基于蛋白芯片研究LDGs通过形成NETs激活肺纤维母细胞诱发多发性肌炎/皮肌炎相关间质性肺病的机制

Polymyositis (PM) and Dermatomyositis (DM) are systemic autoimmune myopathies. The most frequently observed organ involvement is interstitial lung disease (ILD), which is closely related to mortality. In our previous research, it was first to report that abnormally increased LDGs, abnormal formation of NETs and impaired activity of DNaseⅠ were closely related to ILD in PM/DM. However, the etiology and pathogenesis of PM/DM complicated ILD are still unclear. In this project, human lung fibroblasts (LF) and self-sustaining myositis mice with up-regulation of MHC class I will be used to elucidate the following questions: 1) if LDGs from BALF and peripheral blood are main responding cells of abnormally forming NETs in PM/DM, 2) whether NETs are related to LF activation in BALF and lung tissue of PM/DM-ILD to search for the direct evidence of NETs activating LF and infiltrating in lung tissue, 3) how NETs activate LF to develop PM/DM-ILD, validating the mechanism by using the myositis mice models, 4) how DNaseⅠactivity is impaired and whether DNaseⅠcan be a potential therapeutic target to myositis mice model with ILD. Based on proteinchips, these findings will demonstrate not only the molecular mechanism of LDGs activating LF to develop PM/DM-ILD, but also the therapeutic potential targets for the prevention and treatment of PM/DM-ILD.

多发性肌炎/皮肌炎(PM/DM) 是一组常见的自身免疫性疾病，间质性肺病(ILD)是其最常见且严重影响预后的合并症。我们在前期研究中发现低密度粒细胞（LDGs）、中性粒细胞胞外网状陷阱（NETs）及DNaseⅠ异常与PM/DM相关性ILD(PM/DM-ILD)密切相关，但其机制不清。因此拟使用人肺纤维母细胞（LF）和MHC-Ⅰ上调的自发性肌炎小鼠模型，围绕LDGs-NETs-LF激活-肺纤维化，基于信号通路探索抗体芯片技术，探讨：1）BALF来源和外周血来源的LDGs是否是PM/DM患者NETs形成异常的关键效应细胞；2）肺组织中NETs浸润并激活LF的直接病理证据；3)NETs激活LF的分子机制并用小鼠模型进行机制验证；4) DNaseⅠ活性受损的机制及潜在的治疗肺纤维化的机制。本项目将系统研究LDGs通过形成NETs激活LF造成ILD的分子机制，为PM/DM-ILD的防治提供新思路。

多发性肌炎/皮肌炎（PM/DM）是一组常见的自身免疫性疾病，间质性肺病（ILD）是其最常见且严重影响预后的合并症。我们在前期研究中发现中性粒细胞胞外网状陷阱（NETs）调控异常与PM/DM-ILD密切相关，但其机制不清。本研究中首先发现DM患者外周血单个核细胞（PBMCs）中低密度粒细胞（LDGs）的比例显著异常且与ILD相关；在动物实验中，发现肌炎小鼠模型给予NETs可显著促进ILD的进展和肺纤维化的发生，肌炎模型组和NETs干预组肺组织中TLR9,Smad2表达显著增高，而miR-7表达显著减少，而且使用miR-7类似物抑制了NETs促进肌炎小鼠ILD进展的作用；细胞实验中，NETs促进肺纤维母细胞（LF）的增殖及向肌成纤维细胞分化，NETs成分MPO和组蛋白3促进LF增殖和分化，经NETs刺激的LF中，TLR9上调、miR-7表达下调、Smad2表达上调，进一步证实TLR9通过Smad2及miR-7而参与LF的增殖和调控；TLR9-miR-7-Smad2参与NETs对LF增殖和分化的调控。由于LDGs对PBMCs中T细胞功能的影响，发现离心后贴壁法可显著降低PBMCs中LDGs的比例并已申请国家发明专利。项目已发表文章6篇，SCI文章3篇（二区一篇）。