小鼠IL-1β原核表达及其与在布鲁氏杆菌侵染巨噬细胞中差异表达的初步研究  

Prokaryotic expression of mouse IL-1βand preliminary study on its differential expression in Brucella infection

作　　者：杨馨 邹德颖[1] 张凯[1] 常江[1] 常恒祯 战俊澎 郭珣 刘益辛 程梦妍 韩成 胡盼[1] 卢士英[1] 李岩松[1] 柳增善[1] 任洪林[1] YANG Xin;ZOU De-ying;ZHANG Kai;CHANG Jiang;CHANG Heng-zhen;ZHAN Jun-peng;GUO Xun;LIU Yi-xin;CHENG Meng-yan;HAN Cheng;HU Pan;LU Shi-ying;LI Yan-song;LIU Zeng-shan;REN Hong-lin(Key Laboratory of Zoonosis Research,Ministry of Education/Institute of Zoonosis/College of Veterinary Medicine,Jilin University,Changchun 130062,China)

机构地区：[1]吉林大学人兽共患病研究教育部重点实验室/人兽共患病研究所/动物医学学院,长春130062

出　　处：《湖北农业科学》2022年第6期112-116,121,共6页Hubei Agricultural Sciences

基　　金：吉林省科技发展计划资助项目(20200402059NC);中央高校基本科研业务费专项资金资助项目(45121031B007)。

摘　　要：为研究IL-1β在布鲁氏杆菌(Brucella)侵染小鼠巨噬细胞过程中的表达情况,首先构建重组小鼠IL-1β原核稳定表达系统,免疫新西兰兔获得多克隆抗体,通过ELISA检测血清效价,并通过Western blot分析其活性,再以感染复数(multiplicity of infection,MOI)为1∶100(细菌∶细胞)建立猪种布鲁氏杆菌S2株侵染小鼠巨噬细胞RAW264.7的模型,利用RT-PCR检测细菌侵染过程中IL-1β和MyD88 mRNA的变化水平;Western blot分析IL-1β在蛋白水平的变化。结果表明,重组蛋白IL-1β分子质量31 ku,多克隆抗体效价为1∶256000,并且具有良好的特异性;与对照组相比,感染后IL-1βmRNA和IL-1β蛋白表达上调,胞内细菌数减少。猪种布鲁氏杆菌S2株侵染巨噬细胞IL-1β的表达量随时间变化并影响胞内活菌数。To explore the expression of IL-1βduring the infection of host cells by Brucella,the prokaryotic expression of IL-1βin mouse was constructed,polyclonal antibodies was obtained by immunization,and its activity was detected by ELISA and Western blot.The 1∶100(bacterial∶cell)multiplicity of infection was then used to model the infection of RAW264.7 macrophages by Brucella suis S2.Expression of IL-1βmRNA and MyD88 mRNA was detected by RT-PCR;IL-1βprotein was analyzed by Western blot.The molecular weight of the recombinant protein was about 31 ku.And the titer of the polyclonal antibody prepared by ELISA was 1∶25600,with good immunoreactivity and specificity.The expression of IL-1βmRNA and IL-1βprotein increased after infection.And the number of intracellular bacteria decreased.The expression of IL-1βin macrophages infected with Brucella suis S2 varied over time and affected the intracellular bacteria.

关 键 词：布鲁氏杆菌(Brucella) 原核表达 多克隆抗体 IL-1Β MYD88 

分 类 号：R392[医药卫生—免疫学]