Innate Immune Mechanisms in Emphysema

项目总结/摘要 Toll样受体（TLR）已经在病原体挑战的背景下被广泛研究，但它们在病原体挑战中的作用仍然存在。 未感染的肺不清楚。阐明了肺维持结构完整性的机制， 面对复杂的外部和内部环境，对我们理解肺部疾病至关重要。 肺气肿的特征是肺弹性逐渐丧失和不可逆的空域扩大， 在一定程度上，在正常的老化过程中，但病理加速与慢性暴露于 污染物或香烟烟雾。事实上，只有10%到20%的吸烟者会出现临床上显著的 肺气肿表明其他因素对其发病机制也很重要。最近的人体研究表明， TLR4功能降低与COPD的严重程度相关，这使我们的发现与人类相关 TLR4-/-小鼠在2个月大后发生自发性肺气肿，此时肺的发育是 完成.我们发现TLR4缺乏诱导了一种新的NADPH氧化酶（Nox），Nox 3和过量的氧化剂 这也是人类肺气肿的一个特征。用Nox处理TLR4-/-小鼠或肺内皮细胞 抑制剂或Nox3 siRNA逆转表型。我们建立了可诱导的肺靶向Nox3转基因小鼠， 在COPD患者中也检测到Nox3水平升高，这表明 Nox 3在体内肺气肿发病机制中的重要作用。为了研究细胞类型 负责这些TLR4介导的反应，我们产生了骨髓嵌合体，发现TLR4 在非造血细胞上的表达是维持正常肺结构所必需的。TLR4的重建 对气道上皮细胞TLR4-/-的作用仅部分阻止肺气肿，并不抑制Nox3的表达 或氧化剂的产生，表明TLR4在替代结构细胞中也参与调节肺 氧化剂我们使用原代肺内皮细胞的数据指出内皮TLR4的重要作用， 我们会在这个建议中探讨。我们的假设是，内皮细胞TLR4通过抑制 Nox3介导的氧化剂生成。我们将在以下目标中测试这一假设：1）确定特定的 内皮细胞TLR4在体内预防氧化剂介导的肺气肿中的作用，2）阐明 TLR4抑制内皮细胞中Nox 3基因表达的机制，以及3）表征TLR4 参与抑制Nox 3表达和预防体内氧化剂介导的肺气肿的配体， 肺内皮细胞

PROJECT SUMMARY/ABSTRACT Toll-like receptors (TLRs) have been studied extensively in the context of pathogen challenges, yet their role in the unchallenged lung is unclear. Elucidating the mechanisms whereby the lung maintains structural integrity in the face of complex external and internal environments is essential to our understanding of lung disease. Emphysema is characterized by gradual loss of lung elasticity and irreversible airspace enlargement, occurring to a certain extent in the normal aging process but pathologically accelerated with chronic exposure to pollutants or cigarette smoke. The fact that only 10 to 20% of smokers develop clinically significant emphysema indicates that other factors are important to its pathogenesis. Recent human studies indicate that depressed TLR4 function is correlated with the severity of COPD, which lends human relevance to our finding that TLR4-/- mice develop spontaneous emphysema after 2 months of age, when lung development is complete. We found that TLR4 deficiency induces a novel NADPH oxidase (Nox), Nox3, and excessive oxidant generation, also a feature of human emphysema. Treatment of TLR4-/- mice or lung endothelial cells with Nox inhibitors or Nox3 siRNA reversed the phenotype. We generated inducible, lung-targeted Nox3 transgenic mice that develop emphysema and also detected elevated Nox3 levels in patients with COPD, indicating an important role for Nox3 in the pathogenesis of emphysema in vivo. In order to investigate the cell type responsible for these TLR4-mediated responses, we generated bone marrow chimeras and found that TLR4 expression on non-hematopoietic cells is required to maintain normal lung architecture. Reconstitution of TLR4 to the airway epithelium only partially prevented emphysema in TLR4-/- and did not suppress Nox3 expression or oxidant generation, indicating that TLR4 in an alternative structural cell is also involved in regulating lung oxidants. Our data using primary lung endothelial cells point to an important role for endothelial TLR4, which we will explore in this proposal. Our hypothesis is that endothelial TLR4 maintains lung integrity by suppressing Nox3-mediated oxidant generation. We will test this hypothesis in the following Aims: 1) Determine the specific contribution of endothelial TLR4 in preventing oxidant-mediated emphysema in vivo, 2) Elucidate the mechanisms whereby TLR4 inhibits Nox3 gene expression in endothelial cells, and 3) Characterize the TLR4 ligand(s) involved in inhibiting Nox3 expression and preventing oxidant-mediated emphysema in vivo and in lung endothelial cells.