过敏原卵清白蛋白诱导慢性哮喘小鼠模型的建立及其免疫应答特征的动态分析  

Establishment of chronic asthma mouse model induced by allergen ovalbumin and dynamic analysis on its immune response characteristics

作　　者：严皎 白红妹[1] 杨旭[1] 孙文佳[1] 龙琼[1] 马雁冰[1] YAN Jiao;BAI Hongmei;YANG Xu;SUN Wenjia;LONG Qiong;MA Yanbing(Institute of Medical Biology,Chinese Academy of Medical Sciences&Peking Union Medical College,Kunming 650118,Yunnan Province,China)

机构地区：[1]中国医学科学院,北京协和医学院,医学生物学研究所,云南昆明650118

出　　处：《中国生物制品学杂志》2024年第4期420-425,432,共7页Chinese Journal of Biologicals

基　　金：国家自然科学基金(81072399);云南省重大科技专项(202002AA100009);云南省应用基础研究项目(2011FA023)。

摘　　要：目的 建立过敏原卵清白蛋白(ovalbumin,OVA)诱导的慢性哮喘小鼠模型,并对该模型免疫应答特征进行动态分析。方法 以OVA经腹腔致敏22只雌性BALB/c小鼠,并经鼻腔反复滴注刺激气道反应,于不同时间点取样进行分析。收集小鼠支气管肺泡灌洗液,进行炎性细胞计数;取灌洗液离心上清,ELISA法检测多种相关细胞因子水平。取小鼠肺组织,制备切片,苏木精-伊红(hematoxylin-eosin,HE)染色法检测肺组织炎性细胞浸润情况,高碘酸-希夫(periodic acid-Schiff,PAS)染色法检测气道上皮杯状细胞增生情况;马松三色(Masson′s Trichrome)及Sircol染色法检测肺组织胶原积累情况。采集小鼠静脉血,分离血清,ELISA法检测血清中OVA特异的IgE、IgG1、IgG2a水平。应用FlexiVent小动物肺功能仪的强制低频振荡模式分析小鼠气道反应性。结果 OVA的反复刺激成功诱导了小鼠气道及肺组织炎性细胞应答、炎性细胞因子积累、气道杯状细胞增生、肺组织纤维化、过敏原特异IgE抗体应答、气道高反应性等哮喘典型指标,且呈现不同应答特征。随着OVA反复刺激,炎性细胞应答、杯状细胞增生有所下降,但胶原沉积增加,而气道的高反应性在末次刺激4周后仍存在。转化生长因子-β1(transforming growth factor-β1,TGF-β1)伴随炎性应答的持续呈高表达,TGF-β2及白介素-33(interleukin-33,IL-33)在末次刺激4周后才呈高水平表达。结论 本研究采用OVA持续刺激诱导建立了慢性哮喘小鼠模型,并展现了该模型的免疫应答特征,为慢性哮喘机制的研究、治疗靶点及药物研发奠定了基础。Objective To establish a chronic asthma mouse model induced by allergen ovalbumin(OVA),and analyze the immune response dynamic characteristics.Methods Twenty⁃two female BALB/c mice were sensitized intraperitoneally with OVA,and the airway response was stimulated by repeated infusion through nasal cavity.Samples were taken at different time points for analysis.The bronchoalveolar lavage fluid(BALF)of mice was collected and inflammatory cells were counted;the supernatant of lavage fluid was taken and detected for the levels of various related cytokines by ELISA.The lung tissue was taken to prepare sections,which was detected for the inflammatory cell infiltration by hematoxylin⁃eosin(HE)staining,for the goblet cell hyperplasia in airway epithelium by periodic acid⁃Schiff(PAS)staining,and for the collagen deposition by Masson′s Trichrome and Sircol staining.The venous blood of mice was collected,the serum was separated,and the levels of OVA⁃specific IgE,IgG1 and IgG2a in serum were detected by ELISA.The airway reactivity was measured using a FlexiVent small animal ventilator with low⁃frequency forced oscillation.Results Repeated challenges of OVA successfully induced typical asthma features such as inflammatory cell response in airway and lung tissues,inflammatory cytokine accumulation,airway goblet cell hyperplasia,lung fibrosis,allergen⁃specific IgE antibody response and airway hyperresponsiveness in mice,and different dynamic responsive characteristics appeared.With the repeated challenges of OVA,inflammatory cell response and goblet cell hyperplasia reduced,but collagen deposition developed,while airway hyperresponsiveness re⁃mained 4 weeks after the last stimulation.The expression of TGF⁃β1 was high during the inflammatory response,while the expression of TGF⁃β2 and IL⁃33 did not increase until 4 weeks after the last challenge.Conclusion This study successfully developed a chronic asthma model in mice,showing the immune response characteristics,laying a foundation for investigating immun

关 键 词：哮喘 卵清白蛋白 过敏原 免疫应答 

分 类 号：R392.8[医药卫生—免疫学]