cGAS介导牛分枝杆菌诱导小鼠髓源树突状细胞成熟

doi:10.11843/j.issn.0366-6964.2017.05.016

畜牧兽医学报 ›› 2017, Vol. 48 ›› Issue (5): 914-921.doi: 10.11843/j.issn.0366-6964.2017.05.016

cGAS介导牛分枝杆菌诱导小鼠髓源树突状细胞成熟

李强¹, 刘春法², 何小丽¹, 李凡飞¹, 魏凡华¹, 赵德明², 周向梅^2*, 许立华^1*

1. 宁夏大学农学院, 银川 750021;
2. 中国农业大学动物医学院国家动物传染性海绵状脑病实验室, 北京 100193

收稿日期:2016-09-21 出版日期:2017-05-23 发布日期:2017-05-23
通讯作者: 许立华,教授,E-mail:littlezhe99@163.com;周向梅,教授,E-mail:zhouxm@cau.edu.cn
作者简介:李强(1991-),男,河南郑州人,硕士生,主要从事预防兽医学研究,E-mail:qiangweitianxia@126.com
基金资助:
国家自然科学基金（31560687；31572487）；宁夏自然科学基金（NZ12104；NZ15031）

Cyclic GMP-AMP Synthase-mediated Maturation of Murine Bone Marrow Derived Dendritic Cells after Infection with Mycobacterium bovis

LI Qiang¹, LIU Chun-fa², HE Xiao-li¹, LI Fan-fei¹, WEI Fan-hua¹, ZHAO De-ming², ZHOU Xiang-mei^2*, XU Li-hua^1*

1. College of Agriculture, Ningxia University, Yinchuan 750021, China;
2. National Animal Transmissible Spongiform Encephalopathy Laboratory, College of Veterinary Medicine, China Agricultural University, Beijing 100193, China

Received:2016-09-21 Online:2017-05-23 Published:2017-05-23

摘要/Abstract

摘要：

旨在深入研究小鼠髓源树突状细胞（BMDC）感染牛分枝杆菌（M.bovis）后环磷酸鸟苷-磷酸腺苷合酶（cGAS）的调控作用。通过体外诱导C57BL/6骨髓细胞分化为树突状细胞，建立牛分枝杆菌感染模型，运用siRNA技术沉默cGAS基因的表达，分别设置对照组、感染组、沉默组。通过流式细胞仪检测BMDC表型变化，Western blot和免疫荧光技术检测cGAS信号通路中STING、TBK1、p-TBK1及IRF3的表达情况， ELISA检测细胞上清液中细胞因子分泌水平。结果表明，牛分枝杆菌可刺激BMDC引起细胞表面标志物CD86、CD80、CD40、MHC-Ⅱ表达升高，相关细胞因子IFN-β、TNF-α、IL-12p70、IL-6分泌量提高，BMDC的抗原提呈能力增强。同时cGAS信号通路被激活，下游STING、p-TBK1、IRF3蛋白活化，而沉默cGAS基因后相关指标均显著下调。这表明牛分枝杆菌感染BMDC可激活cGAS信号通路，且cGAS有助于BMDC的成熟和活化。

Abstract:

The study was performed to investigate the regulation of cyclic GMP-AMP synthase (cGAS) on Mycobacterium bovis infected murine bone marrow derived dendritic cell (BMDC). The murine bone marrow cells were cultured and induced into DCs in vitro. Then BMDCs were infected by Mycobacterium bovis to establish infection models. Three groups were designed for the in vitro study. They were control group (BMDC control), infection group (M. bovis infected BMDC) and silence group. In silence group, BMDC cGAS gene was knocked down via siRNA and then cells were infected similar to infection group. Flow cytometry was used to analyze the expression of surface markers. In cGAS pathway, STING, TBK1, p-TBK1 and IRF3 were determined by Western blot and Immunofluorescence, and the cytokine production in cell supernatant was assessed by ELISA. The data showed that M. bovis induced higher expression levels of the surface markers CD86, CD80, CD40, MHC-Ⅱ and cytokine production IFN-β, TNF-α, IL-12p70, IL-6 in BMDC. Meanwhile, the cGAS pathway was activated and induced higher expression of related protein, however corresponding data reduced significantly in silence group. Our research indicated that cGAS pathway can be activated by Mycobacterium bovis when BMDC was infected, and cGAS contribute to maturation and activation of BMDC.

中图分类号:

S852.618

李强, 刘春法, 何小丽, 李凡飞, 魏凡华, 赵德明, 周向梅, 许立华. cGAS介导牛分枝杆菌诱导小鼠髓源树突状细胞成熟[J]. 畜牧兽医学报, 2017, 48(5): 914-921.

LI Qiang, LIU Chun-fa, HE Xiao-li, LI Fan-fei, WEI Fan-hua, ZHAO De-ming, ZHOU Xiang-mei, XU Li-hua. Cyclic GMP-AMP Synthase-mediated Maturation of Murine Bone Marrow Derived Dendritic Cells after Infection with Mycobacterium bovis[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2017, 48(5): 914-921.

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cGAS介导牛分枝杆菌诱导小鼠髓源树突状细胞成熟

Cyclic GMP-AMP Synthase-mediated Maturation of Murine Bone Marrow Derived Dendritic Cells after Infection with Mycobacterium bovis

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