Modulating an Astrocyte Hemichannel to Delay Spatial and Temporal Progression in ALS.

调节星形胶质细胞半通道以延迟 ALS 的空间和时间进展。

• 批准号: 10183356
• 负责人: NICHOLAS J MARAGAKIS
• 金额: $ 38.48万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10183356
• 关键词: ALS patients; Affect; Amyotrophic Lateral Sclerosis; Anatomy; Astrocytes; Astrocytosis; Autopsy; Behavioral; Biological Markers; Biology; Blood - brain barrier anatomy; Blood Vessels; Body part; Brain; Buffers; Cell model; Cells; Cessation of life; Chronic; Clinical; Clinical Trials; Coculture Techniques; Complement; Connexin 43; Connexins; Coupled; Data; Development; Diagnosis; Disease; Disease Progression; Dose; Electrophysiology (science); Environment; Event; Eye; Functional disorder; Gap Junctions; Glutamates; Human; Hyperactivity; In Situ; In Vitro; Knock-out; Learning; Libraries; Link; Measures; Mediating; Membrane; Metabolic; Modeling; Motor Neurons; Mus; Nerve Degeneration; Neuraxis; Neurodegenerative Disorders; Neurons; Oral; Pathologic; Pathway interactions; Patient observation; Patients; Pharmacology; Physiology; Play; Probability; Property; Proteins; Proteomics; Publications; Publishing; Regulation; Reporting; Rodent Model; Role; Safety; Spinal Cord; Symptoms; Synapses; Testing; Therapeutic; Time; Tissues; Toxic effect; Translating; Treatment Efficacy; clinical phenotype; design; disease phenotype; epigenomics; extracellular; frontal lobe; human tissue; in vitro Model; in vivo; induced pluripotent stem cell; insight; metabolomics; mouse model; multi-electrode arrays; neuron loss; neuroprotection; neurotoxic; neurotoxicity; novel; patient population; release factor; repository; superoxide dismutase 1; transcriptomics

Project Summary/Abstract Understanding why disease progression in the majority of patients with Amyotrophic Lateral Sclerosis (ALS) occurs in contiguous anatomic regions over time is one of the fundamental limitations to designing disease modifying therapies that can be utilized after a diagnosis. Several studies in ALS rodent models suggest that astrocytes play a role in disease propagation after onset. However, astrocyte dysfunction is not an observation merely limited to ALS rodent models but, importantly, has been one of the most consistent observations in humans with ALS when examined in situ as well as using human cells in vitro. Astrocytes form a highly coupled intercellular network in the central nervous system (CNS) through gap junctions (GJs) and hemichannels (HC) composed of 6 connexin subunits arranged around a central pore. Connexins in astrocytes have key roles: homeostatic buffering, synchronization of astrocyte networks, metabolic support for neurons, and regulation of vascular physiology. They can also propagate Ca2+ waves and modulate synaptic events or release gliotransmitters, including glutamate and ATP, through hemichannels. Connexin 43 (Cx43) is the predominant connexin in astrocytes and is expressed ubiquitously in the CNS. Our recently published studies have demonstrated that astrocyte expression of Cx43 is increased in the frontal cortex and spinal cords of ALS patients, an observation mirrored in the SOD1 mouse model of ALS. This phenomenon is not merely a non-specific effect of reactive astrocytosis as we were also able to show, using in vitro and in vivo mouse modeling, that SOD1 astrocyte-mediated motor neuron toxicity was, at least in part, mediated through Cx43 hemichannels. This proposal builds upon these initial observations by using a fully humanized, spinal cord-specific, ALS iPSC-astrocyte/motor neuron platform to investigate Cx43 HC localization at the hiPSC-astrocyte membrane, examine mechanisms by which Cx43 HC opening is modulated in the context of ALS, and understand how this affects Cx43 HC-mediated ALS hiPSC-astrocyte toxicity to motor neurons. Using newly reported specific blockers of HC activity we will now be able to dissect the specific contributions of Cx43 HC to motor neuron death. As we think about translational potential, we will test these Cx43 HC blockers that can penetrate the blood brain barrier, are well tolerated and orally available, for efficacy in human iSPC-derived astrocyte mediated MN toxicity platforms and in two ALS rodent models. What we learn from this proposal will allow us to develop a systematic approach to ALS therapeutics that combines human tissue data with novel in vitro modeling, and finally to in vivo translational applications.

项目摘要/摘要 了解为什么大多数肌萎缩性侧性硬化症患者的疾病进展（ALS） 随着时间的流逝，发生在连续的解剖区域是设计疾病的基本局限性之一 修改诊断后可以使用的疗法。 ALS啮齿动物模型中的几项研究表明，星形胶质细胞在疾病传播中起作用 发作。但是，星形胶质细胞功能障碍并不仅限于ALS啮齿动物模型，而是一个观察结果 重要的是，当原位检查ALS的人类中，一直是ALS的最一致观察之一 以及在体外使用人类细胞。 星形胶质细胞通过间隙形成了中枢神经系统（CNS）中高度耦合的细胞间网络 连接（GJS）和半通道（HC）由6个连接蛋白亚基组成，该亚基在中央孔周围排列。 星形胶质细胞中的连接素具有关键作用：体内缓冲，星形胶质细胞网络的同步，代谢 对神经元的支持和血管生理的调节。他们还可以传播Ca2+波并调节 突触事件或通过半通道（包括谷氨酸和ATP）释放神经闪光灯。连接蛋白43 （CX43）是星形胶质细胞中的主要连接素，在中枢神经系统中泛表达。 我们最近发表的研究表明，CX43的星形胶质细胞表达在 ALS患者的额叶皮质和脊髓，在ALS的SOD1小鼠模型中反映了一种观察结果。这 现象不仅是反应性星形细胞增多症的非特异性作用，因为我们也能够使用 体外和体内小鼠建模，SOD1星形胶质细胞介导的运动神经元毒性至少部分是 通过CX43半通道介导。 该提案通过使用完全人性化的脊髓特异性ALS建立在这些最初的观察基础上 IPSC-ASSTROCYTE/运动神经元平台，以研究HIPSC-ASSTROCYTE膜的CX43 HC定位， 检查在ALS的背景下调制CX43 HC开口的机制，并了解如何 影响CX43 HC介导的ALS HIPSC-腹腔细胞对运动神经元的毒性。使用新报告的特定 HC活动的阻滞剂我们现在能够剖析CX43 HC对运动神经元的特定贡献 死亡。当我们考虑转化潜力时，我们将测试可以穿透的CX43 HC阻滞剂 血脑屏障，耐受性且口服良好，可在人ISPC衍生的星形胶质细胞中介导的功效 MN毒性平台和两个ALS啮齿动物模型。我们从该建议中学到的东西将使我们能够发展 对ALS治疗剂的系统方法，将人类组织数据与新型体外建模相结合，并 最后到体内翻译应用。