Effect Of Inhibiting Survivin Expression With Antisense Oligodeoxynucleotides Compound On Apoptosis Of Hepatocellular Carcinoma Cell Line SSMC-7721 Induced With Quercetin

Objective:Survivin Antisense Oligodeoxynucleotides(ASODN)compound was used to inhibit the expression of Survivin,and then to study the effect of apoptosis of hepatocellular carcinoma cell line SSMC-7721 induced with Quercetin.Methods:Human hepatocellular carcinoma cell line SSMC-7721 was cultured in vitro,and took the logarithmic growth phase cells with experiment.The whole experiment was divided in two parts-â… andâ…¡.The firstâ… part was divided in empty control group,Lipofectamine^TM2000group,SODN compound group,ASODN compound group,and the ODN compound was transfected into SSMC-7721 cells mediated by Lipofectamine^TM2000for 48h;The secondâ…¡part was continued on the firstâ… part,and then to study the effect of apoptosis of SSMC-7721 cells induced with Quercetin for 24h. The second part was relatively divided in empty control group,40umol/L Quercetin group,Lipofectamine^TM2000group,SODN compound group,ASODN compound group,ASODN compound combined with Quercetin group.The changes of survivin protein and Survivin mRNA expression after the firstâ… part were assessed by cell immuneohisto- chemistry and reverse transcription-polymerase chain reaction(RT-PCR) method.After the secondâ…¡part,The changes of morphology of SSMC-7721 cells were observed with AO staining,and the growth inhibition rate was measured by MTT,and the apoptotic rate of SSMC-7721 cells was measured by the flow cytometry with Annexi-V/PI staining.The cell cycle change was detected by the flow cytometry with PI staining.Results:The expression of Survivin mRNA and its protein had been significantly down-regulated by transfecting ASODN compound for 48h(P＜0.01);After the SSMC-7721cells that had been efficiently down-regulated Survivin gene expression were exposed to Quercetin for 24h,and controling with other group,theASODN compound combined with Quercetin group:the typical morphological changes of cell apoptosis and the occurring apoptotic cell numbers that had increased were observed very clearly with AO staining,the growth inhibition rate or the apoptotic rate of SSMC-7721 cells of combining group were higher significantly than other group(P＜0.01).The ASODN compound blocked cell cycle in G₂/M stage and the quercetin blocked cell cycle in S stage.When ASODN combined with quercetin could block the cell cycle in G₂/M and S stange.Conclusion:Survivin is highly expressed in SSMC-7721 cells and the ASODN compound can efficiently inhibit expression of Survivin gene.Inhibiting expression of Survivin in SSMC-7721 cells with ASODN compound could promote the cell growth inhibition rate and the cell apoptotic rate.The effect may be related to its influence on inhibiting expression of cell Survivin gene.