Effects of shear stress on regulation of epithelial permeability

这个K08应用程序的目标是促进基本技能的发展，使PI成为 一名成功的院士，并取得独立科学调查员的地位。呼吸道上皮 作为一种屏障，调节呼吸道表面液体的含量，以及细胞因子和其他 调节呼吸道生理的产品。除了循环伸展和收缩，呼吸道上皮细胞 细胞暴露在管腔切应力下，定义为单位表面积的摩擦力，由 气流。在生理和生理状态下，呼吸道上皮细胞感受到的剪应力是连续变化的。 病理情况。我们有令人兴奋的新数据表明，呼吸道上皮细胞中的剪切力 导致呼吸道上皮通透性改变。因此，呼吸道上皮细胞的动态调节 屏障可能是上皮细胞将管腔信息传递到 上皮下反应。我们的初步研究表明，切应力通过以下方式诱导TRPV4的激活 随后L型电压门控通道激活，随后细胞内钙离子升高。这个 呼吸道上皮细胞内钙升高导致上皮细胞旁的协同变化 通过调节肌动蛋白重排和一氧化氮的产生以及跨膜的通透性 通过调节AQP5的丰度来提高通透性。我们建议研究剪应力在改变两者中的作用。 细胞旁通透性和跨膜通透性，并剖析调节这些通透性的潜在机制。 效果。在SA#1中，我们将使用初级实验研究切应力对呼吸道上皮通透性的影响 培养的人呼吸道上皮细胞(NHBE)和体外分离的小鼠气管。在SA#2中，我们将定义 剪切性钙通道的作用及机制(TRPV4、L型通道) 使用药物抑制和基因操作的上皮通透性。在SA#3中，我们将研究 剪切性钙升高对NO产生的作用及其对通透性的影响。通过这些 作为技术和课程的补充，PI将开发新的技能并生成关于以下方面的新数据 机械切应力对呼吸道上皮细胞调节和功能的影响--一个较少研究的领域 可能对正常的肺功能以及剪切力改变的情况非常重要，包括 运动和哮喘。

The goal of this K08 application is to facilitate development of essential skills that will allow the PI to become a successful academician and achieve independent scientific investigator status. The respiratory epithelium serves as a barrier, a regulator of the content of airway surface liquid, and a source of cytokines and other products that regulate airway physiology. In addition to cyclic extensions and contraction, airway epithelial cells are exposed to lumenal shear stress, defined as the frictional force per unit surface area, generated by airflow. The shear stress sensed by airway epithelia is continuously changing under both physiologic and pathologic conditions. We have exciting novel data suggesting that shear stress in airway epithelial cells leads to changes in airway epithelial permeability. Therefore, dynamic modulation of the airway epithelial barrier could be an important mechanism by which epithelial cells transduce lumenal information into subepithelial responses. Our preliminary studies indicate that shear stress induces TRPV4 activation with subsequent L-type voltage gated channel activation with subsequent increases in intracellular calcium. The increase in intracellular calcium in airway epithelial cells leads to concerted changes in epithelial paracellular permeability by modulating actin rearrangement and NO production, as well as in transmembrane permeability by modulating AQP5 abundance. We propose to study the role of shear stress in altering both paracellular and transmembrane permeability as well as dissect the underlying mechanisms regulating these effects. In SA#1we will investigate the effects of shear stress on airway epithelial permeability using primary cultured human airway epithelial cells (NHBE) and.ex vivo isolated mouse trachea. In SA#2 we will define the role for and the mechanisms (TRPV4, L-type channel) of shear-induced calcium flux in modulating epithelial permeability using pharmacologic inhibition and genetic manipulation. In SA#3 we will study the role od shear-induced increases in calcium on NO production and its effect on permeability. Through these complimentary techniques and coursework, the PI will develop new skills and generate novel data regarding the effects of mechanical shear stress on airway epithelial cell regulation and function, a little-examined area likely to be of immense importance to normal lung function, as well as conditions of altered shear including exercise and asthma.