Immune Privilege, CNS Autoimmunity, and Clostridium perfringens Epsilon Toxin

免疫特权、中枢神经系统自身免疫和产气荚膜梭菌 Epsilon 毒素

基本信息

批准号：
10754021
负责人：
TIMOTHY VARTANIAN
金额：
$ 67.97万
依托单位：
WEILL MEDICAL COLL OF CORNELL UNIV
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-06-01 至 2028-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10754021
关键词：
Active Immunization Animal Model Autoimmunity Binding Bioinformatics Biological Assay Blood Blood - brain barrier anatomy Blood Vessels Brain Brain Stem CNS autoimmunity Candidate Disease Gene Catalogs Cell Separation Cell Surface Receptors Cells Cerebellum Clinical Clostridium perfringens Clostridium perfringens epsilon toxin Confocal Microscopy DNA Sequence Alteration Demyelinations Disease Endothelial Cells Endothelium Endotoxins Environmental Risk Factor Event Experimental Autoimmune Encephalomyelitis Experimental Models Flow Cytometry Frequencies Functional disorder Gene Deletion Gene Transfer Genes Genetic Risk Goals Human Immune Immunohistochemistry Individual Inflammatory Knockout Mice Lesion Leukocyte Trafficking Leukocytes Localized Lesion Location Lymphocyte Mediating Meningeal Messenger RNA Modeling Molecular Molecular Target Multiple Sclerosis Mus Mutant Strains Mice Myelin Neuroanatomy Neurologic Pathogenicity Pathology Pathway interactions Persons Pertussis Toxin Phenotype Prevalence Process Prosencephalon Proteins RNA Role Sampling Spinal Cord T-Lymphocyte Techniques Testing Time Tissues Toxin Transcriptional Regulation Wild Type Mouse Work adaptive immune response autoreactivity cellular targeting clinical phenotype clinically relevant diagnostic tool disability gain of function gene function gene induction gut microbiome high dimensionality immune cell infiltrate immunopathology innovation loss of function loss of function mutation migration multiple sclerosis patient neuropathology novel diagnostics postcapillary venule receptor single-cell RNA sequencing therapeutic evaluation therapeutic target transcriptome sequencing young adult

Active Immunization Animal Model Autoimmunity Binding Bioinformatics Biological Assay Blood Blood - brain barrier anatomy Blood Vessels Brain Brain Stem CNS autoimmunity Candidate Disease Gene Catalogs Cell Separation Cell Surface Receptors Cells Cerebellum Clinical Clostridium perfringens Clostridium perfringens epsilon toxin Confocal Microscopy DNA Sequence Alteration Demyelinations Disease Endothelial Cells Endothelium Endotoxins Environmental Risk Factor Event Experimental Autoimmune Encephalomyelitis Experimental Models Flow Cytometry Frequencies Functional disorder Gene Deletion Gene Transfer Genes Genetic Risk Goals Human Immune Immunohistochemistry Individual Inflammatory Knockout Mice Lesion Leukocyte Trafficking Leukocytes Localized Lesion Location Lymphocyte Mediating Meningeal Messenger RNA Modeling Molecular Molecular Target Multiple Sclerosis Mus Mutant Strains Mice Myelin Neuroanatomy Neurologic Pathogenicity Pathology Pathway interactions Persons Pertussis Toxin Phenotype Prevalence Process Prosencephalon Proteins RNA Role Sampling Spinal Cord T-Lymphocyte Techniques Testing Time Tissues Toxin Transcriptional Regulation Wild Type Mouse Work adaptive immune response autoreactivity cellular targeting clinical phenotype clinically relevant diagnostic tool disability gain of function gene function gene induction gut microbiome high dimensionality immune cell infiltrate immunopathology innovation loss of function loss of function mutation migration multiple sclerosis patient neuropathology novel diagnostics postcapillary venule receptor single-cell RNA sequencing therapeutic evaluation therapeutic target transcriptome sequencing young adult

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项目摘要

Why some people develop Multiple Sclerosis and others do not, despite similar genetic risk and quantities of circulating autoreactive lymphocytes, is not known. Our long-term goal is to identify environmental triggers of MS, define the molecular and cellular basis of their action, and in doing so, propose new diagnostic tools and therapeutic targets. The objectives of this proposal are to determine mechanistically how Clostridium perfringens epsilon toxin (ETX) and Bordetella pertussis toxin (PTX) overcome CNS immune privilege to trigger autoimmunity in the context of myelin autoreactive lymphocytes and to understand why ETX causes lesions to develop in the forebrain, cerebellum, brainstem, and spinal cord in contrast to PTX where lesions are more commonly localized to the spinal cord. The central hypothesis of this project is that ETX and PTX trigger CNS autoimmunity by inducing critical dysfunction at CNS barriers necessary for entry of pathogenic lymphocytes. The central hypothesis will be tested by pursuing two aims: 1) Determining the effect of cell specific deletion or introduction of the ETX receptor MAL (Myelin and Lymphocyte Protein) in active immunization models of experimental autoimmune encephalomyelitis (EAE), compare the neuroanatomical location, phenotype, and activation state of immune infiltrates between PTX- and ETX-induced EAE, and explore the effect of ETX on human lymphocytes, and 2) Determine the genes induced and suppressed in CNS-endothelial cells by ETX and PTX and define their function in overcoming CNS immune privilege through loss-of-function strategies. We will pursue these aims using an innovative combination of targeted genetic mutations to isolate cellular and molecular targets of ETX required to induce disease. We will use confocal microscopy, immunohistochemistry, high dimensional flow cytometry, and unbiased sampling of the entire CNS to compare the effects of ETX with PTX on immune phenotype, demyelination, and neuroanatomic localization of lesions. To determine toxin induced genes functioning to overcome CNS immune privilege, we will apply a combination of unbiased mRNA profiling techniques to CNS endothelial cells isolated from different neuroanatomic regions, advanced bioinformatics to define relevant gene modules, immunohistochemistry to validate localization of these induced proteins within individual post-capillary venules, and conditional loss-of-function mutations in endothelial cells to determine function. The rationale underlying this proposal is that completion will define the role by which a toxin, clinically associated with MS, functions in the multi-step process of autoimmunity, and will identify key molecular targets that can be tested therapeutically. This work will also help establish an experimental model that has greater clinical relevance to MS and more closely resembles MS neuropathology than experimental autoimmune encephalomyelitis models reliant on pertussis toxin.

为什么有些人出现多发性硬化症而其他人则没有，尽管遗传风险和数量相似循环自动反应性淋巴细胞尚不清楚。我们的长期目标是确定 MS，定义其作用的分子和细胞基础，并在此过程中提出了新的诊断工具和治疗靶标。该提案的目标是从机械上确定梭子座的方式灌注源含量Epsilon毒素（ETX）和Bordetella百日咳毒素（PTX）克服了CNS免疫特权触发在髓磷脂自动反应性淋巴细胞中自身免疫性，并了解为什么ETX会导致病变与PTX相比，在前脑，小脑，脑干和脊髓中发展，病变更多通常位于脊髓。该项目的中心假设是ETX和PTX触发CNS 自身免疫性通过在进入致病性淋巴细胞所需的中枢神经系统屏障处引起严重功能障碍。中心假设将通过追求两个目的来检验：1）确定特定细胞缺失的影响或在主动免疫模型中引入ETX受体MAL（髓磷脂和淋巴细胞蛋白）实验性自身免疫性脑脊髓炎（EAE），比较神经解剖位置，表型和 PTX-和ETX诱导的EAE之间免疫浸润的激活状态，并探索ETX对人类淋巴细胞和2）确定ETX和 PTX并通过功能丧失策略来克服CNS免疫特权方面的功能。我们将使用靶向遗传突变的创新组合来追求这些目标，以分离细胞和诱导疾病所需的ETX的分子靶标。我们将使用共聚焦显微镜，免疫组织化学，高维流式细胞术和整个CNS的无偏抽样，以将ETX的影响与 PTX关于病变的免疫表型，脱髓鞘和神经解剖学定位。确定毒素诱导的基因功能以克服CNS免疫特权，我们将采用无偏mRNA的组合从不同神经解剖区域分离的CNS内皮细胞的分析技术，先进生物信息学以定义相关基因模块，免疫组织化学来验证这些诱导的定位单个毛细血管后静脉内的蛋白质，以及内皮细胞中有条件的功能丧失突变确定功能。该提议的基本原理是完成将定义一个作用毒素，临床上与MS相关的毒素，在自身免疫的多步骤过程中起作用，并将识别关键可以通过治疗测试的分子靶标。这项工作还将有助于建立一个实验模型与实验性相比，它与MS具有更大的临床相关性，并且与MS神经病理学更相似自身免疫性脑脊髓炎模型依赖于百日咳毒素。