Modulation of Immune Defenses in Pathobiology of CNS Infection

中枢神经系统感染病理学中免疫防御的调节

• 批准号: 10084210
• 负责人: Michal A Olszewski
• 金额: --
• 依托单位: VETERANS HEALTH ADMINISTRATION
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10084210
• 关键词: Address; Adoptive Cell Transfers; Adoptive Transfer; Affect; Anti-Inflammatory Agents; Antifungal Agents; Antifungal Therapy; Antiinflammatory Effect; Autoimmune; Biological Markers; CD4 Positive T Lymphocytes; CXCR3 gene; Cells; Central Nervous System Infections; Cerebral Cryptococcosis; Cessation of life; Coculture Techniques; Collaborations; Cryptococcosis; Cryptococcus; Cryptococcus neoformans; Data; Development; Disease; Experimental Models; Exposure to; Gene Expression Profiling; General Population; HIV; Histology; Immune; Immune response; Immunocompromised Host; Immunologics; Immunotherapy; In Vitro; Infection; Inflammatory; Inhalation; Interferon Type II; Kinetics; Knowledge; Ligands; Link; Malaria; Measures; Mediating; Medical; Meningeal Tuberculosis; Meningoencephalitis; Methods; Modeling; Molecular; Mononuclear; Morbidity - disease rate; Mus; Myeloid Cells; Neurologic Symptoms; Nitric Oxide; Outcome; Pathogenesis; Pathologic; Pathology; Patients; Phagocytes; Pharmacotherapy; Population; Process; Production; Research; Role; Signal Transduction; Steroid therapy; T-Lymphocyte; Testing; Th1 Cells; Therapeutic; Tissues; Tumor-infiltrating immune cells; Veterans; Virus; Work; central nervous system injury; disability; experience; experimental study; fungus; immunomodulatory therapies; immunoregulation; in vivo; in vivo Model; infection risk; insight; monocyte; monocyte chemoattractant protein 1 receptor; mortality; mouse model; neurotoxic; novel; pathogen; pathogenic fungus; recruit; response; tissue injury

Objectives: Cryptococcus neoformans is the most important fungal pathogen causing CNS-related mortality and morbidity world-wide. Treatment of C. neoformans CNS infections is very challenging. There is increasing evidence suggesting that paradoxical responses to therapy are mediated by patients’ own immune response to the fungus supported by beneficial effects of anti-inflammatory therapy. However, the use of broad anti- inflammatory (steroid) therapy is controversial because cryptococcosis is predominantly a disease of immunocompromised population. Using model of CNS- crytptococcosis in mice we will investigate CNS damage during cerebral cryptococcosis which we propose is a result of collaboration between myeloid cells with the CNS recruited T-cells. Our studies will define the mechanisms by which these cells are recruited to the CNS and cause detrimental CNS pathology. Hypothesis: CCR2-axis signaling recruits monocytes into the CNS, which further promotes T-cells recruitment and polarization facilitating fatal CNS damage. We further propose that Nitric oxide (NO) is central effector product of monocyte derived cells (MoC) activated by Th1 cells, which induces CNS pathology. To test distinct parts of this hypothesis in our mouse model we will: Aim 1. Determine whether CCR2-dependent MoC accumulation and activation drive CNS pathology. a) To establish the how CCR2-axis affects the kinetics of MoC accumulation and activation in the CNS during CME; b) To assess the contribution of the CCR2 axis/MoC to the development of CNS damage, neurological symptoms and mortality during CME; c)To provide additional mechanistic proof of concept that CCR2+ monocytes induce CNS injury in CME. Aim 2: Establish that CCR2+ MoC promote a Th1 response in the CNS during CME: a) To determine whether MoC produce CXCR3 ligands and to assess whether T-cell recruitment into the CNS is mediated via CXCR3-signaling; b) To determine whether CCR2+ MoC contribute to T-cell recruitment, activation, and polarization; c) To assess whether the adoptive transfer Th1 cells from the infected CCR2+/+ mice into CCR2-/- mice restores CME-associated damage. Aim 3. Determine whether iNOS/NO from CCR2+ MoC is required to mediate pathological CNS tissue damage observed during CME: a) To specifically test whether iNOS induction in CCR2+MoC mechanistically contribute to CNS pathology observed in CME; b) To determine whether IFNγ/IFNγR signaling in CCR2+ MoC is required to induce iNOS in these cells and the effect of IFNγ/IFNγR signaling on subsequent CNS damage. Research Plan and Methods: This proposal will utilize our novel mouse model of CM, which accurately recapitulates severe paradoxical immune responses experienced by patients with C. neoformans CNS infection. We have previously shown that ultra-polarized IFNγ-producing CD4+ T cells accumulate in the CNS and contribute to CNS pathology and mortality. In proposed work, we will use our in vivo model of C. neoformans CNS infection along with in genetically modified mice to test specific aspects of our hypothesis, in vitro T cell co-cultures, while in vivo adoptive cell transfers will help us to define the types of CNS tissue injury and the role of CCR2+ and myeloid cells and other subsets of myeloid cells in CNS immune pathology. Specifically, these strategies will use genetically deficient mice, including those lacking CCR2 (CCR2-/- - receptor selectively removed from CCR2+ cells (CCR2γR-/-), CXCR3-signaling (CXCR3-/-), and mice unabel to make nitric oxide globally (iNOS-/-) or only in CCR2+ cells CCR2-NOS-/-). The outcomes of our experiment manipulations will be measured by a) analysis of fungal burdens b) analysis of neurological symptoms and mortality c) flow cytometric analysis of infiltrating immune cells, d) analysis of mononuclear phagocyte polarization profile and effector functions, e) gene expression analysis and f) histology. Overall, these studies will provide significant insight on T cell mediated pathology during fungal CNS infections and are likely to identify new biomarkers or therapeutic opportunities.

目的：新生隐球菌是导致中枢神经系统相关死亡的最重要的真菌病原体 以及世界范围内的发病率。新生葡萄球菌中枢神经系统感染的治疗是非常具有挑战性的。有越来越多的 有证据表明，对治疗的矛盾反应是由患者自身的免疫反应介导的 这种真菌得到了抗炎治疗的有益效果的支持。然而，使用广泛的反- 炎症(类固醇)疗法是有争议的，因为隐球菌病主要是一种 免疫功能受损的人群。利用小鼠中枢神经系统隐球菌病模型，对中枢神经系统进行研究 我们认为，脑隐球菌病期间的损害是髓系细胞与 中枢神经系统招募T细胞。我们的研究将确定这些细胞被招募到中枢神经系统的机制 并导致有害的中枢神经系统病理。假设：CCR2轴信号征募单核细胞进入中枢神经系统， 这进一步促进了T细胞的募集和极化，促进了致命的中枢神经系统损伤。我们进一步建议 一氧化氮(NO)是Th1细胞激活的单核细胞衍生细胞(MOC)的中枢效应产物，它 诱导中枢神经系统病理。为了在我们的小鼠模型中测试这个假设的不同部分，我们将：目标1。 确定CCR2依赖的MOC积聚和激活是否推动中枢神经系统的病理改变。A)建立 CCR2轴如何影响CME期间中枢神经系统内MOC积累和激活的动力学；b) 评估CCR2轴/MOC在中枢神经系统损伤、神经症状和 CME期间死亡率；c)为CCR2+单核细胞诱导的概念提供额外的机械证据 CME中的中枢神经系统损伤。目标2：建立CCR2+MOC在CME期间促进中枢神经系统Th1反应：a)以 确定MOC是否产生CXCR3配体，并评估T细胞在中枢神经系统的募集是否 通过CXCR3信号调节；b)确定CCR2+MOC是否有助于T细胞募集， C)评估过继转移的Th1细胞是否来自感染的CCR2+/+ 小鼠进入CCR2-/-小鼠可恢复与CME相关的损伤。目的3.确定iNOS/NO是否来自CCR2+ 需要MOC来调解在CME过程中观察到的病理CNS组织损伤：a)具体测试 CCR2+MOC中iNOS的诱导是否对CME的中枢神经系统病理有机械作用；b) 确定干扰素γ/干扰素γR信号在CCR2+MOC中是否需要诱导这些细胞中的iNOS以及 干扰素γ/干扰素γR信号在随后的中枢神经系统损伤中的作用 研究计划和方法：本方案将利用我们新的CM小鼠模型，该模型准确地 概述了新生葡萄球菌中枢神经系统感染患者经历的严重的矛盾免疫反应。 我们先前已经证明，产生γ的超极化干扰素CD4+T细胞聚集在中枢神经系统和 与中枢神经系统的病理和死亡率有关。在拟议的工作中，我们将使用我们的新生隐球菌体内模型 CNS感染与转基因小鼠一起测试我们假设的特定方面，在体外T细胞 共培养，而体内采用细胞转移将帮助我们确定中枢神经系统组织损伤的类型和 CCR2+和髓系细胞及其他髓系细胞亚群在中枢神经系统免疫病理中的作用具体来说， 这些策略将使用基因缺陷的小鼠，包括那些缺乏CCR2(CCR2-/--受体)的小鼠 选择性地从CCR2+细胞(CCR2γR-/-)、CXCR3-信号转导(CXCR3-/-)和小鼠未标记细胞中去除 一氧化氮(iNOS-/-)或仅CCR2+细胞CCR2-NOS-/-)。我们实验的结果 操作将通过a)真菌负荷分析b)神经症状分析和 死亡率c)浸润性免疫细胞的流式细胞术分析，d)单核巨噬细胞的分析 极化曲线和效应器功能，e)基因表达分析和f)组织学。总体而言，这些研究 将为真菌CNS感染过程中T细胞介导的病理提供重要的洞察力，并可能 确定新的生物标记物或治疗机会。