活化蛋白C通过靶向VLA⁃3⁃中性粒细胞亚群来减轻结核杆菌诱导的人气道上皮细胞炎症反应线粒体能量代谢障碍  

Activated protein C reduces mitochondrial energy metabolism disorder induced by Mycobacterium tuber⁃culosis in human airway epithelial cells by targeting VLA⁃3⁃neutrophil subpopulation

作　　者：陈焰 王岗玲 吴海明 CHEN Yan;WANG Gangling;WU Haiming(Department of Infection,Armed police Fujian Provincial General Team Hospital,Fuzhou 350003,China;不详)

机构地区：[1]武警福建省总队医院感染科,福州350003 [2]厦门市儿童医院检验科,福建厦门361000

出　　处：《实用医学杂志》2020年第23期3217-3221,共5页The Journal of Practical Medicine

摘　　要：目的探究活化蛋白C通过靶向VLA⁃3⁃中性粒细胞亚群来减轻结核杆菌诱导的人气道上皮细胞炎症反应线粒体能量代谢障碍的机制。方法人支气管上皮BEAS⁃2B细胞分为3组:BEAS⁃2B组(正常培养的细胞系),结核杆菌诱导组(用50个细菌/细胞的结合杆菌感染BEAS⁃2B细胞培养物),BEAS⁃2B转染组(携带活化蛋白C的慢病毒载体与BEAS⁃2B进行细胞转染)。通过可溶性整联蛋白结合测定活化蛋白C与VLA⁃3的结合度;逆转录定量PCR(RT⁃qPCR)实时分析IL⁃6、IL⁃8、MCP⁃1的mRNA表达;荧光探针JC⁃1评估线粒体功能;通过Biosciences测量线粒体膜电位;ELISA和TUNEL鉴定细胞凋亡及活力情况。结果VLA⁃3较VLA⁃3和αVβ3结合程度升高(P<0.05)。结核杆菌诱导组较BEAS⁃2B组IL⁃6、IL⁃8、MCP⁃1 mRNA表达升高(P<0.05),BEAS⁃2B转染组较结核杆菌诱导组IL⁃6、IL⁃8、MCP⁃1mRNA表达降低(P<0.05)。BEAS⁃2B组较结核杆菌诱导组线粒体复合物Ⅰ、线粒体复合物Ⅳ活性升高(P<0.05),结核杆菌诱导组较BEAS⁃2B转染组线粒体复合物Ⅰ、线粒体复合物Ⅳ活性降低(P<0.05)。BEAS⁃2B组较结核杆菌诱导组膜电位升高(P<0.05),结核杆菌诱导组较BEAS⁃2B转染组膜电位降低(P<0.05)。BEAS⁃2B组较结核杆菌诱导组细胞凋亡率降低(P<0.05),结核杆菌诱导组较BEAS⁃2B转染组细胞凋亡率升高(P<0.05)。BEAS⁃2B组较结核杆菌诱导组细胞活力升高(P<0.05),结核杆菌诱导组较BEAS⁃2B转染组细胞细胞活力降低(P<0.05)。结论活化蛋白C对整联蛋白VLA3有更强的亲和力,可靶向作用VLA3中性粒细胞亚群降低人气道上皮细胞的炎症反应,提高线粒体复合物Ⅰ、复合物Ⅳ活性,有效抑制结合杆菌诱导的细胞线粒体膜电位下降。Objective To investigate the mechanism of activated protein C to alleviate the mitochondrial energy metabolism disorder induced by Mycobacterium tuberculosis in human airway epithelial cells by targeting VLA⁃3⁃neutrophil subpopulation.Methods Human bronchial epithelial BEAS⁃2B cells were divided into three groups:BEAS⁃2B group(normal culture cell line),Mycobacterium tuberculosis induction group(BEAS⁃2B cell culture was infected with 50 bacteria/cell conjugated bacteria),and BEAS⁃2B transfection group(BEAS⁃2B cells were transfected with lentiviral vector carrying activated protein C).The binding degree of activated protein C to vla⁃3 was determined by soluble integrin binding.RT qPCR was used to analyze the mRNA expression of IL⁃6,IL⁃8 and MCP⁃1.The function of mitochondria was evaluated by fluorescence probe JC⁃1 and mitochondrial membrane potential was measured by biosciences.Cell apoptosis and viability were detected by ELISA and TUNEL.Results VLA⁃3 was more highly expressed than VLA⁃3 andαVβ3(P<0.05).The expression of IL⁃6,IL⁃8 and MCP⁃1 mRNA in the induction group of Mycobacterium tuberculosis was higher than that in BEAS⁃2B group(P<0.05).The BEAS⁃2B transfection group was more sensitive than IL⁃6,IL⁃8 and MCP in the induction group of Mycobacterium tuberculosis.1 mRNA expression was decreased(P<0.05).Compared with the Mycobacterium tuberculosis induc tion group,the mitochondrial complexⅠand mitochondrial complexⅣactivity were significantly increased in the BEAS⁃2B group(P<0.05),and the mitochondrial complex I and mitochondrial complexⅣactivity were decreased in the Mycobacterium tuberculosis induction group compared with the BEAS⁃2B transfection group(P<0.05).The membrane potential of the BEAS⁃2B group was higher than that of the Mycobacterium tuberculosis induced group(P<0.05),and the membrane potential of the tuberculosis induction group was lower than that of the BEAS⁃2B transfection group(P<0.05).The apoptosis rate of BEAS⁃2B group was lower tha

关 键 词：活化蛋白C 整联蛋白VLA⁃3 细胞凋亡 线粒体功能障碍 

分 类 号：R512.91[医药卫生—内科学]