目的：建立慢病毒介导的livin基因沉默系统，探讨其对肺癌细胞凋亡的影响。方法：Livin shRNA慢病毒感染肺腺癌细胞株SPCA1沉默livin基因表达。应用PI染色经荧光镜下观察SPCA1细胞凋亡形态，流式细胞术检测SPCA1细胞凋亡率及亚二倍体峰形成，Realtime PCR及Western blotting方法检测livin和caspase 3表达的改变。结果：livin基因在肺腺癌细胞株SPCA1中持续高表达。经慢病毒介导shRNA使livin基因表达沉默后，镜下可见肺腺癌细胞出现典型凋亡形态特征，流式细胞术检测出现亚二倍体峰，细胞凋亡率较空白对照及阴性病毒对照细胞明显增加（8.21％ vs 0.08％, 0.13％；P<0.05），RTPCR及Western blotting 检测结果显示，caspase 3 mRNA表达无改变，但cleavedcaspase 3蛋白表达上调。结论：慢病毒载体介导的shRNA能抑制肺腺癌细胞株SPCA1中livin基因的表达，从而促进SPCA1细胞凋亡。

Objective:To construct a lentiviral livin shRNA vector to silence livin gene expression, and to study its effect on apoptosis of lung carcinoma cells. Methods:Livin expression in human lung adenocarcina SPCA1 cells was silenced by lentiviral livin shRNA infection. The morphology of apoptotic cells was observed by propidine iodide staining and fluoroscope; apoptosis rate and sublipliod apoptotic peak of SPCA1 cells were assessed by flow cytometry; expression of livin and caspase 3 in SPCA1 cells was examined by realtime PCR and Western blotting analysis. Results: Livin was constitutively expressed in SPCA1 cells. After livin expression was silenced by lentiviral livin shRNA infection, SPCA1 cells showed the characteristic morphology of apoptosis under fluoroscope, and the sublipliod apoptotic peak was identified by flow cytometry. Apoptosis rate in livin shRNA infected SPCA1 cells was significantly higher than that in blank and negative control groups (8.3%  vs 0.08% and 0.13%, P<0.05). caspase 3 mRNA expression in SPCA1 cells had no change but the expression of cleavedcaspase 3 was greatly upregulated after lentiviral livin shRNA infection as showed by RTPCR and Western blotting analysis. Conclusion: Lentiviral livin shRNA can inhibit livin expression in human lung adenocarcina SPCA1 cells and induce cell apopotosis.

福建省自然科学基金资助项目（No. 2008J0074 ）