补体C9缺陷对LPS诱导的小鼠早期肝脏炎症和损伤的保护作用及机制  

Protective effects and mechanisms of complement C9 deficiency in LPS-induced early stage murine hepatic inflammatory response and injury

作　　者：吴艳红 付晓燕[1] 于洋 吴沛恩 许虎 房佩佩 魏涛 梁淑娟[1] WU Yan-Hong;FU Xiao-Yan;YU Yang;WU Pei-En;XU Hu;FANG Pei-Pei;WEI Tao;LIANG Shu-Juan(Key Laboratory for Immunology in Universities of Shandong Province,School of Basic Medical Sciences,Weifang Medical University,Weifang 261053,China)

机构地区：[1]潍坊医学院基础医学院,山东省高校免疫学重点实验室,潍坊261053

出　　处：《中国免疫学杂志》2021年第20期2433-2439,共7页Chinese Journal of Immunology

基　　金：国家自然科学基金(81873883,82000525);山东省医药卫生科技发展计划项目(2018WS065);国家级大学生创新训练计划项目(201910438018)。

摘　　要：目的:探讨补体C9基因缺陷对LPS诱导的小鼠早期肝脏炎症反应和损伤的作用及机制。方法:8~10周龄B6.WT和B6.C9^(-/-)小鼠均随机分为LPS组(n=5)和对照组(n=3),实验重复3次。LPS组腹腔注射LPS(0111:B4)5 mg/kg,对照组注射等体积生理盐水,处理12 h。酶法检测血清ALT、AST水平,HE染色观察肝脏病理学和炎症细胞浸润情况。ELISA检测血清和肝脏组织抽提物中IL-1β、IL-8、TNF-α水平以及血清中可溶型MAC(sMAC)水平。IF观察肝脏组织中巨噬细胞、中性粒细胞浸润及MAC沉积。qRT-PCR检测肝脏组织IL-1β、IL-8、TNF-αmRNA水平。FACS检测肝脏单个核细胞中巨噬细胞及中性粒细胞比例。结果:LPS刺激后,B6.C9^(-/-)小鼠血清中ALT、AST水平显著低于B6.WT小鼠(P<0.001),HE染色显示其肝脏组织中的病理损伤和炎症反应也明显轻于B6.WT小鼠。B6.C9^(-/-)小鼠血清中IL-1β(P<0.01)、IL-8(P<0.0001)、TNF-α(P<0.0001)水平均显著低于B6.WT小鼠。B6.C9^(-/-)小鼠肝脏中MAC沉积(P<0.01)及血清sMAC水平(P<0.05)低于B6.WT小鼠,肝脏组织抽提物中IL-1β(P<0.01)、IL-8(P<0.05)水平显著低于B6.WT小鼠,TNF-α水平无显著性差异。肝脏组织中IL-1β(P<0.05)mRNA水平低于B6.WT小鼠,而IL-8、TNF-αmRNA水平无显著差异。同时,肝脏单个核细胞中巨噬细胞及中性粒细胞比例也显著低于B6.WT小鼠(P<0.01)。结论:补体C9缺陷通过下调MAC沉积抑制巨噬细胞和中性粒细胞浸润对LPS诱导的早期肝脏炎症反应和损伤起保护作用。Objective:To explore the roles and mechanisms of complement C9 deficiency in lipopolysaccharide(LPS)-induced early stage hepatic inflammatory response and injury in mice.Methods:8~10-week-old B6.WT and B6.C9^(-/-)mice were randomly divided into LPS group(n=5)and control group(n=3).All results were representative of three independent experiments.The LPS group and control group mice accepted i.p.5.0 mg/kg of LPS or equal volume of NS for 12 h respectively.The levels of ALT and AST in the serum were detected in an enzyme based method according to the manufacture.Liver histopathological alterations and inflammatory immune cell infiltration were assessed by HE staining.Serum and liver tissue proinflammatory cytokines including IL-1β,IL-8,TNF-αand serum soluble membrane attack(sMAC)were detected by ELISA.IF staining was performed to observe the infiltration of macrophages,neutrophils as well as MAC deposition in liver tissue.Quantitative RT-PCR(qRT-PCR)were applied to detect the expressions of IL-1β,IL-8,TNF-αmRNA in liver tissue.Proportion of macrophage and neutrophil in liver mononuclear cells was evaluated by FACS.Results:When stimulated with LPS,B6.C9^(-/-)mice exhibited significantly decreased serum levels of ALT and AST compared with B6.WT mice(P<0.001).B6.C9^(-/-)mice showed less severe liver pathological injury and inflammation response.Serum IL-1β(P<0.01),IL-8(P<0.0001)and TNF-α(P<0.0001)levels were significantly lower than that in B6.WT mice.Intensity of MAC deposition in liver tissue(P<0.01)and serum sMAC level(P<0.05)were significantly reduced in B6.C9^(-/-)mice.Level of IL-1β(P<0.01)and IL-8(P<0.05)in liver tissue extracts were significantly reduced in B6.C9^(-/-)mice,while no marked difference in level TNF-αwas observed when compared with B6.WT mice.Expression level of IL-1βmRNA decreased apparently in liver tissue from B6.C9^(-/-)mice(P<0.05),however no marked difference of IL-8 and TNF-αmRNA were shown in both groups of mice.The proportion of infiltrated macrophage and neutrophil in liver mono

关 键 词：补体C9 LPS 内毒素休克 肝脏损伤 MAC(C5b-9) 

分 类 号：R392[医药卫生—免疫学]