Expression of Twist in hepatocellular carcinoma and effect of silencing Twist gene on the proliferation and apoptosis of hepatocellular carcinoma cells

Abstract

Objective To investigate the expression of Twist in hepatocellular carcinoma (HCC),the relationship between Twist and hypoxia inducible factor (HIF)-lα,and the effect of silencing Twist and HIF-1 α genes on the proliferation and apoptosis of HCC HepG2 cells by RNA interference.Methods The expression of Twist and HIF-1α was detected in 106 cases of HCC by immunohistochemical staining method.Twist and HIF-1α-small interfering RNA (siRNA) were transfected into HepG2 cells mediated by LipofectamineTM 2000.HepG2 cells were divided into five groups:Twist interference group,HIF-1 α interference group,double interference group,negative control group (transfected with negative-siRNA) and blank control group (transfected with Lipofectamine).Cell apoptosis was examined by flow cytometry and cell growth was measured by methyl-thiazol-tetrazolium (MTT) assay.The direct interactions between Twist and HIF-1α were uncovered by chromatin immunoprecipitation assay (ChIP).Results Twist positive expression rate was high (66.98％) in HCC tissue but low in the corresponding peritumoral tissue (17.92％,P＜0.05).There was a positive relationship between the expressions of Twist and HIF-1α(P ＜ 0.05).The apoptosis rate in blank control group,negative control group,Twist interference group,HIF-1o interference group and double interference group was 3.98％,4.26％,41.62％,43.84％,and 45.12％,respectively.The apoptosis rate in interference groups was significantly higher than that in control groups (P ＜ 0.01),while there was no significant difference among three imerference groups (P ＞0.05).The cell growth inhibitory rate in Twist interference group,HIF-1α interference group and double interference group was 30.2％,34.0％ and 35.8％,respectively (P ＞0.05).There were the direct interactions between Twist and HIF-1α promoter in HepG2 by ChIP.Conclusion There was a positive relationship between the expression of Twist and HIF-1α.Down-regulation of Twist and HIF-1α could induce apoptosis of HepG2 cells and inhibit cell growth.The double knockdown of Twist and HIF-1 α does not show a synergistic effect,and there were direct interactions between Twist and HIF-1α. Key words: Carcinoma, hepatocellular; Twist; Hypoxia inducible factor-1 α; Apoptosis ; Cell proliferation