A critical role for intracellular VEGF receptor translocation in ocular angiogenesis

细胞内 VEGF 受体易位在眼部血管生成中的关键作用

• 批准号: 9920715
• 负责人: Michael Edwin Boulton
• 金额: $ 37.13万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-05-01 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9920715
• 关键词: Adherens Junction; Affect; Angiogenesis Inhibitors; Area; Binding; Cell Nucleus; Cell membrane; Choroidal Neovascularization; Complex; Developed Countries; Diabetes Mellitus; Diabetic Retinopathy; Diffusion; Endothelial Cells; Endothelial Growth Factors Receptor; Endothelium; Event; Eye diseases; Family; Gene Expression; Genetic; Genetic Transcription; Investigation; KDR gene; Membrane; Mus; Neuropilins; Nuclear; Outcome; Pathologic; Pathway interactions; Permeability; Receptor Signaling; Retinal Neovascularization; Role; Route; Signal Pathway; Signal Transduction; Site; Sorting - Cell Movement; Testing; Therapeutic Intervention; Tight Junctions; Vascular Endothelial Growth Factor Receptor-1; Vascular Endothelial Growth Factors; Vascular Permeabilities; Visual impairment; Work; angiogenesis; base; beta catenin; cadherin 5; choroidal angiogenesis; gene therapy; genetic manipulation; mouse model; neovascular; neovascularization; novel; ocular angiogenesis; presenilin; receptor binding; receptor internalization; response; retinal angiogenesis; secretase; subcellular targeting; targeted treatment; therapeutic target; trafficking; transcription factor; treatment strategy

The collective evidence dictates that the vascular endothelial growth factor (VEGF) family is critical for ocular angiogenesis in conditions such as diabetic retinopathy and AMD. Investigation of VEGF action has largely focused on receptor binding events and activation of classical downstream signal transduction cascades. Work from us and others show that VEGF receptor (VEGFR) signaling is much more complex and an alternative pathway involves intracellular trafficking of VEGFRs (Fig 1). We have preliminary evidence that a) endosomal sorting, neuropilin, secretases and sumoylation appear to be key regulators of VEGFR trafficking, b) VEGFRs in the nucleus originate from an intracellular pool, c) translocation of VEGFRs and the relative levels of VEGFRs within subcellular compartments dictates angiogenic outcome and d) the VEGFR1:VEGFR2 ratio at adherens junctions (AJs) and tight junctions (TJs) changes in response to pro- and antiangiogenic factors. Based on these observations we propose the following paradigm shifting hypothesis: VEGF-driven vascular permeability and neovascularization are highly dependent on the targeted subcellular translocation of specific VEGFRs in endothelial cells and that the relative levels of VEGFRs within subcellular compartments dictates vascular permeability and angiogenic outcome. We further postulate that endothelial-specific manipulation of the VEGFR1:2 ratio through gene therapy will reduce vascular permeability and inhibit aberrant retinal and choroidal neovascularization. We will test this hypothesis through the following aims. In Aim 1, we will a) To characterizing the origin and routes of VEGFR trafficking, b) identify the nuclear targets of VEGFR1 and VEGFR2 and assess how these targets contribute to angiogenesis, c) assess how the nuclear level of VEGFRs dictates the angiogenic outcome and d) identify the mechanism by which secretases, and/or sumoylation regulate trafficking of VEGFRs. In Aim 2, we will a) determine if translocation of VEGFRs to AJs and TJs is via membrane diffusion or endosomal trafficking and b) assess how the ratio of VEGFR1 and VEGFR2 at AJs and TJs changes in response to pro- and antiangiogenic factors, the junctional binding partners involved, and how this affects permeability. In Aim 3, we will assess the outcome of endothelial-specific genetic manipulation of intracellular VEGFR levels on vascular permeability and retinal and choroidal angiogenesis in mice. The proposed studies will significantly advance our mechanistic understanding of aberrant ocular angiogenesis and will identify new potential therapeutic targets for the inhibition of pathological ocular angiogenesis.

集体证据表明，血管内皮生长因子（VEGF）家族是 糖尿病性视网膜病和AMD等疾病中的眼血管生成至关重要。 对VEGF作用的研究主要集中在受体结合事件上和激活 经典的下游信号转导级联。来自我们和其他人的工作表明VEGF 受体（VEGFR）信号传导要复杂得多，替代途径涉及 VEGFR的细胞内运输（图1）。我们有初步证据表明a）内体 分类，神经蛋白，分泌酶和sumoylation似乎是VEGFR的关键调节剂 贩运，b）核中的vegfrs起源于细胞内池，c） VEGFR和亚细胞室内VEGFR的相对水平决定了血管生成 结果和d）粘附连接处（AJS）和紧密连接处的VEGFR1：VEGFR2比率 （TJ）响应于促和抗血管生成因素的变化。根据这些观察，我们 提出以下范式转移假设：VEGF驱动的血管渗透性和 新血管形成高度依赖于目标的亚细胞易位 内皮细胞中的特定VEGFR和VEGFR的相对水平 亚细胞室决定血管渗透性和血管生成结果。我们 进一步假设vegfr1的内皮特异性操纵通过 基因疗法将降低血管渗透性，并抑制异常的视网膜和脉络膜 新血管形成。我们将通过以下目标检验这一假设。在AIM 1中，我们 a）将表征VEGFR贩运的起源和路线，b）确定核 VEGFR1和VEGFR2的靶标，并评估这些靶标如何促进血管生成，c） 评估VEGFR的核水平如何决定血管生成结果，并确定 分泌和/或sumoylation调节VEGFR的贩运的机制。在AIM 2中， 我们将a）确定VEGFRS转移到AJ和TJ是否是通过膜扩散或 内体贩运和b）评估AJS和TJS VEGFR1和VEGFR2的比率如何 响应于促和抗血管生成因素的变化，涉及的连接结合伙伴， 以及这如何影响渗透率。在AIM 3中，我们将评估内皮特异性的结果 在血管通透性和视网膜上对细胞内VEGFR水平的遗传操纵 小鼠的脉络膜血管生成。拟议的研究将大大提高我们的机理 了解异常的眼血管生成，并将确定新的潜在治疗靶标 为了抑制病理眼血管生成。