Effect of Smac combined with cisplatin on expressions of apoptosis-regulated factors in hepatocellular carcinoma cells

ObjectiveTo study effects of the cotreatment of second mitochondria-derived activator caspase(Smac)and cisplatin on the expressions of caspase-3,caspase-9,and cytochrome c(Cyt c)in hepatocellular carcinoma cells(SMMC7721).MethodsSMMC-7721 cells were transfected with Smac by lipofectamine-mediated method and then screened by G418.Smac mRNA and protein expressions in SMMC-7721/Smac cells acquired by G418 screen were determined by reverse transcription-PCR(RT-PCR)and western blot and its growth curve was performed by methyl thiazolyl tetrazolium(MTT)assay.After cisplatin treatment,expressions of caspase-3,caspase-9,and Cyt c were evaluated by western blot.Results Expressions of both Smac mRNA and its protein in SMMC-7721/Smac cells increased significantly,and its A₄₉₀ value was significantly lower than that of SMMC-7721 cells in the same time-point(P<0.05,P<0.001).Protein expression detection showed that expressions of caspase-3 and Cyt c increased in SMMC-7721/Smac cells compared with that of SMMC-7721 cells,from 19 939 to 28 347 and 14 924 to 21 017,respectively(P<0.05-P<0.001,with F values of 12.588 and 23.811).Cisplatin treatment increased expressions of caspase-3,caspase-9 and Cyt c in both SMMC-7721 and SMMC-7721/Smac cells,especially in Smac cells.After the treatment of 25μg/ml cisplatin,expressions of caspase-3 and caspase-9 increased from 28 347 to 46 696 and 22 412 to 39 728,respectively(P<0.001,F with values of 12.588 and 12.180).ConclusionCotreatment of stable Smac over-expression and cisplatin could increase cellular expressions of caspase-3,caspase-9,and Cyt c,which could promote cell apoptosis.