Cellular mechanisms of fatal respiratory insufficiency in arboviral encephalitis

描述(由申请人提供)：总体目标是确定虫媒病毒性脑炎致死性呼吸功能不全的细胞机制。呼吸功能不全是虫媒病毒性脑炎，更具体地说是西尼罗河神经疾病的一系列后果，可导致不良预后。我们利用啮齿动物模型建立了神经缺陷导致呼吸窘迫，是西尼罗河病毒(WNV)和其他病毒性脑炎死亡的主要病理生理机制。这些发现代表了这些病毒如何导致呼吸衰竭和死亡的突破，但导致这些病理反应的细胞机制(S)尚不清楚，因此是本应用的主题。需要检验的假设是，趋化因子CXCR4/CXCL12轴推迟了病毒特异性T淋巴细胞进入含有呼吸控制神经元的延髓腹外侧区(VLM)的实质。此外，我们假设这种延迟会导致更高的病毒载量，以及导致致死性呼吸衰竭的抑制性生长抑素(SST)神经肽或其受体的增加。这一假设是基于最近的三项发现。1)我们发现神经呼吸功能不全是WNV和其他虫媒病毒感染的啮齿动物死亡的主要病理生理机制，这可能与人类患者的致死性病毒性脑炎有关。2)值得注意的是，西尼罗河病毒包膜免疫染色的神经元(96%)与SST染色的神经元紧密地共存于VLM内，这表明西尼罗河病毒和/或炎症正在诱发SST。尤其是在呼吸节律发生的关键区域--前Bétzinger复合体内，有许多神经元对SST染色很深。这在WN神经系统疾病中可能具有生物学意义，因为VLM中的SST是一种强大的呼吸负调节因子。SST神经肽在呼吸的产生和控制中发挥作用的证据是基于外源性给SST给VLM引起的深刻的负性呼吸效应，以及通过给SST受体拮抗剂恢复呼吸功能。3)CXCR4/CXCL12轴延迟了西尼罗河病毒特异性T淋巴细胞从血管周围间隙进入大脑非特定区域的实质。不幸的是，西尼罗河病毒等嗜细胞神经病毒需要病毒特异性CD8+T细胞穿透实质以清除病毒。这一延迟导致病毒载量增加，导致炎症反应增强，并降低了感染西尼罗河病毒的小鼠的存活率。因此，在免疫病理条件下，VLM特异性保护性T细胞的延迟进入可能导致SST抑制肽的异常增加，从而导致虫媒病毒引起的呼吸衰竭。旨在验证这一假说的具体目的是：目的1.确定西尼罗河病毒特异性T细胞通过CXCR4/CXCL12轴延迟进入是否通过增加SST和VLM实质中的炎症细胞而导致致命的呼吸功能不全。我们将通过与没有呼吸功能不全的西尼罗河病毒感染的小鼠相比，量化西尼罗河病毒感染的有呼吸功能不全的小鼠的VLM中的炎性细胞和SST来实现这一点。此外，CXCR4的一种拮抗剂将被评估其缓解VLM中这些病理事件的能力。目的2.确定SST及其受体(S)的表达增加是否与西尼罗河病毒诱导的呼吸功能不全有关。这将通过确定SST受体的拮抗剂通过立体定向注射到VLM是否可以减轻病毒抑制呼吸功能的影响来实现。

DESCRIPTION (provided by applicant): The overall goal is to identify the cellular mechanism of fatal respiratory insufficiency in arboviral encephalitides. Respiratory insufficiency is a serius outcome of arboviral encephalitis, and more specifically West Nile neurological disease, and can result in a poor prognosis. We have used rodent models to establish that neurological deficits cause the respiratory distress and are the primary pathophysiological mechanism of death for West Nile virus (WNV) and other viral encephalitides. These findings represent a breakthrough on how these viruses cause respiratory failure and death, but the cellular mechanism(s) causing these pathological responses are unknown, and are the subject of this application. The hypothesis to be tested is that the chemokine CXCR4/CXCL12 axis delays early entry of viral specific T lymphocytes into the parenchyma of the ventrolateral medulla (VLM) containing respiratory-control neurons. Additionally, we hypothesize that this delay results in higher virus load, and elevated inhibitory somatostatin (SST) neuropeptide or its receptor that contribute to lethal respiratory failure. This hypothesis is based on three recent findings. 1) Our finding that neurological respiratory insufficiency is a primary pathophysiologica mechanism of death in rodents infected with WNV and other arboviruses, which might be relevant to fatal viral encephalitides of human patients. 2) Remarkably, WNV envelope-immunostained neurons are tightly co-localized (96%) with SST-stained neurons in the VLM, which implies that WNV and/or inflammation are inducing SST. Specifically within the pre-B¿tzinger complex, an area critical for respiratory rhythmogenesis, there are many neurons heavily stained for SST. This may be biologically significant in WN neurological disease, because SST in the VLM is a potent negative regulator of breathing. Evidence that SST neuropeptide plays a role in the generation and control of respiration is based on profound negative respiratory effects caused by exogenously administered SST to the VLM, and a restoration of respiratory function by administration of antagonists to SST receptors. 3) CXCR4/CXCL12 axis delays entry of WNV-specific T lymphocytes from the perivascular spaces into the parenchyma of non-specified areas of the brain. Unfortunately, cytopathic neurotropic viruses, such as WNV, require penetration of viral specific CD8+ T cells into the parenchyma for clearance of the virus. This delay results in an increased viral load leading to increased inflammatory responses and decreased survival of WNV-infected mice. Consequently, the effect of delayed entry of protective T cells specifically in the VLM may aberrantly increase the SST inhibitory peptide under immune-pathological conditions and contribute to arboviral-induced respiratory failure. The specific aims designed to test the hypothesis are: Aim 1. Determine if delayed entry of WNV- specific T cells via the CXCR4/CXCL12 axis contributes to fatal respiratory insufficiency by increasing SST, and inflammatory cells specifically in the parenchyma of the VLM. We will accomplish this by quantifying inflammatory cells and SST in the VLM of WNV-infected mice with respiratory insufficiency, as compared to WNV-infected mice without respiratory insufficiency. Additionally, an antagonist of CXCR4 will be evaluated for its ability to mitigate these pathological events in the VLM. Aim 2. Determine if increased expression of SST or its receptor(s) is involved with WNV-induced respiratory insufficiency. This will be accomplished by determining if an antagonist to the SST receptor stereotaxically injected into the VLM can mitigate the effects of the virus to suppress respiratory functions.