Investigation of endomucin as a novel regulator of angiogenesis

内粘蛋白作为血管生成的新型调节剂的研究

• 批准号: 9238401
• 负责人: Patricia Ann D'Amore
• 金额: $ 49.25万
• 依托单位: SCHEPENS EYE RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-02-01 至 2020-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9238401
• 关键词: Address; Adult; Age related macular degeneration; Binding; Biological Assay; Birth; Blindness; Blood Vessels; Blood capillaries; Cell Communication; Cell physiology; Cell surface; Cells; Choroidal Neovascularization; Cytoplasmic Tail; Development; Diabetic Retinopathy; Disease; Endothelial Cells; Equilibrium; Event; Extracellular Domain; Eye; Glycoproteins; Growth; Growth Factor; Histology; Human; ICAM2 gene; Impairment; In Vitro; Investigation; KDR gene; Lasers; Lead; Legal Blindness; Life; Light; Macular degeneration; Measures; Mediating; Messenger RNA; Modeling; Molecular; Mucins; Mus; Mutation; Ocular Pathology; Oxygen; Pathologic; Pathologic Neovascularization; Pathology; Permeability; Persons; Phosphoric Monoester Hydrolases; Phosphorylation; Play; Process; Proteins; Regulation; Reporting; Reproduction; Retinal; Retinal Diseases; Retinal Neovascularization; Retinopathy of Prematurity; Role; Signal Transduction; Small Interfering RNA; Subfamily lentivirinae; Surface; System; Testing; Therapeutic; Tube; Tyrosine Phosphorylation Site; Vascular Endothelial Growth Factor Receptor-1; Vascular Permeabilities; Vascularization; Veins; Wound Healing; angiogenesis; antiangiogenesis therapy; capillary; crosslink; density; design; endothelial-specific sialomucin; in vivo; inhibitor/antagonist; intravitreal injection; knock-down; loss of function; migration; mutant; new growth; novel; novel therapeutics; ocular neovascularization; overexpression; postnatal; proliferative diabetic retinopathy; promoter; retina blood vessel structure; retinal angiogenesis; small hairpin RNA

ABSTRACT Angiogenesis, the growth of new blood vessels, is an important component of development, wound healing, and reproduction as well as a number of pathologies. Abnormal retinal angiogenesis leads to vision loss in a range of ocular pathologies including diabetic retinopathy, wet macular degeneration, and the retinopathy of prematurity. Endomucin (EMCN) is a mucin-like glycoprotein specifically expressed by endothelial cells of veins and capillaries. Our preliminary studies reveal a role for EMCN in the regulation of VEGF-induced endothelial functions in vitro and in normal angiogenesis in vivo. Reduction of EMCN expression using siRNA led to decreased VEGF-stimulated endothelial proliferation, migration and tube formation in vitro, and impaired retinal vascularization in vivo. This application proposes to test the hypothesis that EMCN plays a role in VEGF-induced pathologic neovascularization and that EMCN influences VEGF-stimulated functions by modulating its signaling. Studies are outlined to examine the role of EMCN in pathological angiogenesis using models of abnormal retinal and choroidal blood vessel growth as well as to understand the molecular mechanism that underlies EMCN's effects on VEGF-induced endothelial cell functions using in vitro systems. Studies in Aim 1 will examine the role of EMCN in the development of pathologic ocular neovascularization in vivo. The oxygen-induced model of retinal neovascularization and the laser-induced model of choroidal neovascularization will be utilized in conjunction with both gain and loss of function studies, using both siRNA and lentivirally-delivered shEMCN under the control of ICAM2, an endothelial-specific promoter, as well as adEMCN under the control of ICAM2. The effect of EMCN knockdown and overexpression on pathologic vessel growth will be quantified. Aim 2 proposes to elucidate the molecular mechanism of EMCN's role in the regulation of VEGF-induced endothelial functions. Studies using EMCN deletion mutants will be employed to determine if the effects of EMCN are mediated at the cell surface. The role of EMCN at the cell surface will be further investigated by examining its effect on VEGF binding and VEGFR2 stabilization/internalization. Intracellular effects will focus on a possible role of EMCN phosphorylation, the effect of EMCN on VEGFR2 phosphatase activity as well as the identification of potential EMCN intracellular binding partners. Results of these proposed studies will provide a better understanding of the role of EMCN in normal and pathologic angiogenesis and may lead to the development of novel endothelial cell-specific therapies for the suppression of vessel growth. !

抽象的 血管生成，即新血管的生长，是发育、伤口愈合、 和生殖以及许多病理学。异常的视网膜血管生成导致视力丧失 一系列眼部病变，包括糖尿病性视网膜病变、湿性黄斑变性和视网膜病变 早熟。内粘蛋白（EMCN）是一种粘蛋白样糖蛋白，由内皮细胞特异性表达 静脉和毛细血管。我们的初步研究揭示了 EMCN 在 VEGF 诱导的调节中的作用 体外内皮功能和体内正常血管生成。使用 siRNA 减少 EMCN 表达 导致体外 VEGF 刺激的内皮增殖、迁移和管形成减少，并受损 体内视网膜血管化。该应用程序旨在检验 EMCN 在以下方面发挥作用的假设： VEGF 诱导的病理性新生血管形成以及 EMCN 通过以下方式影响 VEGF 刺激的功能 调制其信号。概述了研究 EMC​​N 在病理性血管生成中的作用的研究 异常视网膜和脉络膜血管生长的模型以及了解分子 EMCN 使用体外系统对 VEGF 诱导的内皮细胞功能产生影响的机制。 目标 1 的研究将检验 EMCN 在病理性眼病发展中的作用 体内新血管形成。氧诱导视网膜新生血管模型和激光诱导视网膜新生血管模型 脉络膜新生血管模型将与功能获得和丧失研究结合使用， 在 ICAM2 的控制下使用 siRNA 和慢病毒递送的 shEMCN，ICAM2 是一种内皮特异性细胞因子。 启动子，以及 ICAM2 控制下的 adEMCN。 EMCN 敲低的效果 病理血管生长的过度表达将被量化。目标 2 提出阐明分子 EMCN 在调节 VEGF 诱导的内皮功能中的作用机制。研究使用 EMCN 缺失突变体将用于确定 EMCN 的作用是否是在细胞表面介导的。 将通过检查其对 VEGF 结合和的影响来进一步研究 EMC​​N 在细胞表面的作用。 VEGFR2 稳定/内化。细胞内效应将集中于 EMCN 的可能作用 磷酸化、EMCN 对 VEGFR2 磷酸酶活性的影响以及潜在的鉴定 EMCN 细胞内结合伙伴。这些拟议研究的结果将有助于更好地理解 EMCN 在正常和病理性血管生成中的作用，并可能导致新型内皮细胞的发育 用于抑制血管生长的细胞特异性疗法。 ！