Mesenchymal Cell Dysfunction in Fibroproliferative Lung Disease

摘要 进行性纤维化是主要发病率和死亡率的原因，最好的例子是特发性肺纤维化。 纤维化（IPF）。控制持续性肺纤维化的分子机制仍然知之甚少。期间 在本PO 1的资助期内，我们确定了一种侵袭性成纤维细胞表型，似乎是一种重要的 持续性肺纤维化的特征我们已经确定了导致严重肺损伤的侵袭性成纤维细胞， 在小鼠和人中，侵袭性表型需要HAS 2、CD 44和β-抑制蛋白。成纤维 谱系追踪研究表明，T盒基因4（Tbx 4）谱系间充质祖细胞是 损伤的成年鼠肺中肌成纤维细胞的主要来源。为了进一步分析基因调控， 为了检测成纤维细胞的侵袭性，我们对一组侵袭性和非侵袭性成纤维细胞样本进行了RNA-seq分析， IPF患者。出乎意料的是，我们发现包括检查点PD 1配体， PD-L1（CD 274）和PD-L2（CD 273）在侵袭性人成纤维细胞上优先显著上调。 程序性死亡-1（PD-1，CD 279）途径是免疫系统的稳态机制， 预防自身免疫和不受控制的炎症，它也被癌细胞用来逃避免疫， 监视这是一个有趣的和新颖的发现，特别是鉴于FDA批准的多- 用于IPF的酪氨酸激酶抑制剂尼达尼布，一种最初开发用于癌症的药物。一种范式已经进化 IPF的病理生物学更类似于癌症，而不是慢性炎症。然而，PD的作用- 1/PD-L1在肺纤维化中的作用未知。我们的初步研究表明，PD-L1调节成纤维细胞粘附， 迁移和入侵。我们发现粘着斑成分包括粘着斑激酶（FAK） 和Rho/ROCK显著上调。最近，与加州大学洛杉矶分校的约翰·贝尔佩里奥博士合作， PI在项目3中，我们还确定了慢性肺移植患者中的浸润性成纤维细胞 肺功能障碍（CLAD）是肺移植后发病率和死亡率的主要原因。我们 我们建议在这些细胞中进行类似的方法，希望能找到治疗CLAD的新靶点。 我们已经开始研究PD-L1的调节，发现PD-L1在肺部被p53下调 成纤维细胞与免疫检查点基因相反，p53在侵袭性成纤维细胞中显著下调， 与癌细胞中的相似。上皮p53在纤维化中的表达和功能已受到广泛关注， 但p53在成纤维细胞中的作用尚未被彻底研究。p53可能在这一过程中起作用， 不仅在防止上皮细胞凋亡方面，而且在成纤维细胞侵袭方面，从而提供了两种不同的 肺纤维化的发病机制。综合考虑，我们假设p53的缺失- 调节肺成纤维细胞中的免疫检查点基因，导致PD-1配体介导的逃避激活 免疫检测、粘着斑和细胞侵袭性。

Abstract Progressive fibrosis is a cause of major morbidity and mortality as best exemplified by idiopathic pulmonary fibrosis (IPF). The molecular mechanisms that control unremitting lung fibrosis remain poorly understood. During the funding period of this PO1, we identified an invasive fibroblast phenotype that appears to be an important feature of unremitting pulmonary fibrosis. We have identified invasive fibroblasts that contribute to severe lung fibrosis in mouse and human, and the invasive phenotype requires HAS2, CD44, and beta-arrestins. Fibroblast lineage-tracing studies identified that T box gene 4 (Tbx4)-lineage mesenchymal progenitors are the predominant source of myofibroblasts in injured adult murine lung. To further analyze the genetic regulation of fibroblast invasion, we performed RNA-seq analysis of a set of invasive and noninvasive fibroblast samples from IPF patients. Unexpectedly, we found that several immune molecules including both checkpoint PD1 ligands, PD-L1 (CD274) and PD-L2 (CD273), are significantly upregulated preferentially on invasive human fibroblasts. The Programmed Death-1 (PD-1, CD279) pathway is a homeostatic mechanism of the immune system that prevents autoimmunity and uncontrolled inflammation, and it is also used by cancer cells to escape from immune surveillance. This is an intriguing and novel discovery, particularly in light of the FDA approval of the multi- tyrosine kinase inhibitor nintedanib for IPF, a drug that was initially developed for cancer. A paradigm has evolved that the pathobiology of IPF is more similar to cancer than to chronic inflammation. However, the role of PD- 1/PD-L1 in lung fibrosis is unknown. Our preliminary studies revealed that PD-L1 regulates fibroblast adhesion, migration and invasiveness. We found that the focal adhesion components including focal adhesion kinase (FAK) and Rho/ROCK are significantly up-regulated. More recently, in collaboration with Dr. John Belperio at UCLA, PI on Project 3, we have also identified invasive fibroblasts in patients suffering from chronic lung allograft dysfunction (CLAD), which is the major cause of morbidity and mortality following lung transplantation. We propose to perform a similar approach in these cells with the hope of identifying new targets for therapy of CLAD. We have begun to examine the regulation of PD-L1 and found that PD-L1 was down-regulated by p53 in lung fibroblasts. In contrast to the immune checkpoint genes, p53 is markedly down-regulated in invasive fibroblasts, similar to that in cancer cells. Much attention has been paid to epithelial p53 expression and function in fibrosis, but the role of p53 in fibroblasts has not been thoroughly investigated. It appears that p53 may have a role not only in preventing epithelial apoptosis, but also in fibroblast invasiveness, thereby providing two distinct mechanisms in the pathogenesis of pulmonary fibrosis. Taken together, we hypothesize that loss of p53 up- regulates immune checkpoint genes in lung fibroblasts, leading to the activation of PD-1 ligand-mediated evasion of immune detection, focal adhesion and cell invasiveness.