Liu Bing, Zhang Hong, Duan Xin, Hao Jifang, Xie Yi, Zhou Qingming, Wang Yanling, Tian Yuan, Wang Tao
Key Laboratory of Heavy Ion Radiation Medicine of Gansu Province, Institute of Modern Physics, Chinese Academy of Sciences, 509 Nanchang Road, Lanzhou, China.
Cancer Biother Radiopharm. 2009 Feb;24(1):57-66. doi: 10.1089/cbr.2008.0514.
To determine whether adenovirus-mediated wild-type p53 transfer after radiotherapy could radiosensitize non-small-cell lung cancer (NSCLC) cells to subclinical-dose carbon-ion beam (C-beam), H1299 cells were exposed to a C-beam or gamma-ray and then infected with 5 MOI of AdCMV-p53 or GFP (C-beam or gamma-ray with p53 or GFP). Cell cycle was detected by flow cytometric analysis. The apoptosis was examined by a fluorescent microscope with DAPI staining. DNA fragmentation was monitored by the TUNEL assay. P53 mRNA was detected by reverse-transcriptase polymerase chain reaction. The expression of p53, MDM(2), and p21 was monitored by Western blot. Survival fractions were determined by colony-forming assay. The percentages of G(1)-phase cells in C-beam with p53 increased by 8.2%-16.0%, 5.2%-7.0%, and 5.8%-18.9%, respectively, compared with C-beam only, gamma-ray with p53, or p53 only. The accumulation of G(2)-phase cells in C-beam with p53 increased by 5.7%-8.9% and 8.8%-14.8%, compared with those in gamma-ray with p53 or p53 only, respectively. The percentage of apoptosis for C-beam with p53 increased by 7.4%-19.1%, 5.8%-11.7%, and 5.2 %-19.2%, respectively, compared with C-beam only, gamma-ray with p53, or p53 only. The level of p53 mRNA in C-beam with p53 was significantly higher than that in p53 only. The expression level of p53 and p21 in C-beam with p53 was significantly higher than that in both C-beam with GFP and p53 only. The survival fractions for C-beam with p53 were significantly less than those for the other groups (p < 0.05). The data suggested that AdCMV-p53 transfer could more efficiently radiosensitize H1299 cells to subclinical-dose C-beam irradiation through the restoration of p53 function.
为了确定放疗后腺病毒介导的野生型p53基因转移是否能使非小细胞肺癌（NSCLC）细胞对亚临床剂量的碳离子束（C离子束）产生放射增敏作用，将H1299细胞暴露于C离子束或γ射线，然后用5个感染复数（MOI）的AdCMV-p53或绿色荧光蛋白（GFP）（即p53或GFP与C离子束或γ射线联合处理）进行感染。通过流式细胞术分析检测细胞周期。用DAPI染色的荧光显微镜检查细胞凋亡情况。通过末端脱氧核苷酸转移酶介导的缺口末端标记法（TUNEL法）监测DNA片段化。通过逆转录聚合酶链反应检测p53信使核糖核酸（mRNA）。通过蛋白质免疫印迹法监测p53、小鼠双微体2（MDM2）和p21的表达。通过集落形成试验确定存活分数。与仅接受C离子束、p53与γ射线联合处理或仅p53处理相比，p53与C离子束联合处理组中G1期细胞的百分比分别增加了8.2% - 16.0%、5.2% - 7.0%和5.8% - 18.9%。与p53与γ射线联合处理组或仅p53处理组相比，p53与C离子束联合处理组中G2期细胞的积累分别增加了5.7% - 8.9%和8.8% - 14.8%。与仅接受C离子束、p53与γ射线联合处理或仅p53处理相比，p53与C离子束联合处理组的细胞凋亡百分比分别增加了7.4% - 19.1%、5.8% - 11.7%和5.2% - 19.2%。p53与C离子束联合处理组中p53 mRNA的水平显著高于仅p53处理组。p53与C离子束联合处理组中p53和p21的表达水平显著高于p53与GFP联合处理组和仅p53处理组。p53与C离子束联合处理组的存活分数显著低于其他组（p < 0.05）。数据表明，AdCMV-p53基因转移可通过恢复p53功能更有效地使H1299细胞对亚临床剂量的C离子束照射产生放射增敏作用。
