屋尘螨过敏原对人支气管上皮细胞IL-32和相关炎症因子的影响  

Influence of house dust mite allergen on IL-32 and related inflammatory factors in human bronchial epithelial cell

作　　者：张超[1] 李永帅 章晓联[2] 刘光辉 ZHANG Chao;LI Yong-shuai;ZHANG Xiao-lian;LIU Guang-hui(Key Laboratory of Allergy and Immune Related Diseases of Hubei Province,Department of Allergy,Zhongnan Hospital,Wuhan University,Wuhan 430071;Key Laboratory of Allergy and Immune Related Diseases of Hubei Province,Department of Immunology,School of Medicine,Wuhan University,Wuhan 430071)

机构地区：[1]武汉大学中南医院过敏科,过敏及免疫相关疾病湖北省重点实验室,武汉430071 [2]武汉大学基础医学院免疫学系,过敏及免疫相关疾病湖北省重点实验室,武汉大学医学部,武汉430071

出　　处：《中华临床免疫和变态反应杂志》2021年第3期257-263,共7页Chinese Journal of Allergy & Clinical Immunology

基　　金：武汉大学中南医院科研培育项目(RCYJ20190105)。

摘　　要：目的探究IL-32在气传过敏原诱导的固有免疫反应中的作用。方法将同一批次处在指数生长期的人支气管上皮细胞(16HBECs)随机分为4组,设置空白对照组(A组),不同浓度(B:2μg/mL,C:10μg/mL,D:20μg/mL)屋尘螨(house dust mite,HDM)稀释液组,B、C和D组均分别刺激16HBE细胞6、12和24 h。通过荧光定量PCR技术检测胞内IL-32、IL-8、TNF-α和NF-κB等炎症因子的mRNA表达水平;酶联免疫吸附试验检测细胞上清液中IL-32、IL-6、IL-8和IL-1β的水平;Western blotting法检测p-p38、p-p65、TLR-4等蛋白的表达,探究HDM刺激16HBE细胞后的炎症效应。结果与对照组相比,随着HDM浓度和刺激时长增加,高浓度实验组IL-32、IL-8、IL-1β、TNF-α、NF-κB等炎症因子mRNA表达水平显著升高(1.74±0.03,1.87±0.05,2.62±0.20,2.35±0.14,1.40±0.07,P<0.01);IL-32、IL-6、IL-8、IL-1β水平也出现分泌性上调(P<0.05)。Western blotting结果显示,随着HDM浓度增加,p-p38和p-p65蛋白以及TLR-4表达均上调。结论HDM刺激16HBE细胞,会引起气道固有免疫效应,激发IL-32表达上调。IL-32可能通过TLR4/MD-2信号路径激活NF-κB信号通路,并与p38-MAPK通路发生交互作用产生促炎效应,导致过敏性气道炎症的发生与发展。Objective To investigate the role of IL-32 in innate immune response which induced by airborne allergens.Methods Human bronchial epithelial cells(16HBECs)in exponential growth phase of the same batch were randomly divided into 4 groups,and group A as control group was also set up.16HBECs in different groups were stimulated with HDM diluents in different concentrations(group B:2μg/mL,group C:10μg/mL,group D:20μg/mL)of for 6 hours,12 hours and 24 hours respectively.The mRNA transcription levels of intracellular inflammatory cytokines such as IL-32,IL-8,TNF-αand NF-κB were detected by using fluorescence quantitative PCR.The levels of IL-32,IL-6,IL-8,and IL-1βin supernatants were detected by using ELISA method,and the expression of TLR-4,p-p38,p-p65 was detected by using western blotting,in order to investigate the inflammatory effects of HDM allergen stimulation on 16HBECs.Results Compared to control group,the mRNA expression levels of inflammatory cytokines such as IL-32,IL-8,IL-1β,TNF-α,and NF-κB in high concentration experimental group significantly increased along with increasing of HDM concentration or stimulation duration(1.74±0.03,1.87±0.05,2.62±0.20,2.35±0.14,1.40±0.07,P<0.01),while the secretion of IL-32,IL-6,IL-8,and IL-1βwere also up-regulated(P<0.05).The up-regulation of p-p38,p-p65 proteins and TLR-4 expression were demonstrated by the results of western blotting,which is along with the increasing of allergen concentrations.Conclusions The stimulation of HDM on 16HBECs can induce innate immune effect and up-regulate IL-32.IL-32 may activate NF-κB signal pathway through TLR4/MD-2 signal pathway,and can interact with p38-MAPK pathway to produce pro-inflammatory effects,which resulted in occurrence and aggravation of allergic inflammation.

关 键 词：屋尘螨 白介素32 支气管上皮细胞 过敏性气道炎症 

分 类 号：R73[医药卫生—肿瘤]