Expression of ENPP3 in human cyclic endometrium: a novel molecule involved in embryo implantation

2018 ◽  
Vol 30 (10) ◽  
pp. 1277 ◽  
Author(s):  
Qianqian Chen ◽  
Aijie Xin ◽  
Ronggui Qu ◽  
Wenbi Zhang ◽  
Lu Li ◽  
...  
Keyword(s):  
Embryo Implantation ◽  
Implantation Rate ◽  
Endometrial Receptivity ◽  
Human Endometrium ◽  
Bewo Cell ◽  
Western Blot Assay ◽  
Endometrial Cells ◽  
Positive Effects ◽  
Ishikawa Cells ◽  
Cyclic Changes

Ectonucleotide pyrophosphatase–phosphodiesterase 3 (ENPP3), a protein detected in the human uterus, has been found to play an important role in the development and invasion of tumours. It was recently discovered that ENPP3 was upregulated during the window of implantation in the human endometrium but its functional relevance remains elusive. The objective was to determine ENPP3 expression in human endometrium and its roles in endometrial receptivity and embryo implantation. ENPP3 expression was analysed using immunohistochemistry and western blot assay. The effects of ENPP3 on embryo implantation were evaluated using a BeWo cell (a human choriocarcinoma cell line) spheroid attachment assay and BeWo cells were dual cultured with Ishikawa cells transfected with lentiviral vectors (LV5-NC or LV5-ENPP3) to mimic embryo implantation in a Transwell model. The effects of endometrial ENPP3 on factors related to endometrial receptivity were also determined. The results showed that ENPP3 was expressed in human endometrial epithelial cells and its expression levels changed during the menstrual cycle, peaking in the mid-secretory phase, corresponding to the time of embryo implantation. The overexpression of endometrial ENPP3 not only increased the embryo implantation rate but also had positive effects on the expression of factors related to endometrial receptivity in human endometrial cells. The results indicate that ENPP3 levels undergo cyclic changes in the endometrium and affect embryo adhesion and invasion via altering the expression of implantation factors in the human endometrium. Therefore, ENPP3 may play an important role in embryo implantation and may be a unique biomarker of endometrial receptivity.

scholarly journals Proteomic analysis of extracellular vesicles secreted by primary human epithelial endometrial cells reveals key proteins related to embryo implantation

2022 ◽  
Vol 20 (1) ◽  
Author(s):  
Marina Segura-Benítez ◽  
María Cristina Carbajo-García ◽  
Ana Corachán ◽  
Amparo Faus ◽  
Antonio Pellicer ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Proteomic Analysis ◽  
Embryo Implantation ◽  
Endometrial Receptivity ◽  
Biological Processes ◽  
Endometrial Cells ◽  
Isolation Methods ◽  
Ishikawa Cells ◽  
Endometrial Epithelial Cells ◽  
Implantation Success

Abstract Background Successful implantation is dependent on coordination between maternal endometrium and embryo, and the role of EVs in the required cross-talk cell-to-cell has been recently established. In this regard, it has been reported that EVs secreted by the maternal endometrium can be internalized by human trophoblastic cells transferring their contents and enhancing their adhesive and invasive capacity. This is the first study to comprehensively evaluate three EV isolation methods on human endometrial epithelial cells in culture and to describe the proteomic content of EVs secreted by pHEECs from fertile women. Methods Ishikawa cells and pHEECs were in vitro cultured and hormonally treated; subsequently, conditioned medium was collected and EVs isolated. Ishikawa cells were used for the comparison of EVs isolation methods ultracentrifugation, ExoQuick-TC and Norgen Cell Culture Media Exosome Purification Kit (n = 3 replicates/isolation method). pHEECs were isolated from endometrial biopsies (n = 8/replicate; 3 replicates) collected from healthy oocyte donors with confirmed fertility, and protein content of EVs isolated by the most efficient methodology was analysed using liquid chromatography–tandem mass spectrometry. EV concentration and size were analyzed by nanoparticle tracking analysis, EV morphology visualized by transmission electron microscopy and protein marker expression was determined by Western blotting. Results Ultracentrifugation was the most efficient methodology for EV isolation from medium of endometrial epithelial cells. EVs secreted by pHEECs and isolated by ultracentrifugation were heterogeneous in size and expressed EV protein markers HSP70, TSG101, CD9, and CD81. Proteomic analysis identified 218 proteins contained in these EVs enriched in biological processes involved in embryo implantation, including cell adhesion, differentiation, communication, migration, extracellular matrix organization, vasculature development, and reproductive processes. From these proteins, 82 were selected based on their functional relevance in implantation success as possible implantation biomarkers. Conclusions EV protein cargos are implicated in biological processes related to endometrial receptivity, embryo implantation, and early embryo development, supporting the concept of a communication system between the embryo and the maternal endometrium via EVs. Identified proteins may define new biomarkers of endometrial receptivity and implantation success.

scholarly journals Serum estradiol levels in controlled ovarian stimulation directly affect the endometrium

2017 ◽  
Vol 59 (2) ◽  
pp. 105-119 ◽  
Author(s):  
Kamran Ullah ◽  
Tanzil Ur Rahman ◽  
Hai-Tao Pan ◽  
Meng-Xi Guo ◽  
Xin-Yan Dong ◽  
...  
Keyword(s):  
Embryo Implantation ◽  
Controlled Ovarian Hyperstimulation ◽  
Endometrial Receptivity ◽  
Protein Profiling ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Serum Estradiol ◽  
Endometrial Cells ◽  
Ishikawa Cells ◽  
Estradiol Levels

Previous studies have shown that increasing estradiol concentrations had a toxic effect on the embryo and were deleterious to embryo adhesion. In this study, we evaluated the physiological impact of estradiol concentrations on endometrial cells to reveal that serum estradiol levels probably targeted the endometrium in controlled ovarian hyperstimulation (COH) protocols. An attachment model of human choriocarcinoma (JAr) cell spheroids to receptive-phase endometrial epithelial cells and Ishikawa cells treated with different estradiol (10−9 M or 10−7 M) concentrations was developed. Differentially expressed protein profiling of the Ishikawa cells was performed by proteomic analysis. Estradiol at 10−7 M demonstrated a high attachment rate of JAr spheroids to the endometrial cell monolayers. Using iTRAQ coupled with LC–MS/MS, we identified 45 differentially expressed proteins containing 43 significantly upregulated and 2 downregulated proteins in Ishikawa cells treated with 10−7 M estradiol. Differential expression of C3, plasminogen and kininogen-1 by Western blot confirmed the proteomic results. C3, plasminogen and kininogen-1 localization in human receptive endometrial luminal epithelium highlighted the key proteins as possible targets for endometrial receptivity and interception. Ingenuity pathway analysis of differentially expressed proteins exhibited a variety of signaling pathways, including LXR/RXR activation pathway and acute-phase response signaling and upstream regulators (TNF, IL6, Hmgn3 and miR-140-3p) associated with endometrial receptivity. The observed estrogenic effect on differential proteome dynamics in Ishikawa cells indicates that the human endometrium is the probable target for serum estradiol levels in COH cycles. The findings are also important for future functional studies with the identified proteins that may influence embryo implantation.

scholarly journals Down-Regulation of S100A11, a Calcium-Binding Protein, in Human Endometrium May Cause Reproductive Failure

2012 ◽  
Vol 97 (10) ◽  
pp. 3672-3683 ◽  
Author(s):  
Xin-Mei Liu ◽  
Guo-Lian Ding ◽  
Ying Jiang ◽  
Hong-Jie Pan ◽  
Dan Zhang ◽  
...  
Keyword(s):  
Immune Responses ◽  
Calcium Binding ◽  
Binding Protein ◽  
Embryo Implantation ◽  
Endometrial Receptivity ◽  
Reproductive Failure ◽  
Human Endometrium ◽  
Down Regulation ◽  
Endometrial Cells ◽  
Uptake And Release

Abstract Background: Low expression levels of S100A11 proteins were demonstrated in the placental villous tissue of patients with early pregnancy loss, and S100A11 is a Ca2+-binding protein that interprets the calcium fluctuations and elicits various cellular responses. Objectives: The objective of the study was to determine S100A11 expression in human endometrium and its roles in endometrial receptivity and embryo implantation. Methods: S100A11 expression in human endometrium was analyzed using quantitative RT-PCR, Western blot, and immunohistochemical techniques. The effects of S100A11 on embryo implantation were examined using in vivo mouse model, and JAr (a human choriocarcinoma cell line) spheroid attachment assays. The effects of endometrial S100A11 on factors related to endometrial receptivity and immune responses were examined. Using a fluorescence method, we examined the changes in cytosolic Ca2+ and Ca2+ release from intracellular stores in epidermal growth factor (EGF)-treated endometrial cells transfected with or without S100A11 small interfering RNA. Results: S100A11 was expressed in human endometrium. S100A11 protein levels were significantly lower in endometrium of women with failed pregnancy than that in women with successful pregnancy outcomes. The knockdown of endometrial S100A11 not only reduced embryo implantation rate in mouse but also had adverse effects on the expression of factors related to endometrial receptivity and immune responses in human endometrial cells. Immunofluorescence analysis showed that S100A11 proteins were mainly localized in endoplasmic reticulum. The EGF up-regulated endometrial S100A11 expression and promoted the Ca2+ uptake and release from Ca2+ stores, which was inhibited by the knockdown of S100A11. Conclusions: Endometrial S100A11 is a crucial intermediator in EGF-stimulated embryo adhesion, endometrium receptivity, and immunotolerance via affecting Ca2+ uptake and release from intracellular Ca2+ stores. Down-regulation of S100A11 may cause reproductive failure.

scholarly journals Enhancement of Endometrial Receptivity by Cnidium officinale through Expressing LIF and Integrins

2019 ◽  
Vol 2019 ◽  
pp. 1-9 ◽  
Author(s):  
Tae-Wook Chung ◽  
Mi-Ju Park ◽  
Hoyoung Lee ◽  
Keuk-Jun Kim ◽  
Cheorl-Ho Kim ◽  
...  
Keyword(s):  
Therapeutic Potential ◽  
Embryo Implantation ◽  
Implantation Rate ◽  
Endometrial Receptivity ◽  
Female Infertility ◽  
In Vivo Model ◽  
Cnidium Officinale ◽  
Ishikawa Cells

Improvement of endometrial receptivity is necessary for successful embryo implantation, and its impairment is associated with female infertility. In this study, we investigated the effect of the roots of Cnidium officinale Makino (CoM) on endometrial receptivity in both in vitro and in vivo model of embryo implantation. We found that CoM enhanced the adhesion of JAr cells to Ishikawa cells by stimulating expression of leukemia inhibitory factor (LIF) and integrins. In addition, blocking of LIFR using hLA or neutralization of integrins αV, β3, and β5 using antibodies significantly reduced the enhanced adhesion between JAr cell and CoM-treated Ishikawa cells, indicating that LIF and integrin play an important role in trophoblast-endometrium adhesion for embryo implantation. Furthermore, we identified that CoM significantly improved the implantation rate of blastocysts in the mouse model of RU-induced implantation failure. By collecting these results, here, we suggest that CoM has a therapeutic potential against female infertility associated with decreased endometrial receptivity.

scholarly journals Single-cell RNA Expression of SARS-CoV-2 Cell Entry Factors in Human Endometrium during Preconception

2020 ◽  
Author(s):  
Felipe Vilella ◽  
Wanxin Wang ◽  
Inmaculada Moreno ◽  
Stephen R. Quake ◽  
Carlos Simon
Keyword(s):  
Single Cell ◽  
Rna Sequencing ◽  
Viral Entry ◽  
Embryo Implantation ◽  
Human Endometrium ◽  
Cell Entry ◽  
Rna Expression ◽  
Endometrial Cells ◽  
Single Cell Rna Sequencing ◽  
Low Efficiency

AbstractWe investigated potential SARS-CoV-2 tropism in human endometrium by single-cell RNA-sequencing of viral entry-associated genes in healthy women. Percentages of endometrial cells expressing ACE2, TMPRSS2, CTSB, or CTSL were <2%, 12%, 80%, and 80%, respectively, with 0.7% of cells expressing all four genes. Our findings imply low efficiency of SARS-CoV-2 infection in the endometrium before embryo implantation, providing information to assess preconception risk in asymptomatic carriers.

scholarly journals Increased METTL3-mediated m6A methylation inhibits embryo implantation by repressing HOXA10 expression in recurrent implantation failure

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Pingping Xue ◽  
Wenbo Zhou ◽  
Wenqiang Fan ◽  
Jianya Jiang ◽  
Chengcai Kong ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Embryo Implantation ◽  
Endometrial Receptivity ◽  
Mrna Levels ◽  
Implantation Failure ◽  
Quantitative Real Time Pcr ◽  
Recurrent Implantation Failure ◽  
Ishikawa Cells ◽  
Embryo Attachment

Abstract Background Recurrent implantation failure (RIF) is a major limitation of assisted reproductive technology, which is associated with impaired endometrial receptivity. Although N6-methyladenosine (m6A) has been demonstrated to be involved in various biological processes, its potential role in the endometrium of women with RIF has been poorly studied. Methods Global m6A levels and major m6A methyltransferases/demethylases mRNA levels in mid-secretory endometrium from normal and RIF women were examined by colorimetric m6A quantification strategy and quantitative real-time PCR, respectively. The effects of METTL3-mediated m6A modification on embryo attachment were evaluated by an vitro model of a confluent monolayer of Ishikawa cells co-cultured with BeWo spheroids, and the expression levels of homeo box A10 (HOXA10, a well-characterized marker of endometrial receptivity) and its downstream targets were evaluated by quantitative real-time PCR and Western blotting in METTL3-overexpressing Ishikawa cells. The molecular mechanism for METTL3 regulating HOXA10 expression was determined by methylated RNA immunoprecipitation assay and transcription inhibition assay. Results Global m6A methylation and METTL3 expression were significantly increased in the endometrial tissues from women with RIF compared with the controls. Overexpression of METTL3 in Ishikawa cells significantly decreased the ration of BeWo spheroid attachment, and inhibited HOXA10 expression with downstream decreased β3-integrin and increased empty spiracles homeobox 2 expression. METTL3 catalyzed the m6A methylation of HOXA10 mRNA and contributed to its decay with shortened half-life. Enforced expression of HOXA10 in Ishikawa cells effectively rescued the impairment of METTL3 on the embryo attachment in vitro. Conclusion Increased METTL3-mediated m6A modification represents an adverse impact on embryo implantation by inhibiting HOXA10 expression, contributing to the pathogenesis of RIF.

182 CALBINDIN-D28k IS PREDOMINANTLY EXPRESSED AND REGULATED BY ESTROGEN IN HUMAN ENDOMETRIUM DURING MENSTRUAL CYCLE

2011 ◽  
Vol 23 (1) ◽  
pp. 192
Author(s):  
K.-C. Choi ◽  
H. Yang ◽  
E.-B. Jeung
Keyword(s):  
Menstrual Cycle ◽  
Calcium Binding ◽  
Potential Role ◽  
Embryo Implantation ◽  
Endometrial Receptivity ◽  
Human Endometrium ◽  
Ishikawa Cell ◽  
Proliferative Phase ◽  
Calbindin D28k ◽  
The Brain

The human endometrium resists embryo implantation except during the window of receptivity. A change in endometrial gene expression is required for the development of receptivity. Uterine calbindin-D28k (CaBP-28k) has been shown to be involved in the regulation of endometrial receptivity by intracellular Ca2+. Nowadays, this protein has been mainly linked to the brain, kidneys, and pancreas, but potential role(s) of CaBP-28k remain to be clarified in the uterus of humans during the menstrual cycle. Thus, we demonstrated in this study that the expression of CaBP-28k in the human endometrium in more divided in the menstrual phases. During the menstrual cycle of humans, uterine expression levels of CaBP-28k mRNA and protein increased at the proliferative phase and fluctuated in these tissues, compared with other phases. We assessed the effects of the sex-steroid hormones E2 and P4 on the expression of CaBP-28k in the Ishikawa cell line. A significant increase in the expression of CaBP-9k mRNA was observed at the concentration of 17β-oestradiol (E2; 10–9 to 10–7 M). In addition, spatial expression of CaBP-28k was detected by immunohistochemistry. CaBP-28k is abundantly localised in the cytoplasm of the luminal and glandular epithelial cells during the menstrual cycle. Taken together, these results indicate that CaBP-28k, a uterine calcium-binding protein, is abundantly expressed in the human uterus, suggesting that uterine expression of CaBP-28k may be involved in reproductive functions during the menstrual cycle of humans.

Osteopontin in Human Endometrium: A Role in Endometrial Receptivity and Embryo Implantation?

1999 ◽  
pp. 141-148 ◽  
Author(s):  
Christos Coutifaris ◽  
Akinyinka Omigbodun ◽  
Piotr Ziolkiewicz ◽  
John Hoyer
Keyword(s):  
Embryo Implantation ◽  
Endometrial Receptivity ◽  
Human Endometrium

scholarly journals Value of endometrial echo pattern transformation after hCG trigger in predicting IVF pregnancy outcome: a prospective cohort study

2019 ◽  
Vol 17 (1) ◽  
Author(s):  
Zhaojuan Hou ◽  
Qiong Zhang ◽  
Jing Zhao ◽  
Aizhuang Xu ◽  
Aihua He ◽  
...  
Keyword(s):  
Cohort Study ◽  
Prospective Cohort Study ◽  
Prospective Cohort ◽  
Embryo Implantation ◽  
Implantation Rate ◽  
Endometrial Receptivity ◽  
Roc Curve Analysis ◽  
Vitro Fertilization ◽  
Hcg Trigger

Abstract Background There is much value in identifying non-invasive ways of measuring endometrial receptivity, as it has the potential to improve outcomes following in vitro fertilization (IVF). It has been suggested that endometrial echogenicity on the day of hCG administration was a good marker of endometrial receptivity. In the daily practice, we notice that patients with non-homogeneous hyperechoic endometrium on the embryo transfer day usually have lower pregnancy rates. We therefore extended the research onward transformation of echo pattern after hCG trigger to analyze the relationship between endometrial echogenicity transformation and IVF outcomes. Methods A total of 146 infertile women undergoing their first IVF cycle were recruited in the prospective cohort study from August 2017 through August 2018. A series of endometrial echo pattern monitoring was carried out in these patients after hCG trigger: hCG day, from 1 through 3 days after ovum pick-up (OPU + 1, OPU + 2, OPU + 3). Results The endometrial echogenicity value was calculated as the ratio of the hyperechogenic endometrial area over the whole endometrial area. Clinical pregnancy rate and embryo implantation rate had positive relationship with echogenicity value. The ROC curve analysis of endometrial echogenicity showed the area under curve was greatest on the second day after oocyte retrieval (OPU + 1, 2, 3 were 0.738, 0.765, 0.714 respectively) versus pregnancy. Endometrial echogenicity value on OPU + 2 had a higher predictive efficiency, and the cutoff value was 76.5%. The sensitivity was 61.3% and specificity was 82.0%. When putting the cut-off at <60%, the sensitivity was 93.8% and the specificity was 23.1%. Conclusions The endometrial echogenicity value on OPU + 2 was recommended to evaluate endometrial receptivity. It seemed appropriate for clinicians to provide a ‘freeze all’ IVF cycle and transfer in a subsequent frozen-thawed embryos cycle when echogenicity value <60% on OPU + 2. Trial registration The registration number was ChiCTR-OOC-17012214 and the registration date was August 1st, 2017.

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