The Gateway Hypothesis: A new framework for unraveling diverse leukodystrophies

网关假说：解开多种脑白质营养不良的新框架

• 批准号: 8514423
• 负责人: JOHN E RASH
• 金额: $ 43.44万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-05-01 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8514423
• 关键词: Accounting; Action Potentials; Acute; Adult; Astrocytes; Axon; Binding; Biochemical; Biological Assay; Blood capillaries; Brain; Cell Culture Techniques; Cell membrane; Cells; Central Nervous System Diseases; Connexins; Corpus Callosum; Coupled; Coupling; Cytoplasm; Data; Disease; Electrophysiology (science); Exercise; Failure; Functional disorder; Future; Gap Junctions; Giant Cells; Homeostasis; In Vitro; Intervention; Knockout Mice; Knowledge; Lead; Light; Link; Local Anesthetics; Medical; Microscopic; Modeling; Molecular; Monitor; Mus; Mutate; Mutation; Myelin; Nerve Fibers; Oligodendroglia; Optic Nerve; Pathway interactions; Patients; Pharmaceutical Preparations; Physiological; Potassium; Potassium Channel; Process; Production; Proteins; Ranvier' s Nodes; Rattus; Reducing Agents; Replacement Therapy; Research; Resolution; Resources; Rest; Role; Sclerosis; Site; Slice; Sodium Channel; Spinal; Stem cells; Surface; Swelling; Therapeutic Intervention; Water; Work; Xeno; aquaporin 4; base; capillary; immunocytochemistry; insight; leukodystrophy; novel; novel therapeutic intervention; particle; protein K; public health relevance; repaired; sciatic nerve; spinal cord white matter; stem cell therapy; voltage; voltage gated channel; water channel; white matter; white matter damage

DESCRIPTION (provided by applicant): Previously, there was no conceptual framework for linking the diverse molecular and cellular disruptions seen in the leukodystrophies into a coherent model for the production of the generalized CNS white matter destruction that is the hallmark of these diseases. This lack of a coherent model was particularly problematic for molecular disruptions in astrocytes, which are far removed from the oligodendrocyte myelin that becomes sclerotic. We recently introduced the "Gateway Hypothesis" to account for the widespread destruction of CNS myelin that characterizes these diseases. We proposed that generalized myelin sclerosis is caused by mutation or immunological disruption of proteins comprising the primary transport pathways for K+ and water within and between cells of the panglial syncytium. We and others had identified proteins of the K+ and water transport pathway that, when mutated or destroyed, disrupt ionic homeostasis, primarily because K+ and water continue to enter the panglial syncytium but are blocked before they can exit. The Gateway Hypothesis suggests that pharmacological agents that reduce K+ entry into the panglial syncytium may provide new therapeutic approach to treating these diseases. Two abundant protein molecules of that pathway - KV1 (the voltage-gated channels of myelinated axons) and Cx29 (a poorly-understood oligodendrocyte connexin that does not form gap junctions) - are proposed to represent the "gateway" for entry of water and K+ into the panglial syncytium. As such, these tightly-associated proteins, as well as voltage-gated sodium channels at nodes of Ranvier, are proposed as potential targets for pharmacologic intervention in many of the leukodystrophies. The rationale is that by reducing axonal Na+ influx and/or K+ efflux and its coupled influx into the surrounding myelin, the osmotic burden that causes myelin swelling and sclerosis can be reduced sufficiently to allow undamaged K+ and water transport pathways to redistribute the pharmacologically-reduced osmotic load, thereby permitting normal cellular repair mechanisms to partially restore myelin function, allowing for longer-term treatments, potentially including glial stem cell replacement therapies. We propose to use: 1) ultrastructural and super-resolution light microscopic immunocytochemistry; 2) molecular pull-down assays; 3) expression of Cx29 and KV1 channels in cell culture, with monitoring by intracellular recording electrophysiology; and 4) recording from oligodendrocytes in acute slices of mouse corpus callosum of normal and exercised wildtype and Cx29 knockout mice. We will thus characterize KV1 channels in axon plasma membranes and Cx29 channels in the adjacent innermost layer of myelin and establish functional interaction of those molecules as the K+ "gateway" into the panglial syncytium. With these new data, the Gateway Hypothesis will provide the framework for identifying which of the leukodystrophies are immediately amenable to pharmacologic intervention and/or stem cell therapies, allowing medical resources to be delivered to the patients who can be most effectively treated.

描述（由申请人提供）：以前，没有概念性框架将白质营养不良症中所见的各种分子和细胞破坏联系到产生广泛性中枢神经系统白质破坏的连贯模型中，这是这些疾病的标志。星形胶质细胞的分子破坏与硬化的少突胶质细胞髓磷脂相去甚远，缺乏一致的模型尤其成问题。我们最近引入了“通道假说”来解释这些疾病特征的中枢神经系统髓磷脂的广泛破坏。我们提出，广泛性髓磷脂硬化是由包括K+和水的主要运输途径的蛋白质突变或免疫破坏引起的。我们和其他人已经确定了K+和水运输途径的蛋白质，当突变或破坏时，会破坏离子稳态，主要是因为K+和水继续进入盘状合胞体，但在它们离开之前被阻止。门户假说提示，减少K+进入盘状合胞体的药物可能为治疗这些疾病提供新的治疗方法。该途径的两个丰富的蛋白质分子- KV1（髓鞘轴突的电压门控通道）和Cx29（一种不形成间隙连接的少突胶质细胞连接蛋白）-被认为是水和K+进入盘状合胞体的“门户”。因此，这些紧密相关的蛋白，以及Ranvier淋巴结上的电压门控钠通道，被认为是许多白质营养不良的药物干预的潜在靶点。其基本原理是，通过减少轴突Na+流入和/或K+流出及其耦合流入周围髓鞘，可以充分减少导致髓鞘肿胀和硬化的渗透负担，从而允许未受损的K+和水运输途径重新分配药理学上减少的渗透负荷，从而允许正常的细胞修复机制部分恢复髓鞘功能，从而允许长期治疗。可能包括神经干细胞替代疗法。我们建议使用：1)超微结构和超分辨率光镜免疫细胞化学；2)分子下拉试验；3)细胞培养中Cx29和KV1通道的表达，并通过细胞内记录电生理监测；4)正常小鼠、运动野生型小鼠和Cx29敲除小鼠胼胝体急性切片少突胶质细胞的记录。因此，我们将表征轴突质膜中的KV1通道和相邻髓鞘最内层的Cx29通道，并建立这些分子作为K+“门户”进入pangligal合胞体的功能相互作用。有了这些新数据，网关假说将为确定哪些白质营养不良症可以立即进行药物干预和/或干细胞治疗提供框架，从而使医疗资源能够提供给最有效治疗的患者。