chemical synthesis. The corresponding [6,7-3H]-labelled conjugate was prepared and the cross-reactions of the antiserum against other glucuronides and free steroids were examined. Application of RIA to menstrual cycle urines and pregnancy urine is discussed. The results of our own studies carried out throughout the menstrual cycle from seven women showed the mean and range for the follicular phase were 3.8 ± 0.67 (3.0 – 5.4) μmol/24 h and for the luteal phase 17.7 ± 6.9 (6.9 – 29.0) μmol/24h.
Article
The WHO Expert Committee on Biological Standardization has established CR119 HCG, CR119 HCGα, and CR119 HCGβ3 as reference preparations for the immunoassay of human chorionic gonadotrophin (HCG) and its subunits. CR119 HCG has interstitial cell stimulating activities of 13 450 IU/mg (bioassay, Second International Standard for HCG) and proportional thyroid stimulating and follicle stimulating activities when tested in vivo. In the subunit preparations, the potencies of these activities are uniformly less than 0.02 relative to the undissociated hormone, and approximately 75% of the activity of each is recovered after recombination. The chemical, physical, and antigenic properties of each preparation are consistent with those reported for preparations of equivalent purity.
Article
Human CG (hCG) was administered to three groups of four normally cycling women in the early luteal phase (LH +4 to +5, group I), the midluteal phase (LH +8 to +9, group II), and the late luteal phase (LH +11 to +12, group III). Two hundred and fifty IU hCG were given im followed in half of the subjects by 750 IU hCG 24 h later. Serial blood samples were then taken at 15- or 30-min intervals following either the first or second hCG injection and continued for 12 or 24 h. The samples were stored frozen at -20 C until assayed for LH, progesterone, estradiol, and hCG concentrations. Treatment with 250 IU hCG at each stage of the luteal phase did not result in any marked change in hormone concentrations. Further hCG administration (750 IU 24 h later) in both the early and the midluteal phases elicited a clear increase in progesterone concentrations. Further hCG treatment in the late luteal phase did not evoke any rise in progesterone levels. Further hCG administration in the midluteal phase resulted in a sharp decline in LH concentrations, brought about mainly by a decrease in LH pulse frequency, this response was not apparent at any other stage of the luteal phase. Despite the lack of any pulsatile steroidogenic stimulus at this time, progesterone was clearly secreted in a pulsatile manner. The decline in LH levels following hCG administration in the midluteal phase resembled that seen in spontaneous conception cycles following implantation. The restriction of this response to the time of normal implantation may suggest a role for the pituitary in the establishment of the "implantation window." The importance of this pituitary response and the mechanisms involved are currently unknown. Its absence in the early luteal phase would suggest that it cannot be directly attributed to either progesterone or hCG. It is possible that some other luteal factor may be responsible for the midluteal decline in LH concentrations.
Article
Excerpt Sheffield Fertility Centre, 26 Glen Road, Sheffield, S7 1RA, UK and University Department of Obstetrics and Gynaecology, Jessop Hospital for Women, Leavygreave Road, Sheffield, S3 7RE, UK Keywords: implantation; in-vitro fertilization; pregnancy; pregnancy loss; human chorionic gonadotrophin; human Introduction The endocrine characteristics of normal human pregnancy have been difficult to establish, chiefly because spontaneous pregnancies occur unpredictably. More reliable sources of early pregnancy data are conceptions following various assisted reproductive technologies although, unfortunately, many of these may not be useful for determining normal physiology, firstly, because there is multiple follicle development resulting from the use of exogenous gonadotrophins and, secondly, because human chorionic gonadotrophin (hCG) given to induce luteinization masks hCG from the implanting embryo. Furthermore, the practice, at least for in-vitro fertilization (IVF), of replacing up to 3 embryos renders assessment of the number of implantation sites uncertain. In-vitro fertilization in the natural or spontaneous cycle may provide
Article
Loss of a conceptus early in development can be detected by very sensitive assays specific for hCG. We examined 20 menstrual cycles ending in early loss of a conceptus in order to identify hormonal correlates of loss. Each loss cycle was compared to a successful conception cycle in the same woman, using daily concentration of urinary estrone-3-glucuronide and pregnanediol-3-glucuronide (PdG). The estrone-3-glucuronide and PdG profiles in cycles of early pregnancy loss were very similar to those in successful conception cycles until late in the luteal phase. Early pregnancy loss was not related to a midluteal deficiency in PdG. hCG tended to be detected later in cycles of early pregnancy loss than in successful conception cycles, presumably indicating later implantation. Ten of the early pregnancy losses implanted after luteal-day-10; only one of the successful pregnancies implanted that late. The corpus luteum responded to the conception in only 2 of the 10 loss cycles with late implantation. In contrast, the corpus luteum responded in 8 of 10 loss cycles with normally timed implantation. The similarity of preimplantation hormonal profiles in cycles of early pregnancy loss and in cycles with successful conceptions suggests that most early losses in reproductively normal women do not result directly from deficiencies in ovarian steroid production.
Article
Human in vitro fertilization is characterized by a low efficiency of implantation. Possible mechanisms for pregnancy loss are discussed. Embryo viability or quality, abnormal implantation, and delayed or absent corpus luteum rescue may all play a role in pregnancy wastage. Defining the possible mechanism for these losses may allow hormonal treatment to correct specific abnormalities.
Article
We looked at risk of early pregnancy loss among 171 women who conceived while participating in study. Twenty-five percent of biochemically detected pregnancies ended within six weeks of the last menstrual period; all but two of these losses were clinically unrecognized. While our sample is small, it is the first to allow description of possible associations between risk of early pregnancy loss and maternal characteristics or exposures. We looked at risk in relation to a woman's age, pregnancy history, weight, education, prenatal DES exposure, cigarette smoking, use of caffeinated and alcoholic beverages, marijuana, cigarette smoking by baby's father, and other variables. None of these factors was definitely associated with early pregnancy loss. Still, the possibility of real effects cannot be excluded and deserves further study.
Article
Pregnancy rates vary considerably with the type of ovarian stimulation used for in vitro fertilization and embryo transfer (IVF-ET). The window of implantation may represent one of the rate-limiting steps in IVF success. We therefore investigated estimated implantation times of 10 consecutive IVF singleton pregnancies, achieved using pituitary suppression with gonadotropin-releasing hormone agonist (GnRH-a) before and during ovarian stimulation with human menopausal gonadotropins (hMG), and compared those with 9 consecutive IVF pregnancies achieved by hMG stimulation only. Estimated implantation times were calculated by regression analysis of serial human chorionic gonadotropin (hCG) measurements between days 7 and 16 after ET. The GnRH-a/hMG pregnancies implanted between days 7 and 11, whereas hMG pregnancies implanted between days 7 and 9 after ET. The hCG regression curve for the GnRH-a/hMG pregnancies revealed a delay of 1.5 days in estimated implantation time compared with the hMG only group. There were no significant differences in pretransfer in vitro embryos development between the two groups. Thus, the delay in hCG rise probably reflects a delay in embryo implantation. We therefore conclude that a GnRH-a/hMG stimulation protocol appears to widen the implantation window in comparison with a hMG only protocol. This observation may at least in part explain the improved IVF pregnancy success with GnRH-a/hMG stimulation protocols.
Article
Paired blood and urine samples were obtained from patients between the sixth and 14th weeks of normal pregnancy. The levels of intact human chorionic gonadotropin (hCG), and of the free alpha and beta subunits, were measured by specific immunoassays. There was a close association between blood and urine levels of intact hCG and of the alpha subunit of hCG, but no relation between the levels of beta subunit in these sites. These findings suggest that the use of "beta subunit" assays may give discrepant results according to the fluid examined. By contrast, measurement of intact hCG appears to give similar results in blood and urine.
Article
Human chorionic gonadotrophin beta (HCG-beta) is a trophoblast marker. Its expression is normally limited to syncytiotrophoblast cells of chorionic villi, although it is known to be secreted from the human embryo as early as 7 days post-fertilization. To examine the onset of embryonic transcriptional activity of the gene encoding this polypeptide we have performed in-situ hybridization to cellular RNAs of human tripronucleate preimplantation embryos. We see expression of HCG-beta RNA at the 6-8-cell stage, before morphological differentiation between trophectoderm and inner cell mass is apparent. We believe that this RNA is the product of de novo transcription from the embryonic genome, since transcripts are only observed in embryos of at least 2 days post-fertilization in age.
Article
Following in vitro fertilization, the criteria commonly used to select human embryos for transfer are the cleavage rate and gross morphology, the contention being that those embryos which divide more rapidly and have regular, spherical blastomeres are more likely to lead to a pregnancy. In order to assess the validity of this assumption, the development in vitro of spare embryos was investigated. Eggs and embryos were cultured in Earle's balanced salt solution containing 10% heat-inactivated patient's serum, and insemination was performed at 40 hr post human chorionic gonadotropin (hCG). At 82-90 hr post hCG, up to four embryos were transferred. Any spare embryos were cultured in the same medium for up to 6 days and scored daily for cell number and morphology using a "quality" scale of 4-1 according to degree of fragmentation and shape of the blastomeres. Of 317 fertilized eggs, 55 (17%) developed to the fully expanded blastocyst stage. The remaining embryos ceased development at the one-cell (6; 2%), two-cell (49; 15%), four-cell (110; 35%), eight-cell (61; 19%), and cavitating morula (36; 11%) stages. The relationship between developmental arrest and gross morphology is discussed.
Article
The receptivity for blastocyst implantation is controlled by progesterone and in some species by the synergistic action of progesterone and estrogen. The duration of the receptive phase, the so-called "window," is short in rodents (less than 24 hours) and may be three days in the primate. Once the uterus becomes receptive, it automatically becomes refractory at the end of the receptive phase. The uterus in the refractory phase can be toxic to the blastocyst in small laboratory animals. The endometrium of the receptive uterus may be characterized by the following parameters: (1) Formation of bulbous protrusions on the apical surface of the luminal epithelium; (2) Secretion of the stage-specific glycoproteins by the luminal epithelium; (3) Readiness of stromal cells to decidualize when appropriate stimulation is applied; and (4) Reorganization and changes of stromal extracellular matrix components so that stromal cells are conditioned for decidualization, and after decidualization the appearance of basement membrane components in the matrix.
Article
We studied the risk of early loss of pregnancy by collecting daily urine specimens from 221 healthy women who were attempting to conceive. Urinary concentrations of human chorionic gonadotropin (hCG) were measured for a total of 707 menstrual cycles with use of an immunoradiometric assay that is able to detect hCG levels as low as 0.01 ng per milliliter, with virtually 100 percent specificity for hCG in the presence of luteinizing hormone. Our criterion for early pregnancy--an hCG level above 0.025 ng per milliliter on three consecutive days--was determined after we compared the hCG levels in the study group with the levels in a comparable group of 28 women who had undergone sterilization by tubal ligation. We identified 198 pregnancies by an increase in the hCG level near the expected time of implantation. Of these, 22 percent ended before pregnancy was detected clinically. Most of these early pregnancy losses would not have been detectable by the less sensitive assays for hCG used in earlier studies. The total rate of pregnancy loss after implantation, including clinically recognized spontaneous abortions, was 31 percent. Most of the 40 women with unrecognized early pregnancy losses had normal fertility, since 95 percent of them subsequently became clinically pregnant within two years.
Article
We intensively studied 30 women attempting pregnancy in order to lay groundwork for larger studies of early pregnancy loss. These women collected first morning urine specimens for up to 6 months after discontinuing use of birth control. Urine specimens were successfully collected for 98% of the woman-days in the study. Three assays for human chorionic gonadotropin (hCG) were performed on each urine specimen. An immunoradiometric assay (IRMA) specific to the carboxyterminal peptide of the hCG beta-chain proved to be more sensitive and more specific than two radioimmunoassays (RIAs). Using the IRMA, we found four cases in which hCG rose and fell over successive days, consistent with early pregnancy loss. For three of these four cases, the level of hCG was too low to be detectable with the RIAs. Among the control group of five women with tubal ligations, there was no detectable hCG above threshold with the IRMA. Thus, the enhanced sensitivity and specificity of the IRMA allows very early pregnancy losses to be identified that would otherwise be undetectable. Furthermore, its effectiveness with small quantities of first morning urine makes the IRMA a useful tool for epidemiologic studies.
Article
A highly sensitive and specific two-site immunoradiometric assay (IRMA) for hCG has been developed and applied to the detection of the hormone in the urine of normal nonpregnant and pregnant individuals. The IRMA uses a solid phase coupled monoclonal antibody to the hCG beta-subunit for extraction of hormone from urine. The hCG extracted is then directly quantified by the binding of an affinity purified and radiolabeled rabbit antibody that reacts with the unique COOH-terminal peptide region of the hCG beta-subunit. The assay is capable of reliably and accurately measuring as little as 0.01 ng hCG/ml urine without interference from hLH. Assays of urine from normal men and nonpregnant women of reproductive age indicated that most individuals did not have detectable levels of hCG immunoreactivity, although a minority had minute amounts, with a mean value of approximately 0.01 ng hCG/mg creatinine. In contrast, all normal menopausal women studied had easily detectable levels of hCG immunoreactivity in their urine, with a mean value of 0.123 ng hCG/mg creatinine. A study of the excretion of hCG from three men injected with hormone for treatment of infertility indicated that after the first 24 h, hCG was cleared with a single exponential rate and was detectable to a level of 0.01 ng/ml. Application of the IRMA to measurements of hCG in the urine of two artificially inseminated patients indicated that the method was capable of detecting pregnancy as early as 9 days postovulation. The extreme sensitivity and specificity of the IRMA for urinary hCG in conjunction with the simplicity of assay performance and specimen collection should provide a substantial advantage over currently available methods for detection of early pregnancy and tumor monitoring.
Article
The preimplantation effects of progesterone antagonists on the cell biology of the endometrium, corpus luteum function and interactions between these two organs have been studied. The antagon ists lilopristone (ZK 98.734) and onapristone (ZK 98.299) were initially given per os to rabbits early or late in pseudopregnancy in combination with human chorionic gonadotrophin (HCG). These protocols were then modified to include hysterectomy or luteotrophic support with 17β-oestradiol. Given alone, the antagonists gave rise to endometrial regression (inhibition of epithelial proliferation and differentiation, increase of apoptosis). The simultaneous addition of oestradiol did not alter these findings. A rapid luteolysis occurred when the antagonists were given in late pseudopregnancy, but not if combined with oestradiol or hysterectomy. The endometrium was capable of renewal and of sustaining implantation if the corpora lutea survived or oestradiol was administered, and transferred blastocysts displayed normal implantation and normal embryonic development. These events did not occur when the antagonists were given during late pseudopregnancy without any steroid supplement. Progesterone antagonists can evidently exert a direct inhibitory effect on the endometrium, possibly with a later indirect luteolytic effect via endometrial mediators. Simultaneous addition of a proper luteotrophic signal results in corpora lutea which are refractory to lysis, so revealing a potential functional dissociation between endometrium and corpus luteum. The endometrium has the capacity to differentiate normally after an interrupted transformation and becomes receptive and sustains normal pregnancy, due to an expanded lifespan of the corpora lutea and a transposition of the implantation window. Uterine secretions from patients undergoing in-vitro fertilization, collected at the onset of the luteal phase, were evaluated by SDS-PAGE densitometry. The protein profiles gave indications of an adequate luteal phase pattern and of a receptive preimplantation phase. These results open the prospect of manipulating the human implantation window.
Article
To examine whether opening of the zona pellucida (i.e., assisted hatching) accelerates implantation. In a controlled, randomized trial, patients were assigned to control and assisted hatching groups. All patients studied were of the Center for Reproductive Medicine at Cornell University Medical College. All patients underwent stimulation with gonadotropins after luteal phase GnRH down regulation. Assisted hatching with zona drilling using acidic Tyrode's solution was performed on the assigned embryos. Luteal E2, P, and hCG on days +5, +6, +7, +8, +9, +11, +13, and +15 were measured. The implantation time, peak midluteal E2 and intervals between these two values were studied. Implantation occurred significantly earlier in the assisted hatching group. The interval between implantation and peak midluteal E2 was also significantly shorter in the assisted hatching group than in the controls. However, there was no significant difference in the day of the peak midluteal E2 between the assisted hatching and control groups. Assisted hatching may enhance embryo implantation not only by mechanically facilitating the hatching process but also by allowing earlier embryo-endometrium contact. Such early contact may enhance embryonic development potential and may optimize synchronization between embryo and endometrium, resulting in improved implantation efficiency.
Article
Of 48 spare human pre-embryos achieving the expanded blastocyst stage, 22 (45.6%) secreted significant amounts of human chorionic gonadotrophin (HCG) (> 5 IU/l/day). Of these, nine remained intrazonal, seven partially hatched and six fully hatched. Embryonic production of HCG in vitro appeared to be time-dependent, starting after a certain minimum time (approximately 160 h post-insemination) and rising exponentially, with maximal HCG production around day 10. Hatching was not a prerequisite for HCG secretion, since similar amounts were produced by intrazonal blastocysts. Blastocysts derived from abnormally fertilized oocytes also began secreting HCG exponentially but secretion was delayed and the upper limit of maximum HCG secretion rate was comparatively low. The actual amount of HCG is thought to reflect the number of viable trophectoderm cells producing the hormone. HCG doubling times for blastocysts in vitro were rapid when compared to implanting blastocysts of a similar age in vivo, with 19/22 (86.4%) blastocysts having a doubling time of < 10 h. Provided a pre-embryo can secrete HCG and maintain an adequate doubling time, sufficient HCG should be produced for initial stages of embryonic recognition in vivo. Since intrazonal blastocysts are capable of fulfilling both of these criteria, the limiting factor in realizing their full potential may be escaping from the zona pellucida.
Article
Objective: We determined the ovarian response to human chorionic gonadotrophin (hCG) in terms of relaxin and progesterone secretion during the peri-implantation period of normal and failing pregnancies. We wished to test the hypotheses that relaxin production in failing pregnancies is different from that in normal pregnancies, that relaxin is a reliable, quantitative indicator of the biological activity of endogenous hCG, and that relaxin is a useful predictor of peri-implantation spontaneous abortions. Design: Daily blood samples were collected in a prospective longitudinal study from insemination patients. Patients: Women undergoing artificial insemination in natural cycles with non-frozen donor semen at a University clinic. Measurements: Serum LH, hCG, relaxin and progesterone were measured and the relationship between hCG and the ovarian hormones was evaluated in the peri-implantation period of normal pregnancies and spontaneous abortions. Results: Nine of 23 conceptive cycles resulted in a spontaneous abortion between 16 and 70 days after the LH peak. In all normal and failing pregnancies there was a close qualitative relationship between hCG secretion and relaxin production. Six of nine failing pregnancies were associated with abnormally low hCG secretion. Six of the spontaneous abortions were associated with rates of relaxin secretion which were higher than the mean of 14 normal pregnancies. No such alterations in progesterone concentrations were observed. In cases where hCG was extremely low, the quantitative relationship between hCG and relaxin was different from that in cases of normal hCG concentrations. Conclusions: There is a close temporal relationship between the secretion of trophoblastic hCG and ovarian secretion of relaxin in the peri-implantation period of normal and failing pregnancies. In failing pregnancies there is substantial variability in the quantitative relationship between relaxin and hCG, indicating that relaxin is not a reliable quantitative indicator of hCG bioactivity. Contrary to previous reports, relaxin concentrations in failing pregnancies tended to be higher than or equal to concentrations in normal pregnancies until the loss was imminent. Because of this relaxin is not a useful predictor of peri-implantation spontaneous abortions.
Article
Longitudinal epidemiologic studies of menstrual and reproductive function are more informative if one can identify day of ovulation. We previously developed a method for estimating day of ovulation that is feasible for epidemiologic studies. The method relies on the relative concentrations of estrogen and progesterone metabolites in daily first-morning urine specimens and does not require creatinine adjustment. This paper describes results of applying this method to a large study with 724 menstrual cycles from 217 women. The method estimated a credible day of ovulation in 88% of cycles. Missing data accounted for most of the failures. When we excluded anovulatory cycles (1%) and cycles with missing data, the method estimated a day of ovulation in 97% of cycles. Variance in luteal phase length was small for our sample, suggesting that this method of identifying a day of ovulation introduces no more measurement error than when day of ovulation is determined by plasma luteinizing hormone (LH), the standard clinical method.
Article
Embryo implantation involves a series of complex interactions between the developing embryo and the maternal endometrium. Results of studies with animal models suggest that the uterus must undergo a series of morphological and biochemical changes, mediated primarily by oestrogen and progesterone, before it becomes receptive for successful implantation. At present there is little understanding of the endometrial changes required to achieve endometrial receptivity for implantation in the human. It appears that control of receptivity is not as stringent in the human as in some other species, with IVF data suggesting that the duration of receptivity is at least 4 days, and that successful implantation can occur under a relatively wide range of morphological and ultrastructural conditions. Research on the later stages of implantation, including embryo positioning within the uterus, attachment and invasions, has been almost non-existent in the human. Further studies are critical for a better understanding of this complex process, although human studies will always be limited by ethical constraints.
Article
To gain insight into the physiology of human endometrial development after artificial preparation with estrogen (E) and P, before oocyte donation. Review and analysis of relevant studies published in the last decade, identified through the literature and Medline searches. Oocyte donation represents a unique in vivo experimental model in the human that permits the study of endometrial development under controlled variable conditions. Early studies have shown that adequate endometrial preparation can be achieved by sequential E and P only. The successful implementation of the simplified approach to oocyte donation demonstrated that satisfactory endometrial receptivity is not dependent on incremental administration of E and P and similarly can be achieved by fixed dosages of these steroids. Moreover, numerous clinical oocyte donation studies have shown that both physiologic and supraphysiologic levels of E and P have resulted in good endometrial development and pregnancy rates, underlining the relative insensitivity of the endometrium to extreme hormonal conditions. In addition, it has been clarified that the endometrium is tolerant of some manipulations during the follicular phase. Contrary to morphological studies that demonstrated preservation of endometrial preparation after luteal E depletion, preliminary evidence suggests that the functional capacity of the endometrium could be affected adversely. In contrast to early oocyte donation studies, which indicated a correlation between morphologic integrity and functional capacity of the endometrium, some evidence presented in this review demonstrates that adequate endometrial morphology does not always imply normal endometrial receptivity.
Article
Integrins belong to a family of cell adhesion molecules that are present on virtually all cells. The temporal and spatial expression of these important proteins on the human endometrium suggests that certain integrins may participate in the cascade of molecular events leading to successful implantation. Using immunohistochemistry, we studied the expression of 12 different integrins in up to 600 samples of human endometrium throughout the menstrual cycle. Intensity and distribution of staining was determined using the semiquantitative HSCORE, with specific focus on the differences between glandular and luminal expression. We noted that the glandular and luminal epithelium undergo independent alterations in integrin expression throughout the menstrual cycle. Specifically, glandular epithelium express certain integrins only during the window of implantation, while luminal epithelium down-regulate certain integrins during this time. The expression of one integrin (the alpha v beta 3 vitronectin receptor) on both luminal and glandular epithelium coincides with the time of embryo attachment; aberrant expression of this integrin is associated with infertility. It appears that the endometrium is a unique tissue with regard to the number of integrins that undergo temporal and spatial changes during the menstrual cycle. These data may offer new directions for the development of a novel contraceptive approach targeted to the endometrium as well as a better understanding of occult causes of infertility in women.
Article
As implantation approaches the different endometrial components demonstrate certain anatomical and molecular changes. Molecules with an effect on uterine receptivity for blastocyst implantation include integrins, leukaemia inhibitory factor (LIF), interleukin-1 (IL-1) and colony stimulating factor-1 (CSF-1). Although over the past 5 years our understanding of the implantation process has improved dramatically, there are still many unanswered questions. Undisputed morphological, biochemical and molecular markers of uterine receptivity that permit quantification of the human implantation window are still unknown. The factors which regulate the expression of integrins, LIF, CSF-1 and IL-1, the mode of action of these molecules and their role in human implantation remain unclear. Only when these questions are answered shall we be able to apply the molecular aspects of implantation to clinical practice.
Article
From a study of 34 early human ova (24 normal and 10 abnormal) recovered from a series of 107 patients known to be fertile whose conditions for conception were optimal it appears that the maximum fertility rate at implantation is 58%; the maximum normal fertility rate after the twelfth day of ovular development is 42%; the probable maximum fertility rate during the preimplantation stages is from 80 to 90%; the greatest ovular loss is in the preimplantation stage; the next greatest loss is during the week after implantation; the ovular loss after the first missed period may be as great as 28.6%, either with or without clinical signs and is therefore comparable to the clinical abortion rate; these defective human fertilized ova rise because of intrinsic defects rather than from defects of the local or endocrine environment, and, finally, the fertility rates and fate of fertilzed ova are roughly comparable in man and other mammals.
Endometrial preparation: lessons from oocyte donation Klentzeris LD. The role of endometrium in implantation
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Younis JS, Simon A, Laufer N. Endometrial preparation: lessons from oocyte donation. Fertil Steril 1996;66:873-84. 24. Klentzeris LD. The role of endometrium in implantation. Hum Reprod 1997;12:Suppl:170-5.