免疫抑制剂环磷酰胺对小鼠流感相关性脑病的影响  

Effect of immunosuppressant cyclophosphamide on influenza-associated encephalopathy in mice

作　　者：张香港 赵澄 陈纯静 胡珏[1] 卢芳国[1] ZHANG Xiang-gang;ZHAO Cheng;CHEN Chun-jing;HU Jue;LU Fang-guo(Hunan University of Chinese Medicine,Changsha 410208,China)

机构地区：[1]湖南中医药大学,湖南长沙410208

出　　处：《中国病理生理杂志》2022年第11期2063-2071,共9页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.82074250);湖南省自然科学基金资助项目(No.2020JJ4063);2019年湖南省研究生优秀教学团队立项项目(No.2019-118);湖南中医药大学一流学科《基础医学》资助项目(No.1);湖南中医药大学中西医结合一流学科开放基金项目(No.2020ZXYJH13)。

摘　　要：目的:观察免疫抑制剂环磷酰胺对小鼠流感相关性脑病的影响,为流感相关性脑病防治研究奠定基础。方法:BABL/c小鼠随机分为正常对照组、环磷酰胺组、病毒组和环磷酰胺+病毒组,其中环磷酰胺组、环磷酰胺+病毒组小鼠按照75 mg/kg的剂量腹腔注射环磷酰胺1次,注射后24 h,病毒组、环磷酰胺+病毒组小鼠鼻腔接种A型流感病毒,于滴鼻后第3、7天取材。常规法检测小鼠肺指数、脑指数;HE染色法观察脑组织病理变化;ELISA检测脑组织匀浆中白细胞介素6(interleukin-6,IL-6)和肿瘤坏死因子α(tumor necrosis factor-α,TNF-α)的含量水平;RT-qPCR技术检测肺、脑组织中流感病毒载量水平;免疫组化法检测脑组织中IL-6、TNF-α、闭锁小带蛋白1(zonula occluden-1,ZO-1)的蛋白表达水平;Pearson或Spearman法评估脑指数与脑组织匀浆中IL-6和TNF-α含量的相关性。结果:与正常对照组及病毒组比较,环磷酰胺+病毒组肺指数和脑指数升高(P<0.05),脑组织海马病理损伤严重,肺组织中流感病毒核蛋白基因相对表达量显著增高(P<0.01),脑组织匀浆中IL-6和TNF-α含量显著升高(P<0.01),脑组织中IL-6和TNF-α蛋白表达水平显著增加,ZO-1蛋白表达水平显著降低(P<0.05)。结论:使用免疫抑制剂环磷酰胺有利于小鼠流感相关性脑病模型的建立。AIM:To observe the effect of immunosuppressant cyclophosphamide on influenza-associated encephalopathy in mice,and lay a foundation for the prevention and treatment of influenza-associated encephalopathy.METHODS:BABL/c mice were randomly divided into normal control group,cyclophosphamide group,virus group,cyclophosphamide+virus group.The mice in cyclophosphamide group and cyclophosphamide+virus group were intraperitoneally injected with cyclophosphamide once a day at a dose of 75 mg/kg.At 24 h after injection,the mice in virus group and cyclophosphamide+virus group were inoculated with influenza A virus intranasally.The tissues were harvested on day 3 and day 7 after the nasal instillation.The lung index and brain index of the mice were detected by routine method.HE staining method was used to observe the pathological changes of brain tissues.Enzyme-linked immunosorbent assay was used to detect the levels of interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)in brain tissue homogenates.Realtime fluorescence quantitative polymerase chain reaction was used to detect influenza virus load levels in lung and brain tissues.Immunohistochemistry was used to detect the protein expression levels of IL-6,TNF-α and zonula occluden-1(ZO-1).The Pearson or Spearman method was used to evaluate the correlation between the brain index and the level of IL-6 or TNF-αin brain tissues.RESULTS:Compared with normal control group and virus group,the lung index and brain index of the mice in cyclophosphamide+virus group were increased(P<0.05),and severe pathological damage was observed in the hippocampus.The expression of influenza virus nucleoprotein genes in lung tissue was increased(P<0.01),and the levels of IL-6 and TNF-α in brain tissue homogenates were increased(P<0.01).The protein expression levels of IL-6 and TNF-α in brain tissues were increased,while the protein expression level of ZO-1 was decreased(P<0.05).CONCLU⁃SION:Immunosuppressant cyclophosphamide is beneficial for the establishment of influenza-associated encep

关 键 词：免疫抑制剂 A型流感病毒 流感相关性脑病 环磷酰胺 

分 类 号：R742[医药卫生—神经病学与精神病学] R363.21[医药卫生—临床医学]