模拟临床的中性粒细胞性激素抵抗型支气管哮喘小鼠模型的建立及意义  被引量:9

Establishment of a mouse model mimicking neutrophilic steroid-refractory asthma

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作  者:王楠 陈子 葛林阳 王曦 高蕾 李涛[1] 胡蓉 张明顺 周林福 Wang Nan;Chen Zi;Ge Linyang;Wang Xi;Gao Lei;Li Tao;Hu Rong;Zhang Mingshun;Zhou Linfu(Department of Respiratory and Critical Care Medicine,the First Affiliated Hospital,Nanjing Medical University,Nanjing 210029,China;Department of Immunology,School of Basic Medical Sciences,Nanjing Medical University,Nanjing 211166,China)

机构地区:[1]南京医科大学第一附属医院呼吸与危重症医学科,210029 [2]南京医科大学基础医学院免疫学系,211166

出  处:《中华结核和呼吸杂志》2020年第1期40-46,共7页Chinese Journal of Tuberculosis and Respiratory Diseases

基  金:国家重点研发计划(2018YFC1313600);国家自然科学基金重点国际(地区)合作研究项目(81820108001);国家自然科学基金项目(81670029,81370133,81170018);江苏省医学重点人才项目(ZDRCA2016018);江苏省333高层次人才工程第二层次培养对象(BRA2019078);江苏省社会发展重点研发专项(BE2015651);江苏省研究生科研创新计划(KYCX171285)。

摘  要:目的建立一种模拟临床的中性粒细胞性激素抵抗型支气管哮喘(哮喘)小鼠模型并探讨其意义。方法采用屋尘螨(house dust mite,HDM)和脂多糖(lipopolysaccharide,LPS)混合液,气管内给药建立哮喘小鼠模型。将18只雌性C57BL/6小鼠按照数字表法随机分为对照组、哮喘组(HDM+LPS)和地塞米松(dexamethasone,Dex)组(HDM+LPS+Dex)。肺功能仪测定小鼠气道阻力,HE染色观察肺组织炎症细胞浸润,过碘酸-雪夫染色(PAS)观察杯状细胞增生,瑞氏染色检测BALF炎症细胞总数及分类计数,聚合酶链式反应(PCR)和酶联免疫吸附试验(ELISA)检测肺组织和BALF炎症因子。流式细胞术检测肺组织Th17细胞分化。结果HE染色显示,哮喘组肺组织炎症较对照组显著增高(P<0.05),地塞米松组肺组织炎症较哮喘组稍减轻(P>0.05);BALF细胞分类计数显示,哮喘组炎症细胞(除外嗜酸粒细胞)浸润较对照组显著增加[炎症细胞总数分别为(2797±400)×106/L和(105±75)×106/L,中性粒细胞计数分别为(1151±395)×106/L和(12±6)×106/L,淋巴细胞计数分别为(897±135)×106/L和(11±5)×106/L,巨噬细胞计数分别为(215±51)×106/L和(34±16)×106/L,均P<0.05],地塞米松组肺组织炎症细胞总数及巨噬细胞计数较哮喘组显著下降[炎症细胞总数(1140±418)×106/L vs(2797±400)×106/L,巨噬细胞计数(117±31)×106/L vs(215±51)×106/L,均P<0.05],但淋巴细胞和中性粒细胞计数无统计学意义[淋巴细胞计数(587±208)×106/L vs(897±135)×106/L,中性粒细胞计数(294±134)×106/L vs(1151±395)×106/L,均P>0.05];免疫组织化学检测结果同样证实,地塞米松组肺组织中性粒细胞浸润较哮喘组不能被有效抑制;哮喘组和地塞米松组气道阻力较对照组均显著增高(P<0.05),但两组间气道阻力差异无统计学意义(P>0.05);与哮喘组相比,地塞米松组肺组织Th2炎症指标显著降低(均P<0.05),而Th17细胞炎症指标有升高趋势,Th1炎症指标无显著改善;流式细�Objective To establish a murine model of neutrophilic steroid-refractory asthma for a better exploration of its pathogenesis and treatment.Methods A mouse model of neutrophilic steroid-refractory asthma was established by intratracheal administration of a mixture of house dust mite(HDM)and lipopolysaccharide(LPS).Eighteen female C57BL/6 mice were randomly divided into a control group,an asthma group(HDM+LPS)and a dexamethasone(Dex)group(HDM+LPS+Dex).The airway resistance was measured by pulmonary function test.Airway inflammation,hypersecretion and inflammatory cell recruitment were observed by HE,PAS and Wright′s staining,respectively.Inflammatory mediators in lung and bronchoalveolar lavage fluid(BALF)were detected by PCR and ELISA.Th17 cell differentiation in the lung was analyzed by flow cytometry.Results HE staining showed a significantly increased airway inflammation in the asthma group than that in the control group(P<0.05),and a slightly decreased airway inflammation in the Dex group than that in the asthma group(P>0.05).Cell counts in BALF showed a sharply elevated recruitment of inflammatory cells in the asthma group(except eosinophils)than that in the control group(total number of inflammatory cells(2797±400)×106/L vs(105±75)×106/L,neutrophils(1151±395)×106/L vs(12±6)×106/L,lymphocytes(897±135)×106/L vs(11±5)×106/L,macrophages(215±51)×106/L vs(34±16)×106/L,P<0.05).Compared with the asthma group,Dex group significantly reduced the recruitment of total number of inflammatory cells and macrophages[total number of inflammatory cells(1140±418)×106/L vs(2797±400)×106/L,macrophages(117±31)×106/L vs(215±51)×106/L,all P<0.05]albeit no significances in lymphocytes and neutrophils[lymphocytes(587±208)×106/L vs(897±135)×106/L,neutrophils(294±134)×106/L vs(1151±395)×106/L,all P>0.05].Immunohistochemical staining confirmed that Dex could not effectively alleviate the neutrophilic inflammation in asthmatic mice.Although there was no significant difference in airway hyperreactivity be

关 键 词:哮喘 地塞米松 中性粒细胞 THL7细胞 

分 类 号:R-332[医药卫生] R5

 

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