Immune cell dynamics during central nervous system viral infection

项目主任/主要研究者（最后，第一，中间）：德拉托雷，胡安C。1R01 AI075298-01A2 病毒可在中枢神经系统中诱发多种疾病状态。脑膜炎是一种潜在的致命疾病，由一长串人类病原体（包括病毒）引起，通常伴有发热、头痛、颈部僵硬和癫痫发作等症状。目前，除了缓解症状外，对病毒性脑膜炎患者几乎无能为力。我们建议，在真实的时间详细了解这一致病过程可能会促进新的干预措施的发展，以减轻症状，防止永久性神经功能障碍和死亡。为了实现这一目标，我们建议研究由淋巴细胞性脉络丛脑膜炎病毒（LCMV）引起的脑膜炎，一种非细胞病变的小鼠以及人类病原体。脑内接种LCMV的小鼠在6天内导致致死性脑膜炎，其几乎完全由细胞毒性淋巴细胞（CTL）介导。重要的是，这种疾病可以通过事先接种疫苗或免疫接种来完全预防。在感染控制失败或有效期间，脑膜中细胞相互作用的动力学以前没有研究过。此外，在该模型中介导致命损伤的确切机制尚未完全了解。我们将利用最先进的病毒反向遗传学，荧光标记的免疫细胞，双光子激光扫描显微镜结合原位染色不同的分子种类，以遵循局部免疫细胞动态LCMV感染的小鼠大脑皮层和脑膜空间。我们的假设是，CTL损伤中枢神经系统中的星形胶质细胞网络导致全身性致命癫痫发作急性LCMV诱导的脑膜炎，并在接种疫苗的小鼠中激活的记忆T细胞的快速反应性和使用替代效应机制导致星形胶质细胞网络的有限损伤，维持血脑屏障和生存。这一假设将通过以下两个集中的具体目标来解决：1）完成CTL和被荧光标记的LCMV感染的CNS靶之间的相互作用的第一个真实的时间分析，和2）免疫突触形成和参与CNS CTL靶向和损伤的分子机制的体内评价。

Program Director/Principal Investigator (Last, First, Middle): De La Torre, Juan C. 1R01 AI075298-01A2 Viruses can induce a variety of disease states in the central nervous system. Meningitis is a potentially fatal disorder induced by a long list of human pathogens, including viruses, and is often associated with symptoms that include fever, headache, stiffness of the neck, and seizures. Presently, very little can be done for patients with viral meningitis other than to relieve symptoms. We propose that a detailed understanding of this pathogenic process in real time may foster the development of novel interventions to alleviate symptoms and prevent permanent neurological dysfunction and fatalities. To accomplish this goal we propose to study the well-described meningitis induced by lymphocytic choriomeningitis virus (LCMV), a noncytopathic mouse as well as human pathogen. Intracerebral inoculation of mice with LCMV results in a fatal meningitis within 6 days that is mediated almost entirely by cytotoxic lymphocytes (CTL). Importantly, this disease can be completely prevented by prior vaccination or immunization. The dynamics of cellular interactions in the meninges during failed or effective control of infection have not been studied previously. Moreover, the precise mechanisms that mediate fatal injury in this model are not entirely understood. We will utilize a combination of state-of-the art viral reverse genetics, fluorescently-tagged immune cells, and two-photon laser scanning microscopy in combination with in situ staining for different molecular species to follow the local immune cell dynamics in the LCMV-infected mouse cerebral cortex and meningeal space. Our hypothesis is that CTL damage to astrocyte networks in the CNS leads to generalized fatal seizure during acute LCMV-induced meningitis, and that rapid responsiveness and the use of alternative effector mechanisms by activated memory T cells in vaccinated mice results in limited damage to astrocyte networks, maintenance of the blood brain barrier, and survival. This hypothesis will be addressed by the two focused following specific aims: 1) Completion of the first real time analyses of interactions between CTL and CNS targets infected by fluorescently-tagged LCMV, and 2) in vivo evaluation of immunological synapse formation and molecular mechanisms involved in CNS CTL targeting and damage.