抑制 Livin 基因对肺腺癌 SPC-A1 细胞增殖及顺铂敏感性的影响

目的:Livin是近年来发现的人类凋亡抑制蛋白(inhibitorof apoptosis protein,IAP)家族的新成员,发现在多数肿瘤中表达,与肿瘤发生有密切关系,并可能作为肿瘤诱导凋亡治疗的新靶点。本研究旨在探讨si RNA-Livin和夫拉平度(Flavopiridol,FP)协同凋亡诱导配体(TRAIL)两种方式有效抑制Livin表达,诱导非小细胞肺癌SPC-A1凋亡并增强对化疗药物顺铂的敏感性。方法:si RNA-Livin转染SPC-A1,Real-Time PCR检测Livin基因的表达水平,MTT检测干扰组和干扰加药组肿瘤细胞的增殖及活性;TRAIL、FP单独及联合作用诱导细胞凋亡,蛋白质印迹法检测凋亡抑制蛋白Livin的表达水平,MTT检测各处理组细胞的增殖及活性。结果:100 nmol/LFP处理组(F)细胞存活率为(84.30±1.34)%,100 ng/m LTRAIL处理组(T)为(93.40±1.56)%,FP和TRAIL联合组(F+T)为(48.02±1.35)%,si RNA-Livin处理组为(50.88±1.14)%,1.2μg/m L Cisplatin处理组为(19.30±0.89)%,si RNA-livin+Cisplatin组为(14.37±0.81)%,FP+T+Cisplatin组为(10.86±0.87)%,C组存活率为100%。F+T组对细胞的增殖抑制作用显著高于单独用药组,si RNA-livin+Cisplatin与si RNA-Livin组相比、FP+T+Cisplatin与FP+T组相比都显著增强了化疗药物对SPC-A1的杀伤作用。50μmol/LZ-VAD-FMK预处理后联合用药组细胞的存活率为(88.16±1.64)%,caspase抑制剂能明显抑制F+T联合处理组的凋亡效应。结论:RNA干扰和F+T联合用药都能显著降低凋亡抑制蛋白Livin的表达,有效抑制肿瘤细胞的增殖生长,并增强肿瘤细胞对化疗药物顺铂的敏感性,为肺腺癌的靶向治疗提供新的理论依据。

Inhibition of Livin Expression Suppresses Cell Proliferation and Enhances the Chemosensitivity to Cisplatin in Human Lung Adenocarcinoma Cells

Objective:Livin is a novel member of the inhibitor of apoptosis protein (IAP) family that has been reported to be overexpressed in a variety of human malignancies. In order to investigate how depletion of Livin affects the proliferation of human lung adenocarcinoma cells, we used two quite different ways to down-regulate the livin expression, including RNAi-based approach specifically targeting Livin mRNA and the synergistic inhibitory effect between Flavopiridol and TRAIL.Methods:The siRNA recombinant expression vectors targeting Livin gene were transfected into SPC-A1 cells with Lipofectamine 2000. Real-time PCR was used to detect the expression of Livin mRNA. And the cell proliferation and viability was measured by MTT assay. The apoptosis inhibitory effect of Flavopiridol and TRAIL used singly and combinedly was also studied in SPC-A1 . Western blotting analysis was used to confirm the expression of Livin protein. Cell proliferation and viability was tested by MTT assay.Results:The survival rate of each group was showed as follows: the Flavopiridol treatment group was (84.30± 1 .34) %, the TRAIL treatment was (93.40± 1.56) %, the FP and TRAIL combination treatment group was (48.02± 1 .35) %, the siRNA-Livin treatment group was (50.88± 1 .1 4) %, the Cisplatin treatment group was (19.30± 0.89)%, the siRNA-livin and Cisplatin combination group was (1 4.37± 0.81) %, the FP and TRAIL and Cisplatin combination group was (1 0.86 ± 0.87) % , the Caspase inhibitor Z-VAD-FMK treatment group was (88.16 ± 1.64) % , the negative control group was 100 %. The survival rate of FP and TRAIL combination group was much lower than the FP group and TRAIL goup.Conclusion:Both the RNA interference and FT synergistic could obviously inhibited the expression of Livin, effectively decreased the proliferation of SPC-A1 cells, and promoted the sensitivity of chemotherapeutic medicine cisplatin. All of this provide new theoretical foundation for the targeted therapy of lung cancer.