摘要目的 探讨突变型Rad50蛋白联合放射线照射对人鼻咽癌细胞株CNE1杀伤作用.方法 实验设空白对照组、Ad-Rad50-GFP作用组、Ad-EGFP作用组、单纯照射组、Ad-Rad50-GFP+照射组、Ad-EGFP+照射组,携带突变型Rad50蛋白基因的重组腺病毒载体Ad-Rad50-GFP转染CNE1细胞,免疫印迹实验检测MRN复合体相关组分Mre11、Rad50、Nbs1的表达情况,中性彗星电泳检测突变型Rad50蛋白对细胞损伤修复的影响,细胞生长曲线研究突变型Rad50蛋白联合放射线照射对细胞生长的影响.结果 照射完成时和照射后24h,Ad-Rad50-GFP作用组中Mre11、Rad50、Nbs1的表达量均显著低于空白对照组(照射完成时:0.48比0.62,0.42比0.50,0.53比0.69,照射后24 h:0.41比0.69,0.46比0.58,0.34比0.78,P值均＜0.05);在照射后不同时间点,Ad-Rad50-GFP+照射组的平均尾矩(MTM)均高于单纯照射组(照射完成时:16.06比14.8,照射后24 h:58.23比15.89,照射后48 h:45.12比11.42,P值均＜0.05);照射后第7天,Ad-Rad50-GFP+照射组细胞数量显著均低于空白对照组和单纯照射组(P值均＜0.05).结论 突变型Rad50蛋白细胞内表达可显著增强放射线对鼻咽癌细胞株CNE1的杀伤作用.

abstractsObjective To investigate the killing effects of radiation and mutant Rad50 transfection on human nasopharyngeal carcinoma cell line CNE1.Methods The experimental groups included:control group,Ad-Rad50-GFP group,Ad-EGFP group,irradiation group,Ad-Rad50-GFP combined with irradiation group,and Ad-EGFP combined with irradiation group.CNE1 cells were transfected with recombinant adenoviral vector Ad-Rad50-GFP carrying mutant Rad50 gene.The expressions of Mre11,Rad50,Nbs1,and relevant constituents composing MRN complex were detected by Western Blot.Neutral comet assay was used to detect the effect of mutant Rad50 on restoration process of DNA damage.Cell growth curve was used to evaluate the growth inhibition of CNE1 by mutant Rad50 and radiation.Results Expressions of Mre11,Rad50,and Nbs1 in cells of Ad-Rad50-GFP group were less significantly than those in control group when irradiation was completed (0.48 vs 0.62,0.42 vs 0.5,and 0.53 vs 0.69,respectively,P ＜ 0.05) and 24 hours after irradiation (0.41 vs 0.69,0.46 vs 0.58,and 0.34 vs 0.78,respectively,P ＜0.05).The mean tail moment (MTM) in Ad-Rad50-GFP plus irradiation group was higher than that in irradiation group when irradiation was completed (16.06 vs 14.8,P ＜ 0.05),24 hours after irradiation (58.23 vs 15.89,P ＜ 0.05) and 48 hours after irradiation:(45.12 vs 11.42,P ＜ 0.05).Seven days after irradiation,the cells in Ad-Rad50-GFP plus irradiation group was less than those in control group or irradiation group (both P ＜ 0.05).Conclusion Mutant Rad50 enhances killing effects of radiation on nasopharyngeal carcinoma cell line CNE1.