siRNA-Gli-1联合BCNU抑制胶质瘤细胞U251增殖  

The Inhibition of siRNA-Gli-1combined BCNU on prliferation in Glioma Cell line U251

作　　者：田海龙[1] 李惠武[2] 李卉[2] 段明军[3] 姜孝芳[2] 白靖平[1] 

机构地区：[1]新疆医科大学附属肿瘤医院,新疆乌鲁木齐830000 [2]新疆医科大学基础医学院细胞与分子生物教研室,新疆乌鲁木齐830000 [3]新疆医科大学第一附属医院医学中心,新疆乌鲁木齐830000

出　　处：《基础医学与临床》2012年第5期554-560,共7页Basic and Clinical Medicine

摘　　要：目的探讨siRNA-Gli-1联合卡莫司汀(BCNU)对人脑胶质瘤U251细胞增殖和凋亡的影响和作用机制。方法 U251细胞按处理方式不同分为5组:BCNU组(BCNU)、siRNA-Gli-1组(siRNA-Gli-1)、联合组(siRNA-Gli-1+BCNU)、空转组(转染试剂LipofectamineTM2000,lipo)和空白组(细胞培养液RPMI1640)。U251细胞转染siRNA-Gli-1,半定量PCR和Western blot方法检测Gli-1表达及siRNA-Gli-1联合BCNU对BCL-2、CyclinD1和BAX表达的影响;MTT法检测细胞生长抑制率、流式细胞仪检测细胞周期、Annexin V法检测细胞凋亡,观察siRNA-Gli-1联合BCNU对U251细胞增殖能力的改变。结果 siRNA-Gli-1组和联合组中Gli-1 mRNA表达较其他3组明显下降(分别为40.35±0.17、45.20±0.23、93.30±0.43、98.60±0.06和95.93±0.21,P<0.05);BCL-2、CyclinD1表达显著下降,BAX蛋白表达增加或无变化,联合组较siRNA-Gli-1组更明显(P<0.05);与空白组和空转组相比,BCNU组、siRNA-Gli-1组和联合组细胞凋亡率增加(分别为0.76%±0.70%、0.53%±0.82%、12.87%±0.57%、22.23%±0.39%和28.76%±1.55%,P<0.01),细胞分裂明显阻滞于G0/G1期,S期减少;siRNA-Gli-1组、BCNU组和联合组细胞生长抑制率均随时间的延长而升高(P<0.05)。结论 siRNA-Gli-1联合BCNU抑制胶质瘤U251细胞的增殖机制可能与阻滞细胞周期、增强BAX和抑制BCL-2表达有关。Objective To observe the inhibition effect and mechanism of siRNA-Gli-1 combined BCNU on the proliferation and apoptosis in glioma 251 cells.Methods Five groups were divided： BCNU group（BCNU）,siRNA-Gli-1 group（siRNA-Gli-1）,combination group（siRNA-Gli-1＋BCNU）,transfection reagents group（transfection reagents LipofectamineTM2000,lipo）and blank control group（RPMI1640）.siRNA-Gli-1 was transfected into U251 glioma cells,the mRNA and protein expression of Gli-1gene were assessed by semi-quantitative reverse transcription poly-merase chain reaction（sqRT-PCR） and Western blot.After five groups were treated by BCNU and siRNA-Gli-1,the mRNA and protein expression of Gli-1,BCL-2、CyclinD1 and BAX were detected using sqRT-PCRand Western blot,and the proliferation and apoptosis of U251cells were examined by MTT,Flow Cytometry and Annexin V methods.Results The expression of Gli-1 in siRNA-Gli-1 and combination group declined distinctly,as compared with other three groups;In siRNA-Gli-1 group and combination group,especially in combination group,the expression of BCL-2 and CyclinD1 decreased obviously and the expression of BAX increased or had no change.Flow Cytometry analysis showed that the proliferation and apoptosis of U251 cells in BCNU group、siRNA-Gli-1 group and combination group were inhibited and increased respectively and G0/G1 phase rate increased and S phase rate decreased,as compared with the other two groups;The inhibition rate of siRNA-Gli-1、BCNU and combination group increased and stepped by time.Conclusions The effect that siRNA-Gli-1,combined with BCNU enhances the inhibition of U251 cell,may be related to cell cycle arrest and decreasing the ratio of BAX/BCL-2.

关 键 词：胶质瘤 Gli-1基因 RNA干扰 卡莫司汀 

分 类 号：R739.4[医药卫生—肿瘤]