Role of immunoproteasome in airway viral infection

项目概要/摘要 本研究的目的是确定免疫蛋白酶体（IP）在免疫过程中的调节和功能。 具有2型炎症的气道中的鼻病毒感染，2型炎症的标志（例如，白细胞介素13 [IL-13]）高 哮喘鼻病毒（RV）感染仍然是一个重大的医疗保健负担，因为它有助于加重 哮喘和其他肺部疾病。由于呼吸道RV感染尚无有效的治疗方法， 为了定义新颖的机制（例如，IP）从感染的气道细胞中清除RV（例如，上皮细胞）。我们发现 在伴有气道2型炎症的严重哮喘患者中，气道上皮细胞IP表达减少。 重要的是，我们的初步数据有力地表明，IP在人原代气道上皮细胞中的表达 RV（RV-A和RV-C）感染显著上调，并且敲除或抑制关键IP 组件（例如，LMP 2和LMP 7）增加气道上皮病毒载量。2型细胞因子IL-13显著 减少RV的IP诱导。RV感染的LMP 2敲除小鼠（与野生型小鼠相比）不能清除病毒 有效，并增加肺部炎症。我们将检验RV期间气道IP诱导 感染作为消除病毒感染的关键宿主防御机制，但在呼吸道中受损， 2型炎症，导致持续感染和过度炎症。在目标1中，我们将确定 干扰素（IFN）/IFN受体（IFNR）/STAT 1轴在RV调节IP和RV调节2型炎症中的作用 在人原代气道上皮细胞中进行基因敲除实验，并使用人的精确度- 切割的肺切片或基因（例如，IFNAR或STAT 1）缺陷小鼠。在目标2中， 我们将通过使用IP来确定IP在RV介导的哮喘急性发作中的作用和潜在机制， 缺陷型小鼠（全身或气道上皮条件基因敲除）和人原代气道上皮细胞 IP基因敲除的细胞。我们将测试IP的抗病毒和抗炎功能是否依赖于其调节 RV 3C蛋白酶和A20，一种负性免疫调节剂。在目标3中，我们将确定IP作为 抗病毒药物与2型炎症气道。通过递送单一低剂量的IFN-1或病毒模拟物（即， 将polyI：C）导入培养的人气道上皮或具有2型炎症的小鼠气道，我们将测试 增强IP表达促进气道急性RV感染和炎症的消退。研究成果 将显著提高我们对RV感染的宿主防御机制的理解， 为治疗哮喘和其他疾病患者的RV感染提供了新的机会。

Project Summary/Abstract The goal of this proposal is to determine the regulation and functions of immunoproteasome (IP) during rhinovirus infection in airways with type 2 inflammation, a hallmark of the type 2 (e.g., interleukin 13 [IL-13]) high asthma. Rhinovirus (RV) infection remains a significant healthcare burden due to its contribution to exacerbations of asthma and other lung diseases. Because there is no effective therapy for airway RV infection, it is imperative to define novel mechanisms (e.g., IP) that clear RV from the infected airway cells (e.g., epithelial cells). We found reduced IP expression in airway epithelial cells of severe asthma patients with airway type 2 inflammation. Importantly, our preliminary data strongly suggest that IP expression in human primary airway epithelial cells was significantly up-regulated by RV (RV-A and RV-C) infection, and that knockout or inhibition of key IP components (e.g., LMP2 and LMP7) increased airway epithelial viral load. Type 2 cytokine IL-13 significantly reduced IP induction by RV. RV-infected LMP2 knockout (vs. wild-type) mice were unable to clear the viruses effectively, and increased lung inflammation. We will test the hypothesis that airway IP induction during RV infection serves as a critical host defense mechanism to eliminate viral infection, but is impaired in airways with type 2 inflammation, leading to persistent infection and excessive inflammation. In Aim 1, we will determine the role of the interferons (IFNs)/IFN receptor (IFNR)/STAT1 axis in IP regulation by RV and type 2 inflammation by performing gene knockout experiments in human primary airway epithelial cells, and using the human precision- cut lung slices or gene (e.g., IFNAR or STAT1) deficient mice exposed to IL-13, allergen and/or RV. In Aim 2, we will determine the role of IP in RV-mediated asthma exacerbation and underlying mechanisms by using IP deficient mice (whole body or airway epithelial conditional gene knockout) and human primary airway epithelial cells with IP gene knockout. We will test if IP’s antiviral and anti-inflammatory functions depend on its regulation of RV 3C protease and A20, a negative immune regulator. In Aim 3, we will determine the efficacy of IP as an antiviral agent in airways with type 2 inflammation. By delivering a single low-dose of IFN-l or a viral mimic (i.e., polyI:C) into cultured human airway epithelium or mouse airways with type 2 inflammation, we will test if enhancing IP expression promotes resolution of airway acute RV infection and inflammation. Research findings will significantly improve our understanding about host defense mechanisms against RV infection, and likely provide a new opportunity to treat RV infections in patients with asthma and other diseases.