Effects Of Lentivirus-mediated Silence Of Alpha Enolase Gene On Growth,Invasion And Metastasis Of Human Cervical Cancer Cells

The aerobic glycolysis provides most of the building blocks required for cell proliferation and therefore is an essential character of cancer cells. α-enolase(ENO1) is an important glycolytic enzyme and plays a key role in aerobic glycolysis. ENO1functional is diversity.It can catalyze glycolysis2-phosphoglycerate (PGA) converted to phosphoenolpyruvate (PEP). In addition, the ENO1gene encodes the Myc-binding protein-1(MBP1), which downregulates the activity of c-myc protooncogene. ENO1may play a role in the development of tumor. The purpose of this study is to build two stable Hela and SiHa cell lines with ENO1gene silencing by lentivirus-mediated RNA interference technology. To investigate the effects on human cervical cancer cell growth and invasion and metastasis.We silenced the expression of ENO1in two cervical cancer cell line Hela and Siha cells with lentivirus-mediation. Following ENO1knock-down, The expression of ENO1was confirmed by western-blot. The groups were divided into ENO1gene silencing group (pLKO.1shENO1), negative control group (scr) and normal cell group. The drug sensitivity, invasion, metastasis, tumourigenic abilities of ENO1altered cells were analyzed by wound healing assay, trans-well migration or invasion Assay and immunocytochemistry. Then to investigate the probable mechanisms.The results of this study showed that cervical cancer cell lines with ENO1gene silencing were established successfully.The Inhibition rate of cisplatin and paclitaxel with different concentrations on three groups of cells were detected by MTT after48h.The result showed that the suppression rate of different concentrations of drugs on cells of ENO1gene silencing group (pLKO.1shENO1) is higher than negative control group (scr) and normal cell group. The results were statistically significant. The result of clonogenic assay showed that the number of visible cell clones of the cells with ENO1gene silencing were significantly less than the normal cells and the negative control cells. The number of colony forming of Hela pLKO.1shENO1group and SiHa pLKO.1shENO1groups were5.17%and6.33%, These are significantly lower than the negative control cells(P<0.05). The result of cell scratch assay showed that cell migration rate of pLKO.1shENO1groups were lower than the negative control cells. The ability of migration was detected by transwell chamber without matrigel. The cells of pLKO.1shENO1groups transferred to the lower chamber was significantly reduced. Each group was randomly selected five averages vision for statistical analysis. The difference was statistically significant. The ability of invasion was detected by transwell chamber with matrigel.The result showed that the number of cells with ENO1gene silencing transferred to the lower chamber was significantly less than negative control cells (P<0.05). To detect the expression of invasion and metastasis-related factors VEGF, EGFR, MMP9, COX2, Notch2by immunocytochemistry, SiHa pLKO.1shENO1cells was significantly decreased than negative control cells(P<0.05).Our studies demonstrated that knock-down ENO1enhanced Hela and SiHa cell line cells’ sensitivity to paclitaxel and cisplatin and suppressed the tumorigenicity of them. In addition, the abilities of invasion and metastasis of the two cell lines with silence of ENO1decreased. In conclusion,our study clearly showed that there was a correlation between ENO1and cellular growth. ENO1involved in growth, invasion, and metastasis of human cervical cancer cells.