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.

项目摘要/摘要 精确和动态地调节血管屏障功能，即内皮细胞排列血管的能力 在血流和周围组织之间提供选择性可渗透的屏障是普遍存在的 对维持组织动态平衡很重要。屏障失调的病理后果是显而易见的。 在各种心血管疾病中，如动脉粥样硬化和慢性缺血以及炎症性和 神经退行性疾病。在动态平衡条件下，血流动力学切应力、摩擦阻力 血流作用于内皮细胞，通过以下途径促进血管内稳态和屏障功能 细胞-细胞黏附连接(AJ)和固有的肌动蛋白细胞骨架动力学的重塑和增强。这个 关键的细胞间黏附分子血管内皮细胞(VE-)钙粘附素是AJS的主要成分，调节 连接的稳定性通过其周转和内化，并经历了显著的张力变化 剪切力。此外，肌动蛋白细胞骨架通过维持平衡来调节血管屏障。 维持VE-钙粘附素的动态推力和稳定细胞内AJ复合体的张力力。 然而，将血流动力学切应力与机械应力联系起来的特定分子传感器和传感器 AJs和血管屏障功能的调节仍然知之甚少。激活无处不在的重要 最近发现NOTCH1受体通过以下途径调节血管屏障功能以应对切应力 与VE-钙粘附素络合，稳定AJs。虽然之前的工作已经确定了这种Notch1皮质 目前尚不清楚剪切力是如何激活Notch1受体的。 基于将Notch1与固有的细胞黏附和细胞骨架机制联系起来的初步数据，这项提议 验证Notch1与VE-钙粘蛋白的内在偶联和皮质肌动蛋白细胞骨架调节的假说 剪应力介导的Notch1激活。将询问Notch1对剪应力的响应 通过完成两个具体目标来实现：(1)确定VE-钙粘附素如何在时空上调节 NOTCH1及其配体DLL4，以协调剪应力激活和(2)识别机械 剪应力下Notch1与固有肌动蛋白细胞骨架动力学的相互作用。在整个过程中 在拟议的研究过程中，我将接受人体微血管三维仿生模型的培训， 超分辨率活细胞显微镜和机械分子方法，同时增强 通过科学交流、指导和教学方面的培训实现职业发展。我已经组装了一个 帮助我实现研究和职业目标的卓越、互补的指导团队：马修博士 库蒂斯是器官组织建模和细胞力学方面的专家，他将是我和戴安博士的主要赞助人 细胞细胞骨架动力学领域的领导者巴伯将成为我的共同赞助人。最终，这些发现将确定 Notch1被激活以响应切应力的新机制，并可能识别新的 Notch1涉及的调节屏障功能和其他血管病变的治疗靶点。