Regulation of Lung Fibrosis by Alternatively Activated Macrophages and Arginase-1

描述（由申请人提供）：本提案研究的目标是提供支持和持续指导，以促进申请人发展成为间质性肺病和肺免疫学的独立科学研究者。这将是完成 通过课程设计，以获得一个坚实的基础科学背景，涉及实验室经验，定期教学，经常演示，并通过成功的物理学家，科学家密切指导手。该提案将为申请人提供至少75%的受保护研究时间。利用研究提案作为平台，申请人将获得许多基础科学技术的专业知识，包括：小鼠纤维化肺病的分析，细胞特异性敲除小鼠的产生以及巨噬细胞和成纤维细胞功能的评估。这项工作将涉及一个充满活力的研究小组，在整个大学环境中发挥作用，促进尖端研究。研究项目：肺纤维化疾病是发病率和死亡率的重要原因，缺乏有效的治疗方法。调节持续性肺纤维化的机制尚不完全清楚。我们的实验室最近描述了一个严重的和进行性纤维化的模型，以应对非感染性肺损伤。CXCR 3是IFN-γ诱导的趋化因子CXCL 9（Mig）、CXCL 10（IP-10）和CXCL 11（ITAC）的受体。这些趋化因子被认为是Th 1免疫的重要介质。相对于野生型对照小鼠，CXCR 3缺失小鼠在用气管内博莱霉素治疗后发展严重和进行性纤维化。微阵列分析已经鉴定了在CXCR 3不存在的情况下表达的与Th 2免疫相关的基因的优势。此外，我们确定了一些基因，已与交替激活的巨噬细胞表型。这些包括甘露糖受体，YM 1和清道夫受体。最突出的基因与一个交替激活的巨噬细胞表型是一个36倍的诱导，在β-内酰胺酶-1。虽然交替活化的巨噬细胞已涉及介导免疫介导的组织纤维化，但它们在非免疫调节过程中的作用尚不清楚。精氨酸酶-1由响应于非感染性肺损伤的交替活化的巨噬细胞和成纤维细胞产生。它有可能直接介导纤维化。精氨酸酶-1负责将L-精氨酸转化为脯氨酸和多胺。多胺参与细胞增殖，而脯氨酸是胶原的直接前体。巨噬细胞和成纤维细胞表达β-内酰胺酶-1在进行性纤维化中的作用尚未研究。基于我们的初步数据，我们假设巨噬细胞和成纤维细胞中的α-淀粉酶-1表达对非感染性纤维化肺损伤后纤维化的发展至关重要。我们将在以下具体目标中测试该假设：目的1：使用区分招募和常驻群体的新的流式细胞术方法来定义在非感染性肺损伤后CXCR 3缺失小鼠中招募到肺的特定巨噬细胞表型。目标2：使用floxed arg-1等位基因与靶向巨噬细胞或成纤维细胞的cre重组酶菌株杂交，确定非感染性损伤后巨噬细胞和成纤维细胞中cre-1靶向缺失的影响。总的来说，这种方法将使我们能够剖析巨噬细胞和成纤维细胞在进行性肺纤维化的病理生物学中的作用。

DESCRIPTION (provided by applicant): The goal of this proposal research is to provide support and continued mentorship to facilitate the applicant's development into an independent scientific investigator in interstitial lung disease and lung immunology. This will be accomplished through a curriculum designed to obtain a solid basic science background involving hands on laboratory experience, regular didactic teaching, frequent presentations, and close mentoring by successful physician-scientists. The proposal will allow at least 75% of protected research time for the applicant. Using the research proposal as a platform, the applicant will gain expertise in number of fundamental scientific techniques including: the analysis of murine fibrotic lung disease, the generation of cell specific knockout mice and the assessment of macrophage and fibroblast function. This effort will involve a dynamic research group which functions in an overal university environment fostering cutting edge research. Research Project: Fibrotic lung diseases are a significant cause of morbidity and mortality for which effective therapies are lacking. The mechanisms that regulate unremitting pulmonary fibrosis are incompletely understood. Our laboratory has recently described a model of severe and progressive fibrosis in response to non-infectious lung injury. CXCR3 is the receptor for the IFNg-inducible chemokines CXCL9 (Mig), CXCL10 (IP-10) and CXCL11 (ITAC). These chemokines have been suggested to be important mediators of Th1 immunity. CXCR3 null mice develop severe and progressive fibrosis following treatment with intratracheal bleomycin relative to wild type control mice. Microarray analysis has identified a preponderance of genes associated with Th2 immunity expressed in the absence of CXCR3. In addition, we identified a number of genes that have been associated with the alternatively activated macrophage phenotype. These include Mannose Receptor, YM1 and Scavenger Receptor. The most prominent of the genes associated with an alternatively activated macrophage phenotype was a 36-fold induction in arginase-1. While alternatively activated macrophages have been implicated in mediating immunologically mediated tissue fibrosis, their roles in non-immune regulated processes are less clear. Arginase-1 is produced both by alternatively activated macrophages and fibroblasts in response to non-infectious lung injury. It has the potential to directly mediate fibrogenesis. Arginase-1 is responsible for the conversion of L-arginine to proline and polyamines. Polyamines are involved in cell proliferation while proline is a direct precursor of collagen. The role of arginase-1 expression by macrophages and fibroblasts in progressive fibrosis has not been explored. Based on our preliminary data, we hypothesize that arginase-1 expression in macrophages and fibroblasts is critical to the development of fibrosis after non-infectious fibrotic lung injury. We will test thi hypothesis in the following specific aims: AIM 1: Define the specific macrophage phenotypes recruited to the lung in CXCR3 null mice after non-infectious lung injury using a novel flow cytometric approach that distinguishes recruited and resident populations. AIM2: Determine the effects of targeted deletion of arginase-1 in macrophages and fibroblasts after non-infectious injury using a floxed arg-1 allele crossed with either a cre-recombinase strain that targets macrophages or fibroblasts. Collectively, this approach will allow us to dissect the role of arginase-1 production by macrophages and fibroblasts in the pathobiology of progressive pulmonary fibrosis.