A protective role for endothelial cell tissue nonspecific alkaline phosphatase in ischemic stroke

内皮细胞组织非特异性碱性磷酸酶在缺血性中风中的保护作用

• 批准号: 10025934
• 负责人: Candice Brown
• 金额: $ 26.6万
• 依托单位: WEST VIRGINIA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-08 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10025934
• 关键词: Acute; Address; Alkaline Phosphatase; Behavioral; Blood - brain barrier anatomy; Blood Circulation; Blood Preservation; Brain; Brain Diseases; Centers of Research Excellence; Cerebral Ischemia; Cerebrovascular Disorders; Cerebrum; Coupled; Cyclic AMP; Cytoskeletal Proteins; Data; Development; Endothelial Cells; Enzymes; Experimental Designs; Experimental Models; Functional disorder; Funding; Genetic; Glucose; Homeostasis; Impairment; In Vitro; Inflammatory; Injury; Ischemia; Ischemic Stroke; Knowledge; Laboratories; Link; Mediating; Metabolism; Middle Cerebral Artery Occlusion; Minerals; Modeling; Molecular; Mus; Outcome; Oxidative Stress; Oxygen; Pathologic; Pathway interactions; Permeability; Pharmacology; Phase; Phenotype; Phosphotransferases; Physiological; Recovery of Function; Reperfusion Injury; Reperfusion Therapy; Research; Research Personnel; Rho-associated kinase; Role; Signal Pathway; Signal Transduction; Stroke; Structure; Testing; Therapeutic Intervention; Tissues; Universities; West Virginia; artery occlusion; blood-brain barrier function; blood-brain barrier permeabilization; bone; brain endothelial cell; brain health; cerebral ischemic injury; cerebral microvasculature; deprivation; design; effective therapy; enzyme activity; experimental study; functional loss; improved; in vitro Model; in vivo; inhibitor/antagonist; injured; insight; mouse model; nervous system disorder; new therapeutic target; novel; post stroke; prevent; rho; stroke survivor; targeted treatment; tool; translational impact

PROJECT SUMMARY The objective of this application is to determine how tissue nonspecific alkaline phosphatase (TNAP) enzymatic activity preserves blood-brain barrier (BBB) function in ischemic stroke. Increased BBB permeability and oxidative stress are two potential mechanisms through which cerebral ischemia and reperfusion injury elicit BBB dysfunction and subsequent functional deficits in acute ischemic stroke. TNAP is a highly enriched enzyme in cerebral microvessels whose function in brain microvascular endothelial cells (BMECs) is poorly understood. Our preliminary data generated in a cellular BBB model of ischemia-reperfusion injury demonstrate that TNAP activity stimulates a novel mechanism which enhances cAMP-mediated signaling pathways that suppress Rho kinase (ROCK) activity and preserve BBB function. These intriguing findings led us to propose the novel concept that, in the face of cerebral ischemic injury, TNAP-ROCK signaling mechanisms integrate systemic signals at the BBB to sustain cerebral function and protect against post-stroke behavioral deficits. To address this novel concept, the application will investigate the central hypothesis that protection from ischemic stroke-induced impairment of BBB function is mediated by brain endothelial cell TNAP and its associated signaling cascades. The proposed studies will test this hypothesis in a mouse model of experimental ischemic stroke, transient middle cerebral artery occlusion (tMCAO), and in an ex vivo model of ischemia-reperfusion injury, oxygen-glucose deprivation (OGD). The experimental design will employ a mouse model with conditional deletion of TNAP in endothelial cells (VE-cKO) and its wild type littermates to interrogate TNAP’s effects on BBB permeability, short- term functional recovery, and BMEC signaling pathways post-stroke. Aim 1 will elucidate how TNAP preserves BBB permeability and promotes barrier function during cerebral ischemia and reperfusion injury. Aim 2 will interrogate the role of TNAP-ROCK signaling mechanisms in an OGD model of injury. Aim 3 will integrate the experiments in Aims 1 and 2 to prepare a competitive R01 application that will lead to independence of COBRE funding. Taken together, the studies in this proposal will uncover novel insights into the mechanisms that link TNAP enzyme activity to BBB integrity, barrier function, short-term functional recovery in ischemic stroke. The translational impact of these studies may uncover TNAP and its associated signaling cascades as novel therapeutic targets in ischemic stroke and other cerebrovascular disorders.

项目总结 本应用目的是确定组织的非特异性碱性磷酸酶(TNAP)如何被酶催化 活动保护缺血性卒中血脑屏障(BBB)功能。血脑屏障通透性增加和 氧化应激是脑缺血再灌注损伤诱发血脑屏障的两种潜在机制 急性缺血性卒中的功能障碍和随后的功能缺陷。TnAP是一种高度浓缩的酶 脑微血管在脑微血管内皮细胞(BMECs)中的作用尚不清楚。 我们在缺血-再灌注损伤的细胞血脑屏障模型中产生的初步数据表明，TNAP 活性刺激一种新的机制，增强cAMP介导的抑制Rho的信号通路 激活剂(ROCK)活性和维持血脑屏障功能。这些耐人寻味的发现促使我们提出了新的概念 在面对脑缺血损伤时，TNAP-ROCK信号机制在 血脑屏障可以维持大脑功能，防止中风后的行为缺陷。来写这部小说 概念，该应用程序将调查保护免受缺血性中风诱导的中心假设 血脑屏障功能的损害是由脑内皮细胞TNAP及其相关的信号级联反应介导的。 拟议的研究将在实验性缺血性中风的小鼠模型中检验这一假设，短暂性中枢性中风。 脑动脉闭塞(TMCAO)，以及在体外缺血-再灌注损伤模型中，氧-葡萄糖 剥夺(OGD)。实验设计将采用一种条件缺失TNAP的小鼠模型 血管内皮细胞(VE-CKO)及其野生型乳鼠为研究TNAP对血脑屏障通透性的影响。 卒中后足月功能恢复和BMEC信号通路。目标1将阐明TNAP如何保存 脑缺血再灌注损伤时血脑屏障通透性和促进屏障功能。目标2将 询问TNAP-ROCK信号机制在OGD损伤模型中的作用。目标3将整合 AIMS 1和AIMS 2中的实验，以准备具有竞争力的R01应用程序，从而实现Cobre的独立性 资金问题。综上所述，这项提案中的研究将揭示出对联系机制的新见解 TNAP酶活性对缺血性卒中患者血脑屏障完整性、屏障功能、短期功能恢复的影响。这个 这些研究的翻译影响可能会发现TNAP及其相关的信号级联是新的 缺血性中风和其他脑血管疾病的治疗靶点。