The reinforcement of invasion in epithelial ovarian cancer cells by 17β-Estradiol is associated with up-regulation of Snail

Abstract

Objective. The transcription factor Snail, which is implicated in the triggering of epithelial-mesenchymal transitions (EMT), plays an important role in adhesion, invasion and metastasis of tumor cells. In the present study, we assessed 17β-Estradiol (E 2)'s effect on Snail, E-cadherin and MMP-2 expression of epithelial ovarian cancer cell line ES-2 and SKOV3. Then we induced Snail gene silencing by RNA interference to explore the effect of E 2 on E-cadherin and MMP-2 expression when Snail gene expression was blocked. Methods. Treated by 10 −8 M E 2, Snail, E-cadherin and MMP-2 mRNA expression of the cells was measured by RT-PCR; Snail, MMP-2 protein expression was detected by IHC; and MMP-2 activity was determined by Zymography. E-cadherin protein level was measured by Western blot. We constructed the small interfering dsRNA expression vector (pRNAT-U6.1/Neo-Snail) targeting Snail gene, as well as a negative control vector (pRNAT-U6.1/Neo-Neg). Then the cells were transiently transfected with the vectors. Western blot and zymography were conducted to determine E-cadherin protein level and matrix metalloproteinase activity of the cells transfected with pRNAT-U6.1/Neo-Snail or pRNAT-U6.1/Neo-Neg after treated with E 2 for 24 h. Results. The expression of ERα mRNA and protein was negative in ES-2 cells and positive in SKOV3 cells, and ERβ expression was positive in both cell lines. 10 −8 mol/l E 2 elevated expression of Snail and MMP-2 mRNA and protein in both ES-2 and SKOV3 cells, and reduced expression of E-cadherin mRNA and protein in SKOV3 cells. While in the RNAi group transfected with the small interfering dsRNA expression vector (pRNAT-U6.1/Neo-Snail) targeting Snail gene, E 2 treatment did not have a significant effect on MMP-2 activity or E-cadherin protein in ES-2 and SKOV3 cells. Conclusions. 17β-Estradiol increased Snail expression in both ERα-negative ES-2 cells and ERα-positive SKOV3 cells independent of the existence of ERα. The increase of MMP-2 expression in ES-2 and SKOV3 cells and decrease of E-cadherin expression in SKOV3 cells induced by E 2 were associated with up-regulation of Snail.