REGULATION OF INNATE IMMUNITY IN HUMAN METAPNEUMOVIRUS INFECTION

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 人类偏肺病毒(HMPV)--新发现的人类偏肺病毒 副粘病毒科，介导严重的下呼吸道 幼儿、老年人和儿童的肠道疾病 免疫功能低下的患者(1-3)。HMPV会导致毛细支气管炎， 肺炎和流感样症状，以及哮喘 恶化，它被认为是一种实质性的公共卫生 有问题。自2001年发现以来，许多研究提供了 更好地了解与此相关的流行病学 病毒感染。然而，关于 人乳头瘤病毒所致肺部疾病的发病机制及宿主免疫 人们对此的反应还没有完全理解。此外，没有治疗或 目前已有针对hMPV的疫苗。肺部先天免疫 系统是主机防御的关键组件 对抗病毒，并在细胞水平上通过 转录因子的激活调节蛋白的表达 具有抗病毒和/或炎症活性的可诱导基因产物。 开发治疗hMPV感染的新疗法的关键在于- 深入了解人乳头瘤病毒病的发病机制 导致感染或感染的病毒-宿主细胞相互作用的性质 杀死病毒。在这方面，研究中的互动 HMPV和先天免疫细胞之间的关系是特别有趣的。 I型干扰素是天然干扰素的重要组成部分。 免疫系统不仅负责宽广的细胞 对病毒感染的抗菌活性，也是对其 在连接先天免疫反应和获得性免疫反应中的作用(4)。使用 HMPV感染小鼠实验模型(5，6)及原代 人细胞体外实验(7)，我已经证明了I型干扰素的作用 在调节hMPV致病过程中起着基础性作用。更多 重要的是，hMPV干扰Toll样受体(TLR)- 体外(7，8)和体内(5)诱导产生I型干扰素。 然而，I型纤溶酶原激活剂的激活和调节机制 干扰素由 HMPV感染的机制尚未阐明。我们最重要的是 假设hMPV激活不同的先天信号 抗原提呈细胞中的途径，从而触发免疫 导致有效清除病毒和消除 这种疾病。这一假设是基于hMPV的发现 激活人浆细胞样树突状细胞产生I型干扰素 细胞(PDC)通过TLR7/8/9所在的内体隔室 可以被激活。此外，胞浆蛋白MDA5是 人髓系树突状细胞产生I型干扰素的必要条件 (初步数据)。在这项拨款中，我们建议确定 人巨噬细胞病毒激活TLR依赖的机制(S) 体外和体内的独立细胞信号转导。我们将实现 这些目标通过以下具体目标实现： 目的1.明确hMPV感染的分子机制(S) 诱导人原代细胞产生I型干扰素。我们会 研究hMPV如何激活TLR信号通路诱导 PDC中的I型干扰素及其hMPV激活的途径 MODC和单核细胞。 目的2.探讨肺巨噬细胞和树突状细胞在急性肺损伤中的作用。 HMPV感染时I型干扰素的产生。我们将调查 宏页和DC在生产中的相对贡献 用I型干扰素体内感染的小鼠作为实验模型。 此外，我们还将确定肺脏的贡献 巨噬细胞和树突状细胞在hMPV发病中的作用。 目的3.探讨TLR依赖的生物学相关性 体内hMPV感染的独立途径。我们会 HMPV对小鼠的毒力和致病性研究 TLR7、MyD88、TLR7、MyD88、TLR7基因缺陷转基因小鼠感染模型的建立 和MDA5的表达。我们将评估病毒复制，临床 疾病、肺功能和炎症，以及 细胞因子/趋化因子/干扰素的表达。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Human metapneumovirus (hMPV), a newly-recognized member of the Paramyxoviridae family, mediate serious lower respiratory tract illness in very young children, the elderly and immunocompromised patients (1-3). hMPV causes bronchiolitis, pneumonia and flu-like syndromes, as well as asthma exacerbations, and it is considered a substantial public health problem. Since its discovery in 2001, many studies have provided a better understanding of the epidemiology associated with this viral infection. However, fundamental questions regarding the pathogenesis of hMPV-induced lung disease and the host immune response are not fully understood. Furthermore, no treatments or vaccines for hMPV are currently available. The lung innate immune system represents a critical component of the host defense against viruses and is coordinated at the cellular level by activation of transcription factors that regulate the expression of inducible gene products with antiviral and/or inflammatory activity. Key to the development of novel therapies for hMPV infection is in- depth understanding of hMPV disease pathogenesis and the nature of virus-host cell interactions that lead to either infection or killing of the virus. In this respect, the study in the interaction between hMPV and innate immune cells are of particular interest. Type I Interferon (IFN) is known as key component of the innate immune system responsible not only for broad cellular antimicrobial activity in response to viral infections, but also for its role in linking innate and adaptive immune response (4). Using an experimental mouse model of hMPV infection (5,6) and primary human cells in vitro (7), I have demonstrated that type I IFN plays a fundamental role in regulating hMPV pathogenesis. More importantly, hMPV interferes with the Toll-like receptor (TLR)- induced type I IFN production in vitro (7,8) and in vivo (5). However, the mechanisms of activation and regulation of type I IFN by hMPV infection have not yet been elucidated. Our overarching hypothesis is that hMPV activates distinct innate signaling pathways in antigen presenting cells, thus triggers immune responses that lead to efficient viral clearance and elimination of the disease. This hypothesis is based on the discovery that hMPV activates type I IFN production in human plasmacytoid dendritic cells (pDC) through the endosomal compartment where TLR7/8/9 can be activated. In addition, the cytosolic protein MDA5 is necessary for the production of type I IFN in human myeloid DC (preliminary data). In this grant, we propose to identify the mechanism(s) by which hMPV activates TLR-dependent and - independent cellular signaling in vitro and in vivo. We will achieve these objectives through the following specific aims: Aim 1. To define the molecular mechanism(s) by which hMPV induces type I IFN production in primary human cells. We will investigate how hMPV activates TLR signaling pathway to induce type I IFN in pDC and which pathway is activated by hMPV in moDC and monocytes. Aim 2. To determine the role of lung macrophages and DC in the production of type I IFN in hMPV infection. We will investigate the relative contribution of macropages and DC in the production of type I IFN in vivo using an experimental mouse model of infection. Furthermore, we will determine the contribution of pulmonary macrophages and DC in hMPV pathogenesis. Aim 3. To investigate the biological relevance of TLR-dependent and independent pathways in hMPV infection in vivo. We will characterize the virulence and pathogenicity of hMPV in mouse model of infection using transgenic mice deficient in TLR7, MyD88, and MDA5 expression. We will assess viral replication, clinical disease, lung function and inflammation, and cytokine/chemokine/IFN expression.