Embryo implantation involves the intimate interaction between an implantation-competent blastocyst and a receptive uterus, which occurs in a limited time period known as the window of implantation. Emerging evidence shows that defects originating during embryo implantation induce ripple effects with adverse consequences on later gestation events, highlighting the significance of this event for pregnancy success. Although a multitude of cellular events and molecular pathways involved in embryo–uterine crosstalk during implantation have been identified through gene expression studies and genetically engineered mouse models, a comprehensive understanding of the nature of embryo implantation is still missing. This review focuses on recent progress with particular attention to physiological and molecular determinants of blastocyst activation, uterine receptivity, blastocyst attachment and uterine decidualization. A better understanding of underlying mechanisms governing embryo implantation should generate new strategies to rectify implantation failure and improve pregnancy rates in women.
BACKGROUND Time-lapse observation presents an opportunity for optimizing embryo selection based on morphological grading as well as providing novel kinetic parameters, which may further improve accurate selection of viable embryos. The objective of this retrospective study was to identify the morphokinetic parameters specific to embryos that were capable of implanting. In order to compare a large number of embryos, with minimal variation in culture conditions, we have used an automatic embryo monitoring system. METHODS Using a tri-gas IVF incubator with a built-in camera designed to automatically acquire images at defined time points, we have simultaneously monitored up to 72 individual embryos without removing the embryos from the controlled environment. Images were acquired every 15 min in five different focal planes for at least 64 h for each embryo. We have monitored the development of transferred embryos from 285 couples undergoing their first ICSI cycle. The total number of transferred embryos was 522, of which 247 either failed to implant or fully implanted, with full implantation meaning that all transferred embryos in a treatment implanted. RESULTS A detailed retrospective analysis of cleavage times, blastomere size and multinucleation was made for the 247 transferred embryos with either failed or full implantation. We found that several parameters were significantly correlated with subsequent implantation (e.g. time of first and subsequent cleavages as well as the time between cleavages). The most predictive parameters were: (i) time of division to 5 cells, t5 (48.8–56.6 h after ICSI); (ii) time between division to 3 cells and subsequent division to 4 cells, s2 (≤0.76 h) and (iii) duration of cell cycle two, i.e. time between division to 2 cells and division to 3 cells, cc2 (≤11.9 h). We also observed aberrant behavior such as multinucleation at the 4 cell stage, uneven blastomere size at the 2 cell stage and abrupt cell division to three or more cells, which appeared to largely preclude implantation. CONCLUSIONS The image acquisition and time-lapse analysis system makes it possible to determine exact timing of embryo cleavages in a clinical setting. We propose a multivariable model based on our findings to classify embryos according to their probability of implantation. The efficacy of this classification will be evaluated in a prospective randomized study that ultimately will determine if implantation rates can be improved by time-lapse analysis.
Macrophages are prominent in the uterus and ovary at conception. Here we utilize the Cd11b-Dtr mouse model of acute macrophage depletion to define the essential role of macrophages in early pregnancy. Macrophage depletion after conception caused embryo implantation arrest associated with diminished plasma progesterone and poor uterine receptivity. Implantation failure was alleviated by administration of bone marrow-derived CD11b(+)F4/80(+) monocytes/macrophages. In the ovaries of macrophage-depleted mice, corpora lutea were profoundly abnormal, with elevated Ptgs2, Hif1a, and other inflammation and apoptosis genes and with diminished expression of steroidogenesis genes Star, Cyp11a1, and Hsd3b1. Infertility was rescued by exogenous progesterone, which confirmed that uterine refractoriness was fully attributable to the underlying luteal defect. In normally developing corpora lutea, macrophages were intimately juxtaposed with endothelial cells and expressed the proangiogenic marker TIE2. After macrophage depletion, substantial disruption of the luteal microvascular network occurred and was associated with altered ovarian expression of genes that encode vascular endothelial growth factors. These data indicate a critical role for macrophages in supporting the extensive vascular network required for corpus luteum integrity and production of progesterone essential for establishing pregnancy. Our findings raise the prospect that disruption of macrophage-endothelial cell interactions underpinning corpus luteum development contributes to infertility in women in whom luteal insufficiency is implicated.
To study the role of during embryo implantation in rat. expression in rat early pregnancy was detected by Northern blot. The relation between and predicted and confirmed by bioinformatics method, dual-luciferase activity assay, Western blot and immunohistochemistry. The role of was detected by MTS, Edu and chamber assays. The expression level of on gestation day 5–8 (g.d. 5–8) was higher than on g.d. 3–4 in uteri of pregnant rat. was mainly localized in the superficial stroma/primary decidual zone, luminal and glandular epithelia. The expression of was not significantly influenced by pseudopregnancy, but the activation of delayed implantation and experimentally induced decidualization significantly promoted expression. Over-expression of in human endometrial stromal cells (ESCs) inhibited cell proliferation, migration and invasion. Knockdown of promoted cell proliferation and invasion. The results of recombinant luciferase reporters showed that could bind to the 3¢-untranslated region (UTR) of ( ) to inhibit translation. Uterine may be involved in the successful pregnancy, especially during the process of blastocyst implantation through regulating . This study may have the potential to provide new insights into the understanding of function during embryo implantation.
BACKGROUND Repeated implantation failure (RIF) is a major problem encountered in IVF. We have previously reported that RIF-IVF patients have a different endometrial gene expression profile during the window of implantation. Considering microRNA (miRNA) function in post-transcriptional regulation of gene expression, the aim of the study was to evaluate the involvement of miRNA in defects of endometrial receptivity. METHODS We used TaqMan miRNA array cards to identify the miRNAs differentially expressed in the secretory endometrium of RIF-IVF patients when compared with fertile women, and bioinformatics tools to identify their predicted targets and the molecular networks they may affect. RESULTS Comparing miRNA expression profiles, we identified 13 miRNAs, differentially expressed in RIF endometrial samples, that putatively regulate the expression of 3800 genes. We found that 10 miRNAs were overexpressed (including miR 145, 23b and 99a) and 3 were underexpressed. Using our previous gene expression analysis, we paralleled miRNA–mRNA expression profiling. By this means, we identified novel and previously characterized miRNA-regulated molecular pathways such as adherens junctions, cell adhesion molecules, Wnt-signaling, p53 signaling and cell cycle pathways. Consistent with the miRNA-predicted targets, mRNA levels of N-cadherin, H2AFX, netrin-4 and secreted frizzled-related protein-4, belonging to the cell adhesion molecules, Wnt signaling and cell cycle pathways were lower in RIF-IVF patients. CONCLUSIONS To our knowledge, this is the first study to evaluate the differential expression of miRNAs in the secretory endometrium of RIF-IVF patients. We suggest that the RIF-associated miRNAs could be exploited as new candidates for diagnosis and treatment of embryo implantation failures.
The implantation process is complex, requiring reciprocal interactions between implantation-competent blastocysts and the receptive uterus. Because microRNAs (miRNAs) have major roles in regulating gene expression, we speculated that they participate in directing the highly regulated spatiotemporally expressed genetic network during implantation. Here, we show that two miRNAs, mmu-miR-101a and mmu-miR-199a*, are spatiotemporally expressed in the mouse uterus during implantation coincident with expression of cyclooxygenase-2, a gene critical for implantation. More interestingly, our in vitro gain- and loss-of-function experiments show that cyclooxygenase-2 expression is posttranscriptionally regulated by these two miRNAs. We report on miRNA-mediated regulation of uterine gene expression in the context of implantation. We believe that many other critical genes related to this process are also regulated by miRNAs. Thus, elucidating the physiological roles of uterine miRNAs will help us better understand the genetic control of implantation, the gateway to a successful pregnancy.
Citation Rashid NA, Lalitkumar S, Lalitkumar PG, Gemzell‐Danielsson K. Endometrial Receptivity and Human Embryo Implantation. Am J Reprod Immunol 2011; 66 (Suppl. 1): 23–30 Problem The pre‐requisite of successful implantation involves an intricate cascade of molecular interactions which plays a crucial role in preparing receptive endometrium and implanting blastocyst. Method of study Data are hereby presented for a better understanding of endometrial receptivity in women, hoping to provide a comprehensive picture of the process and identify new areas of basic and translational research in the biology of blastocyst implantation. Results Timely regulation of the expression of a number of complex molecules like hormones, cytokines and growth factors, and their crosstalk from embryonic and maternal endometrial side play a major role in determining the fate of the embryo. The molecular basis of endometrial receptivity and the mechanisms by which the blastocyst first adheres to the luminal epithelium and then penetrates into the stroma are only just beginning to be resolved. Conclusion Advances in the development of implantation models and ‘omics’ technologies, particularly proteomics and metabolomics, are set to have a major impact on the development of this field.
The establishment of a receptive uterus is the prime requirement for embryo implantation. In mice, the E-2-induced cytokine leukemia inhibitory factor (LIF) is essential in switching the uterine luminal epithelium (LE) from a nonreceptive to a receptive state. Here we define the LIF-mediated switch using array analysis and informatics to identify LIF-induced changes in gene expression and annotated signaling pathways specific to the LE. We compare gene expression profiles at 0, 1, 3, and 6 h, following LIF treatment. During the first hour, the JAK-STAT signaling pathway is activated and the expression of 54 genes declines, primarily affecting LE cytoskeletal and chromatin organization as well as a transient reduction in the progesterone, TGFbetaR1, and ACVR1 receptors. Simultaneously 256 genes increase expression, of which 42 are transcription factors, including Sox, Kfl, Hes, Hey, and Hox families. Within 3 h, the expression of 3987 genes belonging to more than 25 biological process pathways was altered. We confirmed the mRNA and protein distribution of key genes from 10 pathways, including the Igf-1, Vegf, Toll-like receptors, actin cytoskeleton, ephrin, integrins, TGFbeta, Wnt, and Notch pathways. These data identify novel LIF-activated pathways in the LE and define the molecular basis between the refractory and receptive uterine phases. More broadly, these findings highlight the staggering capacity of a single cytokine to induce a dynamic and complex network of changes in a simple epithelium essential to mammalian reproduction and provide a basis for identifying new routes to regulating female reproduction.
BACKGROUND: The relationship between early embryo post-implantation development in couples undergoing assisted reproductive techniques (ARTs) and sperm chromatin alterations has not been satisfactorily explained. The aim of this study was to assess the relationship between sperm DNA fragmentation in IVF/ICSI patients, sperm parameters (concentration, motility and morphology) and ART outcome, especially with regard to clinical pregnancy and pregnancy loss (spontaneous miscarriage or biochemical pregnancy). METHODS: DNA fragmentation was evaluated by TUNEL assay, performed on sperm suspensions after density gradient separation, in 132 men undergoing an ART cycle (82 IVF and 50 ICSI) and correlated with sperm parameters and ART outcome. RESULTS: A highly significant negative correlation was found between DNA fragmentation and sperm parameters. There was a close relationship between DNA fragmentation and post-implantation development in ICSI patients: the clinical pregnancy and pregnancy loss rates significantly differed between patients with high and low sperm DNA fragmentation (P = 0.007 and P = 0.009, respectively). CONCLUSIONS: Sperm DNA fragmentation seems to affect embryo post-implantation development in ICSI procedures: high sperm DNA fragmentation can compromise 'embryo viability', resulting in pregnancy loss.
T regulatory (Treg) cells are essential mediators of the maternal immune adaptation necessary for embryo implantation. In mice, insufficient Treg cell activity results in implantation failure, or constrains placental function and fetal growth. In women, Treg cell deficiency is linked with unexplained infertility, miscarriage, and pre‐eclampsia. To devise strategies to improve Treg cell function, it is essential to define the origin of the Treg cells in gestational tissues, and the regulators that control their functional competence and recruitment. Male seminal fluid is a potent source of the Treg cell‐inducing agents TGF β and prostaglandin E, and coitus is one key factor involved in expanding the pool of inducible Treg cells that react with paternal alloantigens shared by conceptus tissues. In mice, coitus initiates a sequence of events whereby female dendritic cells cross‐present seminal fluid antigens and activate T cells, which in turn circulate via the blood to be sequestered into the endometrium. Similar events may occur in the human genital tract, where seminal fluid induces immune cell changes that appear competent to prime Treg cells. Improved understanding of how seminal fluid influences Treg cells in women should ultimately assist in the development of new therapies for immune‐mediated pathologies of pregnancy.