The pivotal role of macrophages in regulating pulmonary fibrosis

项目摘要/摘要 特发性肺纤维化(IPF)是一种进行性和致命性的过度胶原沉积的疾病，通过 成纤维细胞导致肺弹性降低，存活率低。尽管有持续的努力和新的 药物方面，在延缓疾病进展方面只取得了有限的效果。这很有可能 反映了疾病的复杂性质，包括重要的细胞间相互作用。最近，我们确定了危急情况 巨噬细胞和成纤维细胞之间的相互作用导致肺纤维化的发展。我们的 有资料表明，Toll样负调控因子--白细胞介素1受体相关激酶(IRAK)-M 人IPF患者巨噬细胞和小鼠巨噬细胞受体信号转导增强 博莱霉素诱导肺纤维化，并导致成纤维细胞胶原表达增强。使用我们的 在实验性的小鼠纤维化模型中，我们证明了IRAK-M缺陷的小鼠受到保护 肺纤维化，这种作用依赖于巨噬细胞中IRAK-M的表达。这样做的目的是 建议研究巨噬细胞表达IRAK-M调节血管内皮细胞生长的机制。 肺纤维化的发展。为了实现这一目标，我们提出了两个具体目标。在具体目标1中， 我们将研究IRAK-M在调节肺纤维化过程中单核细胞转运到肺中的作用。 肺巨噬细胞要么来源于胚胎的卵黄囊，要么来源于骨髓 循环中的单核细胞。我们将询问IRAK-M在使用领养来规范单核细胞贩运方面的作用 移植、竞争性骨髓移植模型和细胞特异性IRAK-M基因敲除小鼠。此外, 我们将研究IRAK-M在调节促纤维化基因和趋化因子CCR2表达中的作用 参与单核细胞募集的受体，使用博莱霉素攻击小鼠的巨噬细胞作为 以及来自IPF患者和正常供体对照的原代巨噬细胞。为了显示翻译相关性， 我们将使用我们特征化的人类样本来生成人源化的IPF小鼠模型，该模型使用来自IPF的细胞 病人。在特定目标2中，我们将确定IRAK-M在调节巨噬细胞功能中的作用，特别是 在肺纤维化期间，它们识别、吸收和降解肺内胶原的能力。我们将评估 IRAK-M在博莱霉素刺激后胶原碎片摄取中的作用。巨噬细胞和单核细胞 我们的小鼠模型以及从IPF患者和正常供体对照中分离出的细胞将 流式细胞分选，检测IRAK-M、胶原摄取受体和胶原的表达 降解酶的表达和活性。最后，我们将建立巨噬细胞-成纤维细胞共培养体系。 系统来研究这两种细胞类型相互作用的双向作用。这些研究 将为开发新的疾病生物标志物以及新的干预措施提供基础 基于特定的分子靶点，应该改善IPF患者的人类健康。

PROJECT SUMMARY/ABSTRACT Idiopathic Pulmonary Fibrosis (IPF) is a progressive and fatal disorder of excessive collagen deposition by fibroblasts resulting in reduced lung elasticity and poor survival. Despite ongoing efforts and new pharmacologic agents, only limited efficacy has been achieved to delay disease progression. This likely reflects the complex nature of the disease including vital cell-cell interactions. Recently, we identified critical interactions between macrophages and fibroblasts which led to the development of pulmonary fibrosis. Our data demonstrate that interleukin-1 receptor associated kinase (IRAK)-M, a negative regulator of Toll-like receptor signaling, was elevated in macrophages from human IPF patients and murine macrophages after bleomycin-induced pulmonary fibrosis and lead to enhanced collagen expression by fibroblasts. Using our experimental murine model of fibrosis, we demonstrate that mice deficient in IRAK-M are protected from pulmonary fibrosis and this effect is dependent on IRAK-M expression in macrophages. The goal of this proposal is to investigate the mechanism by which macrophage expression of IRAK-M regulates the development of pulmonary fibrosis. In order to accomplish this, we propose two specific aims. In specific aim 1, we will investigate the role of IRAK-M in regulating monocyte trafficking to the lung during pulmonary fibrosis. Lung macrophage are either derived embryonically from the yolk-sac or are bone marrow-derived from circulating monocytes. We will interrogate the role of IRAK-M in regulating monocyte trafficking using adoptive transfer, competitive bone marrow transplantation model, and cell-specific IRAK-M knockout mice. In addition, we will investigate the role of IRAK-M in regulating expression of profibrotic genes and CCR2, the chemokine receptor involved in monocyte recruitment, using macrophages isolated from bleomycin challenged mice as well as primary macrophages from IPF patients and normal, donor controls. To show translational relevance, we will use our characterized human samples to generate a humanized IPF mouse model with cells from IPF patients. In specific aim 2, we will determine the role of IRAK-M in regulating macrophage function, specifically their ability to identify, take up and degrade collagen in the lung during pulmonary fibrosis. We will assess the role of IRAK-M in the uptake of collagen fragments after bleomycin challenge. Macrophage and monocyte populations from our murine models as well as cells isolated from IPF patients and normal donor controls will be flow sorted and we will measure the expression of IRAK-M, collagen uptake receptors, and collagen degradation enzyme expression and activity. Finally, we will establish a macrophage-fibroblast co-culture system to investigate the bidirectional interactions that these two cell types exert on each other. These studies will provide the foundation for the development of novel biomarkers of disease as well as new interventions based on specific molecular targets that should improve the human health of IPF patients.