重组腺病毒Ad5F35-IL-12感染不同单核-巨噬细胞的研究  

Infection of recombinant adenovirus Ad5F35-IL-12 in different kinds of human mononuclear macrophages

作　　者：王秦[1] 成凤[1] 张维理[1] 匡文斌[1] 李朴[1] 董晋豫[1] 梁勤东[1] 涂植光[1] 

机构地区：[1]重庆医科大学检验医学院,临床检验诊断学教育部重点试验室,重庆400016

出　　处：《细胞与分子免疫学杂志》2013年第2期113-117,共5页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(81172016)

摘　　要：目的确定人IL-12基因重组腺病毒Ad5F35-IL-12在不同人单核-巨噬细胞中的表达。方法将人IL-12基因重组腺病毒Ad5F35-IL-12分别感染人外周血单个核细胞、胸水巨噬细胞,以及THP-1和U937单核细胞株及其经佛波酯(PMA)诱导后生成的巨噬细胞;感染48 h后荧光显微镜观察对相应细胞的感染效率,RT-PCR检测IL-12双亚基p35和p40的mRNA表达情况,ELISA检测细胞培养上清中IL-12p70的分泌水平。结果人IL-12基因重组腺病毒Ad5F35-IL-12可成功感染人外周血单个核细胞、胸水巨噬细胞以及THP-1和U937单核细胞株及其PMA诱导后生成的巨噬细胞,并分泌IL-12 p70蛋白,蛋白表达量从高至低依次是胸水巨噬细胞、PMA诱导后U937细胞、PMA诱导后THP-1细胞、U937细胞、外周血单个核细胞、THP-1细胞。结论人IL-12基因重组腺病毒Ad5F35-IL-12可以感染不同的单核-巨噬细胞,并成功表达分泌IL-12 p70蛋白。Objective To investigate the efficiencies of transfection and expression of human recombinant adenovirus Ad5F35-IL-12 in the different kinds of human mononuclear macrophages.Methods The human recombinant adenovirus Ad5F35-IL-12 was used to infect human peripheral blood monocytes,pleural fluid macrophages as well as THP-1,U937 monocyte cell lines and their phorbol myristate acetate（PMA）-induced macrophages.48 h later,green fluorescence was observed under the fluorescence microscope to detect the transfection efficiency.The expressions of IL-12 double-subunits（p35,p40） mRNA were tested by RT-PCR and the level of IL-12p70 protein in the cell culture supernatant was detected with ELISA.Results The human recombinant adenovirus Ad5F35-IL-12 successfully infected the human peripheral blood monocytes,pleural fluid macrophages,THP-1 monocytes,U937 monocytes,and THP-1 and U937 macrophages induced with PMA.All above infected mononuclear macrophages effectively secreted IL-12p70 protein,and they were listed from high to low of IL-12p70 protein level as pleural fluid macrophages,U937 and THP-1 macrophages induced with PMA,U937 monocytes,human peripheral blood monocytes and THP-1 monocytes.Conclusion The human recombinant adenovirus Ad5F35-IL-12 could infect different kinds of mononuclear macrophages,and IL-12 p70 protein could be successfully expressed in cell supernatants.

关 键 词：IL-12 腺病毒Ad5F35 单核巨噬细胞 基因治疗 

分 类 号：R392.33[医药卫生—免疫学] R392.12[医药卫生—基础医学]