Research On Effect Of Expression Of β1,4-GalT-Ⅰ By Hormone Endometrium And Correlation With EGFR Signaling Pathway

Objective:Researching on the express of β1,4-GalT-I on uterus endometrium cells(RL95-2cell) under the role of estrogen and progesterone before implantation, and itseffect of attachment function. Analyzing the effect to EGFR and related signal paths,under different estrogen and progesterone conditions. Analyzing the change of EGFRexpressed when regulating β1,4-GalT-I. Analyzing the molecular mechanismof howβ1,4-GalT-Ion uterus endometrium regulates the embryo implantation, and discuss therelationship with EGFR signal path.Methods:(1) By using the implantation model (human monolayers of uterineepithelial cell (RL95-2) and human trophoblast cell (JAR) are used to form the in vitroimplantation model), changing estrogen and progesterone’s level in the environment ofuterus endometrium: Use Western blot to detect the effect of estrogen and progesteroneon the expression of β1,4-GalT-I on uterus endometrium cells. Use Immune cellchemical dyeing to analyze the changing of Galβ1-4GlcNAc with estrogen andprogesterone. Analyze the status of Galβ1-4GlcNAc forming withestrogen andprogesterone by using RCA-1Lectin-blot technique. Detect the impact of estrogen andprogesterone on the expression level of proteins of EGFR signal path. Use Western blotto analyze the relationship between progesterone level and the expression of p-EGFR.Countthe cell attachment rate of JAR cell on the RL95-2cells, to analyze therelationship of estrogen and progesterone’s level.(2) Over-expressβ1,4-GalTⅠplasmids, interfering plasmids, empty vector plasmids, and interferencecontrol plasmids transfect with RL95-2cells, analyze the changing of JAR cellsattachment on RL95-2cells after regulating the expression of uterineepithelialβ1,4-GalT Ⅰ. Use Western blot to analyze the impact of EGFR’s expressionby regulating the expression of uterine epithelial β1,4-GalT Ⅰ. Results:(1) the expression of uterine epithelial β1,4-GalT Ⅰ. has dose andtime-effect relationships with estrogen and progesterone, and the best functionalcondition of estrogen is10-4ug/ul、48h (P<0.01), the best functional condition ofprogesterone is10-4ug/ul、48h (P<0.01);(2) uterine epithelial estrogen and progesteronecan advance the attachment of JAR cells on RL95-2cells, and the attachment rate is:estrogen group (84.7±0.4%), progesterone group (91.4±0.7%), and estrogen combinesprogesteronegroup (88.0±0.9%), all groups are higher than the normal group(78.3±0.7%)(P<0.01) and the progesterone group has the highest advance function.Progesterone group and estrogen combines progesterone group can improve theexpression of uterine epithelialEGFR, NF-κB, ICAM-1and Galβ1-4GlcNAc(P<0.01)obviously;estrogen group, progesterone group, and estrogen combinesprogesterone group advance the expression of β1,4-GalT-Ⅰ protein and EGF protein.(3)Progesterone improves the level of uterine epithelial EGFR’s phosphorylation (P<0.01);(4)After uterine epithelial cell transfects with β1,4-GalTⅠ regulation plasmids, theattachment rates of JAR cells on RL95-2cells are: Over-express group (90.5±1.3%),which is higher than normal goup (77.9±0.9%), empty vector group (79.0±1.9%),interference control group; however, the attachment rate of interfering group reducesobviously (55.2±0.6%)(P<0.01); also, the expression of EGFR of interfering group isincreased, and the over-express group does not have obvious change (P<0.01).Conclusion:(1) estrogen and progesterone can regulate the expression of uterineepithelialβ1,4-GalT-Ⅰ(have dose and time-effect relationships) and advance JAR cellsattachment on RL95-2cells;(2) progesteronecanincrease molecular expression inrelated signal path of uterine epithelial cells EGFR, and regulate the embryoimplantation.(3) The effect of implantation by controlling and regulating the expressionof β1,4-GalT-Ⅰmay pass through the EGFR signal path.