Shear stress-mediated Notch1 activation by intrinsic cell adhesive and cytoskeletal activity

通过内在细胞粘附和细胞骨架活性剪切应力介导的 Notch1 激活

• 批准号: 10389629
• 负责人: Tania Singh
• 金额: $ 3.85万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-15 至 2025-02-14
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10389629
• 关键词: 3-Dimensional; Ablation; Actins; Adherens Junction; Adhesives; Architecture; Atherosclerosis; Barbering; Binding; Biomimetics; Biosensor; Blocking Antibodies; Blood; Blood Circulation; Blood Plasma Volume; Blood Vessels; Blood flow; C-terminal; Cardiovascular Diseases; Cardiovascular system; Cell Adhesion Molecules; Cell Line; Cell membrane; Cell-Cell Adhesion; Cells; Chronic; Clathrin; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Communication; Complex; Coupling; Cytoskeleton; Data; Edema; Educational process of instructing; Endothelial Cells; Endothelium; Equilibrium; Family; Friction; Genetic Transcription; Goals; Homeostasis; Human; Human Development; Inflammatory; Intervention; Ischemia; Knowledge; Lead; Ligands; Link; Liquid substance; Measures; Mechanics; Mediating; Mentors; Microscopy; Modeling; Molecular; Morphogenesis; Myocardial Infarction; Natural regeneration; Neurodegenerative Disorders; Nutrient; Pathogenesis; Pathologic; Pathway interactions; Permeability; Phosphorylation; Play; Process; Proteins; Proteomics; Receptor Activation; Regulation; Research; Resistance; Resolution; Role; Signal Transduction; Stroke; Testing; Tissue Model; Tissues; Training; Transducers; Ursidae Family; Vascular Cell Adhesion Molecule-1; Vascular Diseases; Vascular Endothelium; Vascular Permeabilities; Work; beta catenin; cadherin 5; career; career development; endothelial dysfunction; experience; hemodynamics; human model; improved; inhibitor; insight; live cell microscopy; mechanical force; mutant; new therapeutic target; notch protein; novel; polymerization; prevent; receptor; repaired; response; sensor; shear stress; spatiotemporal

PROJECT SUMMARY/ABSTRACT Precise and dynamic regulation of vascular barrier function, the ability of endothelial cells that line blood vessels to provide a selectively permeable barrier between the bloodstream and surrounding tissue, is universally important for maintaining tissue homeostasis. The pathological consequences of dysregulated barrier are evident in various cardiovascular diseases like atherosclerosis and chronic ischemia as well as inflammatory and neurodegenerative diseases. In homeostatic conditions, hemodynamic shear stress, the frictional drag force exerted by the flow of blood on endothelial cells, promotes vascular homeostasis and barrier function through remodeling and enhancement of cell-cell adherens junctions (AJs) and intrinsic actin cytoskeletal dynamics. The critical cell-cell adhesion molecule vascular endothelial (VE-) cadherin, the principle component of AJs, regulates junctional stability through its turnover and internalization and experiences significant changes in tension under shear stress. Additionally, the actin cytoskeleton regulates vascular barrier by maintaining a balance between dynamic pushing forces to maintain VE-cadherin and tensile forces which stabilize intracellular AJ complexes. However, the specific molecular sensors and transducers that link hemodynamic shear stress to the mechanical regulation of AJs and vascular barrier function remain poorly understood. Activation of the ubiquitously important Notch1 receptor was recently been found to modulate vascular barrier function in response to shear stress by complexing with VE-cadherin and stabilizing AJs. While previous work has determined how this Notch1 cortical pathway modulates vascular barrier function, it remains unclear how shear stress activates the Notch1 receptor. Building on preliminary data linking Notch1 to intrinsic cellular adhesive and cytoskeletal machinery, this proposal tests the hypothesis that intrinsic coupling of Notch1 to VE-cadherin and the cortical actin cytoskeleton regulates shear stress-mediated Notch1 activation. Interrogation of Notch1 activation in response to shear stress will be approached by completing two specific aims: (1) determine how VE-cadherin spatiotemporally regulates Notch1 and its ligand Dll4 to coordinate activation by shear stress and (2) identify the mechanical interplay between Notch1 and intrinsic actin cytoskeletal dynamics under shear stress. Throughout the course of the proposed research, I will gain training in 3D biomimetic models of the human microvasculature, super-resolution live cell microscopy, and mechanistic molecular approaches, while simultaneously enhancing career development through training in scientific communication, mentoring, and teaching. I have assembled an exceptional, complementary mentoring team to help me achieve my research and career goals: Dr. Matthew Kutys, an expert in organotypic tissue modeling and cell mechanics will be my primary sponsor and Dr. Diane Barber, a leader in cellular cytoskeletal dynamics, will be my co-sponsor. Ultimately, these findings will identify new mechanisms by which Notch1 is activated in response to shear stress and potentially identify new therapeutic targets for modulating barrier function and other vasculopathies where Notch1 is implicated.

项目概要/摘要 精确动态调节血管屏障功能、血管内皮细胞的能力 在血流和周围组织之间提供选择性渗透屏障，是普遍的做法 对于维持组织稳态很重要。屏障失调的病理后果是显而易见的 各种心血管疾病，如动脉粥样硬化、慢性缺血以及炎症和 神经退行性疾病。在稳态条件下，血流动力学剪切应力、摩擦阻力 血流对内皮细胞产生作用，通过促进血管稳态和屏障功能 细胞-细胞粘附连接（AJ）和内在肌动蛋白细胞骨架动力学的重塑和增强。这 关键的细胞间粘附分子血管内皮 (VE-) 钙粘蛋白是 AJ 的主要成分，可调节 通过其周转和内化实现连接稳定性，并在压力下经历显着变化 剪切应力。此外，肌动蛋白细胞骨架通过维持之间的平衡来调节血管屏障。 维持 VE-钙粘蛋白的动态推力和稳定细胞内 AJ 复合物的张力。 然而，将血流动力学剪切应力与机械应力联系起来的特定分子传感器和换能器 AJ 和血管屏障功能的调节仍知之甚少。激活无处不在的重要 最近发现 Notch1 受体可以通过以下方式调节血管屏障功能以响应剪切应力 与 VE-钙粘蛋白复合并稳定 AJ。虽然之前的工作已经确定了 Notch1 皮质如何 虽然剪切力如何调节血管屏障功能，但剪切应力如何激活 Notch1 受体仍不清楚。 该提案基于将 Notch1 与内在细胞粘附和细胞骨架机制联系起来的初步数据 检验 Notch1 与 VE-钙粘蛋白和皮质肌动蛋白细胞骨架的内在耦合调节的假设 剪切应力介导的 Notch1 激活。响应剪切应力的 Notch1 激活的询问将是 通过完成两个具体目标来实现：（1）确定 VE-钙粘蛋白如何时空调节 Notch1 及其配体 Dll4 通过剪切应力协调激活，并 (2) 识别机械 剪切应力下Notch1和内在肌动蛋白细胞骨架动力学之间的相互作用。整个 在拟议的研究过程中，我将获得人体微脉管系统 3D 仿生模型的培训， 超分辨率活细胞显微镜和机械分子方法，同时增强 通过科学传播、指导和教学培训进行职业发展。我已经组装了一个 卓越、互补的指导团队帮助我实现我的研究和职业目标：Matthew 博士 Kutys，器官组织建模和细胞力学方面的专家将是我的主要赞助商，Diane 博士 巴伯是细胞骨架动力学领域的领军人物，他将是我的共同发起人。最终，这些发现将确定 Notch1 响应剪切应力而被激活的新机制，并可能识别新的 调节屏障功能和其他涉及 Notch1 的血管疾病的治疗靶点。