自噬对小鼠巨噬细胞吞噬创伤弧菌能力的影响  

Role of autophagy in regulating phagocytosis in Vibrio vulnificus-infected murine macrophages

作　　者：陈娜 蒋烨琳 谢旦立[2] 黄显辉[2] 楼永良[2] 温超玮 Chen Na;Jiang Yelin;Xie Danli;Huang Xianhui;Lou Yongliang;Wen Chaowei(Department of Laboratory Medicine,the First People′s Hospital of Linping District,Hangzhou 311199,China;Key Laboratory of Laboratory Medicine,Ministry of Education,School of Laboratory Medicine and Life Sciences,Wenzhou Medical University,Wenzhou 325035,China)

机构地区：[1]杭州市临平区第一人民医院检验科,杭州311199 [2]温州医科大学检验医学院(生命科学学院),检验医学教育部重点实验室,温州325035

出　　处：《中华微生物学和免疫学杂志》2025年第2期108-114,共7页Chinese Journal of Microbiology and Immunology

基　　金：浙江省自然科学基金(LY22H190002);国家重点研发计划(2021YFC2300302);浙江省医药卫生科技计划(2022RC045);教育部产学协同育人(220604408244731)。

摘　　要：目的探讨自噬对小鼠巨噬细胞吞噬创伤弧菌能力的影响。方法采用Western blot检测创伤弧菌感染RAW264.7细胞和BMMφ细胞自噬标志物的表达变化;构建带荧光蛋白标记的创伤弧菌(Vv-GFP)感染RAW264.7细胞和BMMφ细胞,共聚焦显微镜观察巨噬细胞吞噬后的创伤弧菌与LC3蛋白共定位情况;采用Bafilomycin A1抑制自噬,Western blot检测自噬标志物的表达变化,流式细胞术检测巨噬细胞吞噬清除创伤弧菌能力的改变。结果创伤弧菌感染诱导RAW264.7细胞和BMMφ细胞Atg7、Atg12、Atg16L1和LC3Ⅱ的表达显著上调,p62蛋白表达显著下调;巨噬细胞吞噬的创伤弧菌与自噬体形成膜标志物LC3B发生共定位;创伤弧菌感染促进了自噬体形成和自噬底物降解,促进巨噬细胞对创伤弧菌的吞噬作用,而自噬抑制剂Bafilomycin A1则通过抑制自噬/溶酶体降解过程,导致Atg7、Atg12、Atg16L1和LC3Ⅱ表达累积,p62表达上调,阻断自噬流,进而减弱巨噬细胞对创伤弧菌吞噬清除的能力。结论创伤弧菌感染诱导巨噬细胞发生自噬,有利于增强巨噬细胞吞噬清除创伤弧菌的能力。Objective To investigate the role of autophagy in the regulatory of phagocytosis in Vibrio vulnificus(V.vulnificus)-infected murine macrophages.MethodsThe expression of cellular autophagy-related proteins in PBS-treated and V.vulnificus-infected RAW264.7 and BMMφcells was detected by Western blot.The co-localization of V.vulnificus-GFP and LC3Ⅱprotein in V.vulnificus-GFP-infected RAW264.7 and BMMφcells were detected using confocal microscopy.The phagocytosis of V.vulnificus in V.vulnificus-GFP-infected RAW264.7 and BMMφcells with or without autophagy inhibition using Bafilomycin A1 was detected by flow cytometry.ResultsThe up-regulated levels of Atg7,Atg12 and Atg16L1 proteins,increased LC3Ⅱ/actin ratio,as well as down-regulated p62 protein levels were significantly detected in V.vulnificus-infected RAW264.7 and BMMφcells.The co-localization of V.vulnificus-GFP and LC3Ⅱprotein was clearly observed in V.vulnificus-GFP-infected RAW264.7 and BMMφcells.Enhanced phagocytosis of V.vulnificus and increased autophagy were exhibited in V.vulnificus-GFP-infected RAW264.7 and BMMφcells,while weakened phagocytosis,accumulation of Atg7,Atg12,Atg16L1,LC3Ⅱand p62 protein levels,as well as blocking autophagy flux were detected in those cells within autophagy inhibition using Bafilomycin A1.ConclusionAutophagy induced by V.vulnificus infection could promote phagocytosis of V.vulnificus in macrophages.

关 键 词：自噬 巨噬细胞 吞噬 创伤弧菌 

分 类 号：R73[医药卫生—肿瘤]