Impact of Airway Epithelium on Innate and Adaptive Immunity in the Lung

研究项目2 呼吸道病毒，例如呼吸道合胞病毒（RSV），是哮喘恶化的主要触发因素 儿童和成人。该项目的首要假设是气道上皮细胞（AEC）协调 对呼吸道病毒感染和空气过敏原的反应。此外，AECs之间的内在差异 哮喘和健康儿童在感染和暴露于 过敏原我们推测，RSV或HRV哮喘患儿的AEC感染， 与过敏原暴露组合，诱导固有细胞，包括树突状细胞（DC）， 先天性淋巴样细胞（ILC），以触发2型反应。我们将使用分离的AEC来测试这个想法， 哮喘和健康儿童，生长在空气/液体界面（ALI）和感染呼吸道病毒， 过敏原激发背景。我们将检测与这些AEC共培养的DC和ILC的反应， 它们通过以下目的在促进2型炎症中的作用： 目标1.检验AEC控制局部DC成熟和功能的假设。由于AECs在 哮喘患者可能由于屏障缺陷而“渗漏”，我们将首先确定是否有更多的抗原通过 哮喘的AEC比健康的AEC对使用修饰的RSV株的DC的敏感性更高。然后，我们将测试哮喘患者的AEC是否 儿童在促进DC分化和功能方面存在差异，以及病毒感染和过敏原暴露如何影响 这种互动。最后，我们将评估暴露于哮喘儿童AEC的DC驱动CD 4 + T细胞的能力。 共培养测定中的T细胞分化和增殖。 目标2.检验假设，过敏原暴露和病毒感染后，哮喘和健康 AEC通过脂质介质和血管活性肠肽对肺ILC 2产生不同的影响。我们将 评估AEC在调节ILC 2中的作用，ILC 2是肺中最丰富的ILCs。我们希望能找到 哮喘儿童的AEC在病毒感染和过敏原攻击期间增强ILC 2功能。在我们 初步数据，我们发现两个不同的人肺ILC 2亚群，差异表达受体， 氧化固醇、视黄酸和血管活性肠肽。因此，我们建议确定这些影响的影响， 和神经肽介质对不同ILC 2群体的激活的影响。 目标3.检验ICOS+ ILC调节肺免疫应答的假设。我们创作了这部小说 观察到肺包含不寻常的ICOS+ ILC亚组。这些细胞不适合ILC 1，ILC 2， ILC 3范例，表达调节性淋巴细胞的标志物，并富含免疫抑制性免疫调节因子。 与环境相关。因此，我们假设这些细胞可能具有调节功能。为了验证这个，我们 提出分离肺ICOS+ ILC并定义其离体转录组谱以及其功能 在体外的能力，包括细胞因子和趋化因子分泌。此外，我们建议测试他们的 对源自暴露于HDM和/或病毒的ALI培养物的刺激的反应性。

PROJECT SUMMARY RESEARCH PROJECT 2 Respiratory viruses, such as respiratory syncytial virus (RSV), are major triggers of asthma exacerbation in children and adults. The overarching hypothesis of this project is that airway epithelial cells (AECs) coordinate responses to respiratory virus infection and aeroallergens. Moreover, intrinsic differences between AECs of asthmatic and healthy children change both innate and adaptive responses during infection and exposure to allergens. We hypothesize that infection of AECs from asthmatic children with RSV or HRV, in combination with allergen exposure, induces resident innate cells, including dendritic cells (DCs) and innate lymphoid cells (ILCs), to trigger type-2 responses. We will test this idea using AECs isolated from asthmatic and healthy children, grown at air/liquid interface (ALI) and infected with respiratory viruses in the context of allergen challenge. We will examine the response of DCs and ILCs co-cultured with these AECs, and their role in promoting type-2 inflammation through the following aims: Aim 1. Test hypothesis that AECs control local DC maturation and function. Since AECs in asthmatics may be “leaky” due to barrier defects, we will first determine whether more antigen is transferred by asthmatic than healthy AECs to DCs using modified RSV strains. Then, we will test whether AECs from asthmatic children differ in promoting DC differentiation and function, and how virus infection and allergen exposure impact this interaction. Finally, we will assess the ability of DCs exposed to AECs from asthmatic children to drive CD4 T cell differentiation and proliferation in co-culture assays. Aim 2. Test hypothesis that after allergen exposure and viral infections, asthmatic and healthy AECs differentially impact lung ILC2 through lipid mediators and vasoactive intestinal peptide. We will assess the role of AECs in regulating ILC2s, which are the most abundant ILCs in the lung. We expect to find that AECs from asthmatic children enhance ILC2 function during viral infection and allergen challenge. In our preliminary data, we found two distinct subsets of human lung ILC2s that differentially express receptors for oxysterols, retinoic acid, and vasoactive intestinal peptide. Thus, we propose to define the impact of these lipidic and neuropeptide mediators on the activation of different ILC2 populations. Aim 3. Test hypothesis that ICOS+ ILCs regulate lung immune responses. We made the novel observation that the lung contains an unusual subset of ICOS+ ILCs. These cells do not fit into the ILC1, ILC2, ILC3 paradigm, express a marker of regulatory lymphocytes and are enriched in an immunosuppressive environment associated. Thus, we hypothesize that these cells may have regulatory functions. To test this, we propose to isolate lung ICOS+ ILCs and define their transcriptome profile ex vivo as well as their functional capabilities in vitro, including cytokine and chemokine secretion. Moreover, we propose to test their responsiveness to stimuli derived from ALI cultures exposed to HDM and/or viruses.