Mitochondrial ROS and Microglia in Rett Syndrome

Rett 综合征中的线粒体 ROS 和小胶质细胞

• 批准号: 9311039
• 负责人: LEE-WAY JIN
• 金额: $ 32.14万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-10 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9311039
• 关键词: Acute; Animals; Area; Autistic Disorder; Automobile Driving; Bioenergetics; Brain; Characteristics; Data; Defect; Disease; Drug Kinetics; Engineering; Enzymes; Epigenetic Process; Erythroid; FDA approved; Foundations; Fumarates; Functional disorder; Gene Mutation; Genes; Impairment; In Vitro; Intervention; Knock-out; Knockout Mice; Lead; Leukocytes; Link; Longevity; Mediating; Metabolic; Methyl-CpG-Binding Protein 2; Microglia; Mitochondria; Modeling; Molecular; Motor; Mus; Neurodevelopmental Disorder; Neurologic Symptoms; Nuclear; Oxidation-Reduction; Oxidative Stress; Pathogenesis; Pathogenicity; Pathologic; Pathology; Pathway interactions; Phagocytes; Phagocytosis; Pharmaceutical Preparations; Phenotype; Play; Positioning Attribute; Production; Reactive Oxygen Species; Regulation; Reporting; Research; Respiratory physiology; Rett Syndrome; Role; Safety; Signal Transduction; Symptoms; Testing; Therapeutic; Therapeutic Effect; Transgenes; Translating; Translations; Work; aerobic glycolysis; antioxidant enzyme; autism spectrum disorder; base; catalase; effective therapy; flexibility; functional disability; girls; improved; in vivo; loss of function mutation; mouse model; novel; novel therapeutics; oxidation; pre-clinical; protective effect; selective expression; success; therapeutic evaluation

Project Summary/Abstract Rett syndrome (RTT), a devastating neurodevelopmental disorder, is largely caused by loss-of-function mutations in the X-linked gene encoding the epigenetic regulator MeCP2. Our group was the first to show a detrimental role of microglial abnormalities in a MECP2 knockout (MECP2-KO) mouse model of RTT. Recent in vivo data suggest that correcting microglial abnormalities is sufficient to rectify major RTT-like symptoms in this model. Therefore, how MeCP2 deficiency causes microglial abnormalities and how the functional states of MeCP2-deficient microglia (RTT-MG) influence RTT pathology is a critical question. Recently we found that MeCP2 deficiency in RTT-MG resulted in enhanced mitochondrial reactive oxygen species (mtROS) production, which we hypothesize would lead to metabolic derangements and abnormal microglial functions. Interestingly, in vitro studies showed that several key RTT-MG abnormalities were rescued by mitochondria- targeted transgene mCAT (the antioxidant enzyme catalase being expressed only in mitochondria, which usually do not harbor catalase) and two FDA-approved mitoactive Nrf2 activators, opening a possibility for intervention. Our preliminary data further showed that global expression of mCAT (GL-mCAT) prolonged the lifespan and improved motor and respiratory functions of the MECP2-KO mice, supporting the promise of our approach in a whole animal setting. Here we propose to extend this line of research to gain a deeper understanding of how MeCP2 is related to functional and pathological states of microglia, and to generate preclinical proof of principle that is instrumental for developing novel therapies for RTT: Aim 1: Determine the pathological role of microglial mtROS in vivo: Several lines of evidence support that microglia abnormalities drive the progression of RTT. We have shown that global expression of mCAT ameliorates the RTT-like phenotype in MECP2-KO mice. Based on this encouraging result, we further hypothesize that quenching mtROS selectively in microglia to rectify microglial abnormalities will also ameliorate RTT-like deficits in MECP2-KO mice. For this aim, we will generate and analyze the phenotype and microglial pathology of MECP2-KO/MG-mCAT mice with microglia-targeted expression of mCAT. The result would support the role of microglial mtROS in the pathogenesis of RTT. Aim 2: Determine the role of mtROS in pathological characteristics and functional responsiveness of RTT microglia: Our previous data, mostly in vitro work, support the hypothesis that mtROS-related metabolic/molecular derangements lead to RTT-MG abnormalities including the loss of the “metabolic flexibility” required to drive their functional responsiveness. Now in this aim we will test this hypothesis in vivo, mainly by analyzing microglia acutely isolated from MECP2-KO and MECP2-KO/mCAT mice. We will determine if mCAT, expressed in vivo, is able to (a) rectify the metabolic/molecular derangements of RTT-MG, and (b) recover the ability of RTT-MG to respond to M1/M2-inducing signals with proper functional differentiation. Key microglial cellular features such as phagocytic function, mitochondrial integrity, aerobic glycolysis, functional polarization, and Nrf2 antioxidation pathway will be analyzed to determine the beneficial effects of mtROS suppression by mCAT. Aim 3: Test the therapeutic effects of two FDA-approved mitoactive/Nrf2 activating drugs in vivo: The success of the mCAT approach provides a rationale for anti-mtROS therapy. To translate this finding to potential therapies, we screened 1,600 FDA-approved drugs to identify mitoactive anti-mtROS compounds, and subsequently screened the hits on a RTT-MG culture platform. We now have two leads able to correct RTT- MG abnormalities in vitro. Interestingly, both are known activators of the Nrf2 antioxidation pathway, supporting our hypothesized pivotal role of Nrf2 in RTT-MG. Because of their known favorable pharmacokinetics/safety profiles, now we will test them in vivo; efficacy shown in this aim would enable rapid translation by re-purposing these FDA-approved drugs for RTT. In summary, these studies will clarify mechanisms of MeCP2-regulated microglial function, the role of microglial mtROS in RTT, and advance novel therapies for RTT, for which no effective treatment is available. We hope that findings in this proposal would set the foundation to explore several new areas, such as how mtROS and metabolic modules such as aerobic glycolysis mediate or modulate microglial function including M1/M2 differentiation and phagocytosis, and the role of Nrf2 in microglial function. Moreover, RTT is one of few Autism Spectrum Disorders (ASDs) whose cause is identified as a single gene mutation and shares important pathogenic pathways with autism. Our studies, therefore, may implicate novel mechanisms and therapeutic approaches as well for autism where mitochondrial and microglia dysfunction could also play a role.

项目摘要/摘要 RETT综合征（RTT）是一种毁灭性神经发育障碍，主要是由于功能丧失引起的 编码表观遗传调节剂MECP2的X连锁基因中的突变。我们的小组是第一个展示 小胶质细胞异常在RTT的MECP2敲除（MECP2-KO）小鼠模型中的有害作用。最近的 体内数据表明，纠正小胶质的异常足以纠正重大的RTT样症状 这个模型。因此，MECP2缺乏如何引起小胶质细胞异常以及如何功能状态 MECP2缺乏小胶质细胞（RTT-MG）影响RTT病理学是一个关键问题。最近我们发现 RTT-MG中的MECP2缺乏导致线粒体活性氧（MTROS）增强 我们假设的生产将导致代谢进化和异常小胶质细胞功能。 有趣的是，体外研究表明，线粒体对几种关键的RTT-MG异常作出了反应 靶向转化MCAT（抗氧化剂酶过氧化氢酶仅在线粒体中表达 通常不要携带过氧化氢酶）和两个FDA批准的有丝丝NRF2激活剂，为 干涉。我们的初步数据进一步表明，MCAT（GL-MCAT）的全球表达延长了 MECP2-KO小鼠的寿命和改善的运动和呼吸功能，支持我们的承诺 在整个动物环境中接近。在这里，我们建议扩展这一研究，以获得更深入的研究 了解MECP2如何与小胶质细胞的功能和病理状态相关，并生成 临床前原理证明，这对于开发RTT的新疗法有用： AIM 1：确定小胶质MTRO在体内的病理作用：几条证据支持，支持这些线 小胶质细胞异常驱动RTT的进展。我们已经证明了MCAT的全球表达 改善MECP2-KO小鼠中的RTT样表型。基于这个令人鼓舞的结果，我们进一步 假设在小胶质细胞中选择性淬灭MTROS以纠正小胶质细胞异常也将 改善RTT样在MECP2-KO小鼠中定义。为此，我们将生成和分析表型和 MECP2-KO/MG-MCAT小鼠的小胶质细胞病理，具有MCAT的小胶质细胞表达。结果 将支持小胶质细胞在RTT发病机理中的作用。 目标2：确定MTRO在病理特征和RTT功能反应性中的作用 小胶质细胞：我们以前的数据，主要是体外工作，支持了与MTROS相关的假设 代谢/分子演变导致RTT-MG异常，包括“代谢丧失” 提高其功能响应性所需的灵活性”。现在，在此目标中，我们将在体内检验这个假设， 主要是通过分析从MECP2-KO和MECP2-KO/MCAT小鼠中急性分离的小胶质细胞。我们将 确定在体内表达的MCAT是否能够（a）纠正RTT-MG的代谢/分子发展， （b）恢复RTT-MG响应具有适当功能的M1/M2诱导信号的能力 分化。关键的小胶质细胞特征，例如吞噬功能，线粒体完整性，有氧运动 将分析糖酵解，功能极化和NRF2抗氧化途径以确定有益的 MCAT抑制MTROS的影响。 AIM 3：测试两个FDA批准的丝裂/NRF2激活药物的治疗作用：成功 MCAT方法为抗MTROS疗法提供了理由。将此发现转化为潜力 疗法，我们筛选了1,600种FDA批准的药物，以鉴定有丝分裂性抗MTROS化合物，并 随后，在RTT-MG文化平台上筛选了命中。现在，我们有两个能够纠正RTT-的潜在客户 Mg体外异常。有趣的是，两者都是NRF2抗氧化途径的已知激活剂，支持 我们假设的NRF2在RTT-MG中的关键作用。由于他们已知有利的药代动力学/安全性 个人资料，现在我们将在体内对其进行测试；此目标中显示的效率将通过重新定位来快速翻译 这些用于RTT的FDA批准的药物。 总而言之，这些研究将阐明MECP2调节的小胶质功能的机制，即 RTT中的小胶质细胞MTROS，并为RTT提供了新的疗法，为此没有有效的治疗方法。 我们希望该提案中的发现将奠定基础，以探索几个新领域，例如如何 MTROS和代谢模块（例如有氧糖酵解）介导或调节小胶质功能，包括 M1/M2分化和吞噬作用，以及NRF2在小胶质细胞功能中的作用。而且，RTT是少数 自闭症谱系障碍（ASD），其原因被识别为单个基因突变并具有重要意义 自闭症的致病途径。因此，我们的研究可能会实施新颖的机制和治疗 对于自闭症的方法，线粒体和小胶质细胞功能障碍也可以发挥作用。