Molecular Control of Astrocytes in CNS Inflammation

多发性硬化症（MS）是一种中枢神经系统（CNS）慢性炎症性神经退行性疾病，是年轻人残疾的主要原因，困扰着约40万美国公民，每年造成约100亿美元的经济负担。MS是由遗传和环境因素之间的不完全理解的相互作用引起的，该相互作用触发针对CNS髓磷脂的自身免疫反应。慢性CNS炎症诱导CNS驻留细胞如星形胶质细胞和小胶质细胞中的促炎程序，其对目前可用于MS的治疗方法不响应。星形胶质细胞是丰富的CNS驻留细胞，其参与健康和疾病中CNS稳态的多个方面，包括MS及其动物模型实验性自身免疫性脑脊髓炎（EAE）背景下的促炎信号传导。因此，调节星形胶质细胞促炎活性的机制的研究可以确定MS中的疾病发病机制，以及新的有效疗法，特别是对于其进行性阶段。在以前的研究集中在MS中的环境因素，我们确定了星形胶质细胞中的信号通路，该信号通路由环境污染物控制，并驱动星形胶质细胞的致病活动，促进EAE和MS中的炎症和神经变性。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，其促进星形胶质细胞中的促炎基因表达。在遗传扰动研究中，我们证明了SigmaR 1驱动的IRE 1a-XBP 1激活增强了EAE期间星形胶质细胞中促炎和神经毒性转录程序如Nos 2、Ccl 2、I16、Csf 2（GM-CSF）和Csf 2 ra（GM-CSF受体）的表达。此外，我们检测到MS病变部位的星形胶质细胞中IRE 1a-XBP 1活化增加。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：AIM 1：指导阶段（K99）。使用单细胞RNA测序（scRNA-seq）在EAE（Aim 1.1）和MS（Aim 1.2）中定义由SigmaR 1-IRE 1a-XBP 1信号传导（XBP 1+星形胶质细胞）驱动的星形胶质细胞亚群。目标2：辅导阶段（K99）。使用EAE小鼠模型（目的2.1-2.2）和scRNA-seq（目的2.3）测试用临床相关SigmaR 1抑制剂抑制XBP 1信号传导的治疗潜力。目标3：独立研究者阶段（R 00）。使用空间转录组学方法（包括NICHE-seq（Aim 3.1）和MERFISH（Aim 3.2））研究XBP 1+星形胶质细胞中GM-CSF信号传导的调节。总之，这些研究将定义一个新的疾病相关的星形胶质细胞群体，确定控制它的分子机制，并评估其药理学操作的治疗价值。

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.