人Id3基因在肺腺癌A549细胞中的表达及其对细胞生长的抑制  

Human Id3 expression in lung adenocarcinoma cell line A549 and its inhibiory effect on cell growth

作　　者：朱传东[1,2] 李晓军[1] 汪萍[1] 贾丽[1] 仲爱芳[1] 陈龙邦[2] 

机构地区：[1]南京大学医学院临床学院,南京军区南京总医院临床中心实验科,南京210002 [2]南京大学医学院临床学院,南京军区南京总医院肿瘤内科,南京210002

出　　处：《中国肿瘤生物治疗杂志》2008年第6期527-531,共5页Chinese Journal of Cancer Biotherapy

基　　金：江苏省六大人才高峰基金资助项目(No.050203D)~~

摘　　要：目的:研究外源性分化抑制因子3(Inhibitor of differentiation 3,Id3)基因表达对人肺腺癌细胞系A549细胞生长的抑制作用。方法:构建Id3和EGFP的融合基因表达载体pEGFP/Id3,采用脂质体转染技术将pEGFP/Id3导入A549细胞。FCM分析转染细胞EGFP表达效率,荧光显微镜观察EGFP表达情况;半定量RT-PCR、免疫细胞化学分析转染后A549细胞Id3 mRNA和蛋白的表达。MTT法检测转染后A549细胞生长抑制情况,FCM分析A549细胞周期进程,Annexin V/7-AAD和Hoechst33258染色检测转染后细胞的凋亡情况。结果:成功构建人Id3与EGFP融合基因表达载体pEGFP/Id3;荧光显微镜和FCM观察到EGFP的有效表达,转染48～72 h时EGFP表达率最高;Id3 mRNA及蛋白在转染后的A549细胞中能有效表达。转染后48 h和72 h,pEGFP/Id3转染组A549细胞生长受到明显抑制(P<0.01);细胞周期分析表明,pEGFP/Id3转染组G_0/G_1期细胞显著高于对照组(P<0.05);Annexin V/7-AAD双染结果显示,pEGFP/Id3转染组细胞的早期凋亡率[(10.67±2.60)%]显著高于pEGFP组[(3.39±2.21)%]和未转染组[(2.35±0.95)%](P<0.05);Hoechst33258染色结果也证实pEGFP/Id3组A549细胞出现明显凋亡形态特征。结论:外源性Id3基因在肺腺癌A549细胞中的表达能抑制细胞增殖,并诱导细胞凋亡。Objective：To investigate the inhibitory effect of inhibitor of differentiation 3 （Id3） on growth of human lung adenocarcinoma cell line A549. Methods： Recombinant eukaryotic expression vector pEGFP/Id3 was constructed and transfected into A549 cells by liposome-mediated method. Expression of pEGFP/Id3 in A549 cells was analyzed by flow cytometry （FCM） , fluorescence microscopy, semi-quantitative RT-PCR and immunocytochemistry. The growth inhibitory rate of A549 cells was examined by MTT assay ; cell cycle change was evaluated by PI （ propidium iodide） staining method. Cell apoptotic rate and nuclear morphology were detected by Annexin V/7-AAD and Hoechst33258 staining. Results ： The recombinant eukaryotic expression vector pEGFP/Id3 was successfully constructed. The expression of EGFP reached the peak 48-72 h after transfection ; the expresion of pEGFP-transfected group was higher than that of the pEGFP/ Id3 group. RT-PCR and immunoeytochemistry staining showed that Id3 mRNA and protein were effectively expressed in pEGFP/Id3-transfected A549 cells. The growth of cells in pEGFP/Id3 transfected cells was significantly inhibited 48-72 h after transfection（P 〈0.01 ）. More cells were blocked in G0/G1 phase in pEGFP/Id3-transfeeted group compared with pEGFP group（P 〈0.05）. Annexin V/7-AAD showed that the apoptotic rate of pEGFP/Id3 group （ E 10.67 ＋2. 601% ）Conclusion：Exogenous/d3 gene expression in A549 cells can induce cell growth inhibition and apoptosis of A549 cells.

关 键 词：分化抑制因子3基因(Id3) 肺癌细胞 增殖抑制 凋亡 

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