Initiation of CD8 T cell-mediated BBB disruption and neuronal involvement in experimental cerebral malaria

项目概要/摘要： 恶性疟原虫被世界卫生组织认为是“人类最致命的寄生虫” 疟疾的病原体疟疾感染每年导致大约50万人死亡。 脑型疟疾（CM）是恶性疟原虫感染的致命神经系统并发症， 高死亡率或严重认知缺陷。耐药性、新疗法疗效低和未知 病理学和宿主免疫相互作用的机制突出了迫切需要为此开发解决方案 巨大的全球性问题。我们实验室和其他实验室的工作表明，白细胞在大脑中的积累是 与CM发病和严重程度相关，确立了免疫系统在病理学中的重要作用。的作用 一些免疫细胞亚群是良好建立的，例如CD 8 T细胞的亚群，其已显示介导 感染后期血脑屏障破坏和血管通透性。然而， 启动这种CD 8 T细胞应答的抗原呈递细胞（APC）仍然难以捉摸。我们的实验室将采用新颖的 在C57 BL/6小鼠中发现的两种MHC I类分子（H-2Kb和H-2Db）的细胞类型特异性缺失， 识别此细胞类型。树突状细胞（DC）上任一I类分子缺陷的小鼠被保护免于 脑型疟疾因此，在本建议中，我们计划采用第I类转移足够的预先分类的DC 以鉴定能够启动CD 8 T细胞应答和血脑屏障的DC (BBB)在CM中看到的中断。识别和表征这种细胞类型将在生物医学领域具有广泛的意义。 从发病机理的基本见解以及疫苗策略的改进。后 我们将描述起始事件的病理结果。CM的临床特征为： 脑中VEGF的上调，以及BBB紧密连接蛋白、血管通透性和 MRI可检测到重度水肿。在我们的研究中，我们采用小动物MRI和RNA原位杂交， 诊断这些生物标志物的存在。我们最近证明神经元是一种细胞类型 在疾病状态下负责VEGF转录上调。为了阐明VEGF在以下两个方面的作用： 支持或否定病理学，我们产生了一种新的能够敲除VEGF的诱导型小鼠系， 尤其是神经元。在建立感染后，我们可以诱导神经元VEGF的敲除，并评估 病理阐述VEGF在CM中的作用将具有治疗意义，并提供 了解疾病进展的方法。实现这些目标将使我们能够解决我们的核心问题。 需要特异性DC亚群来引发细胞毒性效应CD 8 T细胞介导的BBB的假设 渗透性和神经元VEGF上调;这加强了ECM中的病理学。这些发现将证明 新颖的任何能力，并将有助于基础知识的几个研究领域，提供机械 深入了解疟疾的发病机制，并确定未来治疗干预的潜在目标。

Project Summary/Abstract: Plasmodium falciparum, deemed “deadliest parasite in humans” by World Health Organization is the causative agent of the disease known as malaria. Malaria infections result in approximately half a million deaths per year. Cerebral malaria (CM) is a deadly neurological complication of Plasmodium falciparum infection resulting in high mortality or severe cognitive deficit. Drug resistance, low efficacy of new therapeutics, and unknown mechanism of pathology and host immune interactions highlight the urgent need to develop solutions for this tremendous global issue. Work from our lab and others have shown that leukocyte accumulation in the brain is correlated with CM onset and severity, establishing a vital role of the immune system in pathology. The roles of some immune cell subsets are well-established such as that of CD8 T cells which have been shown to mediate blood-brain barrier disruption and vascular permeability in the late stages of infection. However, the role of the antigen-presenting cell (APC) that initiates this CD8 T cell response remains elusive. Our lab will employ novel cell-type specific deletion of the two MHC class I molecules found in C57BL/6 mice (H-2Kb and H-2Db) in order identify this cell type. Mice deficient in either class I molecule on dendritic cells ((DC)s) are protected from cerebral malaria. Therefore, in this proposal we plan to adoptively transfer in class I sufficient, pre-sorted DCs of established subsets to identify the DC capable of initiating the CD8 T cell responses and blood-brain barrier (BBB) disruption seen in CM. Identifying and characterizing this cell type will have broad implications in the field ranging from fundamental insights into pathogenesis as well as improvement of vaccine strategies. After addressing the initiating event we will characterize the pathologic outcome. CM is clinically characterized by VEGF upregulation in the brain, and disruption of BBB tight junction proteins, vascular permeability, and severe edema detectable by MRI. In our studies we employ small animal MRI and RNA in situ hybridization to diagnose the presence of these biomarkers. We recently demonstrated that neurons are the cell type responsible for VEGF transcriptional upregulation in the disease state. To address the role of VEGF in either supporting or negating pathology we generated a new inducible mouse line capable of knocking out VEGF specifically in neurons. After establishing infection we can induce knockout of neuronal-VEGF and assess pathology. Elaborating upon the role of VEGF in CM will have therapeutic implications as well as provide insight into methods of disease progression. Completion of these aims will allow us to address our central hypothesis that a specific DC subset is required to elicit cytotoxic effector CD8 T cell-mediated BBB permeability and neuronal VEGF upregulation; which reinforces pathology in ECM. These findings will prove novel in any capacity and will contribute fundamental knowledge to several fields of study, provide mechanistic insight into pathogenesis of malaria and determine potential targets of therapeutic intervention for the future.