靶向沉默Ku80增强顺铂诱导的人肺腺癌细胞凋亡

Targeted reduction of ku80 enhanced cisplatin-induced apoptosis in lung adenocarcinoma cells

目的:观察沉默Ku80能否增强顺铂诱导的人肺腺癌耐药细胞的细胞凋亡。方法:首先采用RT-PCR、Westernblot方法比较Ku80在人肺腺癌亲代(A549)与耐药细胞系(A549/DDP)的表达水平;将Ku80siRNA及si-Scramble转染至A549/DDP细胞后,加药组转染后48小时加顺铂作用24小时;应用MTT法检测顺铂对各组细胞的抑制率,采用流式细胞仪检测凋亡率,Caspase-3活性检测试剂盒检测Caspase-3活化程度。结果:Ku80 mRNA、蛋白在A549/DDP表达水平显著高于其亲代A549;si-Ku80组A549/DDP细胞的Ku80的mRNA、蛋白表达水平下调;顺铂对si-Ku80组细胞的抑制率明显高于si-Scram-ble组与Control组;在6μg/ml顺铂作用24小时后,si-Ku80组细胞凋亡率及Caspase-3活化程度高于si-Scramble组和Control组(P<0.05)。结论:靶向Ku80siRNA导入人肺腺癌耐药细胞特异性下调Ku80的表达,增强顺铂诱导的人肺腺癌耐药细胞凋亡作用。

Objective：To investigate the potential of Ku80 small-interfering RNA（siRNA） to enhance cisplatin-induced apoptosis.Methods：Ku80 mRNA and protein expression levels were investigated by reverse transcription-PCR and Western blot analysis,respectively.Ku80-specific siRNA was synthesized and transfected into cisplatin-resistant human lung adenocarcinoma cell line A549/DDP prior to treatment with cisplatin.Cell survival was assessed by MTT assay.Apoptosis was assessed using flow cytometry analysis.Caspase-3 activity was detected by spectrophotometer.Results：The Ku80 mRNA and protein expressions were significantly elevated in A549/DDP cells compared with the parental A549 cells.The Ku80 mRNA and protein expressions were significantly downregulated by si-Ku80 in A549/DDP cells.Treated with cisplatin for 24 h,the inhibition rates of si-Ku80 cells decreased significantly compared with in control group and si-Scramble group.After 24 h treatment with 6 μg/ml cisplatin,apoptosis rate in si-Ku80 group was increased compared with the control group and si-Scramble group（P0.05）;activities of Caspase-3 significantly increased in siKu80-A549/DDP cells as compared with control cells and si-Scramble cells（P0.01）.Conclusion：Ku80 may correlate with the chemoresistance to cisplatin in lung cancer.Ku80-specific siRNA enhanced cisplatin-induced apoptosis in the cisplatin-resistant A549/DDP cells.