MMP28 Regulation Of Macrophage Recruitment and Polarization

摘要 基质金属蛋白酶(MMPs)是一类细胞外蛋白，在多种环境中发挥作用。 先天免疫的过程，如白细胞的运输。MMP28(Epetysin)是MMP28最新成员之一 与大多数基质金属蛋白酶不同，基质金属蛋白酶家族具有抑制炎症反应的功能。我们发现了 MMP28由肺上皮细胞、单核细胞表达，并募集CD11b高表达的肺巨噬细胞。 MMP28缺失(MMP28-/-)小鼠增加了早期巨噬细胞对肺和其他组织的募集， 肺和血清中促炎症标志物的增加，表明MMP28广泛地起到抑制作用 发炎。此外，我们还发现，Mmp28-/-巨噬细胞对趋化刺激的迁移速度更快， 在对内毒素或细菌的反应中分泌更多促炎细胞因子，提示MMP28参与了 以细胞自主的方式抑制早期巨噬细胞募集和促炎功能。我们 假设MMP28可能已经进化为巨噬细胞偏向于抗- 炎症表型，在炎症的后期阶段被招募。我们还假设这些 作用是通过调节受体，如趋化因子受体或其他黏附蛋白， 定位于单核/巨噬细胞表面。未发表的研究表明，MMP28表达减少 人类单核细胞中MMP28与吸烟者肺气肿的发生有关，我们发现MMP28-/- 会增加吸烟引起的肺气肿的易感性。在目标1中，我们将检验我们的假设 MMP28的缺失通过促进吸烟小鼠肺气肿的发生 巨噬细胞募集和更大的M1极化。我们将使野生型和Mmp28-/-小鼠暴露于慢性 香烟烟雾，并将在几个时间段表征单核细胞和巨噬细胞的数量和表型- 包括MMP28对弹性蛋白水解酶的调节。在目标2中，我们将测试我们的假设 巨噬细胞来源的MMP28是对肺气肿产生抵抗力的关键来源。要评估 巨噬细胞募集和髓系细胞MMP28表达的相对贡献 为了破坏肺部，我们计划产生骨髓嵌合体，并将它们暴露在长期吸烟的环境中 以及确凿的救援实验。在我们的第三个目标中，我们计划识别和验证新的MMP28 用演绎和无偏向法研究巨噬细胞募集和极化的底物 膜蛋白质组学方法。我们提供了几个候选底物的数据，这些数据将包括在 验证性研究和这些研究的结果将导致限制肺气肿的新机制。

ABSTRACT Matrix metalloproteinases (MMPs) comprise a family of extracellular proteinases that function in various processes of innate immunity, such as leukocyte trafficking. MMP28 (epilysin) is one of the newest members of the MMP family, and unlike most MMPs, it functions to dampen inflammatory responses. We have found MMP28 is expressed by lung epithelium, Ly6chigh monocytes, and recruited CD11bhigh pulmonary macrophages. MMP28-null (Mmp28-/-) mice have increased early macrophage recruitment to the lung and other tissues, with increased pro-inflammatory markers in the lung and serum, suggesting that broadly, MMP28 serves to inhibit inflammation. In addition, we have found that Mmp28-/- macrophages migrate faster to chemotactic stimuli and secrete more pro-inflammatory cytokines in response to LPS or bacteria, suggesting that MMP28 participates in inhibiting early macrophage recruitment and pro-inflammatory function in a cell-autonomous fashion. We hypothesize that MMP28 may have evolved as a mechanism by which macrophages, skewed towards an anti- inflammatory phenotype, are recruited during later stages of inflammation. We also hypothesize that these effects are mediated via modulation of a receptor, such as a chemokine receptor or other adhesion protein, localized to the monocyte/macrophage surface. Unpublished studies suggest that reduced MMP28 expression in human monocytes is associated with development of emphysema in smokers, and we show that Mmp28-/- have increased susceptibility to cigarette-smoke induced emphysema. In aim 1, we will test our hypothesis that absence of MMP28 promotes development of emphysema in smoke-exposed mice via enhanced macrophage recruitment and greater M1 polarization. We will expose wildtype and Mmp28-/- mice to chronic cigarette smoke, and will characterize monocyte and macrophage numbers and phenotypes during several time- points, including regulation of elastinolytic proteinases by MMP28. In aim 2, we will test our hypothesis that macrophage-derived MMP28 is the critical source that confers resistance to emphysema. To evaluate the relative contribution of MMP28-expression by epithelial cells vs. myeloid cells in macrophage recruitment and lung destruction, we plan to generate bone marrow chimeras and expose them to chronic cigarette smoke along with confirmatory rescue experiments. In our third aim, we plan to identify and validate novel MMP28 substrates involved in macrophage recruitment and polarization using deduction and unbiased approaches with membrane proteomics. We present data on several candidate substrates to be included in validation studies, and results of these studies will lead to novel mechanisms to limit emphysema.