滋养细胞胞内双链DNA感受器通过MAPK/IκB信号介导炎性细胞因子分泌  

Trophoblastic Intracellular Double-stranded DNA Sensors Mediate Inflammatory Cytokines Secretion via MAPK/IκB Signaling Pathway

作　　者：蒋锦杏[1] 张慧 任莉莉[1] 高宇 李宁 JIANG Jin-xing;ZHANG Hui;REN Li-li;GAO Yu;LI Ning(Cytotherapy Laboratory,The Second Medical College of Jinan University,Shenzhen People's Hospital,Shenzhen,Guangdong,518020,China;Department of Obstetrics and Gynecology,The Second Medical College of Jinan University,Shenzhen People's Hospital,Shenzhen,Guangdong,518020,China;Department of Obstetrics and Gynecology,The Center for Medical Genetics and Molecular Diagnosis,Shenzhen,Guangdong,518106,China)

机构地区：[1]暨南大学第二临床医学院(深圳市人民医院)生物治疗室,广东深圳518020 [2]暨南大学第二临床医学院(深圳市人民医院)妇产科,广东深圳518020 [3]中国科学院大学深圳医院(光明)医学遗传与分子诊断中心,广东深圳518106

出　　处：《现代生物医学进展》2020年第8期1446-1450,共5页Progress in Modern Biomedicine

基　　金：广东省自然科学基金项目(2017A030310646,2019A1515011068);广东省科技计划项目(2016A020215207);深圳市科技计划项目(JCYJ20180228164515747);中国博士后基金项目(2018M643371)。

摘　　要：目的:探究滋养细胞能否感受胞内双链DNA刺激及其对炎性因子分泌的影响,揭示胎盘的免疫识别和免疫屏障功能,探讨妊娠期感染所致不良妊娠结局的发病机制。方法:利用人工合成的双链DNA模拟物poly(dA:dT)转染人滋养细胞系HTR-8/SVneo,Real-Time PCR方法检测滋养细胞胞内双链DNA识别受体的表达水平;Western Blot检测滋养细胞MAPK和IκB信号的活化情况;ELISA检测HTR-8/SVneo细胞培养上清中IL-6、IL-8、MCP-1、CXCL10的分泌水平。结果:Real-Time PCR结果表明,HTR-8/SVneo能够感受胞内双链DNA刺激并上调包括IFI16、AIM2、DHX36、DHX9、LRRFIP1、KU70、ZBP1/DAI和DDX41在内的多种双链DNA感受器的m RNA表达水平;Western Blot实验结果表明,滋养细胞识别双链DNA后能够促进MAPK和IκB信号通路活化,转染90分钟后ERK、JNK、p38和IκBα的磷酸化水平最高,其后随着时间逐渐减弱;加入IκB、p38和JNK特异性抑制剂能够抑制poly(dA:dT)介导的IL-8、IL-6和CXCL10分泌,但其分泌不受ERK抑制剂的影响;MCP-1的分泌能够被p38和JNK抑制剂阻断,但p38和ERK抑制剂不影响其分泌。结论:人滋养细胞存在功能性胞内双链DNA识别机制,活化的DNA识别信号能够激活MAPK和IκB信号通路,并通过IκB、p38或JNK信号介导IL-8、IL-6、MCP-1及CXCL10等细胞因子和趋化因子的分泌。Objective:The study was preformed to investigate the expression pattern of intracellular double-stranded DNA sensors in human trophoblast cells and its effect on the secretion of inflammatory factors,to reveal the immune recognition and immune barrier function of placenta,and to better understand the pathogenesis of adverse pregnancy outcomes caused by pregnancy infection.Methods:Synthetic double-stranded DNA(dsDNA)mimics,poly(dA:dT),were transfected into human trophoblast cell line HTR-8/SVneo,double-stranded DNA sensors’m RNA were detected by quantitative real-time PCR and the phosphorylation levels of MAPK and IκB signaling were detected by Western Blot.ELISA was used to detect IL-6,IL-8,MCP-1 and CXCL10 secretion.Results:Real-Time PCR results showed that IFI16,AIM2,DHX36,DHX9,LRRFIP1,KU70,ZBP1/DAI and DDX41 m RNA levels were dramatically upregulated in HTR-8/SVneo cells.The phosphorylation levels of ERK,JNK,p38 and IκB were elevated at 90 minutes after poly(dA:dT)transfection,and then gradually decreased.The induction of IL-8,IL-6 and CXCL10 secretion is dependent on IκB,p38 and JNK signaling pathways;MCP-1 secretion is dependent on p38 and JNK signaling pathways.However,these cytokines and chemokines were not affected by ERK inhibitor.Conclusions:Human placental trophoblast cells express functional intracellular double-stranded DNA sensors,which can recognize double-stranded DNA and activate MAPK/IκB signaling pathways that mediate the secretion of IL-8,IL-6,MCP-1 and CXCL10.

关 键 词：胞内双链DNA感受器 滋养细胞 MAPK/IκB信号通路 炎性细胞因子 

分 类 号：R-33[医药卫生] R392.11