CD4+CD25-LAG3+T细胞对结缔组织病相关肺纤维化的抑制作用及机制研究

onnective tissue disease-related pulmonary interstitial disease is an important cause of pulmonary fibrosis, pathological changes of diffuse alveolitis, myofibroblast proliferation which leading to collagen deposition is characterized, and the disease eventually leading to lung structural damage, causing respiratory failure. This disease is a highly harmful irreversible disease for 5 years of survival rate less than 50%. Recent studies have shown that immune disorders play an important role in organ fibrosis, as regulatory T cells (Treg) decreased which lead to reduced immune negative regulation may be the key to the development of the disease. We found that: CD4 + CD25-LAG3 + Tregs in patients with peripheral blood levels were significantly lower than normal; LAG3 + Treg infusion into bleomycin-induced pulmonary fibrosis in mice model can significantly improve the disease, LAG3 + Treg may inhibit the proliferation of peripheral fibroblasts and inhibit the proliferation of myofibroblasts by secreting TGF-beta3 and IL-10. The lungs of ATF3 - / - KO mice develop chronic inflammatory infiltration and different degrees of fibrosis and the level of LAG3 + Treg in the arterial blood was decreased. Thus，we propose that LAG3 + Treg can improve the progression of pulmonary fibrosis by inhibiting inflammatory cell infiltration and collagen formation, and its differentiation is regulated by ATF3. LAG3 + Treg in vivo can be used as potential biomarker. If the hypothesis is established, it will provide new ideas and theoretical basis for the immunotherapy of pulmonary fibrosis, and has great application value prospect.

结缔组织病为肺纤维化的重要病因，病理改变以炎症细胞及因子浸润诱导肌成纤维细胞增殖导致胶原沉积为特征，导致肺结构破坏，是危害性极大的不可逆疾病。研究显示免疫调节紊乱在器官纤维化中起重要作用，调节性T细胞（Treg）减少是导致疾病发展的关键。CD4+LAG3+Treg是近年来新发现起重要免疫负调控作用的Treg亚群，在多种炎症及免疫疾病动物模型中显示出治疗作用，我们初步发现其在患者血中水平较正常人下降；在肺纤维化动物模型中输注LAG3+Treg可显著改善病情，而ATF3-/-KO小鼠出现脾脏中LAG3+Treg水平下降，伴肺部自发性慢性炎症浸润及不同程度的纤维化。据此我们提出：LAG3+Treg通过抑制炎症细胞浸润和胶原生成从而有效改善肺纤维化，其分化受ATF3的调控，患者外周血中的LAG3+Treg可作为潜在生物标志物。如假设成立，将为肺纤维化的免疫疗法提供新的思路及依据，具较大应用价值。

结缔组织病相关肺纤维化分为两个阶段，首先是如巨噬细胞等炎症细胞浸润，巨噬发生表型转化，分泌炎症因子，甚至自身转分化为肌成纤维细胞发挥促纤维化作用；在炎症因子刺激下，肺成纤维细胞在TGF-β1诱导下发生间质转化为肌成纤维细胞，大量分泌胶原导致肺结构破坏。因此，在这项研究中，我们结合生物信息学等手段，首次试图检查CD4+LAG3+T细胞与CTD-ILD的关系，并在体外实验中阐述CD4+LAG3+T细胞对肺成纤维细胞以及炎症细胞的作用，从多方面证明其对肺纤维化过程的影响，最后通过动物实验进一步获得体内的证据。我们的数据证实CD4+LAG3+T细胞在CTD-ILD患者中下降。体外实验中，CD4+LAG3+T细胞可有效抑制肺成纤维细胞的间质转化，以及巨噬的分化和间质转化过程。我们首次阐明了免疫负调节细胞在风湿免疫病相关肺纤维化患者中的变化以及其作为生物标志物的潜在应用价值，且在体内体外实验中进一步完整地阐述了其作用机理，从而提供了确实可信的理论依据。而且，对特发性肺纤维化的生信分析表明LAG3+在其中没有分别，这表明了风湿免疫病驱动的肺纤维化机制的独特性，从而证明，影响免疫负调节细胞有可能提供一种治疗风湿免疫病相关肺纤维化的新治疗方法，切合目前国际上大势所趋的细胞治疗，因此在作为生物标志、以及治疗选择上，都有着广阔的应用前景。