小气道和肺泡成纤维细胞在COPD基质转换中的区域性差异及其机制

慢性阻塞性肺疾病（COPD）是我国最大疾病负担之一，现有疗法不能有效延缓病情进展。COPD的病理特点是小气道壁重构和肺气肿，前者细胞外基质（ECM）沉积过多甚至纤维化，后者ECM修复不足，紧邻部位呈现相反的ECM转换，其成因不明。已知成纤维细胞是生成和调控ECM的重要细胞，我们的前期研究提示COPD外周肺组织成纤维细胞弹性纤维沉积障碍、对TGFβ1的反应异常，因此假设成纤维细胞在小气道壁和肺泡存在ECM转换的区域性差异，其机制在于TGFβ1信号转导通路。本项目拟用显微解剖技术分离COPD患者和大鼠小气道及肺泡成纤维细胞，建立体外细胞模型，从细胞迁徙、增殖、活化，ECM成分的合成、沉积、降解及对刺激的反应研究其区域性差异；用PCR芯片、蛋白磷酸化抗体芯片、双荧光素酶报告基因检测、siRNA及过表达质粒等技术研究TGFβ1信号转导的漂移及TGFβR3在其中的作用，为新的治疗靶点提供理论依据。

Chronic obstructive pulmonary disease (COPD) yields huge disease burden in China. Available therapies cannot effectively slow disease progression. Pathological features of COPD are small airway remodeling and emphysema; the former is caused by excessive extracellular matrix (ECM) deposition or even fibrosis, and the latter is owe to insufficient ECM repair. It is unclear why adjacent tissue in the same lung present opposite ECM turnover. Fibroblasts are important effector cells to generate ECM and regulate its turnover, and preliminary research suggests that fibroblasts isolated from the peripheral lung tissue of patients with COPD showed impaired repair function and weakened response to TGF-β1. Thus we postulate regional differences in ECM turnover of fibroblasts between small airways and alveoli, and that TGF-β1 signaling might be the potential molecular mechanisms. In this project, we plan to use microdissection technique to isolate primary fibroblasts from small airways and alveoli tissue of patients with COPD and in rat COPD models for further culture and study. In vitro cell models are to be set up by fibroblast/airway epithelial cell co-culture and fibroblast/typeⅡalveoli epithelial cell co-culture. The heterogeneity of fibroblasts at different compartments are to be investigated into details from the following aspects: (1) cell migration, proliferation, and activation; (2) synthesis, deposition and degradation of ECM components; and (3) the responses to exogenous stimuli including TGF-β1, tobacco smoke, lipopolysaccharide, interleukin-6, interleukin-8, macrophage conditioned media, theophylline and corticosteroid. TGFβ1 signal transduction axis drift and the role of TGFβR3 are then to be investigated using PCR chip, protein phosphorylation antibody chips, dual luciferase reporter gene assays, siRNA and overexpression plasmid technology. The data from these studies will help to understand the development of COPD better and find new potential therapeutic targets.