Differential expression of genes linked to the leukemia inhibitor factor signaling pathway during the estrus cycle and early pregnancy in the porcine endometrium

The objective of this study was to determine the expression profiles of leukemia inhibitory factor (LIF) and its receptor (LIFR), interleukin 6 receptor (IL6R), tumor protein p53 (TP53) and B-cell CLL/lymphoma 2 (BCL2) in the porcine endometrium on selected days of the estrous cycle and pregnancy. Time- and reproductive status (estrous cycle/pregnancy)-specific patterns of expression were identified for all investigated genes. The most pronounced changes were seen on Days 12 and 14 of pregnancy when maternal recognition of pregnancy and implantation, respectively, occurs in pigs.     

p16INK4a在早孕小鼠子宫内膜的表达及其对胚泡着床的影响

本研究旨在检测肿瘤抑制基因p16INK4a(inhibitor of cyclin-dependent kinase 4a)在早孕小鼠子宫内膜中的表达规律,探讨p16INK4a在小鼠胚胎着床过程中的作用.采用荧光定量PCR(FQ-PCR)和免疫组织化学方法分别检测未孕小鼠及孕小鼠第2、3、4、5、7天子宫内膜p16INK4a mRNA和蛋白的表达;子宫角注射p16INK4a抗体观察胚泡着床数.FQ-PCR结果显示孕小鼠子宫内膜组织p16INK4amRNA的表达高于未孕小鼠,且随着妊娠天数的增加呈现表达逐渐增强的趋势,到妊娠第5天达到最高,后渐降.免疫组织化学分析显示p16INK4a蛋白在子宫内膜的表达规律与mRNA结果一致.子宫角注射p16INK4a抗体后胚泡着床数明显减少.以上结果提示,P161INK4a在妊娠早期子宫内膜持续表达,可能参与胚泡着床.     

PTEN在早孕小鼠子宫内膜的表达及其对胚泡着床的影响

本研究旨存检测肿瘤抑制基因PTEN(phosphatase andtensinhomologdeletedonchromosometen)在早孕小鼠子宫内膜中的表达规律,探讨PTEN在小鼠胚胎着床过程中的作用.采用实时荧光定量聚合酶联反应(real.time fluorescent quantitative PCR.FQ.PCR)和免疫组织化学方法分别检测未孕及孕1、3、4、5、7 d小鼠子宫内膜PTEN mRNA和蛋白的表达;子宫角注射PTEN反义寡核苷酸观察胚泡着床数.FQ-PCR结果显示,妊娠小鼠子宫内膜组织PTENmRNA的表达高于未妊娠小鼠,且随着妊娠天数的增加表达逐渐增强,到妊娠第5天达最高.免疫组织化学分析显示,PTEN蛋白在子宫内膜的表达规律与mRNA结果一致.子宫角注射PTEN反义寡核苷酸后胚泡着床数明显减少.结果提示,PTEN在妊娠早期子宫内膜持续表达,可能参与了胚泡着床.     

Apical plasma membrane-bound enzymes of rabbit uterine epithelium

Summary In order to monitor changes in the apical cell membrane of rabbit uterine epithelium which are postulated to be a precondition for trophoblast attachment, the marker enzymes: alkaline phosphatase, aminopeptidase M, -glutamyl transferase and dipeptidyl peptidase IV were investigated during the periimplantation phase. Endometrium of early pregnancy (implantation chamber, interblastocyst endometrium; 5–8 days post coitum, d p.c.) was compared with specimens obtained at hCG-induced pseudopregnancy (p. hCG) to distinguish between membrane changes regulated by maternal plasma steroid hormones and such which might be induced locally by blastocyst-derived signals.All enzymes tested showed their main activity at 5 d p.c./p. hCG. The weakest reaction in this series of stages was generally found at 8 d p.c. (interblastocyst segments) or at 8 d p. hCG. In contrast to the rest of the epithelium, the implantation chamber retained high activity of dipeptidyl peptidase IV, and the activity of alkaline phosphatase even raised here again at 7 and 8 d p.c. indicating a direct local influence of the blastocyst on the luminal epithelium. The results suggest that 1) considerable changes occur in the composition of the apical plasma membrane of the uterine epithelium when the endometrium enters the receptive state, 2) the overall trend is towards a loss of apical-type characteristics of this membrane domain and 3) the changes are modulated both systemically (by plasma steroid hormone levels) and locally by signals from the implanting blastocyst.Abbreviations d p.c. days post coitum - d. p. hCG days post hCG injection - hCG human chorionic gonadotropin - aP alkaline phosphatase - ATPase adenosine triphosphatase - Ca²⁺-ATPase Ca²⁺-activated adenosine triphosphatase - APM aminopeptidase M - GGT -glutamyl transferase - DPP IV dipeptidyl peptidase IV - PCMB p-chloromercuric benzoate - DFP di-isopropylfluorophosphate - DMF dimethylformamide     

Apical plasma membrane-bound enzymes of rabbit uterine epithelium. Pattern changes during the periimplantation phase

In order to monitor changes in the apical cell membrane of rabbit uterine epithelium which are postulated to be a precondition for trophoblast attachment, the marker enzymes: alkaline phosphatase, aminopeptidase M, gamma-glutamyl transferase and dipeptidyl peptidase IV were investigated during the periimplantation phase. Endometrium of early pregnancy (implantation chamber, interblastocyst endometrium; 5-8 days post coitum, d p.c.) was compared with specimens obtained at hCG-induced pseudopregnancy (p. hCG) to distinguish between membrane changes regulated by maternal plasma steroid hormones and such which might be induced locally by blastocyst-derived signals. All enzymes tested showed their main activity at 5 d p.c./p. hCG. The weakest reaction in this series of stages was generally found at 8 d p.c. (interblastocyst segments) or at 8 d p. hCG. In contrast to the rest of the epithelium, the implantation chamber retained high activity of dipeptidyl peptidase IV, and the activity of alkaline phosphatase even raised here again at 7 and 8 d p.c. indicating a direct local influence of the blastocyst on the luminal epithelium. The results suggest that 1) considerable changes occur in the composition of the apical plasma membrane of the uterine epithelium when the endometrium enters the "receptive state", 2) the overall trend is towards a loss of apical-type characteristics of this membrane domain and 3) the changes are modulated both systemically (by plasma steroid hormone levels) and locally by signals from the implanting blastocyst.     

Potential innate immunity-related markers of endometrial receptivity and recurrent implantation failure (RIF)

The successful implantation of the embryo into a receptive endometrium is essential for the establishment of a viable pregnancy while recurrent implantation failure (RIF) is a real challenge in assisted reproduction. The maternal innate immune system, specifically the Toll-like receptors (TLRs), are involved in maintaining immunity in the female reproductive tract (FRT) required for fertility. In this study, we aimed to investigate the importance of innate immunity-related gene expression in the regulation of human fertility and as a prediction of potential outcome of in vitro fertilization - embryo transfer (IVF-ET), thus, we assessed the gene expression levels of TLR signalling molecules using quantitative real-time PCR between endometrial biopsies of healthy fertile women, and the patients experiencing RIF. Interestingly, our results showed that, TRIB2 and TLR9 genes were differentially expressed between the endometrial biopsies of healthy women and those with RIF. However, comparing expression levels of same genes between pre-receptive and receptive healthy endometrial biopsies showed different genes (ICAM1, NFKBIA, VCAM1, LIF, VEGFB, TLR5) had significantly altered expression, suggesting their involvement in endometrial receptivity. Thus, further investigations will enable us to better understand the role of these genes in the biology of FRT and as a possible target for the improvement of infertility treatments and/or development of non-hormonal contraception.     

妊娠小鼠子宫内膜LIF基因表达的研究

本文对妊娠第4天(Ⅰ组)、第7天(Ⅱ组)、第10天(Ⅲ组)的小鼠(各20只)子宫内膜LIF基因表达进行了研究。Ⅰ组20只小鼠子宫内膜全部存在LIF基因的表达、Ⅱ组有5只小鼠表达、Ⅲ组仅有1只小鼠表达。文中对不同孕期LIF基因的表达程度与胚胎着床的关系进行了讨论。 Abstract:Leukemia inhibitory factor(LIF)is a glycoprotein with multiple activities and is essential for blastocyst implantation in mouse.We have examined LIF gene expression in mice endometrium on day 4(group Ⅰ),day 7(group Ⅱ),day 10(group Ⅲ)of pregnancy.In group Ⅰ all had LIF gene expression,5 mice had LIF gene expression in group Ⅱ,only one mouse had LIF gene expression in group Ⅲ.We discussed the relation between level of LIF gene expression and embryonic implantation.     

Human Endometrial Extracellular Vesicles Functionally Prepare Human Trophectoderm Model for Implantation: Understanding Bidirectional Maternal‐Embryo Communication

Embryo implantation into maternal endometrium is critical for initiation and establishment of pregnancy, requiring developmental synchrony between endometrium and blastocyst. However, factors regulating human endometrial–embryo cross talk and facilitate implantation remain largely unknown. Extracellular vesicles (EVs) are emerging as important mediators of this process. Here, a trophectoderm spheroid‐based in vitro model mimicking the pre‐implantation human embryo is used to recapitulate important functional aspects of blastocyst implantation. Functionally, human endometrial EVs, derived from hormonally treated cells synchronous with implantation, are readily internalized by trophectoderm cells, regulating adhesive and invasive capacity of human trophectoderm spheroids. To gain molecular insights into mechanisms underpinning endometrial EV‐mediated enhancement of implantation, quantitative proteomics reveal critical alterations in trophectoderm cellular adhesion networks (cell adhesion molecule binding, cell–cell adhesion mediator activity, and cell adherens junctions) and metabolic and gene expression networks, and the soluble secretome from human trophectodermal spheroids. Importantly, transfer of endometrial EV cargo proteins to trophectoderm to mediate changes in trophectoderm function is demonstrated. This is highlighted by correlation among endometrial EVs, the trophectodermal proteome following EV uptake, and EV‐mediated trophectodermal cellular proteome, important for implantation. This work provides an understanding into molecular mechanisms of endometrial EV‐mediated regulation of human trophectoderm functions—fundamental in understanding human endometrium–embryo signaling during implantation.     

Gene expression of WNTs, β-catenin and E-cadherin during the periimplantation period of pregnancy in pigs--involvement of steroid hormones

WNTs (wingless-type MMTV integration site family, member) are morphogenes considered as important factors taking part in uterus developmental processes and implantation. β-catenin is a downstream effector of WNTs action within the cell as well as, through E-cadherin, affecting epithelial organization and function. This study was conducted to investigate WNT4, WNT5A, WNT7A, β-catenin (CTNNB1) and E-cadherin (CDH1) gene expression and protein localization in the endometrium during the periimplantation period. Furthermore, the effect of 17β-estradiol (E₂) and progesterone (P₄) on WNTs, CTNNB1 and CDH1 gene expression in the porcine endometrium in vitro was examined. WNT4 protein was localized in the luminal and glandular epithelium as well as in the basal lamina of the uterine mucosa. WNT5A protein was detected only in the luminal epithelium. WNT7A, β-catenin and E-cadherin protein were identified both in the luminal and glandular epithelial cells, however, WNT7A protein immunoreactivity varied during respective days of estrous cycle and/or pregnancy. Despite unchanged expression of WNT4 mRNA in the endometrium of cyclic and early pregnant pigs, the negative influence of E₂ on WNT4 gene during in vitro experiment was observed. WNT4 and CDH1 gene expression was negatively correlated with blood plasma E₂ and P₄ level in uterine luminal flushings (ULFs) on Day 12 of pregnancy. Expression of WNT5A gene was up-regulated in the endometrium on Day 9 of pregnancy when compared to the respective day of the estrous cycle. A significant decrease of WNT7A gene expression and increase of CDH1 mRNA amount was detected on Day 12 of pregnancy. Overall, the results show the spatial localization of WNT4, WNT5A, WNT7A, β-catenin and E-cadherin proteins in porcine endometrium during periimplantation period of pregnancy and indicate significant changes of WNT5A, WNT7A and CDH1 gene expression before implantation in the pig.     

Induction of trophinin in human endometrial surface epithelia by CGbeta and IL-1beta

During embryo implantation, trophinin mediates cell adhesion by homophilic binding at the apical surfaces of trophectoderm and endometrium. Trophinin is expressed on the human endometrial epithelia in rare occasions. We developed hCG-coated agarose beads that mimic the physical and physiological features of an implantation-stage human blastocyst. When hCG-coated beads were applied to human endometrial epithelial cells in the presence of IL-1beta, endometrial cells acquired strong trophinin expression and the ability for apical cell adhesion with trophinin-expressing human trophoblastic cells. These results provide a mechanism for trophinin-mediated adhesion of human blastocyst to endometrium by a spatially and temporally restricted paracrine effect of hCG derived from the blastocyst.