ESAT6和CFP10融合蛋白抑制巨噬细胞自噬体形成的实验研究  

Inhibition of ESAT6 and CFP10 fusion proteins on the autophagosomes formation of macrophages

作　　者：师长宏[1] 毛峰峰[1] 赵勇[1] 张海[1] 张彩琴[1] 白冰[1] 赵善民[1] 

机构地区：[1]第四军医大学实验动物中心,西安710032

出　　处：《中国人兽共患病学报》2011年第5期414-417,共4页Chinese Journal of Zoonoses

基　　金：国家"863"专项课题(2007AA02Z473);国家自然科学基金(30972767);陕西省自然科学基金(SJ08C203;2004C125)联合资助

摘　　要：目的观察ESAT6和CFP10融合蛋白对感染MTB的巨噬细胞自噬体形成的抑制作用。方法雷帕霉素诱导小鼠巨噬细胞自噬体形成后,用MTB毒株H37Rv感染巨噬细胞,再用25μg/mL的ESAT6-CFP10融合蛋白作用于巨噬细胞,电镜观察自噬体相成的变化,计数MTB的菌落数。提取巨噬细胞总RNA和蛋白,以RT-PCR和免疫印迹方法检测自噬相关基因(atg)表达水平的变化。结果 ESAT6-CFP10融合蛋白后可抑制巨噬细胞中自噬体的形成,显著提高CFU指数(P<0.05),并导致atg分子表达水平下降,其中atg8表达量下降最为明显(P<0.05)。结论 ESAT6和CFP10融合蛋白可通过调控atg表达水平影响巨噬细胞自噬功能。The objective of this study was to study the inhibition of ESAT6 and CFP10 fusion protein on the autophagosomes formation of macrophages.Following the infection with M.tuberculosis H37Rv strains,autophagosomes of macrophages were induced by rapamycin and the effects of ESAT6-CFP10 fusion protein on the autophagosomes formation were observed by transmission electron microscope.Macrophages cellular mRNAs and proteins were extracted and the expression of autophagy-related genes（atg） was detected by real-time quantitative PCR and immunoblot method respectively.Results demonstrated that macrophages could form autophagosomes by rapamycin inductio,which having scavenging effect on the M.tuberculosis infected cells.ESAT6-CFP10 fusion proteins could inhibit autophagosome formation in macrophages,significantly increase M.tuberculosis colony forming units（CFU）（P〈0.05） and decrease the expression of atgs,especially changing atg8 expression level obviously（P〈0.05）.The result suggested that ESAT6-CFP10 fusion protein could inhibit the formation of autophagosomes and resist phagocytize by regulating the expression level of atg protein.

关 键 词：结核分枝杆菌 自噬 ESAT6 CFP10 自噬相关基因 

分 类 号：R379[医药卫生—病原生物学]