| 摘要: |
| 目的 通过RNA干扰（RNAi）技术沉默DNA聚合酶polβ的表达，观察其对食管癌细胞顺铂（cDDP）耐药性的影响。方法 构建靶向polβ基因的siRNA重组慢病毒pRNAT-U6.2/Lenti-polβ1、pRNAT-U6.2/Lenti-polβ2及阴性对照载体pRNAT-U6.2/Lenti-polβC，将其感染人食管鳞癌顺铂耐药细胞系EC9706/cDDP，通过RT-PCR、蛋白质印迹分析和免疫荧光实验观察细胞polβ的表达情况，采用MTT法观察不同浓度顺铂对细胞生长的抑制作用并计算50%细胞生长抑制所需的药物浓度（IC50）和耐药指数（RI）。结果 靶向polβ的siRNA重组慢病毒pRNAT-U6.2/Lenti-polβ1和pRNAT-U6.2/Lenti-polβ2感染EC9706/cDDP后均可下调polβ的mRNA和蛋白表达水平，其中前者效果更为显著。顺铂以剂量依赖方式抑制EC9706/cDDP细胞增殖，感染pRNAT-U6.2/Lenti-polβ1、pRNAT-U6.2/Lenti-polβC的EC9706/cDDP细胞及未感染EC9706/cDDP细胞对顺铂的IC50分别为55.71、62.41、63.11 μg/mL，耐药指数分别为13.9、15.5、15.7，其中前者与后二者相比差异有统计学意义（P<0.05）。结论 polβ的表达与食管鳞癌细胞系EC9706/cDDP顺铂耐药性有关，沉默其表达可部分逆转EC9706/cDDP细胞对顺铂的耐药性。 |
| 关键词: 食管肿瘤 肿瘤抗药性 顺铂 polβ RNA干扰 |
| DOI：10.3724/SP.J.1008.2014.00279 |
| 投稿时间：2013-07-18修订日期：2013-09-04 |
| 基金项目: |
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| RNAi silencing of polβ gene partly reversing drug-resistance of esophageal cancer cells to cisplatin |
| TANG Yue1,WANG Li2,LIU Lu-lu1,LI Min1* |
(1. Department of Microbiology and Immunology, College of Basic Medical Sciences, Zhengzhou University, Zhengzhou 450052, Henan, China;
2. Department of Parasitology, College of Basic Medical Sciences, Zhengzhou University, Zhengzhou 450052, Henan, China
*Corresponding author.) |
| Abstract: |
| Objective To silence polβ gene by RNAi technique and to observe its effects on the drug-resistance of esophageal carcinoma cells to cisplatin (cDDP). Methods We constructed recombinant lentivirus containing siRNA targeting polβ genes. The two lentiviral vectors containing siRNA targeting the polβ gene were designated as pRNAT-U6.2/Lenti-polβ1 and pRNAT-U6.2/Lenti-polβ2, and the negative control vector as pRNAT-U6.2/Lenti-polβC; they were used to infect EC9706/cDDP cells, which was resistant to cDDP. The expression of polβ was then examined by RT-PCR, Western blotting and immunofluorescence. MTT was used to observe the inhibitory effect of cDDP; the IC50 of cDDP and resistance index (RI) were calculated. Results Both pRNAT-U6.2/Lenti-polβ1 and pRNAT-U6.2/Lenti-polβ2 down-regulated polβ at mRNA and protein levels, with the effect of former being more prominent. Cisplatin inhibited EC9706/cDDP cell proliferation in a dose-dependent manner. The IC50 values of cDDP in cells infected with pRNAT-U6.2/Lenti-polβ1, pRNAT-U6.2/Lenti-polβC and untreated EC9706/cDDP were 55.71 μg/mL, 62.41 μg/mL, and 63.11 μg/mL, respectively, with the first one being significantly different from the latter two (P<0.05); and the corresponding RI values were 13.9, 15.5, and 15.7, respectively, also with the first one being significantly different from the latter two(P<0.05). Results It is suggested that polβ expression is associated with cDDP resistance in EC9706/cDDP, and silencing polβ by RNAi can partly reverse the drug resistance of EC9706/cDDP to cDDP. |
| Key words: esophageal neoplasms neoplasm drug-resistance cisplatin polβ RNA interference |
