干扰hTERT降低人乳腺癌MCF-7细胞对γ射线引起的DNA损伤反应  

The repressive effects of hTERT interference on γ radiation induced DNA damage response in MCF-7 cells

作　　者：石嵘[1] 马文丽[1] 高洋[1] 周珏宇[1] 丁大鹏[1] 余海浪[1] 郑文岭[1] 

机构地区：[1]南方医科大学基因工程研究所,广东广州510515

出　　处：《基础医学与临床》2010年第8期847-851,共5页Basic and Clinical Medicine

基　　金：广东省科技计划(2009B060700112)

摘　　要：目的研究hTERTRNA干扰对人乳腺癌MCF-7细胞γ射线照射引起的DNA损伤反应的影响。方法通过反转录病毒为载体的hTERT-siRNA,抑制人乳腺癌MCF-7细胞中端粒酶催化亚单位hTERT表达,实时定量RT-PCR及Western blot确认hTERT表达水平。用137Cs放射源以3 Gy剂量γ射线照射细胞,于照射前以及照射后1、2、4、8和12 h收集细胞,采用磷酸化53BP1抗体进行免疫荧光染色,并于照射前、照射后1、4和12 h收集细胞,Western blot检测53BP1蛋白磷酸化水平。结果 hTERT-siRNA处理的细胞hTERT表达水平比对照细胞降低3.20(2-△△Ct)倍,hTERT蛋白水平降低2.56倍。hTERT-siRNA处理的细胞对γ射线引起的DNA损伤反应显著降低。结论 RNA干扰hTERT表达水平,可以降低人乳腺癌MCF-7细胞对γ射线照射引起的DNA损伤反应。Objective To study the effect of hTERT RNA interference on γ irradiation induced DNA damage response in breast cancer cell line MCF-7.Methods A retrovirus carrying hTERT-siRNA was used to repress the expression of telomerase catalyze unit hTERT in breast cancer cell line MCF-7.The hTERT expression was confirmed by real-time RT-PCR and Western blot.3 Gy dosage of γ ray from 137Cs was applied to irradiate the cells,cells were then collected and fixed before and 1,2,4,8 and 12 h after irradiation,phospho-53BP1 antibody was used for immunofluorescent staining.Cells were also collected before and 1,4,12 h after irradiation and phosphorylation level of 53BP1 was detected by Western blot as well.Results The hTERT expression level of cells treated with hTERT-siRNA was repressed by 3.20 folds（2-△△Ct） when compared with the control cells.The hTERT protein level also decreased by 2.56 folds.DNA damage response of cells treated with hTERT-siRNA was repressed obviously.Conclusion DNA damage response induced by γ irradiation in breast cancer cell line MCF-7 can be inhibited through RNA interference of hTERT expression level.

关 键 词：端粒酶 Γ射线 人乳腺癌MCF-7细胞 DNA损伤反应 

分 类 号：R737.9[医药卫生—肿瘤]