Targeting Protease Activated Receptor 1 for Repair of the Injured Spinal Cord

靶向蛋白酶激活受体 1 修复受损脊髓

• 批准号: 10175393
• 负责人: ISOBEL A SCARISBRICK
• 金额: $ 47.75万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10175393
• 关键词: Acute; Address; Appearance; Astrocytes; Axon; Bioavailable; Biological Assay; Bone Marrow; Brain; Cell Surface Receptors; Cell physiology; Central Nervous System Diseases; Chimera organism; Chronic; Clinical; Coculture Techniques; Coupling; Disease; Enzymes; Event; Exhibits; Experimental Models; F2R gene; FDA approved; Fostering; G-Protein-Coupled Receptors; Growth Factor; Hormones; Immune; In Vitro; Individual; Inflammation; Inflammatory Response; Injury; Kininogenase; Knock-out; Knowledge; Laboratories; Link; Locomotor Recovery; LoxP-flanked allele; MAP Kinase Gene; Messenger RNA; Microglia; Modeling; Molecular; Mus; Myelin; Myelin Sheath; Myelinated nerve fiber; Neuroglia; Neurons; Oral; Outcome; PAR-1 Receptor; Pathogenesis; Pathology; Pathway interactions; Peptide Hydrolases; Peripheral; Pharmacology; Physiological; Play; Positioning Attribute; Property; Proteinase-Activated Receptors; Publishing; Recovery; Recovery of Function; Research Design; Ribosomes; Role; STAT3 gene; Sensorimotor functions; Serine Protease; Signal Transduction; Spinal cord injury; Techniques; Testing; Therapeutic; Thrombin; Thrombin Receptor; Time; Translations; Treatment Efficacy; United States; astrogliosis; base; brain health; cell injury; central nervous system injury; clinical translation; combinatorial; design; disability; experimental study; functional improvement; functional outcomes; genetic approach; improved; improved outcome; injury and repair; insight; knockout gene; loss of function; monocyte; nerve injury; neurobehavioral; neuroinflammation; neuroprotection; receptor; relating to nervous system; repair strategy; repaired; response; response to injury; restoration; small molecule inhibitor; targeted treatment; therapeutic target; tool

PROJECT SUMMARY/ABSTRACT Secreted serine proteases are abundant in the intact CNS and become deregulated by injury and disease, yet we lack knowledge regarding their physiological functions and contributions to pathology. Several years ago, the discovery of a set of enzyme-activated G protein-coupled receptors, the Protease Activated Receptors (PARs), led to a new conceptual framework for understanding the physiological impact of proteases. PARs permit activating enzymes to signal in a hormone-like fashion to modulate key cellular functions, but when overactivated can contribute to pathology. The PI’s team recently discovered that mice with global PAR1 gene knockout exhibit significant improvements in locomotor recovery after spinal cord injury (SCI). Functional improvements were accompanied by reductions in inflammation and astrogliosis and improvements in the appearance of myelin and axons, all integral substrates to support restoration of function. We also documented that CNS injury relevant proteases, such as thrombin and kallikrein 6 elicit Ca2+, MAPK and STAT3 signaling linked to neuroinflammation and pro-injury responses across neurons and neuroglia in a PAR1-dependent manner. Together, these studies highlight the likely multifactorial roles played by PAR1 in key cellular and molecular events positioned to govern outcomes after SCI. These findings also highlight the potential to target PAR1 for neural protection and repair. Despite these encouraging findings the cellular mechanisms by which blocking PAR1 improves recovery after SCI have not been defined and this knowledge gap hampers progress towards translation of existing FDA approved and orally bioavailable PAR1 small molecule inhibitors. Additionally, whether blocking PAR1 therapeutically at acute or chronic time points after SCI are both capable of improving neural recovery is unknown. Based on recently published findings, taken with new preliminary results, we propose 3 integrated Aims to test the Central Hypothesis that PAR1 is an essential regulator of reactivity across the microglial- astrocyte compartments and can be selectively blocked to improve glial-neuronal trophic coupling, neuroprotection and repair after SCI. In Aim 1, we will determine the impact of pharmacologic PAR1 inhibition initiated at acute or chronic time points after injury on signs of neuroprotection, neural repair and recovery of sensorimotor function and use ribosomal mRNA capture techniques to document cellular and molecular mechanisms engaged across the astroglial, microglial/monocyte and neuron compartments. In Aim 2, we will determine whether conditional deletion of PAR1 selectively in astrocytes, microglia or peripheral monocytes is sufficient to enhance recovery. In Aim 3, we will use glial-neuron co-cultures as bioassays to establish PAR1- regulated glial-neural trophic coupling mechanisms relevant to neuroprotection and repair. The studies proposed address key mechanistic questions regarding the functional roles of PAR1 in neural injury and will provide new information needed to optimize therapeutic targeting strategies for recovery of function.

项目总结/摘要 分泌的丝氨酸蛋白酶在完整的CNS中是丰富的，并且由于损伤和疾病而变得失调，然而， 我们缺乏关于它们的生理功能和对病理学的贡献的知识。几年前 发现了一组酶激活的G蛋白偶联受体，蛋白酶激活受体（PAR）， 为理解蛋白酶的生理影响提供了一个新的概念框架。PARs许可证 激活酶，以类似信号的方式调节关键的细胞功能，但当过度激活时， 会导致病理学改变PI的研究小组最近发现，PAR 1基因敲除的小鼠表现出 脊髓损伤（SCI）后运动恢复的显著改善。功能改进是 伴随炎症和星形胶质细胞增生的减少以及髓鞘外观的改善， 轴突，所有支持功能恢复的完整基质。我们还记录了CNS损伤相关 蛋白酶如凝血酶和激肽释放酶6引发与神经炎症相关的Ca 2+、MAPK和STAT 3信号传导 以及以PAR 1依赖性方式跨神经元和神经胶质的促损伤反应。这些研究一起 强调PAR 1在关键细胞和分子事件中可能发挥的多因素作用， SCI后的结果这些发现还强调了靶向PAR 1用于神经保护和修复的潜力。 尽管有这些令人鼓舞的发现，但阻断PAR 1改善术后恢复的细胞机制仍有待进一步研究。 SCI尚未定义，这种知识差距阻碍了现有FDA翻译的进展 批准的和口服生物可利用的PAR 1小分子抑制剂。此外，阻断PAR 1 在SCI后的急性或慢性时间点治疗都能够改善神经恢复， 未知根据最近发表的研究结果，采取新的初步结果，我们提出3个综合 目的是检验中心假说，即PAR 1是小胶质细胞反应性的重要调节因子， 星形胶质细胞隔室并且可以被选择性阻断以改善胶质细胞-神经元营养偶联， 脊髓损伤后的神经保护和修复。在目标1中，我们将确定药理学PAR 1抑制的影响 在损伤后的急性或慢性时间点开始，对神经保护、神经修复和恢复的迹象进行评估。 感觉运动功能，并使用核糖体mRNA捕获技术来记录细胞和分子 在星形胶质细胞、小胶质细胞/单核细胞和神经元隔室中参与的机制。在目标2中，我们将 确定选择性地在星形胶质细胞、小胶质细胞或外周单核细胞中条件性缺失PAR 1是否 足以促进复苏。在目标3中，我们将使用神经胶质-神经元共培养物作为生物测定，以建立PAR 1- 与神经保护和修复相关的调节神经胶质-神经营养偶联机制。建议的研究 解决了关于PAR 1在神经损伤中的功能作用的关键机制问题，并将提供新的 优化治疗靶向策略以恢复功能所需的信息。