Molecular Control of Astrocytes in CNS Inflammation

多发性硬化症(MS)是一种慢性中枢神经系统(CNS)炎症性神经退行性疾病，是导致年轻人残疾的主要原因，每年困扰约40万美国公民，产生约100亿美元的经济负担。多发性硬化症是由于遗传和环境因素之间的相互作用尚未完全了解，这种相互作用触发了对中枢神经系统髓鞘的自身免疫反应。慢性中枢神经系统炎症诱导中枢神经系统驻留细胞(如星形胶质细胞和小胶质细胞)的促炎程序，而星形胶质细胞是丰富的中枢神经系统驻留细胞，参与健康和疾病中中枢神经系统动态平衡的多个方面，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。因此，对星形胶质细胞促炎活性调节机制的研究可能会发现MS的发病机制，以及新的有效治疗方法，特别是对其进展期的治疗。在以往对MS环境因素的研究中，我们发现星形胶质细胞中的信号通路受环境污染物的控制，并驱动星形胶质细胞的致病活动，促进EAE和MS中的炎症和神经退变。具体地说，我们发现内质网(ER)定位的受体SigmaR1稳定肌醇需要酶1-α(IRE1a)，导致转录因子X-box结合蛋白1(XBP1)的激活，促进星形胶质细胞中促炎基因的表达。在遗传扰动研究中，我们证明了在EAE过程中，SigmaR1驱动的IRE1a-XBP1激活促进了星形胶质细胞中促炎和神经毒性转录程序的表达，如NOS2、CCL2、IL6、CSF2(GM-CSF)和Csf2ra(GM-CSF受体)。此外，我们还检测到定位于MS损伤的星形胶质细胞中IRE1a-XBP1活性增加。我假设SigmaR1-IRE1a-XBP1信号驱动星形胶质细胞在EAE和MS中的致病活动，因此，我提出以下目标：目标1：指导期(K99)。使用单细胞RNA测序(scRNA-seq)确定EAE(AIM 1.1)和MS(AIM 1.2)中由SigmaR1-IRE1a-XBP1信号(XBP1+星形胶质细胞)驱动的星形胶质细胞亚群。目标2：指导阶段(K99)。使用EAE小鼠模型(AIMS 2.1-2.2)和scRNA-seq(AIMS 2.3)，测试临床相关SigmaR1抑制剂抑制XBP1信号转导的治疗潜力。目标3：独立调查员阶段(R00)。用空间转录方法研究XBP1+星形胶质细胞GM-CSF信号的调控，包括NICE-SEQ(Aim 3.1)和MerFish(Aim 3.2)。综上所述，这些研究将定义一个新的与疾病相关的星形胶质细胞群体，识别控制它的分子机制，并评估其药物操作的治疗价值。

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.