miR-17-5p靶向信号调节基因SIRPα调控巨噬细胞抗结核分枝杆菌的炎症反应  

MiR-17-5p targeting signal regulatory gene SIRPα regulates inflammatory response of macrophages to Mycobacterium tuberculosis

作　　者：张帆 何萌 李元[2] 王淑娥 洪丹彤 王雅蓉 刘宏鹏[2] 姜中佳 安维军[3] 郭乐 ZHANG Fan;HE Meng;LI Yuan;WANG Shu-e;HONG Dan-tong;WANG Ya-rong;LIU Hong-peng;JIANG Zhong-jia;AN Wei-jun;GUO Le(Provincial Key Laboratory of Clinical Pathogenic Microbiology,General Hospital of Ningxia Medical University,Yinchuan 750021,China;School of Clinical Medicine,Ningxia Medical University,Yinchuan 750021,China;Orthopedic Trauma,General Hospital of Ningxia Medical University,Yinchuan 750021,China)

机构地区：[1]宁夏医科大学总医院,临床病原微生物省级重点实验室,宁夏银川750021 [2]宁夏医科大学临床医学院,宁夏银川750021 [3]宁夏医科大学总医院创伤骨科,宁夏银川750021

出　　处：《生物技术》2020年第1期30-37,共8页Biotechnology

基　　金：国家自然科学基金项目(“miR-20a/miR-17相关ceRNAs网络调控巨噬细胞抗结核分枝杆菌炎症反应和细胞自噬的作用机制”,81760359);2018年自治区青年拔尖人才培养工程。

摘　　要：[目的]验证miR-17-5p对TLRs负向调控因子SIRPα的靶向性,探讨其对抗结核分枝杆菌(MTB)炎症反应的调控作用。[方法]利用生物信息学预测miR-17-5p对SIRPα的靶向性,构建SIRPα野生型和突变型报告载体,利用双荧光素酶报告法、Western Blot、激光共聚焦等技术验证miR-17-5p对SIRPα的靶向性;通过H37Ra感染THP-1巨噬细胞,用miR-17-5p mimics及其inhibitor处理细胞。利用Q-PCR检测H37Ra感染后miR-17-5p的表达;通过免疫荧光、Western Blot和ELISA等技术检测SIRPα和细胞因子TNF-α的表达情况。[结果]H37Ra感染可下调miR-17-5p表达,且随感染复数的增加,下调表达愈加显著;荧光素酶报告法、Western Blot等结果证实miR-17-5p可靶向结合SIRPα3’-UTR,下调SIRPα的表达,进而上调细胞因子TNF-α的表达。[结论]MTB可下调miR-17-5p表达,而miR-17-5p可靶向抑制SIRPα的表达,从而调控巨噬细胞抗MTB的炎症反应。[Objective] To investigate the role of miR-17-5 p in targeting the negative regulatory factor SIRPα of TLRs,and its regulation of anti-tuberculosis inflammatory response.[Method]Bioinformatics method was used to predict the targeting relationship between miR-17-5 p and SIRPα,SIRPα wild type and mutant vector were constructed. The targeting relationship between miR-17-5 p and SIRPα was verified by double luciferase reporting system,Western Blot and laser Confocal. The THP-1 macrophage was infected with H37 Ra,and treated with miR-17-5 p mimics and miR-17-5 p inhibitor. The effect of H37 Ra infection on the expression of miR-17-5 p was detected by Q-PCR. Further,the expression of SIRPα and cytokine TNF-α was detected by immunofluorescence,Western Blot and ELISA.[Result]H37 Ra infection could lead to the downregulation of miR-17-5 p,which decreased with the increase of the number of infections. The results of double luciferase reporting system and Western Blot confirmed that miR-17-5 p could bind to SIRPα3’-UTR,down-regulated the expression of SIRPα,and then up-regulated the expression of the cytokine TNF-α. [Conclusion]MTB could down-regulate the expression of miR-17-5 p,while miR-17-5 p targeted binding to SIRPα and inhibited its expression,thereby regulated the inflammatory response of macrophages against MTB.

关 键 词：结核分支杆菌 炎症反应 miR-17-5p SIRPα 

分 类 号：R34[医药卫生—基础医学] R37