Group 1 Metabotropic Glutamate Receptor’s Influence on T Cells During Chronic Toxoplasma Infection

项目摘要/摘要 弓形虫是世界上最有效的传播病原体之一，可感染 大约有20亿人。寄生虫在大脑神经元中的包囊导致终生慢性 感染。在大脑内，促炎反应是防止寄生虫重新激活的关键。感染在 免疫功能低下会导致致命性弓形虫脑炎，而在免疫功能正常的人中，有 持续性低度炎症，无临床症状。这表明，有一个严格监管的 脑部对弓形虫的炎症反应。T细胞是主要的免疫细胞，控制 通过分泌穿孔素和干扰素-γ等效应分子进行复发和寄生虫复制。 然而，关于这些细胞是如何在这个组织中被调控的，人们知之甚少。在慢性感染期间，有一种 细胞外(EC)谷氨酸增加。高EC谷氨酸不是弓形虫感染的特异性，可以发生 在多种病理过程中，但可能是组织特异性免疫细胞的重要环境信号。我们 假设这种富含谷氨酸的环境在T细胞的功能和调节中发挥作用。 初步数据显示，弓形虫感染脑部的T细胞表达G蛋白偶联 代谢性谷氨酸受体(mGluR‘s)、mGluR1和mGluR5。这种表达在T细胞中很丰富 与次级淋巴来源的细胞相比，被招募到大脑的细胞。本项目的目标是：1)定义第一组 感染期间大脑中表达mGluR的T细胞，并确定是否存在功能差异 与外围国家相比。这将通过表征规范T细胞亚群的表面蛋白来完成， 测试效应器分子的分泌，确定它们是否定位于大脑的不同区域，以及 在单细胞水平上表征转录图谱；2)确定第1组mGluR如何调节T 细胞对感染的反应。这将通过跟踪过继转移的mGluR缺陷T细胞来完成 感染野生型受体，并在体外通过激活或抑制这些受体，然后测量 活性、增殖、细胞因子产生和规范的GQ分子的变化；3)决定是否降低 感染大脑中的谷氨酸浓度会改变T细胞向中枢神经的募集情况 系统(CNS)。这将通过表征mGluR表达的T细胞在之后被招募到大脑来完成 头孢曲松治疗，是我们实验室以前使用的一种抗生素，通过以下方式降低大脑中的外源性谷氨酸 增加谷氨酸转运体-1的表达。总体而言，这项提议将帮助我们了解组织如何- 衍生信号影响保护性免疫。此外，我们从我们提议的研究中学到的可能是 扩展到其他领域，如神经退行性研究，因为这些神经疾病中的一些不仅 既有谷氨酸改变的共性，又有T细胞向中枢神经系统的募集。

PROJECT SUMMARY/ABSTRACT Toxoplasma gondii (T. gondii) is one of the most effective transmissible pathogens in the world, infecting approximately two billion people. Encystment of the parasite in neurons in the brain results in a lifelong chronic infection. Within the brain, a pro-inflammatory response is essential to prevent parasite reactivation. Infection in the immunocompromised leads to lethal toxoplasmic encephalitis while in the immunocompetent, there is persistent low-grade inflammation which lacks clinical symptoms. This suggests that there is a tightly regulated inflammatory response to T. gondii in the brain. T cells are the dominant immune cell that control recrudescence and parasite replication through secretion of effector molecules such as perforins and IFN-γ. However, little is known on how these cells are regulated in this tissue. During chronic infection there is an increase in extracellular (EC) glutamate. High EC glutamate is not specific to T. gondii infection and can occur during multiple pathologies but may be an important environmental signal to tissue specific immune cells. We hypothesize that this glutamate-rich environment plays a role in T cell function and regulation. Preliminarily data demonstrate that T cells from the T. gondii -infected brain express the G-protein coupled metabotropic glutamate receptors (mGluR’s), mGluR1 and mGluR5. This expression is enriched in T cells recruited to the brain compared to secondary lymphoid derived cells. This project aims to: 1) define group 1 mGluR expressing T cells in the brain during infection, and determine if there is a functional difference compared to the periphery. This will be done by characterizing surface proteins for canonical T cell subsets, testing the secretion of effector molecules, defining if they localize to distinct regions of the brain, and characterizing the transcriptional profile at a single cell level; 2) determine how group 1 mGluR’s regulate T cells in response to infection. This will be done by tracking adoptively transferred mGluR-deficient T cells in an infected wild type recipient, and by activating or inhibiting these receptors ex vivo, followed by measuring changes in viability, proliferation, cytokine production, and canonical Gq molecules; 3) determine if lowering the glutamate concentrations in the infected brain will alter the T cell recruitment profile to the central nervous system (CNS). This will be done by characterizing mGluR expressing T cells recruited to the brain after ceftriaxone treatment, an antibiotic previously used in our lab to lower exogenous glutamate in the brain by increasing glutamate transporter-1 expression. Overall, this proposal will help us understand how tissue- derived signals influence protective immunity. Furthermore, what we learn from our proposed studies could be broaden to other fields such as neurodegenerative research, as some of these neurological maladies not only have a commonality of glutamate alteration, but also a recruitment of T cells to the CNS.