基因芯片分析RNA干涉MSP58后细胞周期相关基因表达的变化  

Changes of cell cycle related gene expression after MSP58 gene RNA interference

作　　者：林伟[1] 章翔[1] 张璟[2] 费舟[1] 刘卫平[1] 药立波[2] 

机构地区：[1]第四军医大学西京医院神经外科,陕西西安710032 [2]第四军医大学基础部生物化学与分子生物学教研室,陕西西安710032

出　　处：《中华肿瘤防治杂志》2010年第19期1501-1505,共5页Chinese Journal of Cancer Prevention and Treatment

基　　金：国家自然科学基金(39970752);陕西省科技计划项目资助(2008K09-09)

摘　　要：目的:采用基因芯片技术,观察RNA干涉法抑制人脑胶质瘤U251细胞MSP58基因表达后细胞周期相关基因的变化,从而了解MSP58在细胞周期相关基因通路中的可能定位。方法:将特异性的MSP58干涉表达载体pSilencer3.1-MSP58转染人脑胶质瘤细胞系U251,同时构建相应的阴性对照载体pSilenc-er3.1-NC以及空载体pSilencer3.1-H1neo。运用半定量RT-PCR及蛋白质印迹法,检测稳定转染和未转染干涉载体的胶质瘤细胞MSP58的mRNA和蛋白表达水平。运用基因芯片技术分析pSilencer3.1-MSP58转染后细胞周期相关基因的变化。结果:成功建立了具有稳定下调MSP58基因表达的U251-S细胞、阴性对照U251-NC细胞以及含有空载体的U251-H1neo细胞。干涉组细胞MSP58的表达无论在mRNA水平还是在蛋白质水平均较阴性对照组及空载体组细胞显著降低,P<0.01。基因芯片技术共筛选细胞周期相关基因128个,其中上调及下调>2倍的共有33个基因。结论:干涉MSP58基因表达后,胶质瘤细胞增殖的抑制与细胞周期相关基因表达的变化有关。OBJECTIVE：To investigate the changes of cell cycle related gene expression after the suppression of MSP58 by RNA interference in U251 cells using DNA microarray,in order to understand the role of MSP58 in cell cycle related gene pathway. METHODS：Human glioma cells （U251） were transfected with the specific siRNA expression vector （pSilencer3.1-MSP-58） designing to target MSP58 mRNA. A corresponding site-mutated vector was constructed as a negative control （pSilencer3.1-NC）. The expression of MSP58 mRNA and its protein among the stable transfected cells and the untransfected ones were detected by semi-quantitative RT-PCR and Western blotting respectively. RESULTS：Three stable transfected cell lines： U251-S （with pSileneer3.1-MSP8）,U251-NC （with pSilencer3.1-NC） and U251-H1 neo （with empty vector pSilencer3.1-H1 neo） were established. The expression levels of MSP58 gene mRNA and protein in U251-S were significantly lower than those in U251-NC,U25 1-H1 neo and those untransfected U251 cells （P0.01）. A total of 128 genes related with cell cycle were examined by DNA microarray. Totally 33 genes were up-regulated and down-regulated by more than 2 folds.CONCLUSION：The inhibition of human glioma cell proliferation by MSP58 gene RNA interference may be concerned in the changes of cell cycle related gene expression.

关 键 词：基因表达 RNA干扰 神经胶质瘤 细胞周期 印迹法 RNA 

分 类 号：R739.41[医药卫生—肿瘤]