17β雌二醇对卵巢透明细胞癌ES-2细胞系侵袭力及Snail表达的影响

17β Estradiol's Impact on Cell Proliferation,Invasion and Snail Expression of Ovarian Clear Cell Adenocarcinoma Cell Line ES-2

目的研究雌激素对体外培养的卵巢透明细胞癌细胞系ES2增殖、侵袭力的影响和对MTA3、Snail、基质金属蛋白酶2(MMP2)表达的调节作用,以探讨雌激素作用对ES2细胞系的影响。方法体外培养卵巢透明细胞癌ES2细胞系,去激素培养7d后加入17β雌二醇(E2)1×10-7、1×10-8、1×10-9mol/L,MTT法检测细胞增殖情况,流式细胞仪测定细胞周期,改良的Boyden小室检测细胞侵袭力、运动力的变化,RTPCR检测MTA3、Snail、MMP2mRNA表达,免疫组化法检测Snail、MMP2表达,明胶酶谱检测基质金属蛋白酶活性。结果RTPCR、WesternBlot结果示ES2细胞系为ERα阴性表达,ERβ阳性表达,Ecad为阴性表达。17βE21×10-7、1×10-8、1×10-9mol/L均可促进细胞增殖;E21×10-8mol/L可使G0～G1期细胞减少,S期细胞增加,但对细胞凋亡无影响;17βE21×10-8mol/L可显著增强ES2细胞的侵袭力和趋向运动能力,并可降低MTA3mRNA表达,增加Snail、MMP2mRNA及蛋白表达。结论17βE2可显著促进ES2细胞的侵袭能力,此作用可能是通过17βE2对MTA3表达下调和对Snail、MMP2表达的上调实现的。

Purpose To assess 17β estradiol （E2）＇s effect on cell proliferation, cell invasiveness and MTA3,Snail, MMP-2 expression of ovarian clear cell adenocarcinoma cell line ES-2, and to further investigate its mechanisms. Methods Before all experiments,ES-2 cell line was grown in medium depleted of steroid for more than 7 days. Treated by 1 × 10^-7,1 × 10^-8 and 1 × 10^-9 mol/L 17β E2, cell viability of ES- 2 was determined by MTT method,and cell cycle distribution and apoptosis were detected by flow cytometry （FCM）. The invasion and mobility assay was performed by using the modified Boyden chambers. MTA3, snail and MMP-2 mRNA expression was measured by RT-PCR, and Snail, MMP-2 protein expression was detected by IHC, and MMP-2 activity detected by zymography. Results RT-PCR and Western Blot showed that the expression of ERα and E-cad mRNA and protein in ES-2 cell line was negative, while ERβ expression was positive. E2 1 × 10^-7, 1 × 10^-8 and 1 × 10^-9 mol/L all stimulated cell proliferation. E2 1 × 10^-8mol/L reduced the proportion of G0--G1 phase, increased the proportion of S phase, but it had no effect on apoptosis. E2 1 × 10^-8mol/L significantly improved invasion and mobility of ES-2 cell. Cells treated with 1× 10^-8mol/L 17β E2 led to reduced mRNA expression of MTA3, and elevated mRNA and protein expression of Snail and MMP-2. Conclusions 17β E2 improved invasion of ES-2 cell,and the effect maybe mediated by downregulation o{ MTA3 and upreguation of Snail and MMP-2.