RNA干扰C-erbB-2对肺腺癌细胞calu-3增殖的影响  

Effect of RNAi-mediated Gene Silencing of C-erbB-2 on Proliferation of Lung Adenocarcinoma Cell Line calu-3

作　　者：任淑华[1] 张伟[2] 曲平[2] 刘义[2] 王井伟[3] 张林[1] 

机构地区：[1]中国医科大学第一附属医院胸外科,辽宁沈阳110001 [2]中国医学科学院中国协和医科大学肿瘤研究所肿瘤生物学检测中心,北京100021 [3]唐山工人医院骨外科,河北唐山063000

出　　处：《癌症》2005年第10期1173-1178,共6页Chinese Journal of Cancer

基　　金：国家自然科学基金项目(No.30371624)~~

摘　　要：背景与目的:C-erbB-2基因在人乳腺癌、卵巢癌、肺癌中扩增及过量表达,与肿瘤恶性程度增高、转移能力增强、化疗抵抗密切相关,且与预后不良有关。RNA干扰(RNA interfering,RNAi)是一种新的基因治疗技术,能有效、特异性地抑制细胞内基因表达。本实验研究靶向C-erbB-2基因的siRNA(small interferingRNA)对C-erbB-2过表达的肺腺癌细胞calu-3增殖的影响。方法:化学合成的靶向C-erbB-2基因siRNA在脂质体的介导下转染calu-3细胞,观察转染前后细胞表型的改变;通过逆转录聚合酶链反应(reversetranscription-polymerasechainreaction,RT-PCR)和流式细胞术检测转染前后细胞中C-erbB-2mRNA和蛋白水平的变化,四甲基偶氮唑蓝(MTT)法评价对calu-3细胞增殖的抑制作用,流式细胞术检测细胞周期的变化和细胞凋亡情况。结果:靶向C-erbB-2基因siRNA降低了calu-3细胞中C-erbB-2mRNA和蛋白的表达,转染C-erbB-2siRNA48h后calu-3细胞出现C-erbB-2蛋白表达下调。C-erbB-2蛋白阳性率为(25.04±1.56)%,与未转染对照组、空载体组、非特异性siRNA组比较[(98.24±2.23)%、(95.67±1.98)%和(94.79±0.87)%]有显著性差异(P<0.01)。同时C-erbB-2siRNA对calu细胞增殖有抑制作用,使细胞周期阻滞于G0/G1期[C-erbB-2siRNA组(56.6±3.6)%,未转染对照组(45.5±3.2)%,P<0.01],并能诱导细胞凋亡。结论:化学合成的靶向C-erbB-2基因siRNA能有效抑制C-erbB-2基因在calu-3细胞中的表达,并对calu-3细胞增殖有抑制作用。BACKGROUND ＆ OBJECTIVE： C-erbB-2 gene is amplified or overexpressed in breast cancer, ovarian cancer, and lung cancer, and is related with enhanced malignancy and metastatic ability, intrinsic chemoresistance, and poor prognosis of tumors. RNA interfering （RNAi）, a new genetic technique, can efficiently and specifically suppress gene expression. This study was to investigate the effect of small interfering RNA （siRNA） mediated gene silencing of C-erbB-2 on proliferation of human lung adenocarcinoma cell line calu-3. METHODS： C-erbB-2 siRNA was transfected into calu-3 cells; cell morphology was observed under light microscope. The mRNA and protein levels of C-erbB-2 were detected by reverse transcription-polymerase chain reaction （RT-PCR） and flow cytometry （FCM）. The proliferation of calu-3 cells was assessed by MTT assay. Cell cycle and apoptosis were analyzed by FCM. RESULTS： C-erbB-2 siRNA down-regulated the mRNA and protein levels of C-erbB-2 in calu-3 cells; 48 h after transfection of C-erbB-2 siRNA, the protein level of C-erbB-2 was markedly decreased. The positive rate of C-erbB-2 was significantly lower in C-erbB-2 siRNA group than in untransfected group, empty vector group, and nonspecific siRNA group [ （25.04±1.56）% vs. （98.24±2.23）%, （95.67±1.98）%, and （94.79±0.87）%, P 〈 0.01 ]. C-erbB-2 siRNA inhibited proliferation of calu-3 cells： G0/G1 phase proportion of C-erbB-2 siRNA group was significantly higher than that of untransfected group [（56.6±3.6）% vs. （45.5±3.2）%, P 〈 0.01]. C-erbB-2 siRNA also enhanced cell apoptosis. CONCLUSION： Specific siRNA targeting C-erbB-2 can effectively inhibit C-erbB-2 expression and proliferation of calu-3 cells.

关 键 词：肺肿瘤 基因治疗 RNA干扰 C-ERBB-2 SIRNA calu-3细胞 

分 类 号：R734.2[医药卫生—肿瘤]