Norrin Signaling in the Retina: Regulation of the Blood-Retina Barrier

视网膜中的诺里蛋白信号传导：血视网膜屏障的调节

• 批准号: 10020983
• 负责人: Harald Junge
• 金额: $ 45.14万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10020983
• 关键词: Address; Age related macular degeneration; Aging; Agonist; Attenuated; Basic Science; Biological Assay; Biological Models; Biology; Blood Vessels; Blood-Retinal Barrier; Cells; Chronic; Classical Complement Pathway; Complement; Complement Activation; Data; Defect; Diabetic Retinopathy; Disease; Disease model; Drug Delivery Systems; Edema; Electroretinography; Endothelial Cells; Engineering; Extravasation; Familial exudative vitreoretinopathy; Fluorescein Angiography; Functional disorder; Genetic; Goals; Health; Histopathology; Hypoxia; Immunoglobulin G; Immunoglobulins; Impairment; In Vitro; Ionizing radiation; Knowledge; Maintenance; Mediating; Metabolic stress; Modality; Modeling; Mus; Pathologic; Pathology; Pathway interactions; Perfusion; Pericytes; Phenotype; Proteins; Regulation; Retina; Retinal Diseases; Retinoblastoma; Retinopathy of Prematurity; Role; Signal Pathway; Signal Transduction; Stains; Stress; TP53 gene; Testing; Therapeutic; Therapeutic Intervention; Time; Uveitis; Vascular Endothelial Growth Factors; Vascular Permeabilities; angiogenesis; antiangiogenesis therapy; antibody engineering; base; beta catenin; chemotherapy; complement pathway; field study; genetic approach; human disease; in vivo; inhibiting antibody; inhibitor/antagonist; insight; mouse genetics; mouse model; neovascularization; novel; sensor; side effect; transcriptome sequencing

Our overall objective is to understand the role of norrin/frizzled4 signaling as regulator of the blood-retina barrier (BRB) in human disease, using mice as model system. Impairment of the BRB causes edema and is associated with diabetic retinopathy, age-related macular degeneration, retinopathy of prematurity, familial exudative vitreoretinopathy (FEVR), and uveitis. BRB induction and maintenance are controlled by the critical b-catenin-dependent (canonical) norrin/frizzled4 signaling pathway. Disrupted norrin/frizzled4 signaling causes FEVR. How the impairment of norrin/frizzled4 signaling causes pathological changes, how the pathway is regulated under stress, and if activation of norrin/frizzled4 signaling is a valid therapeutic approach to restore BRB dysfunction, are all not understood. These knowledge gaps need to be addressed to exploit the enormous basic research progress on norrin/frizzled4 signaling for therapeutic intervention. Aim1: We showed that endothelial cell-specific inactivation of norrin signaling in developed mice provides a new model to study pathological consequences of BRB breakdown. The new model does not display compound pathologies observed in existing models of BRB disruption characterized by pericyte defects, perfusion defects, and hypoxia, allowing us to study pathological consequences of BRB defects per se. Endothelial cell-specific disruption of norrin/frizzled4 signaling in developed mice causes immunoglobulin extravasation, complement activation, cystoid edema, and reduced b-wave in electroretinography (ERG). We will use mouse genetics to test the role of the classical complement pathway in mediating long-term pathological consequences of BRB breakdown in an otherwise intact vasculature. Fluorescein angiography and ERG will be performed at multiple time points, and histopathology at the 1-year end-point. Successful completion of this aim will determine roles of the classical complement pathway in retinal disease beyond the known roles in age-related macular degeneration. Aim2: Preliminary data show that increased levels of a stress sensor protein in endothelial cells strongly inhibit canonical signaling and negatively regulate BRB function. We will use conditional mouse genetics to activate the stress sensor in endothelial cells, determine BRB function, and test for genetic interactions with increased or reduced levels of b-catenin signaling. Cell-based assays will be performed to determine the mechanistic basis of this powerful regulation. Successful completion of this aim will provide novel insights into the regulation of the BRB under stress. These mechanisms may be exploited for modulating the BRB, e.g., in the context of restoring the barrier in disease, or for drug delivery. Aim3: Current efforts to target norrin/frizzled4 signaling for therapeutic intervention have focused on anti-angiogenesis approaches in proliferative retinopathies. We will determine if the activation of b-catenin in endothelial cells restores BRB function in a transient hypoxia disease model. Successful completion of this aim is expected to provide a model to test agonists of the pathway for their ability to restore a dysfunctional BRB (aim3).

我们的总体目标是了解 Norrin/frizzled4 信号作为血视网膜调节剂的作用 人类疾病中的屏障（BRB），使用小鼠作为模型系统。 BRB 受损会导致水肿 与糖尿病性视网膜病变、年龄相关性黄斑变性、早产儿视网膜病变、家族性视网膜病变相关 渗出性玻璃体视网膜病变（FEVR）和葡萄膜炎。 BRB 感应和维护由关键控制 b-连环蛋白依赖性（规范）norrin/frizzled4 信号通路。 Norrin/frizzled4 信号传导中断的原因 FEVR。 Norrin/frizzled4信号受损如何引起病理变化，通路如何 在压力下受到调节，并且如果激活 Norrin/frizzled4 信号是一种有效的恢复治疗方法 BRB功能失调，大家都不明白。需要解决这些知识差距，以利用巨大的潜力 Norrin/frizzled4 信号传导治疗干预的基础研究进展。目标1：我们证明了 发育小鼠内皮细胞特异性诺里蛋白信号失活提供了新的研究模型 BRB 崩溃的病理后果。新模型不显示复合病理学 在现有的 BRB 破坏模型中观察到，其特征是周细胞缺陷、灌注缺陷和 缺氧，使我们能够研究 BRB 缺陷本身的病理后果。内皮细胞特异性 发育小鼠中 Norrin/frizzled4 信号传导的破坏会导致免疫球蛋白外渗、补体 激活、囊样水肿和视网膜电图 (ERG) 中 b 波减少。我们将利用小鼠遗传学来 测试经典补体途径在介导 BRB 长期病理后果中的作用 原本完好的脉管系统崩溃。荧光素血管造影和 ERG 将在多个 时间点和 1 年终点的组织病理学。成功完成这一目标将决定角色 经典补体途径在视网膜疾病中的作用超出了年龄相关性黄斑的已知作用 退化。目标2：初步数据表明，内皮细胞中应激传感器蛋白的水平增加 强烈抑制经典信号传导并负向调节 BRB 功能。我们将使用条件鼠标 遗传学激活内皮细胞中的压力传感器、确定 BRB 功能并测试遗传 与β-连环蛋白信号传导水平升高或降低的相互作用。将进行基于细胞的测定 确定这种强有力的调节的机制基础。成功完成这一目标将提供 对压力下 BRB 调节的新颖见解。这些机制可用于调节 BRB，例如在恢复疾病屏障或药物输送的情况下。目标 3：当前的努力 用于治疗干预的目标诺里蛋白/卷曲4信号传导主要集中在抗血管生成方法上 增殖性视网膜病。我们将确定内皮细胞中 β-连环蛋白的激活是否可以恢复 BRB 在短暂缺氧疾病模型中发挥作用。成功完成这一目标预计将提供 模型来测试该途径的激动剂恢复功能失调的 BRB 的能力（目标 3）。