下呼吸道菌群通过p38MAPK信号通路介导肺泡巨噬细胞极性在慢性阻塞性肺疾病中的作用机制

As an important chronic airway disease, previous studies have focused on the airway inflammation, oxidative stress and epigenetics in chronic obstructive pulmonary disease (COPD), however, there was few research on the respiratory tract microbiome. We have previously shown that there was difference in the lower respiratory tract microbiome between healthy non-smokers and COPD patients; the number of alveolar macrophage also increased significantly in aged lungs with COPD; meanwhile, the activation of p38MAPK was significantly increased in alveolar macrophages in COPD patients. Further, alveolar macrophage polarity plays a key role in the development of COPD. Herein, we hypothesize that microbial community of the lower respiratory tract is implicated in the mechanism of regulating alveolar macrophage polarity through p38MAPK signaling pathway in the occurrence and development of COPD. To address this, we will observe microbial community of the lower respiratory tract and alveolar macrophage polarity in COPD patients compared with control subjects, and analyze the relationship between microbial community and lung function. Using a murine model of chronic exposure to cigarette-smoke, the relationship between microbial community of the lower respiratory tract and alveolar macrophage polarity will be detected at different time points. Furthermore, using primary human alveolar macrophage or RAW264.7 cells culture, it will be studied that microbial community of the lower respiratory tract affects alveolar macrophage polarity through p38MAPK signaling pathways, contributing to the occurrence and development of COPD. We believe that the present study will contribute to enrich the pathogenesis of COPD, and benefit for innovative treatment of COPD.

慢性阻塞性肺疾病（慢阻肺）作为一种重要的慢性气道疾病，既往研究较为关注气道炎症、氧化应激、表观遗传学等方面，对呼吸道微生物群关注较少。我们前期研究发现非吸烟健康者与慢阻肺患者下呼吸道菌群存在差异；并且，肺泡巨噬细胞在慢阻肺患者肺脏中数量明显增多，磷酸化p38MAPK表达增加。而肺泡巨噬细胞极性与慢阻肺病情进展密切相关。因此，我们假设下呼吸道菌群调控p38MAPK信号通路介导肺泡巨噬细胞极性参与慢阻肺的发生发展。为了验证假说，首先观察慢阻肺患者下呼吸道菌群、肺泡巨噬细胞极性，并探讨其与疾病严重程度的相关性；利用小鼠慢性烟雾暴露模型，动态观察下呼吸道菌群与肺泡巨噬细胞极性间的关系；借助人原代肺泡巨噬细胞/RAW264.7巨噬细胞系培养，验证下呼吸道菌群通过p38MAPK信号通路影响肺泡巨噬细胞极性，参与慢阻肺的发生发展。该项目的实施将有助于丰富对慢阻肺发病机制的认识，探寻新的治疗靶点。

随着测序技术的发展，研究者们发现下呼吸道有大量微生物菌群定植。研究显示，吸烟对下呼吸道微生物菌群结构及组成有显著影响，且其变化可能与肺部疾病有关。慢性阻塞性肺疾病（慢阻肺）作为一种重要的慢性气道疾病，既往研究较为关注气道炎症、氧化应激、表观遗传学等方面，对呼吸道微生物群关注较少。通过痰液微生物16S rRNA测序技术分析稳定期慢阻肺患者下呼吸道微生物菌群结构的变化，并探讨其与慢阻肺患者临床症状及生物学指标的关系。选取2019年12月至2020年11月于中日友好医院门诊招募的稳定期慢阻肺患者 32例为研究对象。采用16S rRNA测序技术分析痰液微生物丰度与菌群结构组成，并分析其与慢阻肺患者临床特征的关系。慢阻肺患者的痰液中共检测到58个门水平的基因序列分类单元（OTUs）。其中，前3位分别是厚壁菌门（36%±22%）、变形菌门（23%±24%）和拟杆菌门（21%±14%），其次是放线菌门（10%±14%）和梭菌门(6%±7%)。按照菌群丰度将患者分为两组（聚类1和聚类2）(n=27, 5)。与聚类2相比，聚类1中拟杆菌门和梭杆菌门的比例更高(p=0.002和p=0.035)；聚类2中厚壁菌门多于聚类1(p<0.001)。运用CAT和mMRC问卷评估慢阻肺患者症状，FEV1评估患者病情严重程度。呼吸道菌群不同的慢阻肺患者，症状及肺功能严重程度亦不同。通过对微生物群落菌群和结构分析，可为前瞻性研究提供有用的临床指标，以描述正常定植中异常的菌群负荷，进而为慢阻肺患者临床治疗提供新的思路及依据。