Impact of versican deficiency on the innate immune response to influenza virus

项目总结 我们发表的工作和使用小鼠模型和体外细胞培养的初步数据表明 细胞外基质(ECM)分子，如通西康，暴露于病毒后在肺内积聚 和细菌。在入侵和保留的同时，还会产生一种专门的富含杂西卡的细胞外基质。 在对流感病毒的免疫反应过程中，肺内白细胞的数量增加。我们发现有两个细胞 Toll样受体(TLR)激动剂：巨噬细胞和 成纤维细胞。研究发现，在这两个细胞中，有两条不同的信号通路指导着Verscan的合成 类型-β-连环蛋白/T细胞因子(Tcf)途径在间质细胞中，如平滑肌细胞和 巨噬细胞中的成纤维细胞和TLR/Trif/I型干扰素受体途径。然而， 这些信号通路的细胞特异性尚不清楚。使用两个新的遗传菌株 我们发表的基因工程小鼠在细胞和时间特定的方式中删除了变态基因 初步数据显示，凡西康对白细胞表型有显著影响。在接受治疗时 使用多肌苷：多胞苷(Poly(I：C))，凡西卡缺乏的小鼠的数量减少 支气管肺泡液中白细胞的恢复表明维西康具有促炎和抗炎作用 是白细胞迁移到肺部所必需的。相比之下，巨噬细胞中缺乏维西康的小鼠 BAL液中白细胞回收率增加，提示巨噬细胞来源的Versican具有抗 发炎，抑制白细胞向肺部迁移。两个品系的维西卡基因缺陷小鼠都有 肺组织或BAL液中I型IFN和IL-10的回收率显著低于 控制。我们的体外研究将此归因于这些抗炎细胞因子的减少 缺乏维斯康星的巨噬细胞。在这些新的杂交种缺陷小鼠身上观察到的差异是 我们的中心假设的基础是巨噬细胞和 基质细胞为先天性免疫反应提供关键的上下文线索和细胞外控制 病毒性肺部感染。我们提出了三个目的来检验这一假设。目标1定义了凡西嘉人的角色 来源于巨噬细胞和/或I型干扰素信号，提供细胞外对固有的控制 流感病毒感染小鼠肺组织的免疫反应。目标2确定基质细胞和/或β- 连环蛋白/TCF信号促进富含万西肯的细胞外基质的产生，从而提供对 对流感病毒的先天免疫反应。目标3确定机制和信号通路 因此，巨噬细胞与成纤维细胞来源的Versican提供了上下文细胞外控制 先天免疫反应。了解Verscan提供精细控制的上下文设置 对流感病毒的先天免疫反应对于设计治疗策略至关重要 提高对病毒性肺部感染的免疫反应。

PROJECT SUMMARY Our published work and preliminary data using mouse models and in vitro cell cultures show that extracellular matrix (ECM) molecules such as versican accumulate in lungs following exposure to viruses and bacteria. The creation of a specialized versican-enriched ECM coincides with invasion and retention of leukocytes in lungs during the immune response to influenza virus. We have found that two cellular sources synthesize versican in response to Toll-like receptor (TLR) agonists: macrophages and fibroblasts. Studies identify two distinct signaling pathways directing versican synthesis in these two cell types - the β-catenin/T-cell factor (TCF) pathway in stromal cells such as smooth muscle cells and fibroblasts and the TLR/Trif/Type I interferon (IFN) receptor pathway in macrophages. However, the cellular specificity of these signaling pathways is not known. Using two novel strains of genetically engineered mice in which the versican gene is deleted in a cell- and time-specific manner, our published and preliminary data shows that versican has a dramatic effect on leukocyte phenotype. When treated with polyinosinic:polycytidylic acid (poly(I:C)), mice with a global deficiency in versican have decreased recovery of leukocytes in bronchoalveolar (BAL) fluid suggesting that versican is pro-inflammatory and required for leukocyte migration into lungs. In contrast, mice lacking versican in macrophages have increased recovery of leukocytes in BAL fluid suggesting that macrophage-derived versican is anti- inflammatory and suppresses leukocyte migration into lungs. Both strains of versican-deficient mice had significantly decreased recovery of Type I IFNs and IL-10 in lung tissue or BAL fluid when compared to controls. Our in vitro studies attribute this to decreased production of these anti-inflammatory cytokines by versican-deficient macrophages. The differences observed in these novel versican-deficient mice are the basis of our central hypothesis that the formation of a versican-enriched ECM by macrophages and stromal cells provides critical contextual cues and extracellular-control of the innate immune response to viral lung infection. We propose three aims to test this hypothesis. Aim 1 defines the role of versican derived from macrophages and/or Type I IFN signaling in providing extracellular-control of the innate immune response in lungs of mice exposed to influenza virus. Aim 2 determines if stromal cells and/or β- catenin/TCF signaling promote the generation of a versican-enriched ECM that provides fine-control of the innate immune response to influenza virus. Aim 3 identifies the mechanisms and signaling pathways whereby macrophage- versus fibroblast-derived versican provide contextual extracellular-control of the innate immune response. Understanding the contextual settings in which versican provides fine-control of the innate immune response to influenza virus is critical for the design of therapeutic strategies for improving the immune response to viral lung infection.