Nanocrystal Quantum Dot Biomimetics of SARS-CoV-2 to Interrogate Neutrophil-Mediated Neuroinflammation at the Blood-Brain Barrier

SARS-CoV-2 的纳米晶量子点仿生学研究中性粒细胞介导的血脑屏障神经炎症

• 批准号: 10510611
• 负责人: HARRIS A GELBARD
• 金额: $ 42.35万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-15 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10510611
• 关键词: 2019-nCoV; Acute; Alveolar; Behavior; Biological Assay; Biological Models; Biomimetics; Blood - brain barrier anatomy; Bradykinin; Bradykinin B2 Receptor; Bronchoalveolar Lavage Fluid; COVID-19; COVID-19 complications; COVID-19 long hauler; COVID-19 patient; Capsid Proteins; Cells; Central Nervous System Infections; Chronic; Complement; Coupled; Data; Diffuse; Dilatation - action; Disease; Electrical Resistance; Encapsulated; Endothelium; Event; Exhibits; Future; Gene Expression; Germany; Health; Heterogeneity; Image; Immune; Incubated; Individual; Infection; Inflammation; Inflammation Mediators; Inflammatory; Inflammatory Response; Intervention; Ions; Kallikrein-Kinin System; Kininogenase; Kininogens; Kinins; Label; Lead; Ligands; Liquid substance; Long COVID; Maps; Measurement; Measures; Mediating; Medical; Micelles; Microscope; Modeling; Mus; Nature; Neuraxis; Neurologic Deficit; Neurologic Symptoms; Neutrophil Activation; Particle Size; Pathway interactions; Patients; Permeability; Population; Proteins; Proxy; Quality of life; Quantum Dots; Renin-Angiotensin System; Reporting; Research; Resolution; Role; SARS coronavirus; SARS-CoV-2 infection; SARS-CoV-2 spike protein; Scanning; Scheme; Seminal; Signal Transduction; Structure; Symptoms; System; Testing; Tight Junctions; Tracer; Up-Regulation; Virion; Visualization; antagonist; blood damage; blood-brain barrier permeabilization; brain endothelial cell; confocal imaging; coronavirus disease; cytokine; design; experimental study; fluorescence imaging; immune activation; in vitro Model; inhibitor; innovative technologies; monolayer; nanocrystal; nanoscale; nervous system disorder; neuroinflammation; neurotransmission; neurovascular unit; neutrophil; persistent symptom; public health relevance; receptor; response; therapy design; transcytosis; virus development

PROJECT SUMMARY/ABSTRACT Public/health/relevance: Chronic, or recurring, neurological deficits in 60% of recovered COVID-19 patients are now an unmet medical need to treat the aftermath of SARS-CoV-2 infection of the central nervous system (CNS). A recent study from Germany suggests that these symptoms persist beyond a year, similarly to patients suffering from chronic symptoms due to SARS-CoV-1 infection. Thus, there is clear need for interventions against chronic neurologic symptoms after COVID. Elucidating the mechanism for SARS-CoV-2 impact on the CNS is essential to inform the design of such interventions. Objective: This proposal aims to identify a pathway for SARS-CoV-2’s effects on the CNS through a dysregulated blood-brain barrier (BBB) mediated by a neutrophil-dependent “storm” of bradykinin (BK). We hypothesize that this storm induces neuroinflammation that ultimately disrupts normal neuronal signaling, providing the substrate for enduring neurological symptoms. Research Plan: Recent studies have reported altered integrity of the BBB in response to the spike (S) protein of SARS-CoV-2, thereby suggesting a neuroinvasive pathway for SARS-CoV-2 or inflammatory immune cells through the BBB. In line with these observations, this proposal will investigate how pro-inflammatory mediators associated with COVID infection activate neutrophil-mediated upregulation of BK; this leads to an increased permeability through paracellular gaps across the BBB due to dysregulated tight junctions (TJs). Such a model aligns with the upregulated levels of BK observed in bronchoalveolar fluid taken from COVID-19 patients coupled with the ability of neutrophils to engage the kinin system to remodel endothelial barriers in acute inflammation. As a proxy for native SARS-CoV-2, we will construct S protein coated quantum dots as high fidelity biomimetics of SARS-CoV-2 to investigate the size and structural constraints regulating SARS-CoV-2 permeability across the BBB. These constructs will be used to bias neutrophils to a pro-inflammatory state in the presence of relevant kallikrein-kinin factors to increase the permeability of cultured bEnd.3 monolayers, a high-fidelity in vitro model system for murine BBB. A leakier BBB will be indicated by increased permeability of our fluorescent SARS-CoV-2 biomimetic and corroborated with complementary measurements of global barrier health, as measured by transendothelial electrical resistance (TEER). Lastly, we will construct a correlated scanning ion conductance and confocal microscope system to examine the heterogeneity of dysregulated barrier function and the specific nanoscale changes in TJ expression and localization that regulate it.

项目摘要/摘要 公共/健康/相关性：60％的COVID-19患者中有60％的慢性或经常性神经系统防御 现在是未满足的医疗需要治疗中枢神经系统的SARS-COV-2感染的后果 （CNS）。德国最近的一项研究表明，这些症状持续了一年，与患者类似 患有SARS-COV-1感染引起的慢性症状。那显然需要干预 反对慢性神经系统症状。阐明SARS-COV-2的机制对 中枢神经系统对于告知此类干预措施的设计至关重要。 目的：该提案旨在通过A来确定SARS-COV-2对CNS的影响的途径 由中性粒细胞依赖性的Bradyinin（BK）介导的血脑屏障（BBB）失调。我们 假设这场风暴会诱导神经炎症，最终破坏正常的神经元信号传导， 提供持久神经符号的基板。 研究计划：最近的研究报道了BBB对峰值蛋白的完整性的改变 SARS-COV-2，因此提出了SARS-COV-2或炎性免疫细胞的神经侵染途径 通过BBB。根据这些观察结果，该提案将研究促炎性调解人 与共同感染有关激活中性粒细胞介导的BK上调；这导致增加 由于紧密连接（TJS）失调，通过跨BBB的细胞细胞间隙渗透性。这样的模型 与从COVID-19患者取的支气管肺泡液中观察到的更新水平的BK级别对齐 嗜中性粒细胞使Kinin系统在急性炎症中重塑内皮屏障的能力。 作为天然SARS-COV-2的代理，我们将构建S蛋白涂层量子点作为高保真仿生型 SARS-COV-2研究调节整个SARS-COV-2渗透性的大小和结构约束 BBB。这些结构将在存在的情况下将中性粒细胞偏向促炎状态 kallikrein-kinin因子增加了培养弯曲的渗透性。3单层，高保真的体外模型 鼠BBB的系统。 Leafier BBB将通过我们的荧光SARS-COV-2仿生型的渗透性提高，并通过全球屏障健康的完整测量来证实 跨内皮电阻（TEER）。最后，我们将构建相应的扫描离子电导 和共聚焦显微镜系统，以检查屏障功能失调的异质性和特定 纳米级变化的TJ表达和调节它的定位。