| 庄亮,于世英,黄晓园,等.Ku80表达抑制细胞模型的建立及其功能检测[J].中华放射医学与防护杂志,2007,27(5):447-450.ZHUANG Liang,YU Shi-ying,HUANG Xiao-yuan,et al.Construction of cell model of silenced Ku80 and the radiobiology change of HeLa cell[J].Chin J Radiol Med Prot,2007,27(5):447-450 |
| Ku80表达抑制细胞模型的建立及其功能检测 |
| Construction of cell model of silenced Ku80 and the radiobiology change of HeLa cell |
| 投稿时间：2006-09-04 |
| DOI： |
| 中文关键词: Ku80 RNA干扰 稳定转染 放射敏感性 细胞周期 |
| 英文关键词:Ku80 RNA interference Stable transfection Radiosensitivity Cell cycle |
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| 中文摘要: |
| 目的 建立利用RNAi技术抑制Ku80表达的HeLa细胞模型,探讨Ku80在放射生物学方面的功能。方法 构建靶向抑制Ku80的siRNA表达质粒,转染HeLa细胞,筛选稳定表达的转化克隆;Western blotting检测Ku80表达变化;6 MV X线照射6 Gy后,流式细胞术检测细胞凋亡率和细胞周期;克隆形成实验检测细胞SF2、D0等值。结果 构建的质粒转染HeLa细胞获得3个稳定转染克隆,Western blotting分析表明2个阳性克隆Ku80蛋白抑制率分别达到89.3%和96.4%;Ku80表达抑制细胞受X线照射后48及72 h的凋亡率高于对照细胞(P＜0.05),但4株细胞的细胞周期变化无统计学意义(P＞0.05);2个阳性克隆细胞株的D0和SF2值明显降低,在D10剂量时的增敏比分别为1.315和1.365。结论 运用RNAi技术建立的Ku80表达抑制克隆细胞可以成为简单实用的细胞模型;Ku80-siRNA可以抑制Ku80的表达,从而促进HeLa细胞对X线的敏感性。 |
| 英文摘要: |
| Objective To construct the cell model of Ku80 with expression inhibited by siRNA and to explore the role of Ku80 in radiobiology. Methods Ku80-siRNA expression plasmids were constructed and HeLa cells were transfected with these plasmids by lipofectamine. Western blotting was used to measure the expression of Ku80. After irradiation with 6 MV X ray,cells were collected and analyzed by flow cytometry for apoptosis and cell cycle at 24, 48 and 72 h; The radiobiology parameters of four cell lines were acquired by clone formation array. Results Three stable transfected cell clones were obtained, and the inhibition rates of Ku80 protein expression of two positive clones were 89.3% and 96.4%; The apoptosis rates of HeLa cells Ku80 inhibited were higher than control cells at 48 and 72 after X ray irradiation (P＜0.05), but the differences of cell cycle change after irradiation were not significant (P＞0.05). HeLa cells of silenced Ku80 had lower SF2 and D0 than control cells, and their SER (sensitization enhancement ratio) based on D10 were 1.315 and 1.365, respectively. Conclusions The HeLa cell models with Ku80 expression suppressed were successfully established; the inhibition of Ku80 by siRNA could enhance the radiosensitivity of HeLa cells. |
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