Research on Mouse Endometria Proteome of Different Stages in Blastocyst Implantation
|School||Chongqing Medical University|
|Keywords||Proteome Annexin A1 Embryo implantation|
Objective: embryonic invasive process called endometrial implantation or implantation (implantation -). Embryo implantation is a complex and delicate process, involving embryos in endometrial adhesion, invasion, development and differentiation, and synchronize changes in the endometrium. Embryonic development to a specific period of time, a complex series of changes must occur in the endometrium, and reached receptivity state embryo to successfully implant, but so far its mechanism is not yet clear. The present study is limited to a single protein, can not explain the entire complex process. In this study, proteome analysis of mouse embryonic protein differences in the different periods of endometrial implantation, the molecular mechanisms that contribute to the overall level of the proteome of embryos implanted. Methods: 1, NIH mice animal models of pregnancy, separation pregnancy d3, d5, d7 endometrial tissue in mice, were stored at -80 ℃. 2, two-dimensional gel electrophoresis (2-dimensional gel electrophoresis, 2-DE) were isolated pregnancy d3, d5, d7 the mouse endometrial tissue protein, Coomassie blue staining, PDQuest software for image analysis. 3 of 12 differentially expressed proteins with trypsin gel digestion, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis of digested fragments, peptide mass fingerprinting the diagram (PMF) to use the Internet the the PMF identification procedures on Matrix Science (Mascot: Peptide Mass Fingerprint) queries, each query to get a score (Mowse Score), score calculation of the probability of each search result to evaluate whether it makes sense. Usually acceptable domain values ??are: the random probability of occurrence of an event is less than 5%, then the event is meaningful (p LT; 0.05). 4, collecting pregnant d3, d5, d7 mouse endometrium by RT-PCR, Western blotting Determination of Annexin A1 mRNA expression levels of Annexin A1 using immunohistochemistry and in situ hybridization qualitative detection of Annexin A1 and distribution of Annexin A1 mRNA in mouse endometrium, and it a semi-quantitative analysis. The results of the 1,2-DE map, the mouse endometrial tissue protein molecular weight (Mw) are mainly concentrated in the range of 14.4 ~ 116 kDa, isoelectric point (pI) is mainly distributed in the pH 4.0 ~ 8.0. Comparing pregnant 3d, 5d, 7d the different periods endometrial protein test dye the map (PDQuest7.3 software analyzes the image), results show pregnant d3, d5, d7 endometrial protein expression profiling broadly consistent with the 713 protein spots were ( d3), 736 (d5), 673 (D7). Pregnant d5 endometrial 2-DE maps for reference, pregnant d3, d7 2-DE map their matching rates were 68%, 61%. 3 times more differentially expressed protein in a total of 105: pregnant d3 endometrial expression of progesterone d5 24 raised more than three times the protein expression of the protein spots down more than 3 times 7; with the pregnant d7 uterus endometrium compared pregnant d5 expression raised more than three times the protein 32 expression down more than 3 times the protein 19; compared with pregnant d7 endometrium, progesterone d3 raised more than three times the protein expression 15 expression protein spots were cut three times more than the 8. 2 MALDI-TOF-MS identification of 12 differentially candidate protein spots, database queries, five protein spots get meaningful results (p lt; 0.05), respectively, apolipoprotein AI (Apo-AI) Annexin A1 (Annexin A1), glutathione transferase (Glutathionetransferase omega-1, GSTO1-1), a serine protease inhibitor (Serine protease inhibitor A3M), enolase (Alpha-enolase). 3, RT-PCR, in situ hybridization, Western blotting and immunohistochemistry method analysis showed Annexin A1 gene expression and protein expression in pregnant d3 strong, the most pregnant d5, d7 weakest pregnant, its main distribution in pregnant mice endometrial glandular epithelium and luminal epithelium. Conclusion Apo-AI in pre-implantation can be detected, and lasted late into the implant, suggesting that Apo-AI has to promote the role of the mouse embryo implantation; Annexin A1 d3 and d5 endometrial, organization was significantly higher than d7, suggesting that the protein may promote endometrial through multiple pathways of apoptosis, inflammation response regulator reach receptivity state to promote embryo implantation; GSTO1-1 may be through the local immune endometrial the regulation to promote embryo implantation; serine protease inhibitor A3M preimplantation, is followed by the expression d5 strongest, prompting may prevent matrix degradation over to participate in embryo implantation by inhibiting the activity of the protease, into the regulation; the enolase in pregnant d3 highest expression, suggesting that the protein may be involved in the plasmin degradation of extracellular matrix, trophoblast cell infiltration ready. 2, Annexin A1 in embryonic implantation period in mice endometrial epithelial luminal epithelium were expressed, and has a very strong period of pregnant d5 (implantation window period) to express the strongest. Thus speculated the Annexin A1 may be involved in the early embryo implantation endometrial adhesion and embryonic trophoblast invasion decidualization change. 3, the results of this study show that in mouse embryos around the bed of endometrial tissue differentially expressed proteins are mostly involved in the immune response and uterine stromal degradation of protein molecules, these proteins in regulating endometrial optimal receptivity state and plays a very important role in promoting embryo implantation. The peak Annexin A1 expression in mouse endometrium with mouse embryonic implantation time highly consistent, suggesting that Annexin A1 is closely related to embryo implantation.