凋亡抑制基因Livin对肺腺癌细胞A549增殖与耐药的作用

Effect of Livin on Proliferation and Multi-drug Resistance of Lung Cancer Cells A549

目的研究凋亡抑制Livin对肺腺癌细胞A549增殖与耐药的作用。方法通过脂质体将真核表达载体pc DNA3.1-Livin转染至人肺腺癌细胞A549中,经过G418筛选获得稳定表达Livin的A549细胞克隆;采用RT-PCR和Western blot法检测Livin在转染细胞中基因和蛋白的表达,通过流式细胞仪分析细胞周期的分布,应用MTT比色法检测细胞在不同化疗药物作用下的细胞耐药性,并通过RT-PCR和Western blot法检测转染细胞中P-糖蛋白(P-gp)基因和蛋白的表达。结果在pc DNA3.1-Livin转染后的A549细胞中,Livin基因的m RNA和蛋白表达显著增加,G0/G1期细胞比例降低,而S期细胞比例增高;与对照组比较,转染pc DNA3.1-Livin的A549细胞中P-gp基因的表达明显增高,对ADM、MTX、CTX和DDP等多种化疗药物的耐药性也明显增强(P<0.05)。结论 Livin基因的表达升高能增强肺腺癌细胞A549的增殖能力,并且可以通过增加P-gp的表达降低细胞对多种化疗药物的敏感性。

Objective To study the effect of the inhibitor of apoptosis protein, Livin on proliferation and multi-drug resistance of lung adenocareino- ma cells A549. Methods A549 cells were transfected with the eukaryofie expression vector pcDNA3.1 -Livin. A549 cell clone with stable expres- sion of Livin was obtained through G418 screening. Expressions of Livin mRNA and protein in the transfected cells were respectively measured by re- verse transcription polymerase chain reaction （RT-PCR） and Western blot. The distribution of cell cycle phase was determined using flow eytometry. The level of P-gp mRNA and protein in A549 cells transfected with pcDNA3.1-Livin was detected by RT-PCR and Western blot. The analysis of multi-drug resistance of A549 treated with different chemotherapeutics was performed by MTY. Results The mRNA and protein expressions of Liv- in were both significantly increased in the transfected A549 cells. The flow eytometry analysis showed there was higher percentage of S phase and low- er percentage of C-n/G1 phase in A549 cells transfected with pcDNA3.1 -Livin. Compared with control groups, the expression of P-gp mRNA and pro- tein was increased in A549 cells transfected with pcDNA3.1- Livin, which showed a higher drug resistance and lower sensitivity to chemothempic drugs such as ADM, MTX, CTX, and DDP （P 〈 0.05）. Conclusion Overexpression of Livin could enhance the proliferation of A549 cells, and high expression of P-gp caused by Livin could serve as one of the causes for multi-drug resistance in lung adenccarcinoma against chemotherapies