Nicotine-induced fibronectin expression in lung injury and repair

描述(由申请人提供)： 摘要：烟碱型乙酰胆碱受体(NAChRs)是由神经细胞和非神经细胞表达的具有信号转导功能的五聚体通道。虽然关于nAChRs功能的大部分已知与神经系统有关，但关于它们在非神经系统中的作用的信息正在迅速涌现。NAChRs在肺中表达，在非人类灵长类动物中进行的有趣的工作表明，尼古丁激活nAChR会导致肺结构/功能异常。我们探索了nAChR激活导致基质表达的机制，发现这种作用是通过nAChRs的一个特定亚群介导的，称为17个nAChRs。当这些受体在肺成纤维细胞中被激活时，我们观察到MEK-1/Erk通路的诱导和转录因子CREB的磷酸化。反过来，CREB刺激纤维连接蛋白的表达，纤维连接蛋白是一种与肺损伤和修复有关的基质糖蛋白。这些事件被17个nAChR拮抗剂阻断，而在17个nAChR KO原代肺成纤维细胞中不存在。此外，我们还发现nAChR的激活通过MEK-1/Erk依赖的途径刺激肺成纤维细胞的增殖。在长期受尼古丁刺激的小鼠中也有类似的观察，但在发育中的和成年的17只nAChR KO动物中则没有。最近，我们记录了暴露于香烟烟雾提取物的成纤维细胞中纤维连接蛋白的增殖和表达增加，并观察到烟草烟雾暴露的小鼠肺中纤维连接蛋白蛋白沉积增加，证实了这一过程与烟草相关疾病的相关性。我们还发现，在长期接触尼古丁的背景下，博莱霉素诱导的肺损伤后，肺修复机制似乎受到了损害。这些数据揭示了非神经性nAChRs在控制肺组织重塑中的新作用。在此基础上，我们推测肺组织中17nAChR的激活通过MEK-1/Erk依赖的信号刺激成纤维细胞的增殖及其纤维连接蛋白等基质成分的过度表达，从而促进组织重塑。在基线时，这会导致基质成分的细微变化，而不会导致纤维化等明显后果。相反，在肺损伤的背景下，这会导致组织重建的夸大，从而促进低效修复和纤维化。这将在特定的目标下进行测试，目的是：1)检测17nAChRs在肺成纤维细胞的增殖、肌成纤维细胞转分化和基质表达中的作用，以及MEK-1/Erk在野生型和17nAChR KO肺成纤维细胞中介导这些事件的作用；2)在两种肺损伤模型(博莱霉素模型和异种气管移植模型)中，检测17 nAChRs在肺损伤和修复中的作用；3)检测17 nAChR持续激活在肺损伤后修复中的作用。 公共卫生相关性： 项目简介本项目将探索损伤后肺组织重塑的细胞机制。组织重塑是以结缔组织成分(也称为细胞外基质)的表达、沉积和降解的变化为特征的过程。这对现役战斗人员很重要，但对退伍军人也很重要，因为许多肺部疾病(如慢性阻塞性肺病、肺纤维化)通常出现在这一人群中，与组织重塑的变化有关。有趣的是，烟草及其成分被认为与许多这些疾病有关，因此，人们认为它们促进了组织重塑的改变。但是，导致这些事件的机制尚不清楚。该项目将通过重点研究尼古丁受体来探索这些过程中涉及的机制，以及这些受体的激活如何促进肺细胞外基质相对组成的变化。

DESCRIPTION (provided by applicant): ABSTRACT: Nicotinic acetylcholine receptors (nAChRs) are pentameric channels expressed by neuronal and non-neuronal cells that are capable of signal transduction. Although much of what is known about the function of nAChRs relates to the nervous system, information about their roles in non-neuronal systems is rapidly emerging. nAChRs are expressed in lung and interesting work performed in non-human primates suggests that nAChR activation by nicotine leads to pulmonary structural/functional abnormalities. We explored the mechanisms by which nAChR activation leads to matrix expression and found that this effect is mediated through a specific subgroup of nAChRs termed 17 nAChRs. Upon activation of these receptors in lung fibroblasts, we observed induction of the MEK-1/Erk pathway and phosphorylation of the transcription factor CREB. In turn, CREB stimulated the expression of fibronectin, a matrix glycoprotein implicated in lung injury and repair. These events were blocked by 17 nAChR antagonists and were not present in 17 nAChR KO primary lung fibroblasts. In addition, we found that nAChR activation stimulates lung fibroblast proliferation through MEK-1/Erk-dependent pathways. Similar observations were made in mice chronically stimulated with nicotine, but not in developing and adult 17 nAChR KO animals. More recently, we documented an increase in the proliferation and expression of fibronectin in fibroblasts exposed to cigarette smoke extract, and observed an increase in fibronectin protein deposition in the lungs of mice exposed to tobacco smoke confirming the relevance of this process to tobacco-related disease. We also found that lung repair mechanisms appeared to be impaired after bleomycin-induced lung injury in the setting of chronic exposure to nicotine. These data unveil a novel role for non-neuronal nAChRs in control of lung tissue remodeling. Based on the above, we hypothesize that 17 nAChR activation in lung promotes tissue remodeling by stimulating the proliferation of fibroblasts and their overexpression of matrix components like fibronectin through MEK-1/Erk-dependent signals. At baseline, this leads to subtle alterations in matrix composition without causing overt consequences such as fibrosis. In contrast, in the setting of lung injury, this leads to exaggerated tissue remodeling that promotes inefficient repair and fibrosis. This will be tested in specific aims designed to: 1) Examine the role of 17 nAChRs in the proliferation, myofibroblast transdifferentiation, and matrix expression of lung fibroblasts, and the role of MEK-1/Erk in mediating these events in wildtype vs. 17 nAChR KO lung fibroblasts, 2) Examine the role of 17 nAChRs in lung injury and repair using wildtype and 17 nAChR KO mice in two models of lung injury (bleomycin model and the model of heterologous tracheal transplantation), and 3) Examine the effect of persistent 17 nAChR activation in lung repair after injury. PUBLIC HEALTH RELEVANCE: Project Narrative This project will explore the cellular mechanisms involved in tissue remodeling of the lung after injury. Tissue remodeling is a process characterized by alterations in the expression, deposition, and degradation of connective tissue components (also called extracellular matrices). This is important to active combatants but also to veterans in view that many pulmonary disorders typically seen in this population (e.g., COPD, lung fibrosis) that are associated with alterations in tissue remodeling. Interestingly, tobacco and its components have been implicated in many of these disorders and, therefore, it is thought that they promote alterations in tissue remodeling. But, the mechanisms responsible for these events are unknown. This project will explore the mechanisms involved in these processes by focusing on nicotinic receptors and how activation of these receptors promotes alterations in the relative composition of the lung extracellular matrix.