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: 8841025
• 负责人: JOHN E RASH
• 金额: $ 39.07万
• 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 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8841025
• 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; Health; 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; aquaporin 4; base; capillary; immunocytochemistry; insight; leukodystrophy; novel; novel therapeutic intervention; particle; protein K; 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(申请人提供)：以前，没有概念性框架将脑白质营养不良中的各种分子和细胞破坏联系起来，形成一个连贯的模型，以产生作为这些疾病标志的广泛性中枢神经系统白质破坏。对于星形胶质细胞的分子破坏来说，缺乏连贯的模型尤其成问题，因为星形胶质细胞远离形成硬化的少突胶质髓鞘。我们最近引入了“门户假说”来解释表征这些疾病的中枢神经系统髓鞘的广泛破坏。我们认为泛发性髓鞘硬化症是由组成脑室合体细胞内和细胞间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K+和水被突变或破坏时，会扰乱离子的动态平衡，主要是因为K+和水继续进入苍白球合体，但在它们可以退出之前被阻止。Gateway假说表明，减少K+进入苍白球合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(有髓轴突的电压门控通道)和Cx29(一种知之甚少的不形成缝隙连接的少突胶质细胞连接蛋白)被认为是水和K+进入苍白球合体的“门户”。因此，这些紧密相关的蛋白质，以及兰维尔结节上的电压门控钠通道，被认为是许多脑白质营养不良的潜在药物干预靶点。其基本原理是，通过减少轴突Na+内流和/或K+外流及其与周围髓鞘的耦合内流，可以充分减少导致髓鞘肿胀和硬化的渗透负荷，使未受损的K+和水运输途径重新分配药物降低的渗透负荷，从而允许正常的细胞修复机制部分恢复髓鞘功能，从而允许更长期的治疗，可能包括神经胶质干细胞替代疗法。我们建议使用：1)超微结构和超分辨光镜免疫细胞化学；2)分子下拉实验；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学进行监测；4)从正常、运动野生型和Cx29基因敲除小鼠的急性脑片中记录少突胶质细胞。因此，我们将描述轴突质膜上的KV1通道和邻近最内层髓鞘中的Cx29通道，并建立这些分子之间的功能相互作用，作为进入苍白球合体的K+“门户”。有了这些新的数据，Gateway假说将为确定哪些脑白质营养不良症立即适用于药物干预和/或干细胞疗法提供框架，从而使医疗资源能够提供给能够得到最有效治疗的患者。