Mechanisms Regulating Vascular Homeostasis

调节血管稳态的机制

• 批准号: 10299286
• 负责人: Christopher D Kontos
• 金额: $ 59.09万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10299286
• 关键词: ANGPT1 gene; Address; Adhesions; Angiopoietins; Apolipoprotein E; Atherosclerosis; Binding; Binding Sites; Biology; Blood Vessels; CRISPR/Cas technology; Calmodulin; Cardiovascular Diseases; Cell Survival; Cell physiology; Cessation of life; Co-Immunoprecipitations; Coronary heart disease; Defect; Endothelial Cells; Endothelium; Event; Gene Expression; Genetic; Genetic Transcription; Goals; Health; Homeostasis; Hypertension; In Vitro; Inflammation; Lead; Ligands; Link; Liquid substance; Malignant Neoplasms; Mediating; Modeling; Molecular; Mus; Myography; NOS3 gene; Nitric Oxide; Oxygen; Pathologic Neovascularization; Perfusion; Permeability; Phenotype; Phosphorylation; Play; Proteins; Public Health; Receptor Signaling; Recombinant Proteins; Regulation; Regulatory Pathway; Retinal Diseases; Role; Scaffolding Protein; Septic Shock; Signal Pathway; Signal Transduction; Testing; Transcription Factor AP-1; Vascular Diseases; Vascular Permeabilities; Vascular remodeling; angiogenesis; atheroprotective; base; cardiovascular disorder prevention; density; endothelial dysfunction; in vivo; induced pluripotent stem cell; loss of function; monocyte; mutant; novel; pressure; prevent; receptor; response; rho; shear stress; transcription factor; vascular inflammation; western diet

PROJECT SUMMARY Regulation of normal endothelial cell (EC) function plays a critical role in vascular homeostasis and the prevention of cardiovascular disease (CVD). Endothelial dysfunction in CVD is characterized by loss of endothelial nitric oxide synthase (eNOS)-derived nitric oxide (NO). The Tie receptors and their ligands, the angiopoietins (Ang), are important regulators of vascular homeostasis and the transition from EC quiescence to "activation" and vascular remodeling. Whereas Ang-1/Tie2 signaling promotes EC survival and quiescence, in part through activation of Akt and eNOS, the homologous Tie1 receptor has been shown to promote EC activation, inflammation, and atherosclerosis. However, Tie1 is also required for Tie2-mediated angiogenesis, and Tie1's overall functional effects appear to be context-dependent and linked, in part, to the presence of inflammation. Unlike Tie2, Tie1 has no known activating ligands, therefore its signaling pathways and mechanisms of action have been difficult to study and remain poorly understood. We identified a novel and previously uncharacterized molecular scaffolding protein, Caskin2, as a Tie1-interacting protein. Although few studies have specifically addressed Caskin2's function, it has been identified as a strong marker of ECs, and numerous molecular studies have linked Caskin2 to phenotypes associated with defects in vascular homeostasis. Importantly, our studies indicate that Caskin2 is a critical molecular link between Tie receptor signaling, eNOS biology, and vascular homeostasis. Our preliminary studies in this proposal demonstrate that Caskin2 is highly expressed in ECs in vitro and in vivo and that Caskin2 promotes EC quiescence and survival in vitro. Caskin2 binding to Tie1 is disrupted by phosphorylation of T794 on Tie1, an event that is mediated by PAK in a Rac-dependent manner and that is required for angiogenesis. Moreover, Caskin2 is required for Ang- 1/Tie2-mediated signaling through Akt, suggesting that Caskin2 integrates signals from both Tie1 and Tie2 to regulate vascular homeostasis. Furthermore, Caskin2 binds eNOS and its activator calmodulin (CaM) and increases eNOS expression and activity and is upregulated by fluid shear stress and by KLF2. Finally, we show that Caskin2–/– mice have increased vascular permeability, atherosclerosis, pathological angiogenesis, and are hypertensive, all disorders of vascular homeostasis characterized by abnormal eNOS signaling. Based on these findings, we hypothesize that Caskin2 is a critical integrator of Tie receptor signaling that regulates vascular homeostasis in part through eNOS activation. To test this hypothesis, the Specific Aims of this proposal are to: 1) Determine the molecular mechanisms by which Caskin2 regulates Tie receptor-mediated endothelial function and eNOS activity; 2) Determine the mechanisms by which Caskin2 responds to and senses changes in fluid shear stress to regulate EC phenotypes in vitro; and 3) Determine the effects of Caskin2 deficiency on vascular homeostasis in vivo.

项目摘要 正常内皮细胞（EC）功能的调节在血管稳态中起着至关重要的作用 预防心血管疾病（CVD）。 CVD中的内皮功能障碍的特征是丢失 内皮一氧化氮合酶（ENOS）衍生的一氧化氮（NO）。领带受体及其配体， 血管生成素（ANG）是血管稳态的重要调节剂，并且是静态的过渡 要“激活”和血管重塑。而ang-1/tie2信号传导促进了EC的生存和静止 部分通过AKT和ENOS的激活，已显示同源TIE1受体可促进EC 激活，炎症和动脉粥样硬化。但是，TIE1也需要TIE1介导的血管生成， TIE1的总体功能效应似乎与上下文有关，并部分链接到存在 炎。与TIE2不同，TIE1没有已知的激活配体，因此其信号通路和 作用机制很难研究，并且仍然了解不足。我们确定了一部小说， 以前未表征的分子脚手架蛋白Caskin2作为拉皮1相互作用蛋白。虽然很少 研究专门针对Caskin2的功能，已被确定为EC的强标记，并且 许多分子研究将Caskin2与与血管缺陷相关的表型联系起来 稳态。重要的是，我们的研究表明caskin2是领带接收器之间的关键分子联系 信号传导，eNOS生物学和血管稳态。我们在该提案中的初步研究表明 caskin2在体外和体内高度表达 体外。 caskin2结合与TIE1的结合被TIE1上T794的磷酸化破坏了，这一事件是由 PAK以RAC依赖性方式，这是血管生成所必需的。此外，ang-需要caskin2 1/TIE2介导的信号通过AKT，这表明Caskin2集成了TIE1和TIE2的信号 调节血管稳态。此外，caskin2绑定eNOS及其激活剂钙调蛋白（CAM）和 增加eNOS的表达和活性，并通过流体剪切应力和KLF2进行更新。最后，我们 表明caskin2 - / - 小鼠的血管渗透性增加，动脉粥样硬化，病理血管生成， 并且是高血压的，所有的血管稳态疾病都以异常的eNOS信号传导为特征。基于 在这些发现中，我们假设Caskin2是TIE受体信号传导的关键积分器， 通过ENOS激活部分调节血管稳态。为了检验这一假设， 该建议的目的是：1）确定Caskin2调节领带的分子机制 受体介导的内皮功能和eNOS活性； 2）确定caskin2的机制 对流体剪切应力的反应和感觉变化以在体外调节EC表型。 3）确定 Caskin2缺乏对体内血管稳态的影响。