LUNG COLLAGEN CROSSLINKING--BIOSYNTHESIS AND MATURATION

在这笔赠款下进行的以前的工作已经证明 胶原蛋白生物合成与成熟研究的可行性 正常大鼠皮肤、骨、腱、肺的活体标记 技巧。本申请建议将这些扩展 对有充分证据的肺纤维化动物模型的研究， 大鼠气管内注射博莱霉素，研究胶原蛋白 在受损的(前)纤维化肺中合成和成熟。我们 将专门测试键的羟化作用的假设 参与胶原交联反应的赖氨酸残基 是纤维化肺胶原蛋白生物合成的关键步骤。 几种动物模型肺组织的平行研究 人类纤维化肺表现为羟化增加 赖氨酸和增加的双官能席夫碱的比例- 衍生、交联化DHLNL到HLNL在(前)纤维化(急性纤维化)中的作用 肺部。同样，我们观察到 慢性纤维化中的完全羟化三官能团交联物 无论是动物模型还是人类肺部。 因此，我们建议在本文中检验以下假设 建议：(I)增加赖氨酸羟基化区别于 肺内“正常胶原蛋白”中的“纤维化胶原蛋白”； 这种差异是由赖氨酸酶活性增加所调节的。 纤维化前肺中的羟基酶；和(Iii)曾经 交联型、“纤维性胶原”在肺内不受 周转或崩溃，而是将持续一生 主持人。体内标记和组织切片的结合 将进行匀浆实验来检查这些 问题。与人肺组织的相关性研究 适用于急性(ARDS、IRDS)和慢性(IPF)患者 肺纤维化将研究胶原的潜在机制 在人类肺部疾病中的积聚，也许会发现 合理治疗这些疾病的潜在新靶点。

Previous work performed under this grant has demonstrated the feasibility of studying collagen biosynthesis and maturation in skin, bone, tendon, and lungs of normal rats by in vivo labelling techniques. The present application proposes to extend these studies into a well documented animal model of pulmonary fibrosis, intra-tracheally injected bleomycin in the rat, to study collagen synthesis and maturation in the damaged, (pre)fibrotic lung. We will specifically test the hypothesis that hydroxylation of key lysine residues that take part in collagen crosslinking reactions is a critical step in the biosynthesis of fibrotic lung collagen. Parallel studies in lung tissue from several animal models and in human fibrotic lungs have demonstrated increased hydroxylation of lysine and an increased ratio of the difunctional Schiff base- derived, crosslink DHLNL to HLNL in (Pre)fibrotic (acute fibrosis) lungs. Similarly, we have observed an increased content of the fully hydroxylated trifunctional crosslink OHP in chronic fibrosis of both animal models and human lungs. Thus, we propose to examine the following hypotheses in this proposal: (i) that increased lysine hydroxylation distinguishes "fibrotic collagen" from "normal collagen" in the lung; (ii) that this difference is mediated by increased activity of lysyl hydroxylase in the (pre)fibrotic lung; and (iii) that once crosslinked, "fibrotic collagen" in the lung is not subject to turnover or breakdown, but rather will persist for the lifetime of the host. A combination of in vivo labelling and tissue slice and homogenate experiments will be performed to examine these questions. Correlative studies with human lung tissue, as available, from patients with acute (ARDS, IRDS) and chronic (IPF) lung fibrosis will examine the underlying mechanisms of collagen accumulation in human lung disease, and will perhaps identify potential new targets for rational therapy of these diseases.