中文摘要：

目的 构建稳定的XRCC2基因沉默大肠癌细胞系及阐明沉默XRCC2基因表达的大肠癌细胞对电离辐射损伤的反应。方法 采用Westernblot法检测不同肿瘤细胞系人大肠癌细胞系（T84、HT29、Lovo）、食管癌EC9706细胞、乳腺癌MCF-7细胞和人正常胚肾HEK293细胞中XRCC2蛋白的表达水平。大肠癌T84细胞经0、2、4、8、12GyX射线照射后和8GyX线照射后0、6、24、48h,用Western blot法和实时定量PCR法测定XRCC2蛋白和mRNA的表达水平。将XRCC2shRNA质粒转染T84细胞,用嘌呤霉素进行细胞筛选,采用Westernblot法和实时定量PCR法,检测沉默XRCC2蛋白和mRNA表达的效率。将XRCC2shRNA质粒转染T84细胞,经X线照射后,采用单细胞凝胶电泳测定沉默XRCC2基因表达的T84细胞的DNA损伤修复。结果 与人正常胚肾HEK293细胞相比,在不同肿瘤细胞系中XRCC2蛋白均呈不同程度的高表达,其中在大肠癌T84细胞中XRCC2蛋白表达最高。随着X线照射剂量的增加,T84细胞中XRCC2蛋白的表达水平逐渐升高,8Gy照射后XRCC2蛋白表达水平在48h内随着时间的延长而逐渐升高;XRCC2mRNA表达也表现出随剂量增加和时间延长而逐渐升高。与未处理的细胞相比,转染XRCC2shRNA质粒的细胞中XRCC2蛋白和mRNA表达明显下降,约分别减少了70%和60%。照射后的T84细胞DNA损伤修复中shRNA-XRCC2组TDNA%、TM和OTM均显著高于未处理组和shR-NA-SC组,差异有统计学意义（t=15.12、11.36、13.15,P〈0.01）。结论 成功建立了稳定表达XRCC2基因沉默的大肠癌T84细胞系,XRCC2基因沉默导致辐射诱导的DNA损伤修复能力下降。

英文摘要：

Objective To establish a colorectal cancer cell line with silenced XRCC2 gene expression and de- scribe the damage response of these cells induced by ionizing radiation. Methods XRCC2 protein expression in various human tumor cell lines including colorectal cancer cell lines （T84, HT29, Lovo）, esophageal carcinoma cell line （EC9706） and breast cancer cell line （ MCF - 7 ） and human embryo kidney cell line （ HEK293 ） was determined by west- ern blotting. XRCC2 protein and mRNA expression in T84 cells irradiated with 0 Gy, 2 Gy, 4 Gy, 8 Gy, 12 Gy X- ray and at 0 h, 6 h, 24 h, 48 h after exposure to 8 Gy radiation was determined by western blot and real - time quantitative PCR analyses. T84 cells were transfected with the vector - based shRNA plasmid （ shRNA - XRCC2） and selected with puromycin. The efficiency of XRCC2 gene silencing was determined by western blotting and real - time PCR. DNA dam- age repair of T84 cells transfeeted with shRNA - XRCC2 plasmid after exposure to X - ray radiation was determined by single cell gel electrophoresis. Results XRCC2 protein expression in various tumor cell lines was at different high levels compared with normal HEK293 cells and XRCC2 protein expression in colorectal cancer T84 cells was the highest. XRCC2 protein expression in T84 ceils increased with increasing dosage of X - ray exposure and with increasing duration within 48 h after exposure to 8 Gy radiation. XRCC2 mRNA expression of T84 cells also increased with increasing radiation dose and time after radiation. XRCC2 protein and mRNA expression in T84 cells transfected with shRNA -XRCC2 showed signifi- cant decline, down by 70% and 60%, respectively, compared with control cells. The value of TDNA%, TM and OTM in T84 cells irradiated with 8 Gy of the shRNA - XRCC2 group were significantly higher than that of the control group and shRNA - SC group （ t = 15.12,11.36 and 13.15, P 〈 0. 01 ）. Conclusion Colorectal cancer T84 cell line with silenced XRCC2 expression was established successfully. The