Osteopontin Increases The Expression Of β1,4-Galactosyltrans Ferase I And Affects Cellular Adhesion In Human RL95-2Cells

Objective: To explore the expression of Osteopontin (OPN) and β1,4-Galactosyltransferase-I (β1,4-GalT-I) in endometrium in early pregnant mice．To analyze theexpression and positioning of OPN and β1,4-GalT-I in the human uterus endometrium.Analyze the relationship between β1,4-GalT-I and OPN in vitro, to detect the humanrecombinant OPN protein (rhOPN) how to influence the β1,4-GalT-I expression andanalysis related pathways molecules. To research the effect of β1,4-GalT-I siRNA andrhOPN in blastocyst adhesion in easy implantation model in vitro．Methods: Edometrium samples were collected from D1-D5early pregnant mouse,used western blot and Real-time PCR to detecte the expression β1,4-GalT-I and OPN.Clinical endometrial specimen of proliferative phage and secretory phase wasassemble, to analyze allocation and expression of β1,4-GalT-I and OPN by Immunohistochemistry technique.Cell line human endometrial carcinoma RL95-2was used as a model of receptiveendometrium, rhOPN protein was used to stimulate RL95-2cells, detect the expressionof β1,4-GalT-I by Western-blot and Real-time PCR.Western-blot and Real-time PCR were used to investigate the signaling moleculesinvolved in the process of rhOPN regulated the β1,4-GalT-I.An adhesion assay was performed to investigate the adhesion of JAR cells to theRL95-2cells when rhOPN and β1,4-GalT-I siRNA exist or not.Results: In early pregnant mouse, β1,4-GalT-I and OPN were slightly increasedfrom D1to D4, and show a strong expression on D4, which was followed by a declinethereafter.In human endometrium section, both β1,4-GalT-I and OPN were expressedsurrounding gland, and in secretory phase endometrium the expression was significantlyhigher than proliferative phase endometrium. rhOPN could induce β1,4-GalT-I expression in time-and does-dependent mannerin RL95-2cells. At the point of12hours，200ng/ml rhOPN was the optimumcondition for RL95-2cells to express β1,4-GalT-I. In this condition rhOPN could activethe ERK1/2、AKT(thr308)、p38and degrade IκBα. When p38phosphorylation wasinhibited, there was no effection on rhOPN to the expresson of β1,4-GalT-I. Bothβ1,4-GalT-1protein and RNA expression were inhibited when ERK1/2, AKT andNFκB pathway were inhibited.In adhesion assay, rhOPN could significantly facilitate the adhesion ability of JARcells to the RL95-2cell monolayer. But no difference was seen between the adhesion ofJAR cells in the group which treated with rhOPN, at the same time, down-regulating thesynthesis of β1,4-GalT-1by β1,4-GalT-1siRNA.Conclusion:(1) β1,4GalT-Ⅰand OPN are participate in the process of embryo implantation.(2) OPN can induce β1,4-GalT-I expression in RL95-2cells and it might throughactivating ERK1/2, KT(thr308) and degrading IκBα.(3) OPN may regulate the expression of β1,4GalT-I to involve in the process ofadhesion...