Interferon regulation of the blood brain barrier during West Nile encephalitis

总结/摘要 本提案的目的是阐明I型干扰素（IFN）在血脑中的调节作用， 西尼罗河病毒（WNV）脑炎期间的血脑屏障（BBB）。西尼罗河病毒是一种蚊子传播的黄病毒，现在流行于 北美在过去的十年中，西尼罗河病毒在非洲造成了致命病毒性脑炎的日益流行， 美国的西尼罗河病毒的发病机制，特别是病原体如何浸润中枢神经系统 (CNS)，仍然知之甚少。在正常情况下，中枢神经系统受到保护，免受病原体的侵害。 血脑屏障是一个由脑微血管内皮细胞（BMEC）组成的界面，通过紧密的连接， 连接并由相邻的星形胶质细胞终足支持[1]。然而，西尼罗河病毒能够通过 未知的机制，并在神经元和其他实质CNS组织中建立感染[2，3]。CNS 感染导致免疫激活，其失调BBB，促进必需的抗病毒药物的进入。 白细胞运输到CNS;然而，受感染的白细胞运输到CNS也可能提供 西尼罗河病毒的神经侵袭的机会[4，5]。此外，虽然trans-BBB免疫运输是必要的， CNS病毒清除、CNS对损伤的脆弱性及其有限的修复能力可导致 在抗病毒免疫应答期间显著的旁观者损伤和免疫病理学。因此，BBB必须 在CNS感染期间受到严格调控，在促进抗病毒白细胞进入部位 感染和保护中枢神经系统免受免疫介导的损伤和潜在的病原体暴露， 循环病毒和受感染的宿主免疫细胞。 抗病毒细胞因子信号传导是实现这种平衡的可能机制。作为主要 WNV的全身免疫应答的组分，I型IFN家族的先天免疫细胞因子 在保护CNS和外周组织免受感染方面至关重要， 活性和促进适应性免疫应答[6-8]。有趣的是，I型干扰素还被 显示在CNS自身免疫的情况下在BBB发挥抗炎特性，增强 内皮屏障的完整性和阻碍白细胞结合内皮细胞和迁移到 CNS [9，10]。然而，在病毒性脑炎期间，I型IFN在BBB中的作用尚未被证实。 研究了我们实验室使用体外血脑屏障模型的初步研究表明， WNV感染后BMEC和星形胶质细胞中I型IFN信号的诱导导致增强的 内皮屏障功能和减少的跨内皮病毒和免疫运输。在此基础上， 体外研究结果，我们建议使用一个建立的小鼠模型，WNV脑炎的小鼠细胞特异性 内皮细胞或星形胶质细胞中I型IFN受体（IFNAR）的缺失，以便更好地了解 BBB处的I型IFN信号传导影响神经侵袭和CNS病毒发病机制。此外，我们将 研究I型IFN对CNS抗病毒免疫运输和随后的病毒免疫运输的潜在调节作用。 清除和/或免疫病理学。我们假设I型干扰素是血脑屏障的关键调节因子， WNV感染，保持屏障完整性，限制WNV进入CNS，并限制CNS 免疫浸润和损伤。

Summary/Abstract The goal of this proposal is to elucidate a regulatory role for type-I interferon (IFN) at the blood brain barrier (BBB) during West Nile virus (WNV) encephalitis. WNV is a mosquito-borne flavivirus, now endemic in North America. Over the past decade, WNV has caused a growing epidemic of lethal viral encephalitis in the United States. West Nile pathogenesis, particularly how the pathogen infiltrates the central nervous system (CNS), remains poorly understood. Under normal conditions, the CNS is protected from pathogens in the circulation by the BBB, an interface consisting of brain microvascular endothelial cells (BMEC's) joined by tight junctions and supported by adjacent astrocyte endfeet [1]. However, WNV is able to cross the BBB through unknown mechanisms, and establishes infection in neurons and other parenchymal CNS tissues [2, 3]. CNS infection results in immune activation which dysregulates the BBB, facilitating the access of essential antiviral leukocytes to the CNS during infection; however, trafficking of infected leukocytes to the CNS may also provide an opportunity for neuroinvasion by WNV [4, 5]. Moreover, while trans-BBB immune trafficking is necessary for CNS viral clearance, the vulnerability of the CNS to damage and its limited capacity for repair can result in significant bystander injury and immunopathology during antiviral immune responses. Thus, the BBB must be tightly regulated during CNS infection, with a balance between facilitating access of antiviral leukocytes to sites of infection and protecting the CNS from immune-mediated damage and potential pathogen exposure from circulating virus and infected host immune cells. Antiviral cytokine signaling is a likely mechanism by which this balance is achieved. As a major component of the systemic immune response to WNV, the innate immune cytokines of the type-I IFN family are of critical importance in protecting both CNS and peripheral tissues from infection, via both direct antiviral activity and promotion of adaptive immune responses [6-8]. Interestingly, type-I IFN's have additionally been shown to exert anti-inflammatory properties at the BBB in the context of CNS autoimmunity, strengthening endothelial barrier integrity and impeding the ability of leukoyctes to bind endothelium and migrate into the CNS [9, 10]. However, the actions of type-I IFN at the BBB during viral encephalitis have not yet been investigated. Preliminary studies in our laboratory using in vitro models of the BBB have shown that the induction of type-I IFN signaling in both BMEC's and astrocytes after WNV infection results in enhanced endothelial barrier function and decreased transendothelial viral and immune trafficking. Building on these in vitro findings, we propose to use an established murine model of WNV encephalitis in mice with cell-specific deletions of the type-I IFN receptor (IFNAR) in endothelial cells or astrocytes in order to better understand how type-I IFN signaling at the BBB impacts neuroinvasion and CNS viral pathogenesis. In addition, we will examine potential regulatory effects of type-I IFN on CNS antiviral immune trafficking and consequent viral clearance and/or immunopathology. We hypothesize that type-I IFN is a key regulator of the BBB during WNV infection, preserving barrier integrity, restricting access of WNV to the CNS, and limiting CNS immune infiltration and damage during WNV encephalitis.