Shear Stress Modulation of Wnt Signaling

Wnt 信号传导的剪切应力调制

• 批准号: 6537972
• 负责人: ARLENE R WECHEZAK
• 金额: $ 18.05万
• 依托单位: THE HOPE HEART INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-04-01 至 2004-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6537972
• 关键词: atherosclerotic plaque; biological signal transduction; cardiovascular disorder; cytoskeletal proteins; disease /disorder etiology; fluorescence microscopy; gene expression; genetic regulation; immunocytochemistry; intercellular connection; mechanical stress; molecular pathology; phosphorylation; protein localization; protein tyrosine kinase; protooncogene; reporter genes; shear stress; tissue /cell culture; vascular endothelium; western blottings

DESCRIPTION (Applicant's abstract): Cardiovascular disease is responsible for over 50 percent of the deaths in the United States. Plaque formation, a hallmark of atherosclerosis, predictably occurs at arterial sites characterized by shear stress that is low, high or alternating. The overall objective of this research is to determine the molecular mechanisms by which shear stress participates in endothelial injury and, thus, contributes to the pathogenesis of atherosclerosis. As mediators of cell-cell cohesion and communication, endothelial adherens junctions represent a potential vulnerability to shear stress perturbations. This investigation centers on the junctional role(s) of intra-and intercellular signaling, focusing on the Wnt pathway. The theory that adherens junctions and Wnt signaling are linked stems from the functional roles of beta-catenin as a pivotal protein in cell cohesion and as a key regulatory molecule in the Wnt pathway. Preliminary studies have demonstrated that in subconfluent endothelial cells, about-catenin is translocated to the nucleus, an event associated with Wnt pathway activation. In addition, pulsatile but not steady shear stress results in a marked decrease in Wnt-1 cell labeling. The specific aims are: (1) to determine the phosphorylation state and a subcellular compartmentalization of beta-catenin in endothelial cells for compatibility with an operative Wnt signaling pathway, (2) to test the hypothesis that shear stress modulates Wnt signaling in endothelial cells utilizing a TCF/LEF reporter gene construct and (3) to identify the mechanisms by which shear stress may modulate Wnt signaling in endogenous endothelium. Methods to accomplish these aims will include the use of an in vitro shear stress apparatus, Western blotting, RT-PCR, reporter gene constructs, cell transfection, high resolution 3-D microscopy and immunocytochemistry. These proposed studies would establish a linkage between shear stress as a controlling influence on adherens junctions in endothelium and that a consequence of this regulation is modulation of Wnt siqnaling.

描述（申请人摘要）：心血管疾病导致 超过50%的死亡病例。菌斑形成，a 动脉粥样硬化的标志，可预见地发生在动脉部位， 通过低、高或交替的剪切应力。本报告的总体目标 研究的目的是确定剪切应力 参与内皮损伤，从而导致发病 动脉粥样硬化作为细胞间凝聚和通讯的介质， 内皮粘附连接代表了对剪切的潜在脆弱性 应力扰动本研究的中心是 细胞内和细胞间信号传导，重点是Wnt途径。的理论 粘附连接和Wnt信号传导是联系在一起的功能性作用 β-连环蛋白是细胞凝聚的关键蛋白， Wnt通路中的分子。初步研究表明， 亚融合的内皮细胞，β-连环蛋白易位到细胞核， 与Wnt通路激活相关的事件。此外，搏动但不 稳定的剪切应力导致Wnt-1细胞标记的显著减少。的 具体目的是：（1）确定磷酸化状态和亚细胞 内皮细胞中β-连环蛋白的区室化以与 一个有效的Wnt信号通路，（2）测试剪切的假设， 应激利用TCF/LEF调节内皮细胞中的Wnt信号传导 报告基因构建体和（3）确定剪切的机制 应激可调节内源性内皮细胞Wnt信号转导。方法 实现这些目的将包括使用体外剪切应力 仪器，蛋白质印迹，RT-PCR，报告基因构建，细胞 转染、高分辨率3-D显微镜和免疫细胞化学。这些 建议的研究将建立剪切应力之间的联系， 控制对内皮细胞粘附连接的影响， 这种调节的结果是Wnt信号的调节。