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，特别是西尼罗河神经系统疾病的严重后果，可导致预后不良。我们使用啮齿类动物模型来确定神经功能缺损引起呼吸窘迫，这是西尼罗病毒和其他病毒性脑病的主要病理生理机制。这些发现代表了这些病毒如何引起呼吸衰竭和死亡的突破，但引起这些病理反应的细胞机制尚不清楚，并且是本应用的主题。待验证的假设是趋化因子CXCR4/CXCL12轴延迟病毒特异性T淋巴细胞早期进入含有呼吸控制神经元的腹外侧延髓（VLM）的实质。此外，我们假设这种延迟会导致更高的病毒载量，以及抑制性生长抑素（SST）神经肽或其受体的升高，从而导致致命性呼吸衰竭。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啮齿动物感染西尼罗河病毒及其他虫媒病毒死亡的主要病理生理机制，这可能与人类致命病毒性脑炎有关。2)值得注意的是，在VLM中，WNV包膜免疫染色的神经元与SST染色的神经元紧密共定位（96%），这表明WNV和/或炎症正在诱导SST。特别是在对呼吸节律发生至关重要的pre-B¿tzinger复合体内，有许多神经元被严重染色。这在神经系统疾病中可能具有重要的生物学意义，因为VLM中的SST是一种有效的呼吸负调节因子。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药SST对VLM造成的深刻的负呼吸效应，以及通过给药SST受体拮抗剂恢复呼吸功能。3) CXCR4/CXCL12轴延缓了wnv特异性T淋巴细胞从血管周围间隙进入脑非特定区域的实质。不幸的是，细胞病变嗜神经病毒，如西尼罗河病毒，需要穿透病毒特异性CD8+ T细胞进入薄壁组织以清除病毒。这种延迟导致病毒载量增加，导致炎症反应增加，西尼罗河病毒感染小鼠的存活率降低。因此，保护性T细胞特异性延迟进入VLM的作用可能在免疫病理条件下异常增加SST抑制肽，并导致虫媒病毒诱导的呼吸衰竭。为验证这一假设而设计的具体目标是：目标1。确定WNV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性呼吸功能不全。我们将通过量化呼吸功能不全的西尼罗河病毒感染小鼠VLM中的炎症细胞和SST来实现这一目标，并将其与没有呼吸功能不全的西尼罗河病毒感染小鼠进行比较。此外，将评估CXCR4拮抗剂减轻VLM中这些病理事件的能力。目标2。确定SST或其受体的表达增加是否与西尼罗河病毒引起的呼吸功能不全有关。这将通过确定将SST受体拮抗剂立体定向注射到VLM中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.