目的： 探讨单独及联合转染survivin、 bcl-2反义寡核苷酸（AsODN）对胆囊癌细胞系GBC-SD的凋亡促进作用。方法： 设计并合成特异性靶向survivin及bcl-2的AsODN。免疫组织化学法观察胆囊癌细胞中Survivin及Bcl-2蛋白的表达情况；实验分为4组：空白对照组、survivin AsODN转染组、bcl-2 AsODN转染组、联合survivin及bcl-2 AsODN转染组。转染24 h后，采用逆转录 聚合酶链反应（RT-PCR）检测survivin基因表达的变化，电镜下观察细胞形态变化，流式细胞仪技术检测细胞凋亡率，四甲基偶氮唑蓝（MTT）法检测AsODN对细胞的生长抑制率。结果：Survivin及Bcl-2蛋白在胆囊癌细胞中均有较高表达；单独及联合转染survivin及bcl-2 AsODN后，胆囊癌细胞内survivin基因mRNA表达下调47.8%；，电镜下胆囊癌细胞呈典型凋亡样改变；凋亡率分别为11.38％±3.91％（survivin AsODN组）、9.26％±4.15％（bcl-2 AsODN组）、28.45％±6.34％（联合转染组）；分别与对照组相比差异有显著性（P<0.01），而联合转染组同单独转染survivin AsODN组及bcl-2 AsODN组相比差异也有显著性（P<0.05）。各转染组相对于空白对照组的AsODN抑制率分别为54.3％,47.6％,76.5％。结论：Survivin及Bcl-2蛋白在胆囊癌细胞中呈高表达，单独及联合转染survivin和bcl-2反义寡核苷酸可促进胆囊癌细胞凋亡，而联合转染更具显著性

Objective:To investigate the effect of survivin and bcl-2 antisense oligodeoxynucleotides (AsODN) combined transfection on the human gallbladder carcinoma cell line GBC-SD in vitro.Methods: Survivin and Bcl-2 protein expressions were detected by immunohistochemical method; Cultured cells were divided into 4 groups: Nomal control group, survivin antisense observed group, bcl-2 antisense observed group and combined group. After transfected for 24 h, the expression of survivin mRNA was detected by reverse transcription polymerase chain reaction(RT-PCR). Cell morphological changes were observed under electron microscopy. Apoptosis index (AI) was examined by flow cytometry; Inhibitory rate (IR ) was determined by the colorimetri MTT cell viability and proliferation assay.Results:Survivin and Bcl-2 protein were highly expressed in gall bladder carcinoma cells; The expression of survivin mRNA was decreased 47.8%. Abnormal morphological changes of cells were observed in the three AsODN transfection groups; The AI in survivin antisense observed group,bcl-2 antisense observed group,and combined group was 11.38％±3.91％, 9.26％±4.15％, 28.45％±6.34％ respectively and significantly higher than the nomal control group (P<0.01), and that in the combined group was significantly higher than the other two observed groups(P<0.05). The IR was 54.3％,47.6％,76.5％ respectively in the three AsODN transfection groups. Conclusion:Survivin and bcl-2 protein were highly expressed in gallbladder carcinoma cells. Survivin AsODN and bcl-2 AsODN can induce GBC-SD cell apoptosis and the effect was more markedly by combining the two AsODNs.