Viral and immune-mediated CNS pathology during SARS-CoV-2 infection

项目摘要 新冠肺炎疫情肆虐全球，对世界各国人民产生持久影响 全球社会的经济、健康和心理。尽管它与SARS-CoV-1有相似之处，但完整的 SARS-CoV-2引起的病理生理程度尚不清楚。尤其是肺外表现 SARS-CoV-2感染的影响仍然知之甚少。中国和欧洲的案例系列表明 中枢神经系统参与了至少一部分患者的疾病过程，有报道称 据估计，多达30%的新冠肺炎患者有神经系统症状，包括癫痫，难以治愈 头痛，嗅觉和味觉受损。尽管有报道称新冠肺炎中有神经系统疾病 目前尚不清楚SARS-CoV-2是否会侵入中枢神经系统(CNS)。对其他的研究 冠状病毒，包括SARS-CoV-1，表现出明显的神经嗜性和神经炎症 与这一病毒家族的其他成员有关。这些研究提出了SARS-CoV-2病毒 可能通过侵犯中枢神经系统或通过增加炎症反应而导致神经症状 中枢神经系统内的细胞因子。我们假设SARS-CoV-2感染具有神经侵袭潜力并导致 与感染者中枢神经系统内的免疫状态改变和高度炎症有关。我们进一步假设 中枢神经系统感染通过ACE2表达的直接毒性加重呼吸功能障碍 对心肺功能起关键调节作用的神经元。我们的调查将结合 与那些利用小鼠模型进行的人体研究，在这些模型中，我们可以很容易地管理病毒并评估 病理生理学。目的1、检测新冠肺炎患者中枢神经系统免疫反应。 神经症状。使用单细胞RNA测序、细胞因子分析、病毒 测序和抗体验证，我们将充分剖析中枢神经系统内的炎症反应 新冠肺炎患者的脑室与体循环的比较。使用小鼠模型，在目标2中，我们 将调查SARS-CoV-2的脑炎潜力。使用几种互补的方法来 用SARS-CoV-2感染小鼠，我们会将病毒引入小鼠的中枢神经系统。vbl.使用 耗尽抗体和各种基因敲除小鼠，我们将确定哪些免疫细胞是需要的 通过存活研究、流式细胞术和免疫荧光染色观察这些小鼠的神经病理改变。最后， 在目标3中，我们将评估中枢神经系统感染对呼吸结局的影响。因为已知的 血管紧张素转换酶2在脑干的表达以及脑干在调节心肺功能中的关键作用， 我们怀疑SARS-CoV-2感染中枢神经系统会加重SARS-CoV-2呼吸道疾病。 这三个目标将有助于支持我们的假设，即SARS-CoV-2感染如何影响中枢神经系统和 呼吸隔间。我们期望我们的发现将发现新的策略来治疗被诊断为 与新冠肺炎的合作，有助于对SARS-CoV-2的生物学病理生理学有新的了解。

Project Summary The COVID-19 pandemic has rampantly affected the population of the world and created lasting effects on the economy, health and psyche of the global community. Although it shares similarities with SARS-CoV-1, the full extent of the pathophysiology caused by SARS-CoV-2 is unclear. In particular, extrapulmonary manifestations effects of SARS-CoV-2 infection remain poorly understood. Case series from China and Europe suggest that the central nervous system is involved in the disease process in at least a subset of patients, with some reports estimating up to 30% of COVID-19 patients having neurological symptoms, including seizure, intractable headache, and impaired smell and taste. Although there are reports of neurological disease in in COVID-19 patients, it is unclear if SARS-CoV-2 invades the central nervous system (CNS). Studies of other coronaviruses, including SARS-CoV-1, demonstrate clear neurotropism as well as neuroinflammation associated with other members of this family of viruses. These studies raise the possibility that SARS-CoV-2 may cause neurological symptoms either through invasion of the CNS or through an increase in inflammatory cytokines within the CNS. We hypothesize that SARS-CoV-2 infections have neuroinvasive potential and lead to altered and hyperinflammatory immune states within the CNS of infected individuals. We further hypothesize that infection of the CNS exacerbates respiratory dysfunction through direct toxicity of ACE2 expressing neurons that are critical regulators of cardiopulmonary function. Our investigations will combine the power of human studies with those utilizing mouse models in which we can readily administer virus and assess for pathophysiology. Aim 1, we will determine the CNS immune responses in COVID-19 patients with neurological symptoms. Using a combination of single cell RNA-sequencing, cytokine profiling, viral sequencing and antibody validations, we will fully dissect out the inflammatory responses within the CNS compartment compared to the systemic circulation in COVID-19 patients. Using mouse models, in Aim 2, we will investigate the encephalitic potential of SARS-CoV-2. Using several complementary approaches to infect mice with SARS-CoV-2, we will introduce the virus into the central nervous system of mice. Using depletion antibodies and various knockout mice, we will identify which immune cells are required for neuropathology in these mice through survival studies, flow cytometry and immunofluorescent staining. Finally, in Aim 3, we will evaluate the effects of CNS infection on respiratory outcomes. Because of the known expression of ACE2 in the brainstem, and the brainstem’s critical role in regulating cardiopulmonary functions, we suspect that CNS infection with SARS-CoV-2 will exacerbate SARS-CoV-2 respiratory disease. These three aims will help support our hypotheses of how SARS-CoV-2 infections can affect the CNS and respiratory compartments. We expect that our findings will uncover new strategies to treat patients diagnosed with COVID-19 and help gain new insight to understanding the biology of SARS-CoV-2 pathophysiology.