过表达miR-320e抑制呼吸道合胞病毒感染支气管上皮细胞炎症反应的机制研究  

Overexpression of miR-320e inhibits inflammatory response of bronchial epithelial cells infected by respiratory syncytial virus

作　　者：许振浪 邝向东 谢景臣 秦玉春 XU Zhenlang;KUANG Xiangdong;XIE Jingchen;QIN Yuchun(Department of Respiratory and Critical Care Medicine,Hainan Western Central Hospital,Danzhou 571700,China)

机构地区：[1]海南西部中心医院呼吸与危重症医学科,儋州571700

出　　处：《中国免疫学杂志》2024年第12期2506-2512,共7页Chinese Journal of Immunology

基　　金：2020年度海南省卫生健康行业科研项目(20A200304)。

摘　　要：目的:探讨过表达miR-320e抑制呼吸道合胞病毒(RSV)感染支气管上皮细胞炎症反应的机制。方法:体外培养人支气管上皮细胞16HBE,用RSV感染,将细胞分为Con组、RSV组、RSV^(+)miR-NC组、RSV^(+)miR-320e组、RSV^(+)miR-320e^(+)vector组、RSV^(+)miR-320e^(+)TLR4组。RT-qPCR检测miR-320e、TLR4 mRNA表达水平;MTT检测细胞增殖变化;流式细胞术检测细胞凋亡;Western blot检测Bcl-2、Bax、TLR4、IκBα、p-IκBα、NF-κB、p-NF-κB蛋白表达;ELISA检测TNF-α、IL-6、IL-1β和IFN-α、IFN-β表达;双荧光素酶实验验证miR-320e和TLR4的靶向关系。结果:与Con组相比,RSV组miR-320e表达水平、存活率、Bcl-2和IκBα蛋白表达明显降低,凋亡率、Bax蛋白表达、TNF-α、IL-6、IL-1β和IFN-α、IFN-β表达、TLR4 mRNA和蛋白表达,以及p-IκBα蛋白表达和p-NF-κB/NF-κB明显增加。与RSV^(+)miR-NC组相比,RSV^(+)miR-320e组miR-320e表达水平、存活率、IFN-α、IFN-β表达、Bcl-2和IκBα蛋白表达明显增加,凋亡率、Bax蛋白表达、TNF-α、IL-6、IL-1β表达、TLR4 mRNA和蛋白表达,以及p-IκBα蛋白表达和p-NF-κB/NF-κB明显降低。miR-320e靶向负调控TLR4表达,上调TLR4可部分回复过表达miR-320e对RSV感染支气管上皮细胞16HBE凋亡和炎症反应的影响。结论:miR-320e通过靶向负调控TLR4表达抑制RSV感染支气管上皮细胞16HBE凋亡和炎症反应。Objective:To investigate the mechanism of overexpression of miR-320e in inhibiting inflammatory response of respiratory syncytial virus(RSV)infected bronchial epithelial cells.Methods:Human bronchial epithelial cells 16HBE were cultured in vitro and infected with RSV,and cells were divided into Con group,RSV group,RSV^(+)miR-NC group,RSV^(+)miR-320e group,RSV^(+)miR-320e^(+)vector group,RSV^(+)miR-320e^(+)TLR4 group.RT-qPCR was used to detect expression levels of miR-320e and TLR4 mRNA;MTT to detect cell proliferation changes;flow cytometry to detect cell apoptosis;Western blot was used to detect Bcl-2,Bax,TLR4,IκBα,p-IκBα,NF-κB and p-NF-κB protein expressions;ELISA to detect TNF-α,IL-6,IL-1βand IFN-α,IFN-βexpressions;dual luciferase experiment to verify the tageting relationship between miR-320e and TLR4.Results:Compared with Con group,miR-320e expression level,survival rate,Bcl-2 and IκBαprotein expressions were significantly reduced,apoptosis rate,Bax protein expression,TNF-α,IL-6,IL-1βand IFN-α,IFN-βexpressions,TLR4 mRNA and protein expression,and p-IκBαprotein expression and p-NF-κB/NF-κB were increased significantly in RSV group.Compared with RSV^(+)miR-NC group,miR-320e expression level,survival rate,IFN-α,IFN-βexpressions,Bcl-2 and IκBαprotein expressions were significantly increased,apoptosis rate,Bax protein expression,TNF-α,IL-6,IL-1βexpressions,TLR4 mRNA and protein expression,and p-IκBαprotein expression and p-NF-κB/NF-κB in RSV^(+)miR-320e group were significantly reduced.miR-320e targets and negatively regulates the expression of TLR4.Upregulation of TLR4 can partially restore the effect of overexpression of miR-320e on apoptosis and inflammatory response of RSV-infected bronchial epithelial cells 16HBE.Conclusion:miR-320e inhibits 16HBE apoptosis and inflammation in RSV-infected bronchial epithelial cells by targeting and negatively regulating TLR4 expression.

关 键 词：miR-320e 呼吸道合胞病毒 支气管上皮细胞 炎症反应 

分 类 号：R562.25[医药卫生—呼吸系统]