aVB3 Activation and Phosphorylation in Angiogenesis

血管生成中的 aVB3 激活和磷酸化

• 批准号: 7062112
• 负责人: Tatiana V Byzova
• 金额: $ 30.54万
• 依托单位: CLEVELAND CLINIC LERNER COM-CWRU
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7062112
• 关键词: angiogenesis; biological signal transduction; cell adhesion; cell aggregation; cell migration; cell proliferation; flow cytometry; growth factor receptors; integrins; laboratory mouse; matrigel; phosphorylation; platelets; polymerase chain reaction; protein protein interaction; protein structure function; vascular endothelial growth factors; vascular endothelium

The beta3 integrins mediate cellular functions that underlie a number of physiological and pathophysiological processes including angiogenesis which in turn are critical during atherosclerosis, myocardial injury, wound healing, tumor development and inflammatory diseases. Angiogenesis is triggered by interaction of vascular endothelial growth factors (VEGFs) with their receptors on endothelial cells which, in turn, leads to the cell activation and the changes of integrin functions. Despite an importance of the process of angiogenesis, its mechanisms are poorly understood. The overall objective of this proposal is to determine molecular mechanisms of communication between VEGFRs and alpha-v-beta3 integrin in angiogenesis and lymphangiogenesis. The major focus of this proposal is to assess the role of beta3 integrin cytoplasmic domain and its phosphorylation in the regulation of alpha-v-beta3 activity during the process of VEGF-induced angiogenesis, The elucidation of these mechanisms would create the basis for the development of novel therapeutic strategies in cardiology and vascular medicine. Proposed specific aims are: Aim I. To evaluate the role of beta3 integrin phosphorylation in VEGF-induced angiogenesis in vivo. The angiogenic responses will be assessed in mice expressing a mutated beta3 subunit that impairs its capacity to undergo tyrosine phosphorylation (DiYF) using matrigel and tumor-induced angiogenesis models. These studies will be complemented by unique adenovirus-induced subcutaneous and muscle models developed in our previous studies. We will compare responses initiated by different sets of VEGF receptors, VEGFR-2 and VEGFR-3, and we will utilize two VEGFs, VEGF-A165 and VEGF-D. Through these analyses, we will establish a physiological role of beta3 integrin phosphorylation and dissect how different communication pathways from VEGFRs to integrins influence angiogenesis. Aim II. To assess the molecular mechanism for the role of beta3 integrin phosphorylation in angiogenesis. In these studies we will utilize endothelial cells of different origin isolated from normal and DiYF mice. We will characterize the adhesive, migratory and proliferative responses of endothelial cells of different origin from normal and DiYF animals to VEGF as compared to other agonists. We will compare normal and DiYF endothelial cells for their ability to form capillary-like structures ex vitro. Aim III. To further determine the molecular requirements for beta3 integrin subunit to interact with VEGF receptors. In this set of studies, we will focus on the role of beta 3 cytoplasmic domain and its phosphorylation in the VEGF-induced functional responses. We will assess whether beta3 integrin phosphorylation plays a role in VEGF-induced integrin activation. The role of specific intermediates in the signal transduction from different VEGFRs will be considered (including role of talin and skelemin in VEGFRs-induced activation).

β3整联蛋白介导许多生理和病理生理过程的基础的细胞功能，包括血管生成，而血管生成又在动脉粥样硬化，心肌损伤，伤口愈合，肿瘤发育和炎症性​​疾病中至关重要。血管生成是由血管内皮生长因子（VEGF）与其受体在内皮细胞上的相互作用引起的，这反过来导致细胞激活和整联蛋白功能的变化。尽管具有血管生成过程的重要性，但其机制还是很少的理解。该提案的总体目的是确定VEGFRS和α-V-Beta3整合蛋白在血管生成和淋巴管生成中的通信的分子机制。该提案的主要重点是评估β3整联蛋白细胞质结构域的作用及其在VEGF诱导的血管生成过程中α-V-BetA3活性调节中的磷酸化中，这些机制的阐明将为发展新颖的治疗策略在心脏病学和血管学上的新治疗策略开发提供基础。提出的具体目的是：目标I。评估β3整联蛋白磷酸化在VEGF诱导的体内血管生成中的作用。将在表达突变的beta3亚基的小鼠中评估血管生成反应，该小鼠使用Matrigel和肿瘤诱导的血管生成模型损害其经历酪氨酸磷酸化（DIYF）的能力。这些研究将与独特的腺病毒诱导的皮下和肌肉模型相辅相成。 在我们以前的研究中发展。我们将比较不同的VEGFR-2和VEGFR-3的不同集合受体启动的响应，我们将利用两个VEGFS，即VEGF-A165和VEGF-D。通过这些分析，我们将建立beta3的生理作用 整联蛋白磷酸化并剖析从VEGFRS到整联蛋白的不同通信途径如何影响血管生成。目标II。评估β3整联蛋白磷酸化在血管生成中的作用的分子机制。在这些研究中，我们将利用从正常和DIYF小鼠分离的不同来源的内皮细胞。与其他激动剂相比，我们将表征从正常和DIYF动物到VEGF的不同来源的内皮细胞的粘合性，迁移和增殖反应。我们将比较正常和DIYF内皮细胞，以便将其形成毛细血管样结构的能力。目标三。进一步确定β3整合素亚基与VEGF受体相互作用的分子需求。在这组研究中，我们将重点关注β3细胞质结构域的作用及其在VEGF诱导的功能反应中的磷酸化。我们将评估β3整联蛋白磷酸化是否在VEGF诱导的整联蛋白激活中起作用。特定中间体在信号中的作用 将考虑来自不同VEGFR的转导（包括Talin和Skelemin在VEGFRS诱导的激活中的作用）。