| Liver cancer is a serious disease to threat human health with high morbidity and mortality.China is a high incidence area of liver cancer,and the mortality of liver cancer is the highest in the world.Early diagnosis of Liver cancer has been difficult because of lacking typical clinical symptoms.Plenty of patients lost the chance of surgical treatment when firstly diagnosed in an advanced stage.The efficacy of clinical treatments besides the surgery,such as radiotherapy and biological therapy are not very satisfied.The malignant degree of liver cancer is high and it is easy to recur and metastasize,so that the treatment of advanced hepatocellular carcinoma is still a chanllenge worldwide.Therefore,the development of effective drug therapy is of great significance.Synthetic small molecule purine analogues reversine had been proved to have anti-tumor effects in a lot of human carcinoma cells including human prostate cancer cells,thyroid cancer cells,breast cancer cells,non-small cell lun cancer cells and human urothelial cell tumors.Besides,reversine had been proved to be a ATP competitive inhibitors of Aurora kinase inhibition.There have not been sisitematic reporting about reversine on hepatocellular carcinoma up to the present time,in this study,we try to explore the effect of reversine on the growth of human hepatocellular carcinoma cell HepG2 and its related mechanisms,which may help to provides theoretical foundation on new targets of hepatocellular carcinoma treatment.This study investigated the effects of reversine on the proliferation,clone formation,DNA content and apoptosis in human hepatocellular carcinoma cell HepG2.HepG2 cells were treated with different concentrations of reversine(0.1?M-1.6?M),and the effect of reversine on cell proliferation ability was detected by MTS.The long-term effect of reversine on human hepatocellular carcinoma HepG2 cells was evaluated by clone formation assay.Besides,after staining with Annexin-V-FITC/PI,flow cytometry was used to measure the apoptosis rate of human hepatocellular carcinoma cells HepG2 after reversine treatment.PI staining was used to test the HepG2 cell DNA content with flow cytometry.Moreover,Western blot methods to detect the expression of apoptosis-related protein PARP,Bax,Bcl-2,Bcl-xL and cell-cycle-related protein cyclinB1 and p21.Datas showed that reversine significantly suppressed the proliferation of HepG2 cell in a time and dosage-dependent manner(P<0.05);The IC50 of reversine in HepG2(72h)were 0.94?M.After incubation with reversine for 10 days,the colony formation of HepG2 was also inhibited.Furthermore,the macroscopic cell colony was virtually impossible to form with high-concentration of reversine treatment(0.2?M-1.6?M).Moreover,Reversine could induce the polyploid formation(P<0.05)in HepG2 cells in a dosage-dependent manner.Western blot results showed that,after 48 hours of incubation with reversine,the expression level of cyclinB1 decreased,meanwhile,the expression level of p21 increased.In addition,results also showed that reversine could also induce cell apoptosis in a dosage-dependent manner(P<0.05),and Western blot results showed that the expression level of PARP and Bax significantly increased after reversine treatment,and the expression level of Bcl-2 and Bcl-xL decreasedat the same time.In conclusion,Reversine can inhibit the growth of human hepatocellular carcinoma cell HepG2 through interfering the cell cycle and inducing polyploid formation.Moreover,it can induce apoptosis via a mitochondria-dependent endogenous apoptosis pathway. |
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