Regulation and Function of the Matricellular Protein CCN1 in Ischemic Retinopathy

基质细胞蛋白CCN1在缺血性视网膜病变中的调控和功能

• 批准号: 8220689
• 负责人: BRAHIM CHAQOUR
• 金额: $ 40.39万
• 依托单位: SUNY DOWNSTATE MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-12-01 至 2015-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8220689
• 关键词: Affect; Apoptosis; Astrocytes; Automobile Driving; Behavior; Binding; Biological; Biological Process; Blindness; Blood Vessels; Cell Adhesion; Cell Differentiation process; Cell Surface Receptors; Cell Therapy; Cell physiology; Cells; Childhood; Cues; Data; Development; Disease; Elements; Endothelial Cells; Environment; Event; Extracellular Matrix Proteins; Fostering; Future; Gelatinase A; Gene Expression; Gene Targeting; Genes; Goals; Growth; Growth Factor; Growth Factor Receptors; Healthcare Systems; Human; Hyperoxia; Immediate-Early Genes; Impairment; In Vitro; Integrins; Ions; Ischemia; Knowledge; Laboratories; Ligands; Modeling; Molecular; Morphogenesis; Mus; Mutant Strains Mice; Oxygen; Parents; Pathologic Neovascularization; Pathologic Processes; Pathology; Pattern Formation; Peptide Hydrolases; Pericytes; Pharmacotherapy; Phase; Phenotype; Physiological; Physiological Processes; Prevention; Proliferating; Protein Isoforms; Proteins; Regulation; Retina; Retinal; Retinal Diseases; Retinopathy of Prematurity; Rodent Model; Role; Screening procedure; Signal Transduction; Specificity; Stress; TNF gene; Testing; Therapeutic; Vascular Diseases; Vascular Endothelial Growth Factors; Vascularization; Vision; Visual impairment; Wnt proteins; age group; angiogenesis; base; cell behavior; cell type; cyr61 protein; cytokine; density; design; effective therapy; extracellular; gain of function; gene repression; improved; in vivo; insight; loss of function; migration; neovascularization; notch protein; novel; prevent; promoter; protein expression; protein function; receptor; response; retina blood vessel structure; small molecule

DESCRIPTION (provided by applicant): Disorders of retinal vessel growth and function are responsible for vision loss in retinopathy of prematurity (ROP), a leading cause of vision impairment and blindness in childhood. ROP develops as a result of excessive growth of abnormal pre-retinal blood vessels, a compensatory mechanism that overcomes an earlier phase of hyperoxia-induced vaso-obliteration. Elucidation of the molecular bases of angiogenic cell function and behavior in physiological and pathological processes will have important therapeutic implications for the treatment of human retinal vascular diseases. The long term goal of our laboratory is to uncover the in vivo functions of the matricellular protein CCN1, also known as cysteine-rich protein 61, and the functional consequences of its expression, or lack thereof, in development and ischemic retinopathy. The CCN1 protein is an inducible immediate-early gene-encoded extracellular matrix (ECM) protein required for proper vascular development. In our preliminary studies, we have found that over-expression of CCN1 in the retina via either gene- or cell-based therapy, enhanced normal retinal vascularization and reduced pathological angiogenesis in the model of oxygen-induced retinopathy (OIR). In vitro data showed that CCN1 functions primarily through direct binding to specific integrins and ECM proteins, and/or indirectly through modulation of growth factor and Wnt protein expression and/or activity, thereby triggering signaling events that culminate in the regulation of cell adhesion, migration, proliferation, gene expression, differentiation, and survival. Our hypothesis is that CCN1 normalizes the biological mechanisms of retinal vessel formation during development and following OIR and overrides those leading to abnormal vessel formation. In Specific Aim 1, we will use mutant mice with inducible conditional inactivation of the CCN1 gene to determine how loss of CCN1 in endothelial cells (ECs) causes defective retinal vessel growth. We will identify interactions with Wnt- and Notch-derived signals known to influence functional specialization of ECs (tip and stalk cell phenotypes) and sprouting angiogenesis. We will further determine whether and how forced expression of CCN1 in ECs only, allows normal retinal vessel formation in OIR. In Specific Aim 2, we will define the molecular interactions of CCN1 with astrocytes, the primary proangiogenic cells responsible for retinal vessel formation and patterning during development, and we will determine the functional significance of CCN1 loss in astrocytes (and/or in ECs) in mutant mice on astrocyte activation state and behavior (e.g., migration, density and ensheathment) during development and in OIR. In Specific Aim 3, we will define the dynamics of the CCN1 promoter activity and identify the functional elements responsible for CCN1 gene modulation both in cultured retinal ECs subjected to hyperoxic stress and in retinas of OIR mice. This proposal will provide new insights into the molecular mechanisms of CCN1 activities in vivo and may foster future safer, less destructive, and more effective therapies to harness ischemia-induced neovascularization in ROP. PUBLIC HEALTH RELEVANCE: Pathological angiogenesis is the hallmark of ischemic retinopathy, a leading cause of visual impairment in all age groups and a major financial burden for health care systems. Our project investigates the in vivo regulation and function of a specific component of the extracellular environment referred to as CCN1/Cyr61 in retinal vessel development and determines how this protein normalizes the mechanism of retinal vessel formation and prevents abnormal vessel growth in response to oxygen-induced retinopathy. Knowledge gained from these studies is also a promising opportunity to develop therapeutic applications using this protein or small molecules that control its expression to enhance the formation of normally functioning retinal blood vessels and improve the pharmacotherapy of retinal ischemic diseases.

描述（由申请人提供）：视网膜血管生长和功能障碍是导致早产儿视网膜病变（ROP）视力丧失的原因，这是儿童期视力损伤和失明的主要原因。 ROP 是异常视网膜前血管过度生长的结果，这是一种克服高氧引起的血管闭塞早期阶段的补偿机制。阐明生理和病理过程中血管生成细胞功能和行为的分子基础将对人类视网膜血管疾病的治疗具有重要的治疗意义。我们实验室的长期目标是揭示基质细胞蛋白 CCN1（也称为富含半胱氨酸的蛋白 61）的体内功能，以及其表达或缺乏在发育和缺血性视网膜病变中的功能后果。 CCN1 蛋白是一种诱导型立即早期基因编码的细胞外基质 (ECM) 蛋白，是血管正常发育所需的。在我们的初步研究中，我们发现通过基因或细胞治疗，CCN1在视网膜中过度表达，可以增强氧诱导性视网膜病变（OIR）模型中的正常视网膜血管化并减少病理性血管生成。体外数据表明，CCN1 主要通过直接结合特定整合素和 ECM 蛋白，和/或通过调节生长因子和 Wnt 蛋白表达和/或活性间接发挥作用，从而触发信号事件，最终调节细胞粘附、迁移、增殖、基因表达、分化和存活。我们的假设是，CCN1 使发育期间和 OIR 之后视网膜血管形成的生物机制正常化，并覆盖导致异常血管形成的机制。在具体目标 1 中，我们将使用 CCN1 基因可诱导条件失活的突变小鼠来确定内皮细胞 (EC) 中 CCN1 的缺失如何导致视网膜血管生长缺陷。我们将确定与 Wnt 和 Notch 衍生信号的相互作用，这些信号已知会影响 EC 的功能特化（尖端和茎细胞表型）和萌芽血管生成。我们将进一步确定 CCN1 仅在 EC 中强制表达是否以及如何允许 OIR 中正常的视网膜血管形成。在具体目标 2 中，我们将定义 CCN1 与星形胶质细胞（负责发育过程中视网膜血管形成和模式形成的主要促血管生成细胞）的分子相互作用，并且我们将确定突变小鼠星形胶质细胞（和/或 EC）中 CCN1 缺失对发育过程和 OIR 中星形胶质细胞激活状态和行为（例如迁移、密度和鞘）的功能意义。在具体目标 3 中，我们将定义 CCN1 启动子活性的动态，并确定在经受高氧应激的培养视网膜 EC 和 OIR 小鼠视网膜中负责 CCN1 基因调节的功能元件。该提案将为 CCN1 体内活性的分子机制提供新的见解，并可能促进未来更安全、破坏性更小、更有效的治疗方法，以利用 ROP 中缺血诱导的新生血管形成。 公共卫生相关性：病理性血管生成是缺血性视网膜病变的标志，是所有年龄段视力损害的主要原因，也是医疗保健系统的主要经济负担。我们的项目研究了视网膜血管发育中细胞外环境的特定成分（称为 CCN1/Cyr61）的体内调节和功能，并确定该蛋白质如何使视网膜血管形成机制正常化并防止因氧诱导的视网膜病变而导致的异常血管生长。从这些研究中获得的知识也是一个很有前途的机会，可以利用这种蛋白质或控制其表达的小分子来开发治疗应用，以增强正常功能的视网膜血管的形成并改善视网膜缺血性疾病的药物治疗。