ABSTRACT
Objective To establish a cisplatin (DDP)-resistant HepG-2 cell line,and to explore the role of Wnt/β-catenin signaling pathway on multidrug resistance in human hepatocellular carcinoma.Methods The HepG-2 cells were exposed in a gradually increasing dose of DDP to establish a cisplatin ( DDP)-resistant HepG-2 cell line.MTT assay was used to detect the cytotoxic activity of DDP against HepG-2 and HepG-2/DDP cells.The mRNA expression of β-catenin was determined by Real-time PCR assay.The small interfering RNA was used to specifically knockdown β-catenin expression in HepG-2/DDP cells.The protein expression was detected by western blot analysis.Results The DDP-resistant cell line HepG-2/DDP was established by gradient DDP induction successfully.The IC50 values of DDP against HepG-2 and HepG-2/DDP cells were (2.29 ± 0.14) μmol/L and ( 20.51 ± 0.84 ) μmol/L,respectively ( t=95.68,P＜0.01 ),HepG-2/DDP cells was 8.96 times than HepG-2 cells on the resistance of cisplatin.The result of real time PCR showed that 2-△Ct value of β-catenin in HepG-2 cells and HepG-2/DDP cells were (0.323±0.065) and (0.674 ±0.097) (P＜0.01 ).And the protein expression of cisplatin in HepG-2/DDP cells was also significantly higher than that in the HepG-2 cells.The expresssion of β-catenin was significantly and specifically depleted by siRNA duplexes(P＜0.01 ).The IC50 values of cisplatin against HepG-2/DDP cells were (21.02 ± 1.64) μmol/L in cisplatin control group,(6.23 ± 0.68 ) μmol/L in SiRNA targeting interference group and ( 20.44 ± 1.26 ) μmol/L in SiRNA negative interference group,and there was significant difference between control group and SiRNA targeting interference group ( P＜0.01 ).Conclusion The Wnt/β-catenin signaling pathway was activated on the cisplatin(DDP)-resistant HepG-2 cell line and down regulation of β-catenin increased the chemosensitivity of HepG-2/DDP cells against cisplatin.It provided a theoretical basis for finding the new targets of multidrug resistance in liver cancer.