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。

基本信息

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
项目类别：
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

项目摘要

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.

项目概要/摘要了解大多数肌萎缩侧索硬化症 (ALS) 患者疾病进展的原因随着时间的推移发生在连续的解剖区域是设计疾病的基本限制之一修改诊断后可以使用的疗法。 ALS 啮齿动物模型的多项研究表明，星形胶质细胞在疾病传播中发挥作用发病。然而，星形胶质细胞功能障碍并不仅仅限于 ALS 啮齿动物模型，而是，重要的是，这是对 ALS 患者进行原位检查时最一致的观察结果之一以及在体外使用人体细胞。星形胶质细胞通过间隙在中枢神经系统（CNS）中形成高度耦合的细胞间网络连接点 (GJ) 和半通道 (HC) 由围绕中心孔排列的 6 个连接蛋白亚基组成。星形胶质细胞中的连接蛋白具有关键作用：稳态缓冲、星形胶质细胞网络同步、代谢支持神经元和调节血管生理学。它们还可以传播 Ca2+ 波并调制突触事件或通过半通道释放胶质递质，包括谷氨酸和 ATP。连接蛋白43 (Cx43) 是星形胶质细胞中主要的连接蛋白，在中枢神经系统中广泛表达。我们最近发表的研究表明，星形胶质细胞 Cx43 的表达在 ALS 患者的额叶皮层和脊髓，这一观察结果反映在 ALS 的 SOD1 小鼠模型中。这正如我们也能够证明的那样，这种现象不仅仅是反应性星形细胞增多症的非特异性效应，使用体外和体内小鼠模型表明，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。该提案建立在这些初步观察的基础上，使用完全人性化、脊髓特异性的 ALS iPSC-星形胶质细胞/运动神经元平台用于研究 Cx43 HC 在 hiPSC-星形胶质细胞膜上的定位，检查在 ALS 背景下调节 Cx43 HC 打开的机制，并了解这是如何调节的影响 Cx43 HC 介导的 ALS hiPSC 星形胶质细胞对运动神经元的毒性。使用新报告的具体 HC 活性的阻断剂我们现在能够剖析 Cx43 HC 对运动神经元的具体贡献死亡。当我们考虑转化潜力时，我们将测试这些 Cx43 HC 阻断剂，它们可以穿透血脑屏障，耐受性良好，可口服，对人类 iSPC 衍生的星形胶质细胞介导的疗效 MN 毒性平台和两种 ALS 啮齿动物模型。我们从这个提案中学到的东西将使我们能够发展 ALS 治疗的系统方法，将人体组织数据与新颖的体外模型相结合，以及最终进入体内转化应用。