幽门螺杆菌毒力因子CagA通过靶向SHARPIN促进炎症反应的机制研究  

Mechanisms of Helicobacter pylori virulence factor CagA in promoting inflammatory response by targeting SHARPIN

作　　者：苏娜贇 王廷义 左钱飞 路倩 赵喆 梅浩 王斌 陈东风 兰春慧 SU Nayun;WANG Tingyi;ZUO Qianfei;LU Qian;ZHAO Zhe;MEI Hao;WANG Bin;CHEN Dongfeng;LAN Chunhui(Department of Gastroenterology,Chongqing Key Laboratory of Digestive Malignancies,Daping Hospital,Army Medical University,Chongqing 400042,China;National Engineering Research Center of Immunological Products,Department of Microbiology and Biochemical Pharmacy,College of Pharmacy,Army Medical University,Chongqing,400038,China)

机构地区：[1]陆军军医大学陆军特色医学中心(大坪医院)消化内科、消化系统肿瘤精准防治重庆市重点实验室,重庆400042 [2]陆军军医大学院微生物与生化药学教研室、国家免疫工程中心,重庆400038

出　　处：《免疫学杂志》2023年第12期1021-1027,共7页Immunological Journal

基　　金：国家自然科学基金(82072253)

摘　　要：目的幽门螺杆菌(H.pylori)引起的慢性炎症是胃癌的主要原因之一,本研究旨在探讨幽门螺杆菌的重要毒力因子细胞毒素相关蛋白A(CagA)促炎的作用及其调控机制。方法采用质谱鉴定CagA在AGS细胞内的相互作用蛋白,并通过免疫共沉淀、免疫荧光验证其相互作用。通过使用siRNA敲低后,免疫印迹法检测通路蛋白表达、qRT-PCR检测炎症因子mRNA水平、ELISA检测蛋白水平。使用血红素-伊红染色法(H&E)检测临床样本中CagA诱导的炎症反应。结果CagA与SHARPIN具有相互作用。与CagA敲除株相比,CagA激活了NF-κB信号通路并上调了炎症因子IL-6、IL-8、TNF-α的mRNA水平和蛋白表达(P均<0.05)。通过siRNA敲低SHARPIN后,炎症水平降低、NF-κB信号被部分抑制。临床样本中CagA^(+)样本炎症反应显著增加。结论CagA促进宿主炎症反应,干扰SHARPIN可部分抑制CagA所致炎症反应,表明CagA通过与SHARPIN结合这一新机制激活NF-κB信号通路促炎。Chronic inflammation induced by Helicobacter pylori is considered to be one of the main causes of gastric cancer,and CagA is a main virulence factor of H.pylori.The study aimed to investigate the role and mechanism of CagA in host inflammatory response.Mass spectrometry was used to identify the interacting proteins of CagA in AGS cells.By immunoprecipitation and immunofluorescence,the interaction was validated.Pathway expression was detected by immunoblotting after knockdown by using siRNA,and mRNA levels of inflammatory cytokines were detected by quantitative PCR.CagA-induced inflammatory responses were detected in clinical samples using hemoglobin-eosin staining(H&E).Data showed that CagA interacted with SHARPIN.And CagA activated the NF-κB signaling pathway and upregulated the mRNA and protein levels of the inflammatory cytokines IL-6,IL-8,and TNF-α,as compared with the CagA knockout strain(all P<0.05).Knockdown of SHARPIN by siRNA reduced inflammation levels and partially inhibit NF-κB signaling.In clinical samples,CagA-positive samples exhibited stronger inflammatory responses.To sum up,CagA promoted the host inflammatory response,and CagA-induced inflammatory response was reduced when SHARPIN was partially inhibited,suggesting that CagA activates the NF-κB signaling pathway through binding to SHARPIN.

关 键 词：幽门螺杆菌 细胞相关蛋白A 炎症 

分 类 号：R573.3[医药卫生—消化系统]