In Vivo Study Of E2/ GPER/Sphk Pathway In The Proliferation Of Endometrial Carcinoma

Objective:Determine the correlation between development of endometrial cancer and estradiol(E2),GPER by detection of GPER,ER expression in cancer tissues of patients.And study the mechanism of E2 induced endometrial carcinoma proliferation via GPER nongenomic activity.Methods: 1.The expression of GPER,ER in endometrial cancers.Tumor tissue specimens of patients with endometrial cancer were collected.And speciments from postmenopausal women without hourmone related disorder were choosed as control group.The expression of GPER,ER in cancer tissues was detected by immunohistochemistry.Plasma concentration of E2 and activity of SphK in cancer tissue were calculated.2.The study of GPER/SphK/ERK pathway in E2 induced endometrial carcinoma proliferation in nude mice.(1)Establish ovariectomized nude mice model.And establish xenograft tumor nude mice model by HEC-1A endometrial carcinoma cells injection.(2)Groups(1)Control group,(2)Estradiol(E2)group,(3)E2 +DMS group,(4)E2+G15 group,(5)E2+PD98059 group(3)Measured the diameter of tumor every three days,and calculated their tumor volumes.After three weeks,all mice were killed via cervical dislocation.Removed tumors and measured tumor weight.(4)Phosphorylation of ERK1/2 and expression of CyclinD1,E1,MMP-9,SphK1,SphK2 were observed via western blot method.(5)Activities of tumor SphK were measured.Results: 1.The expression of GPER,ER in endometrial cancers.Patients in endometrial cancer gourp had higher level of plasma E2.The GPER expression in ER positive patients and ER negative patients had no significant difference.GPER expression was correlated with FIGO stage,grade,myometrial invasion.And the activity of SphK in GPER positive patients is higher.2.The study of GPER/SphK/ERK pathway in E2 induced endometrial carcinoma proliferation in nude mice.(1)The effects of E2 on tumor growth in xenograft tumor nude mice.After 21 days of administration,E2 can efficiently increase tumor weights which are inhibited by G15,DMS and PD98059.(2)The effects of E2 on SphK activity of tumor tissue in xenograft tumor nude mice.E2 increase the activity of SphK of tumor,and either G15 or DMS can abrogate the change of SphK,whereas PD98059 had no effect of SphK activity.(3)The effects of E2 on SphK1 and SphK2 expression of tumor tissue in xenograft tumor nude mice.E2,G15,DMS and PD98059 had no effect on expression of SphK1 and SphK2 in tumor tissue of mice.(4)The effects of E2 on phosphorylation of ERK1/2 and expression of Cyclin D1,E1 and MMP-9 of tumor tissue in xenograft tumor nude mice.E2 can promote phosphorylation of ERK1/2.On the contrary,PD98059 can inhibit increased ERK1/2 phosphorylaiton induced by E2.While G-15 and DMS can block the change of p-ERK.E2 can increase expression of Cyclin D1,E1 and MMP-9,these effects can be inhibited by G-15,DMS and PD98059.Conclusion:E2 can promote the development of endometrial carcinoma cells via GPER.And GPER/SphK/ERK1/2 pathway can mediate E2 induced HEC-1A proliferation,which is related to increased expression of Cyclin D1,Cyclin E1,MMP-9.