Collagen Oxidation, Myofibroblast Activation and Age-Associated Pulmonary Fibrosis

胶原蛋白氧化、肌成纤维细胞激活和年龄相关性肺纤维化

• 批准号: 10445737
• 负责人: Yvonne M. W. Janssen-Heininger
• 金额: $ 31.98万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10445737
• 关键词: Address; Affect; Age; Aging; Attenuated; Autophagocytosis; Bleomycin; C-terminal; Calnexin; Cells; Characteristics; Cicatrix; Collagen; Cysteine; Data; Deposition; Environment; Enzymes; Extracellular Matrix; Fibroblasts; Fibrosis; Glutathione; Human; Hydrogen Peroxide; Knock-out; Laboratories; Link; Lung; Mass Spectrum Analysis; Modeling; Mus; Myofibroblast; Nature; Oxidation-Reduction; Pathogenesis; Patients; Preparation; Process; Production; Progressive Disease; Proliferating; Protein Disulfide Isomerase; Proteins; Proteomics; Pulmonary Fibrosis; Quality Control; Recombinants; Resistance; Role; Signal Transduction; Solubility; Stimulus; Stress; Structure of parenchyma of lung; System; TGFB1 gene; Testing; Tissues; aged; collagenase; design; effective therapy; glutaredoxin; idiopathic pulmonary fibrosis; improved; insight; novel; oxidation; pulmonary function; repair enzyme; trafficking

PROJECT SUMMARY Idiopathic pulmonary fibrosis (IPF) is a devastating, progressive disease characterized by scarred lung tissue containing excessive extracellular matrix (ECM). Changes in the oxidative (redox) environment have long been known to accompany IPF, yet the mechanisms whereby redox perturbations affect IPF pathogenesis remain incompletely understood. Our laboratory has discovered that a specific type of protein oxidation, termed protein glutathionylation, is increased in the lungs of IPF patients and inversely correlates with lung function. Importantly, the activity of glutaredoxin (GLRX), an enzyme that reverses protein glutathionylation, was strongly decreased in IPF patient lungs. New studies from our laboratory showed that collagen 1A1 (COL1), a major component of the fibrotic ECM, is a target for glutathionylation (COL1-SSG) that is increased in IPF. Glutathionylation of COL1- SSG in the c-terminal pro-domain caused partial resistance to degradation by multiple collagenases. We also discovered that COL1-SSG is a potent activator of fibroblasts and causes oxidative signals that promote further glutathionylation. Collagens are the most abundant proteins produced in the ER and are challenging to properly assemble and process, requiring oxidative processes. Collagen has its own autophagy system consisting of calnexin (CANX) and FAM134B to remove aberrantly processed collagen via ER-linked autophagy. Autophagy is decreased in aging, and both CANX and FAM134B are decreased in lungs from IPF patients. These collective observations led us to hypothesize that in the aging lung, increases in ER oxidation and diminished collagen autophagy result in the secretion of COL1-SSG to promote the progression of pulmonary fibrosis by the activation myofibroblasts via a self-propagating stimulus that can be diminished by GLRX. In Specific Aim 1 we will address whether the activity of protein disulfide isomerase A3 which is important in the oxidative folding of COL1, and attendant increases in ER oxidoreductin 1-derived hydrogen peroxide contribute to COL1-SSG and the increases in age-associated persistent fibrosis. In Specific Aim 2 we will examine whether COL1-SSG is secreted by fibroblasts from fibrotic lung and whether this is linked to alterations in ER redox stress and collagen autophagy. In Specific Aim 3 we will elucidate whether GLRX status affects COL1-SSG, myofibroblast activation and age- associated lung fibrosis. Completion of these proposed studies that utilize a combination of targeted knockout models in aged mice and isolated (myo)fibroblasts from patients with IPF will provide novel insights into the mechanisms by which ECM dysregulation via oxidation (specifically glutathionylation) affects COL1 stiffness and promotes the progression of pulmonary fibrosis. Completion of the workplan will begin to unravel the potential role of aberrant collagen autophagy in IPF. Finally, completion of the proposed studies will also further elucidate the mechanisms of action whereby GLRX exerts anti-fibrotic activity and identify the potential improved anti- fibrotic action of a newly created oxidation-resistant version of GLRX.

项目概要 特发性肺纤维化 (IPF) 是一种破坏性的进行性疾病，其特征是肺组织出现疤痕 含有过量的细胞外基质（ECM）。长期以来，氧化（氧化还原）环境的变化 已知伴随 IPF，但氧化还原扰动影响 IPF 发病机制的机制仍然存在 不完全理解。我们的实验室发现了一种特定类型的蛋白质氧化，称为蛋白质 IPF 患者肺部的谷胱甘肽水平升高，与肺功能呈负相关。重要的是， 谷氧还蛋白 (GLRX)（一种逆转蛋白质谷胱甘肽化的酶）的活性大幅下降 IPF 患者肺部。我们实验室的新研究表明，胶原蛋白 1A1 (COL1) 是 纤维化 ECM 是 IPF 中谷胱甘肽化 (COL1-SSG) 增加的靶标。 COL1- 的谷胱甘肽化 C 端前结构域中的 SSG 导致对多种胶原酶降解的部分抵抗。我们也 发现 COL1-SSG 是成纤维细胞的有效激活剂，并引起氧化信号，进一步促进 谷胱甘肽化。胶原蛋白是内质网中产生的最丰富的蛋白质，并且具有挑战性。 组装和加工，需要氧化过程。胶原蛋白有自己的自噬系统，包括 calnexin (CANX) 和 FAM134B 通过 ER 相关的自噬去除异常加工的胶原蛋白。自噬 随着年龄的增长而减少，IPF 患者肺部的 CANX 和 FAM134B 均减少。这些集体 观察结果使我们推测，在老化的肺中，内质网氧化增加，胶原蛋白减少 自噬导致COL1-SSG的分泌，通过激活促进肺纤维化的进展 肌成纤维细胞通过自我传播的刺激，可以被 GLRX 减弱。在具体目标 1 中，我们将解决 蛋白质二硫键异构酶 A3 的活性是否对 COL1 的氧化折叠很重要，以及 ER 氧化还原素 1 衍生的过氧化氢的随之增加有助于 COL1-SSG 的增加 与年龄相关的持续性纤维化。在具体目标 2 中，我们将检查 COL1-SSG 是否由 来自纤维化肺的成纤维细胞，以及这是否与内质网氧化还原应激和胶原自噬的改变有关。 在具体目标 3 中，我们将阐明 GLRX 状态是否影响 COL1-SSG、肌成纤维细胞激活和年龄 相关肺纤维化。完成这些利用靶向敲除组合的拟议研究 老年小鼠模型和 IPF 患者分离的（肌）成纤维细胞将为研究提供新的见解 ECM 通过氧化（特别是谷胱甘肽化）失调影响 COL1 硬度和 促进肺纤维化的进展。工作计划的完成将开始揭示潜力 异常胶原自噬在 IPF 中的作用。最后，拟议研究的完成也将进一步阐明 GLRX 发挥抗纤维化活性的作用机制，并确定潜在的改进抗纤维化活性 新创建的抗氧化版本 GLRX 的纤维化作用。