Effect Of GPER Specific Antagonist G15 On Invasion And Migration Of Endometrial Carcinoma And Its Mechanism

Objective:To explore the role of GPER specific antagonist G15 on ER positive Ishikawa cells,low ER expression of HEC-1 a and ER negative KLE cell.Then to investigate the effect of G15 on invasion and migration of three types of endometrial cancer cells,and the related protein level changes.Methods:Based on the preliminary experiment,the study confirmed that G15 had a significant inhibitory effect on the proliferation of endometrial carcinoma.This experimental stage focused on tumor invasion and migration.1 Cell culture:endometrial cancer cell line Ishikawa and HEC-1A were cultured in DMEM medium containing 10%fetal bovine serum.The KLE cell line was cultured with DMEM-F12 medium containing 10%fetal bovine serum.2 The changes of three cell migration capacities were detected by scratch healing test after different concentrations of G15.3 Transwell migration and invasion experiments were conducted to detect the changes in the invasion and migration ability of the three cells after different concentrations of G15.4 Western Blot method was used to detect the changes of GPER,MMP9and ERK protein expression levels in three endometrial cancer cells.Results:1.Scratches healing experiment showed:G15 inhibited the migration of HEC-1A、KLE cells with the increase of concentration and time.While G15had a weaker inhibition migration ability of Ishikawa cells.2.Transwell little chamber experiment showed:E2 promoted the invasion and migration ability of three types of endometrial carcinoma.The migration ability of HEC–1A and KLE cells was decreased under different concentration of G15(0mol/L、10^-9mol/L、10^-7mol/L、10^-5mol/L).The difference was significant（P<0.05）.The inhibition of G15 to migration by E2 was low in Ishikawa cells,different concentration was statistical significance（P<0.05）.3.Western Blot method was used to detect the effect of G15 on the expression of GPER,ERK and MMP9 in the three cells under E2.Three types of cells were treated for 48h with blank,E2（10-6mol/L）,E2（10-6mol/L）+G15（10-5mol/L）.The results showed that GPER,ERK and MMP9 were expressed in all three cells.E2 increased the expression of GPER,ERK and MMP9 protein in HEC-1A、KLE cells,compared with the control.G15 inhibited the up-regulation of GPER,ERK and MMP9 by E2 in HEC-1 A and KLE cells（P<0.05）.G15 reduced the expression of GPER in Ishikawa cells,no effect on the expression of ERK and MMP9 protein,significant difference（P<0.05）.Conclusion:1.E2 promotes cell invasion and migration through the activated membrane estrogen receptor GPER,and is evident in HEC-1A and KLE cells,the type II endometrial cancer cells.2.G15 has inhibitory effect on the migration of three type cells by E2,and the inhibitory effect on HEC-1A and KLE cell migration is dose-time dependent.3.In type II endometrial cancer,GPER specific antagonist G15 can significantly inhibit GPER-mediated GPER/MAPK/ERK signaling pathway.Therefore,GPER specific antagonist G15 may be a new targeted drug for the treatment of endometrial cancer.