K1型高毒力肺炎克雷伯菌通过激活NLRP3炎症小体引发肝脓肿  

Hypervirulent Klebsiella pneumoniae serotype K1 induces liver abscess through activating NLRP3 inflammasome

作　　者：汪强 欧茜 罗薇薇 马欣鹏 李雯婷 裘佳灵 陈雪静 方佳琪 Wang Qiang;Ou Qian;Luo Weiwei;Ma Xinpeng;Li Wenting;Qiu Jialing;Chen Xuejing;Fang Jiaqi(Department of Laboratory Medicine,the Second Affiliated Hospital of Zhejiang Chinese Medical University,Hangzhou 310005,China;School of Medicine,Zhejiang University City College,Hangzhou 310015,China)

机构地区：[1]浙江中医药大学附属第二医院医学检验科,杭州310005 [2]浙大城市学院医学院,杭州310015

出　　处：《中华微生物学和免疫学杂志》2022年第10期810-816,共7页Chinese Journal of Microbiology and Immunology

基　　金：浙江省基础公益研究计划项目(LGF22H190004);浙江省自然科学基金探索项目(LQ20H100001);浙江省卫生健康科技计划项目(2021KY840,2022KY927)。

摘　　要：目的探究高毒力肺炎克雷伯菌(hypervirulentKlebsiella pneumoniae,hvKP)通过激活NLRP3炎症小体引发肝脓肿。方法C57BL/6小鼠腹腔分别注射对数期的K1型hvKP(K1-hvKP)和K35型肺炎克雷伯菌(K35-非hvKP)悬液构建小鼠肝脓肿模型。流式细胞术检测CD45^(+)和Gr-1^(+)免疫磁珠分选的人外周血中性粒细胞纯度。总糖试剂盒检测K1-hvKP和K35-非hvKP荚膜多糖含量。实时荧光定量RT-PCR和ELISA分别检测K1-hvKP和K35-非hvKP对人血中性粒细胞IL-18与IL-33的mRNA和蛋白质表达水平的影响。Western blot检测K1-hvKP和K35-非hvKP对人血中性粒细胞NLRP3炎症小体活化的影响。激光共聚焦显微镜观察K1-hvKP和K35-非hvKP对人血中性粒细胞NETosis形成的影响。结果与K35-非hvKP比较,K1-hvKP感染C57BL/6小鼠后产生明显的肝脓肿。分离的人外周血中性粒细胞纯度>95%。K1-hvKP的荚膜多糖含量显著高于K35-非hvKP。与K35-非hvKP比较,K1-hvKP显著促进人血中性粒细胞IL-18和IL-33的表达、NLRP3炎症小体活化和NETosis的形成。结论hvKP通过激活NLRP3炎症小体促进人血中性粒细胞NETosis形成而引发肝脓肿。Objective To investigate whether the hypervirulent Klebsiella pneumoniae(hvKP)induces liver abscess through activating NLRP3 inflammasome.Methods K1-hvKP and K35-non-hvKP bacterial suspensions were intraperitoneally injected into C57BL/6 mice to establish the models of liver abscess.Human peripheral blood neutrophils were sorted by immunomagnetic beads with CD45^(+)and Gr-1^(+),and the purity was detected by flow cytometry.The concentrations of capsular polysaccharide of K1-hvKP and K35-non-hvKP were detected by total carbohydrate assay kit.The expression of IL-18 and IL-33 by neutrophils at mRNA and protein levels was detected by real-time fluorescence quantitative PCR and ELISA,respectively.The activation of NLRP3 inflammasome in neutrophils was detected by Western blot.Neutrophil extracellular trap formation(NETosis)was observed under confocal laser scanning microscope.Results The C57BL/6 mice with K1-hvKP infection had significantly serious liver abscess as compared with the K35-non-hvKP-infected mice.The purity of human neutrophils was more than 95%.The concentration of capsular polysaccharide in K1-hvKP was significantly higher than that in K35-non-hvKP.Compared with K35-non-hvKP,K1-hvKP significantly promoted the neutrophils to express IL-18 and IL-33 at both mRNA and protein levels,enhanced the activation of NLRP3 and induced NETosis.Conclusions This study suggested that hvKP could promote NETosis by activating NLRP3 inflammasome to cause liver abscess.

关 键 词：高毒力肺炎克雷伯菌 肝脓肿 中性粒细胞 NLRP3炎症小体 NETosis 

分 类 号：R575.4[医药卫生—消化系统]