siRNA沉默ATM增强CpG ODN 7909对肺癌A549细胞的放射增敏作用  

Silencing ATM by siRNA enhances the radiosensitization effect of CpG ODN 7909 on lung cancer A549 cells

作　　者：袁素娟[1] 乔田奎[1] 刘小群[1] 陈伟[1] 

机构地区：[1]复旦大学附属金山医院肿瘤科,上海200540

出　　处：《中国肿瘤生物治疗杂志》2013年第3期278-283,共6页Chinese Journal of Cancer Biotherapy

基　　金：上海金山区科技创新基金资助项目(No.2010-316)~~

摘　　要：目的:研究siRNA沉默毛细血管扩张—共济失调突变(atxia-telangiectasia mutated,ATM)基因的表达增强胞嘧啶鸟嘌呤二核苷酸寡脱氧核苷酸(cytosine-phophate-guanine oligodeoxynucleotide,CpG ODN)7909对人非小细胞肺癌A549细胞的放射增敏作用。方法:将ATM-siRNA转染至A549细胞中,Western blotting检测A549细胞中ATM蛋白的表达。A549细胞随机分为6组:对照组、CpG组、X射线(IR)组、CpG+IR组、ATM-siRNA+CpG+IR组和NC-siRNA+CpG+IR组,克隆形成分析法检测各组细胞克隆形成率,Graphpad prism 5.0软件进行单击多靶模型和L-Q线性模型拟合辐射后A549细胞的生存曲线,以D0、Dq、N、α/β及SF2等参数分析A549细胞辐射损伤修复能力,流式细胞术检测A549细胞的凋亡。结果:ATM-siRNA转染可明显抑制A549细胞中ATM蛋白的表达(P<0.01)。X射线可剂量依赖性抑制A549细胞的克隆形成能力(P<0.05);且CpG+IR组A549细胞的克隆形成能力进一步降低(P<0.01);ATM-siRNA转染后,CpG处理的A549细胞克隆形成能力再度降低[10 Gy时,(0.05±0.00)%vs(0.71±0.00)%,P<0.01]。辐射损伤剂量生存曲线结果显示,ATM-siRNA转染后,ATM-siRNA+CpG+IR组较CpG+IR组A549细胞的α/β值明显增大(1.48 vs 0.97,P<0.05),对放射损伤修复能力明显减弱。CpG+IR组较IR组细胞凋亡率显著升高[(9.18±0.16)%vs(6.56±0.33)%,P<0.01];ATM-siRNA+CpG+IR组A549细胞凋亡率进一步升高[(10.45±0.40)%vs(9.18±0.16)%,P<0.05]。结论:siRNA沉默ATM的表达可增强CpGODN 7909对A549细胞的放射增敏作用,ATM可作为肺癌治疗的潜在靶点。Objective：To explore the potentiation of silencing atxia-telangiectasia mutated（ATM） gene expression in the radiosensitization effect of cytosine-phophate-guanine oligodeoxynucleotide（CpG ODN） 7909 on non-small cell lung cancer A549 cell line.Methods： ATM-siRNA was transfected into A549 cells,and the expression of ATM protein in A549 cells was examined by Western blotting.A549 cells were randomly classified into six groups： control group,CpG group,X-ray（IR） group,CpG＋IR group,ATM-siRNA＋CpG＋IR group and NC-siRNA＋CpG＋IR group.Cell colony rates were evaluated by colony formation assay.One-hit multi-target model and linear quadratic model,which generated the radiation dose survival curve of the A549 cells,were fitted with Graphpad prism 5.0 software,and the parameters,including D0,Dq,N,α/β and SF2 were applied to analyze radiation damage repair capacity of A549 cells.The apoptosis of A549 cells was analyzed by flow cytometry.Results： ATM-siRNA transfection remarkably inhibited the expression of ATM protein in A549 cells（P0.01）.X-ray radiation inhibited the colony formation capacity of A549 cells in a dose-dependent manner（P0.05）.Moreover,a further decrease of the colony formation capacity of A549 cells was found in CpG＋IR radiation combined treatment group（P0.01）.With the transfection of ATM-siRNA,the colony formation capacity of CpG-treated A549 cells was further decreased（10 Gy,[0.05±0.00]% vs [0.71±0.00]%,P0.01）.Radiation damage dose survival curves demonstrated that the value of α/β was significantly increased（1.48 vs 0.97,P0.05）and the radiation damage repair capability was significantly decreased in ATM-siRNA＋CpG＋IR group compared with CpG＋IR group.The apoptotic rate was significantly increased in CpG＋IR group compared with IR radiation group（[9.18±0.16]% vs [6.56±0.33]%,P0.01）.The apoptotic rate of A549 cells in ATM-siRNA＋CpG＋IR group was further increased（[9.18±0.16]% vs [10.45±0.40]%,P0.05）.Conclusion： Silencing expression of

关 键 词：非小细胞肺癌 胞嘧啶鸟嘌呤二核苷酸寡脱氧核苷酸7909 A549细胞 放射敏感性 ATM 凋亡 

分 类 号：R734.2[医药卫生—肿瘤] R730.54[医药卫生—临床医学]