高毒力肺炎克雷伯菌引发小鼠肝脓肿致脓毒症研究  

Study on sepsis induced by liver abscess in mice due to hypervirulent Klebsiella pneumoniae

作　　者：谢子文 樊力铭 蒋委余 龚可艺 章兴东 王家栋 江子妍 汪强 方佳琪 Xie Ziwen;Fan Liming;Jiang Weiyu;Gong Keyi;Zhang Xingdong;Wang Jiadong;Jiang Ziyan;Wang Qiang;Fang Jiaqi(Department of Clinical Medicine,School of Medicine,Hangzhou City University,Hangzhou 310015,China;Department of Laboratory Medicine,the Second Affiliated Hospital of Zhejiang Chinese Medical University,Hangzhou 310005,China)

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

出　　处：《中华微生物学和免疫学杂志》2025年第3期231-238,共8页Chinese Journal of Microbiology and Immunology

基　　金：浙江省自然科学基金探索项目(LQ20H100001);浙江省大学生科技创新活动计划暨新苗人才计划(2023R446004)。

摘　　要：目的探究高毒力肺炎克雷伯菌(hypervirulent Klebsiella pneumoniae,hvKP)对小鼠肝脓肿所致脓毒症免疫应答的影响及初步机制。方法C57BL/6小鼠腹腔分别注射对数期hvKP NTUH-K2044与经典肺炎克雷伯菌株(classical Klebsiella pneumoniae,cKP)HS11286菌悬液构建小鼠脓毒症模型,观察小鼠在24 h内的生存率;HE染色检测小鼠肝组织炎性细胞浸润情况;免疫组化法检测小鼠肝组织中性粒细胞标志物淋巴细胞抗原6G(lymphocyte antigen 6G,Ly6G)的水平。活性氧(reactive oxygen species,ROS)试剂盒检测小鼠肝脏巨噬细胞和外周血单核细胞ROS活性。Western blot检测小鼠肝脏巨噬细胞和外周血单核细胞中NF-κB信号通路p105/p50和p65蛋白的表达情况。qRT-PCR和ELISA分别检测小鼠肝脏巨噬细胞和外周血单核细胞中细胞因子IL-1β、IL-6和TNF-α的mRNA和蛋白质表达水平。采用单因素方差分析和t检验进行统计学分析。结果与cKP相比,hvKP感染C57BL/6小鼠后产生明显的肝脓肿,肝组织中有大量炎性细胞浸润,肝组织中性粒细胞Ly6G的水平显著增高(P<0.0001),小鼠存活率显著降低(P<0.0001)。hvKP感染后,小鼠肝脏巨噬细胞和外周血单核细胞中ROS活性显著增强(P<0.0001),且NF-κB信号通路p105/p50和p65的磷酸化水平显著提高(P<0.001)。此外,hvKP显著促进小鼠肝脏巨噬细胞和外周血单核细胞中IL-1β、IL-6和TNF-α的mRNA和蛋白质表达(P<0.001)。结论hvKP显著促进小鼠肝脓肿的发生并引发脓毒症。ObjectiveTo investigate the effect and preliminary mechanism of hypervirulent Klebsiella pneumoniae(hvKP)on the immune response to sepsis induced by liver abscess in mice.MethodsC57BL/6 mice were intraperitoneally injected with hvKP strain NTUH-K2044 or classical Klebsiella pneumoniae(cKP)strain HS11286 suspension to prepare the model of sepsis.The survivals rates of mice within 24 h were recorded.HE staining was used to observed the inflammatory cell infiltration in mouse liver tissues.The levels of neutrophil marker lymphocyte antigen 6G(Ly6G)in mouse liver tissues were detected by immunohistochemistry.The activity of reactive oxygen species(ROS)in mouse liver macrophages and peripheral blood monocytes was measured by ROS assay kit.The activation of p105/p50 and p65 in NF-κB signaling pathway in mouse liver macrophages and peripheral blood monocytes was detected by Western blot.The expression of IL-1β,IL-6 and TNF-αat mRNA and protein levels in mouse liver macrophages and peripheral blood monocytes were detected by qRT-PCR and ELISA,respectively.One-way analysis of variance and t test were used for statistical analysis.ResultsCompared with cKP,hvKP infection could induce C57BL/6 mice to develop obvious liver abscess with massive inflammatory cell infiltration.Moreover,the level of Ly6G in liver tissues was significantly higher in hvKP-infected mice than in cKP-infected mice(P<0.0001),but the survival rate of hvKP-infected mice was significantly lower than that of cKP-infected mice(P<0.0001).hvKP significantly promoted the ROS activity(P<0.0001)and enhanced the phosphorylation of p105/p50 and p65 in NF-κB signaling pathway in mouse liver macrophages and peripheral blood monocytes as compared with cKP(P<0.001).The expression of IL-1β,IL-6 and TNF-αat mRNA and protein levels in mouse liver macrophages and peripheral blood monocytes were significantly higher in hvKP-infected mice than in cKP-infected mice(P<0.001).ConclusionhvKP can promote the development of liver abscess and induce sepsis in mice.

关 键 词：肺炎克雷伯菌 肝脓肿 细胞因子 脓毒症 

分 类 号：R459.7[医药卫生—急诊医学]