肺癌A549细胞中RNAi抑制MeCP2表达的实验研究  

Experimental study of RNA inference targeting MeCP2 in lung carcinoma cell line A549

作　　者：王汉姣[1] 吴建兵[2] 邱小华[2] 郭琳琳[2] 

机构地区：[1]福建医科大学附属三明市第一医院肿瘤内科,福建三明365000 [2]南昌大学第二附属医院,江西南昌330006

出　　处：《现代肿瘤医学》2010年第4期631-634,共4页Journal of Modern Oncology

基　　金：江西省科学技术厅社会发展攻关项目(编号:20041095)

摘　　要：目的:探讨RNA干扰下调MeCP2基因后对人肺癌A549细胞RASSF1A和C/EBPα基因表达及细胞增殖能力的影响。方法:根据MeCP2的碱基序列设计并合成短发夹RNA(shRNA),构建质粒表达载体,用阳离子脂质体法体外转染A549细胞,筛选出稳定转染的细胞株,RT-PCR法检测MeCP2抑制效果及RASSF1A和C/EBPα表达,MTT法检测转染后细胞增殖能力。结果:成功构建了靶向MeCP2的真核表达载体pSilencer-MeCP2/Ⅰ和pSilencer-MeCP2/Ⅱ,与空白组和阴性组相比,pSilencer-MeCP2/Ⅰ和pSilencer-MeCP2/Ⅱ组均可有效抑制MeCP2基因mRNA的表达,抑制率分别为47.6%和70.3%。且在上述两组中C/EBPα表达分别上调约115%和135%、细胞增殖能力明显减弱,与空白组和阴性组比较差异具有统计学意义(P<0.05),而RASSF1A的表达在各组细胞中未见明显变化(P>0.05)。结论:肺癌A549细胞中MeCP2基因的表达可被RNA干扰载体有效抑制,进而导致细胞中部分与增殖和凋亡相关基因的表达改变,细胞增殖能力减弱。Objective：To investgate the expression of RASSF1A and C/EBPα as well as cell proliferation in A549 human lung carcinoma cell by RNA interference inhiibiting expression of MeCP2 gene.Methods：Two siRNAs targeting MeCP2 were designed and synthesized according to its mRNA,and their corresponding expression vectors were constructed and transfected into A549 cell with Lipofectamine^TM 2000.After selection,the stable transfected cells were obtained.The inhibition of MeCP2 expression,together with the expression of RASSF1A and C/EBPα were analyzed by semi-quantitative RT-PCR,MTT method was applied to measure cell growth.Results： Two MeCP2-targeted siRNAs recombinant vectors named as pSilencer-MeCP2Ⅰand pSilencer-MeCP2/Ⅱ were constructed successfully.Compared with the controls,the groups of pSilencer-MeCP2/Ⅰand pSilencer-MeCP2/Ⅱ effectively inhibited the expression of MeCP2 with inhibitory rate of 47.6%and 70.3%,among the two groups,the expression of C/EBPα were up-regulated by 120% and 135% respectively with significiently decreased cell proliferation（P〈0.05）.The expression of RASSF1A had no change in all groups（P〉0.05）.Conclusion： The expression of MeCP2 in A549 cells could be supressed by vector-based RNA interference efficiently.The knockdown of MeCP2 in turn caused changes in gene expression related with cell proliferation and apoptosis,resulted in decreased cell proliferation.

关 键 词：RNA干扰 肺癌 MECP2 细胞增殖 基因表达 

分 类 号：R730.54[医药卫生—肿瘤] R734.2[医药卫生—临床医学]