FOXM1在呼吸道病毒感染致哮喘小鼠急性发作中的机制

doi:10.6040/j.issn.1671-7554.0.2022.1201

摘要/Abstract

摘要： 目的探讨呼吸道病毒感染诱发哮喘小鼠急性发作的致病机制。方法选取6~8周龄BALB/c雌性小鼠,随机分为正常对照组、哮喘组、Poly(I:C)组、RCM1+Poly(I:C)组和Poly(I:C)+RCM1组(后两组分别为提前和推后1 h给予RCM1),每组15只。利用屋尘螨(HDM)构建哮喘小鼠模型,病毒类似物Poly(I:C)经鼻滴入模拟呼吸道病毒感染诱导小鼠哮喘急性发作,以罗伯特·科斯塔纪念药1(RCM1)选择性抑制叉头框M1(FOXM1)表达,留取小鼠肺组织和肺泡灌洗液(BALF)。测定气道狭窄指数评估哮喘小鼠模型,采用Rt-PCR、ELISA法检测小鼠肺组织和BALF的白介素4(IL-4)、白介素13(IL-13)、干扰素γ(IFN-γ)炎症因子的表达及肺组织FOXM1和黏蛋白5AC(MUC5AC)表达;流式细胞分选小鼠BALF中的细胞计数;采用苏木精-伊红染色法和免疫组化染色法观察各组小鼠肺组织和气道上皮细胞形态学改变及FOXM1和MUC5AC表达。结果哮喘组小鼠气道阻力较正常对照组增高,而Poly(I:C)组小鼠气道阻力较哮喘组增加,差异均有统计学意义(P<0.05);与正常对照组相比,哮喘组小鼠肺组织IL-4和IL-13 的mRNA表达水平升高(P<0.05),IFN-γ的mRNA表达水平降低(P<0.05);Poly(I:C)组IL-4和IL-13 的mRNA表达水平高于哮喘组、RCM1+Poly(I:C)组和Poly(I:C)+RCM1组(P<0.05),RCM1+Poly(I:C)组IFN-γ的mRNA表达水平高于Poly(I:C)+RCM1组(P<0.05);与正常对照组相比,哮喘组BALF中细胞总数、嗜酸性粒细胞、巨噬细胞及IL-4、IL-13表达水平升高(P<0.05),IFN-γ下降(P<0.05),Poly(I:C)组BALF中细胞总数、嗜酸性粒细胞、巨噬细胞及IL-4、IL-13、IFN-γ高于哮喘组、RCM1+Poly(I:C)组和Poly(I:C)+RCM1组(P<0.05),且RCM1+Poly(I:C)组IL-4、IL-13表达低于Poly(I:C)+RCM1组(P<0.05);哮喘组小鼠肺组织FOXM1和MUC5AC的mRNA表达水平高于对照组(P<0.05),Poly(I:C)组的表达水平则高于哮喘组、RCM1+Poly(I:C)组和Poly(I:C)+RCM1组(P<0.05),RCM1+Poly(I:C)组肺组织FOXM1 mRNA表达水平高于Poly(I:C)+RCM1组(P<0.05),而MUC5AC的表达水平低于Poly(I:C)+RCM1组(P<0.05);哮喘组小鼠肺组织支气管上皮增厚、排列紊乱并有炎性细胞浸润,FOXM1及MUC5AC蛋白表达较正常对照组增高,差异均有统计学意义(P<0.05),而Poly(I:C)组小鼠上述形态学表现及FOXM1、MUC5AC蛋白表达水平较哮喘组更为显著(P<0.05),RCM1+Poly(I:C)组和Poly(I:C)+RCM1组气道上皮细胞形态学及FOXM1、MUC5AC蛋白表达较Poly(I:C)组明显减轻(P<0.05)。结论呼吸道病毒感染通过激活FOXM1通路,诱发气道上皮杯状细胞过度增生及黏液分泌亢进,同时促进哮喘小鼠肺组织炎症因子分泌,加重肺组织炎症,增加气道阻力,导致哮喘发作。

关键词: 叉头框M1, 病毒, 哮喘, 气道炎症, 杯状细胞增生

Abstract: Objective To investigate the role of FOXM1 in respiratory virus-induced asthmatic attacks in mice. Methods BALB/c female mice(aged 6-8 weeks)were randomly divided into control group, asthma model group, Poly(I:C)group, RCM1+Poly(I:C)group and Poly(I:C)+RCM1 group, with 15 mice in each group. Asthma was induced in mice via nasal administration of house dust mite(HDM)and viral analogue, Poly(I:C). The FOXM1 inhibitor RCM1 was used to investigate the role of FOXM1 in virus-induced asthmatic attacks. The lung tissue and bronchial alveolar lavage fluid(BALF)were collected. The induction of asthma was determined by measuring the airway stenosis index. The expressions of IL-4, IL-13, IFN-γ, FOXM1 and MUC5AC in lung tissue and BALF were detected with real-time PCR(RT-PCR)and ELISA. The cell counts in BALF were determined with flow cytometry. The morphological changes of lung tissue and airway epithelial cells were observed with haematoxylin & eosin(HE)staining. The expressions of FOXM1 and MUC5AC were determined with immunohistochemistry. Results The airway resistance was significantly higher in asthma model group mice than in the control group(P<0.05), and Poly(I:C)administration further increased airway resistance(P<0.05). The IL-4 and IL-13 levels were as follows: control groupP<0.05). The IFN-γ level was as follows: control model>asthma model group>Poly(I:C)group(P<0.05). The administration of RCM1, before or after the administration of Poly(I:C), reversed the cytokine levels(P<0.05). The levels of FOXM1 and MUC5AC were as follows: control groupP<0.05). The administration of RCM1 further decreased the expressions of FOXM1 and MUC5AC(P<0.05). The FOXM1 expression was higher, while MUC5AC expression was lower in RCM1+Poly(I:C)group than those observed in Poly(I:C)+RCM1 group(P<0.05). In the lungs of asthmatic mice, the epithelium was thickened, the cells were disorderly arranged, infiltration of the inflammatory cells was observed, and the protein levels of FOXM1 and MUC5AC were high(P<0.05). After administering Poly(I:C), the histological changes were aggravated, and protein expression was further elevated(P<0.05). The administration of RCM1 reversed the histological changes and FOXM1 and MUC5AC levels in RCM1+Poly(I:C)and Poly(I:C)+RCM1 groups(P<0.05). Conclusion Respiratory virus in mice induces airway goblet cell hyperplasia, increases mucus secretion, promotes the expression of the inflammatory cytokines, and increases airway resistance via the FOXM1 pathway and thus leading to asthmatic attacks.

Key words: Forkhead box M1, Virus, Asthma, Airway inflammation, Goblet cell hyperplasia

中图分类号:

R725.6

步美玲,王金荣,冯梅,孙立锋. FOXM1在呼吸道病毒感染致哮喘小鼠急性发作中的机制[J]. 山东大学学报 (医学版), 2023, 61(6): 1-9.

BU Meiling, WANG Jinrong, FENG Mei, SUN Lifeng. Mechanism of FOXM1 in acute exacerbation of asthma induced by respiratory virus infection in mice[J]. Journal of Shandong University (Health Sciences), 2023, 61(6): 1-9.

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