中性粒细胞胞外诱捕网在肺内源性急性呼吸窘迫综合征中的作用研究  

The role of neutrophil extracellular traps in the pathogenesis of pulmonary endogenous ARDS

作　　者：高飞 惠姣洁 王康 郭赟[3] Gao Fei;Hui Jiao-jie;Wang Kang;Guo Yun(Department of Intensive Care Unit,the Affiliated Wuxi People's Hospital of Nanjing Medical University,Wuxi 214023,China)

机构地区：[1]南京医科大学附属无锡人民医院重症医学科,江苏无锡214023 [2]南京医科大学附属无锡市儿童医院儿科实验室,江苏无锡214023 [3]南京医科大学附属无锡市儿童医院呼吸科,江苏无锡214023

出　　处：《中国急救医学》2020年第9期853-857,共5页Chinese Journal of Critical Care Medicine

基　　金：国家自然科学基金(81401619);江苏省卫生计生委青年医学人才项目(QNRC2016178)。

摘　　要：目的本研究拟通过人群及动物模型探讨中性粒细胞胞外诱捕网(NETs)在肺内源性急性呼吸窘迫综合征(ARDS)中的作用.方法收集21例肺内源性ARDS患者(ARDS组),同期收治21例普通肺部感染患者(非ARDS组),检测外周血浆游离DNA(cf-DNA)含量并进行比较分析,了解NETs的总量有无区别;分离两组患者外周血中性粒细胞(PMNs),使用相同浓度(50 ng/mL)的佛波醇-12-肉豆蔻酯-13-乙酸(phorbol-12-myristate-13-acetate,PMA)进行刺激,评估NETs产生的比例,了解PMNs功能有无变化;采用ARDS组及非ARDS组的肺泡灌洗液(BALF)与正常人PMNs进行共培养,评估NETs产生的比例,了解ARDS组肺部微环境有无异常.以内毒素(LPS,6 mg/kg)经鼻气管滴注法,建立ARDS动物模型,造模后24 h取肺组织,进行苏木素-伊红(HE)染色和免疫组化,在动物模型验证NETs在ARDS发病中的作用.结果ARDS组cf-DNA含量明显高于非ARDS组[(639.11±291.16)ng/mL vs.(169.29±194.98)ng/mL,P=0.000].在ARDS动物模型中,肺组织NETs的表达同样较对照组增加.两组患者外周血提取的PMNs在使用50 ng/mL PMA刺激3 h后,ARDS组产生NETs的比例高于非ARDS组(78.16%vs.19.7%,P=0.000).正常健康人来源的PMN在与BALF共培养后,ARDS-BALF刺激PMNs产生NETs的比例高于非ARDS-NETS(81.4%vs.29.7%,P=0.000).结论NETs的异常增多在肺内源性ARDS的发病中起着作用,ARDS患者PMNs功能异常及肺内微环境紊乱可能是引起NETs异常增多的原因.Objective The purpose of this study is to explore the role of neutrophil extracellular traps(NETs)in pulmonary endogenous ARDS by human and animal models.Methods 21 patients with pulmonary endogenous ARDS were selected as ARDS group and 21 patients with common pulmonary infection as non-ARDS group.The cf-DNA content of peripheral plasma were detected and analyzed to find out whether there was any difference in the total amount of NETs.The peripheral blood polymorphonuclear neutrophils(PMNs)of the two groups were separated and stimulated with phorbol-12-myristate-13-acetate(PMA)of the same concentration(50 ng/mL).The proportion of NETs production was evaluated to know whether the function of PMNs changed.Bronchoalveolar lavage fluid(BALF)from ARDS group and non-ARDS group was co cultured with PMNs of normal people,and the proportion of NETs production was evaluated to know whether there was abnormality in the lung microenvironment of ARDS group.The animal models of ARDS were established by intranasal and tracheal instillation of lipopolysaccharide(LPS,6 mg/kg).The lung tissue was taken at 24th hour after the models were made,and HE staining and immunohistochemistry were carried out to verify the role of NETs in the pathogenesis of ARDS in the animal models.Results The cf-DNA of the ARDS group was significantly higher than that of the non-ARDS group[(639.11±291.16)ng/mL vs.(169.29±194.98)ng/mL,P=0.000).In the ARDS animal models,the expression of NETs in lung tissue was also increased compared with the control group.The PMNs extracted from peripheral blood were likely to produce the higher proportion of NETs in the ARDS group than in the non-ARDS group(78.16%vs.19.7%,P=0.000).The normal human-derived PMNs co-cultured with BALF,and ARDS-BALF stimulated PMNs to produce the higher proportion of NETs than non-ARDS-NETs(81.4%vs.29.7%,P=0.000).Conclusion The abnormal increase of NETs plays a role in the pathogenesis of endogenous ARDS in the lung.The abnormal function of PMNs in the ARDS patients and the disorder of mic

关 键 词：中性粒细胞胞外诱捕网(NETs) 急性呼吸窘迫综合征(ARDS) 中性粒细胞(PMNs) 

分 类 号：R563.8[医药卫生—呼吸系统]