Analyzing Oligodendrocytes-Derived Exosomes in Mouse Models of Multiple Sclerosis

总结 多发性硬化是一种进行性多灶性脱髓鞘疾病， 全球百万人。它是年轻人神经功能障碍的主要原因。的贡献 对这种疾病的免疫系统已经建立。靶向B和T淋巴细胞的多种治疗已经被广泛应用。 针对最流行的复发缓解型疾病（RRMS）开发。然而， 疾病仍然不清楚。人多发性硬化与啮齿类实验性自身免疫性脑脊髓炎的研究 (EAE)模型表明，神经系统来源的髓鞘抗原是重要的发展， 疾病然而，这些抗原信号的形式以及它们如何被转运到外周淋巴细胞， 组织仍然知之甚少。最近的证据表明，成熟细胞分泌的外泌体 少突胶质细胞（OL）富含多种髓鞘抗原。外泌体是分泌的囊泡， 越来越多地被认为在小区-小区通信中发挥重要作用。重要的是， 外泌体已被报道在MS患者外周。OL衍生的外泌体的特定作用， 人类MS或啮齿动物EAE尚未研究。我们假设外泌体是由髓鞘形成 细胞，特别是OL，可以作为中枢和外周髓鞘抗原呈递的早期来源， 最终促进自身反应性淋巴细胞在CNS实质中的致病性积累。具体地说， 我们建议通过首先分析髓鞘形成细胞衍生的原位动力学来研究这一假设。 使用细胞类型特异性外泌体在Aim 1中小鼠EAE的CNS和外周免疫组织中的外泌体 报告员鼠标线，最近开发的MPI实验室。对于这些实验，我们将使用修改后的EAE模型 Cup-EAE包括使用cuprizone短期治疗破坏中央髓鞘，然后使用Cuprizone短期治疗破坏中央髓鞘。 诱导外周免疫应答。然后，我们将检查外泌体到达各种细胞的能力， 外周组织以及对照组与Cup-EAE组CNS内相关细胞类型的摄取 条件我们还计划扩大我们有希望的初步数据，以进一步描述保护， EAE由外泌体释放抑制剂引起。在目标2中，我们将专注于理解 通过对照和MS相关因子（TNF α和谷氨酸）处理的OL体外产生的外来体。各种 将评估这些囊泡的性质，包括大小、货物、抗原呈递细胞（APC）的摄取， 以及这些外泌体在体外支持APC向髓鞘特异性T细胞呈递髓鞘抗原的能力。 总的来说，我们相信我们利用创新的外泌体报告小鼠的提议将提供重要的和有益的结果。 开始理解髓鞘形成细胞产生的外泌体在MS中的作用， 特别强调OL衍生的含髓鞘外泌体。

Summary Multiple Sclerosis is a progressive multifocal demyelinating disease which affects approximately 2.5 million people worldwide. It is the leading cause of neurologic disability in young adults. The contribution of the immune system to this disease is well established. Multiple treatments targeting B and T lymphocytes have been developed against the most prevalent Recurring Remitting form of the disease (RRMS). However, etiology of the disease remains unclear. Studies from human MS and rodent Experimental Autoimmune Encephalomyelitis (EAE) models suggest that nervous system-derived myelin antigens are important in the development of the disease. However, the format of these antigenic signals and how they are transported to the peripheral lymphoid tissues remain poorly understood. Recent evidence suggest that exosomes secreted by mature oligodendrocytes (OLs) are enriched in a variety of myelin antigens. Exosomes are the secreted vesicles that are increasingly appreciated to play important roles in cell-cell communications. Importantly, myelin-containing exosomes have been reported peripherally in MS patients. The specific roles of OL-derived exosomes either in human MS or rodent EAE has not been investigated. We hypothesize that exosomes produced by myelinating cells, especially OLs, may serve as early sources for central and peripheral myelin antigen presentation, ultimately facilitating pathogenic accumulation of autoreactive lymphocytes in the CNS parenchyma. Specifically, we propose to investigate this hypothesis by first analyzing in situ dynamics of the myelinating cell-derived exosomes in CNS and peripheral immune tissues in mouse EAE in Aim 1 using a cell-type specific exosome reporter mouse line, recently developed by the MPI lab. For these experiments we will use a modified EAE model (Cup-EAE) which includes both disruption of central myelin using short treatment with cuprizone followed by the induction of the peripheral immune response. We will then examine both the ability of exosomes to reach various peripheral tissues as well as their uptake by relevant cell types within the CNS under control versus Cup-EAE conditions. We also plan to expand on our promising preliminary data to further characterize the protection from EAE by the exosome release inhibitor. In Aim 2, we will focus on understanding the functional properties of exosomes produced in vitro by control and MS-associated factors (TNFa and glutamate)-treated OLs. Various properties of these vesicles will be assessed including size, cargo, uptake by antigen presenting cells (APCs), and ability of these exosomes to support myelin antigen presentation by APCs to myelin-specific T cells in vitro. Overall, we believe that our proposal utilizing an innovative exosome reporter mouse will provide important and informative data to begin the understanding of the roles of exosomes produced by myelinating cells in MS with specific emphasis on OL-derived myelin-containing exosomes.