Parasite dissemination in Toxoplasmic encephalitis

弓形虫脑炎中的寄生虫传播

• 批准号: 9889871
• 负责人: Melissa Bruckner Lodoen
• 金额: $ 38.63万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-01 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9889871
• 关键词: AIDS/HIV problem; Acquired Immunodeficiency Syndrome; Address; Affect; Animal Model; Biochemical; Biological; Biological Assay; Brain; Cell Adhesion; Cell Adhesion Molecules; Cell surface; Cells; Central Nervous System Diseases; Central Nervous System Infections; Chronic; Clinical; Data; Disease; Encephalitis; Equus caballus; Event; Fluorescence Microscopy; Focal Adhesion Kinase 1; Focal Adhesions; Goals; HIV; HIV Infections; Hand; Human; Image; Immune; Immunosuppression; Impairment; In Vitro; Individual; Infection; Inflammatory; Integrins; Intervention; Label; Leukocytes; Life; Light; Mediating; Microglia; Migration Assay; Molecular; Mus; Myeloid Cells; Neuraxis; Organ; Parasites; Pathogenesis; Patients; Phenotype; Population; Process; Public Health; Reagent; Regulation; Research; Role; Signal Transduction; Site; Talin; Techniques; Testing; Therapeutic; Tissues; Toxoplasma gondii; Toxoplasmosis; Transgenic Mice; Variant; acute infection; base; cell motility; cell type; chronic infection; clinically relevant; combat; foodborne; in vivo; innovation; insight; interdisciplinary approach; intravital imaging; microscopic imaging; migration; monocyte; mutant; novel; pathogen; paxillin; public health relevance; scaffold; therapeutic development; therapeutic target; tool; toxoplasmic encephalitis; two photon microscopy; two-photon

 DESCRIPTION (provided by applicant): Toxoplasma gondii is an opportunistic parasite that infects one-third of the global population and is among the most frequent causes of central nervous system (CNS) infection complicating AIDS. T. gondii establishes a chronic infection in the CNS, and immune compromise due to HIV infection leads to parasite reactivation and fatal encephalitis. This proposal addresses a poorly understood aspect of parasite pathogenesis during clinical AIDS - how does T. gondii migrate through the brain? Evidence suggests that T. gondii infection of migratory leukocytes facilitates the colonization of vital organs such as the brain via a 'Trojan horse' mechanism. Monocytes in particular are highly motile cells and a target for T. gondii infection. Our strong preliminary data demonstrate that T. gondii infection of human monocytes induces a hypermigratory phenotype and dysregulates the function of key adhesion molecules called integrins. The objective of this proposal is to define how T. gondii induces monocyte motility and disseminates through the brain. The central hypothesis is that T. gondii spreads through tissues by co-opting the migratory potential of infected cells. Two aims are proposed to test the hypothesis: 1) Determine how T. gondii enhances the migration of infected cells, and 2) Define how T. gondii disseminates through the infected brain. In the first aim, we will investigate the regulation of integrin signaling and focal adhesion formation in infected primary human monocytes from healthy donors and from HIV patients. We will determine the role of focal adhesion kinase (FAK) by using monocytes expressing mutant variants of FAK that are constitutively active or non-phosphorylatable (inactive). We will also determine the interactions of the scaffolding molecules talin and paxillin with FAK and the dynamics of their assembly and disassembly in infected monocytes during migration. In the second aim, we will use intravital two-photon imaging and an animal model for T. gondii infection to define the myeloid cell type(s) that are infected and potentiate parasite spread through the mouse brain. Transgenic mice that express fluorescently labeled monocytes or microglia will be infected with T. gondii. The dynamics of their migration through the parenchyma will be determined during acute infection and in chronically infected mice undergoing parasite reactivation due to immune suppression. All of the necessary reagents are on hand in the lab or commercially available, and all the techniques, including biochemical and migration assays, total internal reflection fluorescence (TIRF) microscopy, and intravital imaging, are currently employed by our research team. The significance of this research is that understanding T. gondii dissemination may allow for therapeutic targeting of this process to mitigate disease during AIDS. The proposal is innovative because it applies a multidisciplinary approach and novel tools to identify the cellular and molecular mechanisms that mediate T. gondii dissemination in vitro and in vivo, which will ultimately shed light on a key aspect of parasite pathogenesis in HIV/AIDS patients.

 描述（由申请人提供）：弓形虫是一种机会寄生虫，感染全球三分之一的人口，是艾滋病并发中枢神经系统（CNS）感染的最常见原因之一。T.弓形虫在中枢神经系统中建立慢性感染，并且由于HIV感染引起的免疫损害导致寄生虫再激活和致命的脑炎。这一建议解决了临床艾滋病期间寄生虫发病机制的一个知之甚少的方面-T。弓形虫会通过大脑传播吗证据表明T.迁移性白细胞的弓形虫感染通过“特洛伊木马”机制促进重要器官如脑的定殖。特别是单核细胞是高度运动的细胞，是T。弓形虫感染我们强有力的初步数据表明，T。弓形虫感染 人单核细胞诱导超迁移表型并使称为整联蛋白的关键粘附分子的功能失调。本建议的目的是确定如何T。弓形虫诱导单核细胞运动并通过脑传播。中心假设是T.弓形虫通过吸收受感染细胞的迁移潜能在组织中传播。本文提出了两个检验假设的目标：1）确定T.弓形虫感染细胞的迁移能力增强;弓形虫通过受感染的大脑传播。在第一个目标中，我们将研究整合素信号传导和粘着斑形成的调节在感染的原代人单核细胞从健康的捐助者和艾滋病患者。我们将确定粘着斑激酶（FAK）的作用，通过使用单核细胞表达突变体的FAK是组成型活性或非磷酸化（无活性）。我们还将确定支架分子talin和桩蛋白与FAK的相互作用，以及它们在迁移过程中在受感染的单核细胞中组装和拆卸的动力学。在第二个目标中，我们将使用活体双光子成像和T。弓形虫感染，以确定被感染的骨髓细胞类型，并增强寄生虫通过小鼠脑的传播。表达荧光标记的单核细胞或小胶质细胞的转基因小鼠将被T.刚地将在急性感染期间和由于免疫抑制而经历寄生虫再活化的慢性感染小鼠中确定它们通过实质的迁移动力学。所有必要的试剂都在实验室或商业上可用，所有的技术，包括生化和迁移测定，全内反射荧光（TIRF）显微镜和活体成像，目前都由我们的研究团队采用。本研究的意义在于了解T.弓形虫传播可能允许治疗靶向该过程，以减轻艾滋病期间的疾病。该提案是创新的，因为它应用了多学科的方法和新的工具来确定介导T。弓形虫传播的体外和体内，这将最终揭示了艾滋病毒/艾滋病患者的寄生虫发病机制的一个关键方面。