Ufd1基因沉默逆转SW1116/HCPT细胞株对羟基喜树碱耐药的研究  

Study of Ufd1 Gene Silencing on Reverse of SW1116/HCPT Cell Line Resistance to Hydroxycamptothecin

作　　者：陈翔[1] 聂芳[1] 冉志华[1] 童锦禄[1] 朱明明[1] 徐锡涛[1] 萧树东[1] 

机构地区：[1]上海交通大学医学院附属仁济医院消化内科上海市消化疾病研究所,200001

出　　处：《胃肠病学》2011年第10期580-584,共5页Chinese Journal of Gastroenterology

基　　金：上海市重点学科建设项目(No.Y0205);国家自然科学基金项目(No.30770964);上海市科委重点实验室项目(No.06DZ22027);杨森科学基金项目资助

摘　　要：背景:泛素融合降解1样蛋白(Ufd1)在蛋白质逆向转运中发挥一定作用。前期研究发现羟基喜树碱(HCPT)耐药人结肠癌细胞株SW1116/HCPT Ufd1表达增高。目的:探讨Ufd1基因沉默在逆转SW1 116/HCPT细胞株对HCPT耐药性中的作用。方法:干扰组SW1116/HCPT细胞转染Ufd1特异性siRNA以沉默Ufd1基因,对照组转染非特异性siRNA。体外药物敏感性试验检测细胞对HCPT的敏感性,流式细胞术检测细胞周期,比色法测定caspase-3活性,蛋白质印迹法检测p-JNK、p53表达情况和细胞内泛素化蛋白贮留情况。结果:干扰组SW1116/HCPT细胞对HCPT的敏感性显著高于对照组(P<0.05)。经HCPT作用,干扰组和对照组细胞均出现S期阻滞、caspase-3活性增高和p-JNK、p53、泛素表达增加,干扰组更为明显。结论:以RNA干扰技术沉默Ufd1基因可逆转SW1116/HCPT细胞株对HCPT的耐药性,推测该作用可能与活化JNK信号通路、上调p53表达以及致细胞内泛素化蛋白贮留而破坏细胞内蛋白代谢的协调性有关。Ubiquitin fusion-degradation 1 like protein （Ufdl） plays a role in the retro-translocation of proteins. Preliminary studies have shown that Ufdl was up-regulated in hydroxycamptothecin （HCPT）-resistant human colon cancer cell line SW1116/HCPT. Aims： To study the role of Ufdl gene silencing on reverse of SW1116/HCPT cell line resistance to HCPT. Methods： SWlll6/HCPT celIs were transfected with plasmid containing Ufdl-specific siRNA （intervention group） or Ufdl-non-specific siRNA （control group）. In vitro drug sensitivity test was performed to assess the sensitivity of ceils to HCPT. Flow cytometry was used for analysis of cell cycle, and colorimetry was conducted for detection of caspase- 3 activity. Western blotting was performed to assess the expressions of p-JNK, p53 and cytosolic retention of ubiquitinated proteins. Results： SW1116/HCFF cells in intervention group was much sensitive to HCPT than ceils in control group （P〈 0.05）. S-phase arrest, activation of caspase-3, and up-regulation of p-JNK, p53 and ubiquitin were seen in both intervention group and control group when treated with HCPT. The above-mentioned changes were more significant in intervention group than in control group. Conclusions： RNA interference of Ufdl can reverse the resistance of SW1116/ HCPT cell line to HCPT. The mechanism is presumed to be related with activation of JNK pathway, up-regulation of p53, and cellular protein metabolism disturbance induced by cytosolic retention of ubiquitinated proteins.

关 键 词：结直肠肿瘤 羟基喜树碱 多药耐药相关蛋白质类 泛素类 基因 Ufd1 RNA干扰 

分 类 号：R734.2[医药卫生—肿瘤]