The function and mechanisms of voltage-gated proton channel Hv1 in spinal cord injury

电压门控质子通道Hv1在脊髓损伤中的作用及机制

• 批准号: 10398137
• 负责人: Junfang Wu
• 金额: $ 47.57万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-15 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10398137
• 关键词: Acidosis; Acute; Address; Affect; Astrocytes; Attenuated; Brain; Brain Injuries; Brain Ischemia; Cell Death; Charge; Chronic; Clinical; Complex; Contusions; Coupling; Cytoplasm; Data; Development; Electrons; Failure; Financial compensation; Functional disorder; Future; Generations; Genetic; Goals; Hyperesthesia; Immune system; Impairment; In Vitro; Inflammation; Inflammatory; Injury; Interferons; Intervention; Ion Channel; Knockout Mice; Lead; Mediating; Microglia; Modeling; Molecular; Molecular Genetics; Mus; NADP; NADPH Oxidase; Neuraxis; Neuroglia; Neurologic; Neurologic Dysfunctions; Neurological outcome; Neurons; Pathologic; Pathway interactions; Phagocytes; Pharmacology; Phenotype; Process; Production; Protons; Reactive Oxygen Species; Recovery; Recovery of Function; Reporting; Research; Respiratory Burst; Rest; Role; Signal Pathway; Signal Transduction; Spinal Cord Contusions; Spinal cord injury; Testing; Therapeutic Intervention; Transgenic Mice; Up-Regulation; base; effective therapy; functional outcomes; gene therapy; genetic approach; improved; in vivo; inhibitor; innovative technologies; insight; mouse model; neuroinflammation; neuron loss; neuroprotection; neurotoxic; new therapeutic target; novel; novel therapeutics; painful neuropathy; voltage

Project Summary Despite considerable research over the past 30 years, there is still no established effective treatment to improve recovery following spinal cord injury (SCI). In part, this reflects incomplete understanding of the complex secondary pathobiological mechanisms involved. The aim of our research is to understand the cellular and molecular mechanisms responsible for post-injury neuroinflammation in order to allow future development of novel therapies. The voltage-gated proton channel Hv1 is a newly discovered ion channel, highly expressed in resting microglia of the brain. Under pathological conditions, microglial Hv1 is required for NADPH oxidase (NOX)-dependent generation of ROS (reactive oxygen species) by providing charge compensation for exported electrons and relieving intracellular acidosis. Thus, Hv1 is a unique target for controlling multiple NOX activities and ROS production. However, neither the precise signaling mechanisms underlying this finding nor critical role of Hv1 in the pathophysiology of SCI are fully understood. Based on our preliminary data, we will test the hypothesis that microglial Hv1 functions as a key mechanism in neuroinflammation, through altered NOX2/ROS/IFN- signaling that modulates microglia-astrocyte interaction, thus affecting long-term neurological outcomes after SCI. We will use systemic or microglial Hv1 KO, microglial NOX2 KO transgenic mice and in vivo and in vitro innovatively technologies to determine the mechanisms of SCI-triggered Hv1 elevation on post-injury neuroinflammation. Aim 1 will determine the function and mechanisms of the Hv1 in neuroinflammation after SCI. Multiple quantitative assessments of microglia-mediated neuroinflammation will be combined with genetic or pharmacological intervention targeting Hv1 to test the hypothesis that SCI-induced microglial Hv1 activation mediates detrimental neuroinflammation and functional deficits through altered microglial NOX2/ROS signaling. Aim 2 will elucidate the role of microglial NOX2 in post-injury neuroinflammation. We will utilize genetic intervention to delete Hv1-dependent up-regulation of NOX2 in microglia, and evaluate the effects on microglial NOX2 coupling to Hv1 on neuroinflammation after SCI. Aim 3 will identify critical role of Hv1/NOX2-derived ROS/IFN- in SCI-chronic neuroinflammation through microglia-astrocyte interaction. Complimentary cellular, molecular, and genetic approaches will be used to test the hypothesis that Hv1/NOX2-mediated microglial ROS activates pro-inflammatory astrocytes resulting in secreting IFN that in turn reinforces microglial inflammation, thus contributes to astrocytes dysfunction and neuronal damage. Our study will be the first to implicate microglial Hv1/NOX2/ROS/IFN- signaling in the pathophysiology of SCI, leading to novel treatment approaches for SCI. Given the proposed roles for Hv1 in other inflammatory models, Hv1 signaling represents a generic mechanism relevant to other neuroinflammatory states.

项目概要 尽管过去30年进行了大量的研究，但仍然没有确定的有效治疗方法 改善脊髓损伤 (SCI) 后的恢复。这在一定程度上反映了对这一问题的不完全理解。 涉及复杂的继发病理生物学机制。我们研究的目的是了解细胞 和负责损伤后神经炎症的分子机制，以便未来的发展 的新疗法。电压门控质子通道 Hv1 是新发现的离子通道，高表达 在大脑静息的小胶质细胞中。病理条件下，小胶质细胞 Hv1 是 NADPH 氧化酶所必需的 通过提供电荷补偿来产生依赖于 (NOX) 的 ROS（活性氧） 输出电子并缓解细胞内酸中毒。因此，Hv1是控制多种NOX的独特目标 活动和 ROS 产生。然而，这一发现背后的精确信号机制和 Hv1 在 SCI 病理生理学中的关键作用已得到充分了解。根据我们的初步数据，我们将 通过改变小胶质细胞 Hv1 作为神经炎症关键机制的假设 NOX2/ROS/IFN-信号调节小胶质细胞-星形胶质细胞相互作用，从而影响长期 SCI 后的神经系统结果。 我们将使用全身或小胶质细胞Hv1 KO、小胶质细胞NOX2 KO转基因小鼠并进行体内和体外实验 创新技术确定损伤后 SCI 触发 Hv1 升高的机制 神经炎症。目标 1 将确定 Hv1 在神经炎症中的功能和机制 SCI之后。小胶质细胞介导的神经炎症的多重定量评估将与 针对 Hv1 的遗传或药物干预，以检验 SCI 诱导的小胶质细胞 Hv1 的假设 激活通过改变小胶质细胞介导有害的神经炎症和功能缺陷 NOX2/ROS 信号传导。目标 2 将阐明小胶质细胞 NOX2 在损伤后神经炎症中的作用。 我们将利用基因干预来消除小胶质细胞中 Hv1 依赖性的 NOX2 上调，并评估 小胶质细胞 NOX2 与 Hv1 偶联对 SCI 后神经炎症的影响。目标 3 将确定关键 Hv1/NOX2 衍生的 ROS/IFN- 通过小胶质细胞-星形胶质细胞在 SCI 慢性神经炎症中的作用 相互作用。将使用免费的细胞、分子和遗传方法来检验该假设 Hv1/NOX2 介导的小胶质细胞 ROS 激活促炎性星形胶质细胞，导致分泌 IFN 反过来又会加剧小胶质细胞炎症，从而导致星形胶质细胞功能障碍和神经元损伤。 我们的研究将首次将小胶质细胞 Hv1/NOX2/ROS/IFN- 信号传导与病理生理学联系起来 SCI 的研究，导致 SCI 的新治疗方法。鉴于 Hv1 在其他炎症中的拟议作用 模型中，Hv1 信号传导代表了与其他神经炎症状态相关的通用机制。