Anti-fibrotic effects of the Mirlet7/NuRD ribonucleoprotein complexin idiopathic pulmonary fibrosis

特发性肺纤维化（IPF）是一种病因不明、预后不良的慢性、进行性、高致死性间质性肺疾病。IPF患者在诊断后2 - 5年内死亡，主要原因是呼吸衰竭。IPF患者的肺组织显示成纤维细胞病灶，包括扩张的肌成纤维细胞群和过量的细胞外基质（ECM）蛋白沉积。肌成纤维细胞是表达α-平滑肌肌动蛋白（ACTA 2）以促进损伤后伤口闭合的收缩性活化成纤维细胞。然而，这些细胞的持续活化导致ECM在间质空间中过度沉积，从而导致纤维化。基因表达的精确控制对于损伤后活化的成纤维细胞的扩增和伤口愈合后去分化为静息成纤维细胞两者都是必不可少的。在活化的成纤维细胞去分化为静息成纤维细胞期间，谱系特异性基因的转录与促纤维化基因的抑制之间的平衡允许纤维化病灶的消退。基因表达的调控与染色质结构密切相关。核小体重塑和脱乙酰酶（NuRD）复合物是染色质结构的主要调节因子之一。NuRD参与多种核过程，包括基因转录、DNA损伤修复、基因组稳定性维持和染色质组装。另一方面，以前的报告表明，非编码RNA（ncRNA）是必需的，无论是调节染色质调节剂的活性或招募他们到特定的基因位点。有趣的是，尽管ncRNA与IPF相关，但它们在IPF中的分子水平功能仍然难以捉摸。在这里，我们将研究细胞核中微小RNA（miRNAs）的功能，重点是miRNAs致死7（Mirlet 7，也称为let-7），因为它与肺部疾病有关。本项目提案旨在表征Mirlet 7 d介导的NuRD复合物向特定位点的募集，并在IPF背景下活化成纤维细胞去分化为静息细胞期间对其活性进行微调。我们将使用从供体和IPF患者分离的人原代成纤维细胞作为实验系统。我们的研究结果将为IPF治疗方法的发展提供分子机制基础。此外，将使用博来霉素动物模型（最常用的肺纤维化模型）证实Mirlet 7 d对IPF的治疗潜力。我们预期Mirlet 7 d给药将减弱博来霉素诱导的纤维化作用，从而证实我们的体外数据并支持使用Mirlet 7 d开发针对IPF的治疗策略。

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive and highly lethal interstitial lung disease with unknown etiology and with poor prognosis. IPF patients die within 2 to 5 years after diagnosis mostly due to respiratory failure. Lung tissues from IPF patients show fibroblastic foci that consist of expanded myofibroblast population and excessive extracellular matrix (ECM) protein deposition. Myofibroblasts are contractile activated fibroblasts that express α-smooth muscle actin (ACTA2) to facilitate wound closure after injury. However, persistent activation of these cells causes excessive deposition of ECM in the interstitial space that leads to fibrosis. The precise control of gene expression is essential for both expansions of activated fibroblasts after injury and dedifferentiation into resting fibroblasts after wound healing. During dedifferentiation of activated fibroblast into resting fibroblast, the balance between transcription of lineage specific genes and repression of pro-fibrotic genes allows resolution of fibrotic foci. The regulation of gene expression is intimately linked to chromatin structure. The nucleosome remodeling and deacetylase (NuRD) complex is one of the main regulators for chromatin structure. NuRD has been implicated in a wide variety of nuclear processes including gene transcription, DNA damage repair, maintenance of genome stability and chromatin assembly. On the other hand, previous reports suggest that noncoding RNAs (ncRNAs) are required either for modulating the activity of chromatin regulators or for recruiting them to specific gene loci. Interestingly, even though ncRNAs has been related to IPF, their function at molecular level in IPF has remained elusive. Here, we will investigate the function of micro RNAs (miRNAs) in the nucleus, focusing on the miRNA lethal 7 (Mirlet7, also known as let-7) due to its implication in lung diseases. The present project proposal aims to characterize Mirlet7d-mediated recruitment of NuRD complex to specific loci and fine tuning of its activity during dedifferentiation of activated fibroblasts into resting cells within the context of IPF. We will use as experimental system human primary fibroblasts isolated from donor and IPF patients. Our findings will provide the molecular mechanism as basis for the development of therapeutic approaches against IPF. Furthermore, the therapeutic potential of Mirlet7d against IPF will be confirmed using the bleomycin animal model, the most commonly used model of pulmonary fibrosis. We expect that Mirlet7d administration will attenuate the fibrotic effects induced by bleomycin, thereby confirming our in vitro data and supporting the development of therapeutic strategies against IPF using Mirlet7d.