Molecular Control of Astrocytes in CNS Inflammation

项目总结/摘要 多发性硬化（MS）是一种慢性炎症性中枢神经系统神经退行性疾病 (CNS)是导致年轻人残疾的主要原因，影响了大约40万美国公民， 每年约100亿美元的经济负担。MS是由一种不完全理解的 遗传和环境因素之间的相互作用，触发针对CNS的自身免疫反应 髓磷脂慢性CNS炎症诱导CNS驻留细胞（如星形胶质细胞）的促炎程序 和小胶质细胞，它们对目前可用于MS的治疗方法没有反应。 丰富的CNS驻留细胞参与健康和疾病中CNS稳态的多个方面， 包括在MS及其动物模型背景下的促炎信号传导，实验性自身免疫 脑脊髓炎（EAE）。因此，研究调节星形胶质细胞促炎活性的机制 可以确定MS的疾病发病机制，以及新的有效疗法，特别是用于 它的进步阶段。在以前的研究集中在MS的环境因素，我们确定了一个信号， 星形胶质细胞中受环境污染物控制并驱动星形胶质细胞致病活动的途径 在EAE和MS中促进炎症和神经变性。具体地说，我们发现， 内质网（ER）定位的受体SigmaR 1稳定肌醇需要酶1-α（IRE 1a），导致 转录因子X-box结合蛋白1（XBP 1）的激活，其促进促炎基因 在星形胶质细胞中表达。在遗传扰动研究中，我们证明了SigmaR 1驱动的IRE 1a-XBP 1 激活增强星形胶质细胞中促炎和神经毒性转录程序的表达， 如Nos 2、Ccl 2、IL 6、Csf 2（GM-CSF）和Csf 2 ra（GM-CSF受体）。此外，我们发现， 在定位于MS病变的星形胶质细胞中增加IRE 1a-XBP 1活化。我假设SigmaR 1-IRE 1a- XBP 1信号转导驱动EAE和MS中的星形胶质细胞致病活性。因此，我提出以下目的： 目标1：辅导阶段（K99）。定义由SigmaR 1-IRE 1a-XBP 1信号转导驱动的星形胶质细胞亚群 (XBP1使用单细胞RNA测序（scRNA-seq），在EAE（Aim 1.1）和MS（Aim 1.2）中检测星形胶质细胞（+星形胶质细胞）。 目标2：辅导阶段（K99）。测试临床上抑制XBP 1信号传导的治疗潜力- 使用EAE小鼠模型（目的2.1-2.2）和scRNA-seq（目的2.3）的相关SigmaR 1抑制剂。 目标3：独立研究者阶段（R 00）。研究GM-CSF信号在XBP 1+星形胶质细胞中的调控 使用空间转录组学方法，包括NICHE-seq（Aim 3.1）和MERFISH（Aim 3.2）。 总之，这些研究将定义一种新的疾病相关星形胶质细胞群体，确定分子机制， 控制它的机制，并评估其药理学操作的治疗价值。

PROJECT SUMMARY/ABSTRACT Multiple sclerosis (MS) is a chronic inflammatory neurodegenerative disorder of the central nervous system (CNS) and is the leading cause of disability in young adults, afflicting some 400,000 U.S. citizens and generating an economic burden of approximately $10 billion annually. MS results from an incompletely understood interaction between genetic and environmental factors that triggers an autoimmune response against CNS myelin. Chronic CNS inflammation induces pro-inflammatory programs in CNS-resident cells such as astrocytes and microglia, which are not responsive to the therapeutic approaches currently available for MS. Astrocytes are abundant CNS-resident cells which participate in multiple aspects of CNS homeostasis in health and disease, including pro-inflammatory signaling in the context of MS and its animal model, experimental autoimmune encephalomyelitis (EAE). Thus, the study of the mechanisms that regulate astrocyte pro-inflammatory activities may identify mechanisms of disease pathogenesis in MS, as well as novel efficacious therapies, particularly for its progressive phase. In previous studies focused on environmental factors in MS, we identified a signaling pathway in astrocytes that is controlled by environmental pollutants, and drives astrocyte pathogenic activities that promote inflammation and neurodegeneration in EAE and MS. Specifically, we found that the endoplasmic reticulum (ER)-localized receptor SigmaR1 stabilizes the inositol requiring enzyme 1-alpha (IRE1a), leading to the activation of the transcription factor X-box binding protein 1 (XBP1) which promotes pro-inflammatory gene expression in astrocytes. In genetic perturbation studies we demonstrated that SigmaR1-driven IRE1a-XBP1 activation boosts the expression of pro-inflammatory and neurotoxic transcriptional programs in astrocytes such as Nos2, Ccl2, Il6, Csf2 (GM-CSF), and Csf2ra (the GM-CSF receptor) during EAE. Moreover, we detected increased IRE1a-XBP1 activation in astrocytes localized to MS lesions. I hypothesize that SigmaR1-IRE1a- XBP1 signaling drives astrocyte pathogenic activities in EAE and MS. Thus, I propose the following Aims: AIM 1: Mentored phase (K99). Define astrocyte subpopulations driven by SigmaR1-IRE1a-XBP1 signaling (XBP1+ astrocytes) in both EAE (Aim 1.1) and MS (Aim 1.2) using single-cell RNA sequencing (scRNA-seq). AIM 2: Mentored phase (K99). Test the therapeutic potential of suppressing XBP1 signaling with clinically- relevant SigmaR1 inhibitors using EAE mouse models (Aims 2.1-2.2), and scRNA-seq (Aim 2.3). AIM 3: Independent investigator phase (R00). Study the regulation of GM-CSF signaling in XBP1+ astrocytes using spatial transcriptomic approaches including NICHE-seq (Aim 3.1) and MERFISH (Aim 3.2). Taken together, these studies will define a novel disease-associated astrocyte population, identify the molecular mechanisms that control it, and evaluate the therapeutic value of its pharmacologic manipulation.