Role of membrane potential in the regulation of endothelial cell deformability

越来越多的证据表明，血管是醛固酮的靶标。最近的数据表明，在某些情况下，类固醇会使内皮变得僵硬和脆弱。尽管其临床相关性明显，且已成功地使用了醛固酮拮抗剂进行治疗，但其在血管内皮细胞中的作用机制尚不清楚。细胞质膜电势差受多种离子通道的活动控制，在调节内皮细胞的刚性和控制细胞的变形能力方面起着关键作用。醛固酮导致内皮细胞顶膜上皮钠通道的表达，被认为是内皮细胞的去极化作用。为了测试膜电位的作用和潜在的离子通道活动，我们设计了一个项目，利用荧光方法操纵和测量内皮细胞的膜电位差。同时，细胞的硬度和变形性(通过使用原子力显微镜作为纳米传感器)将被量化。最后，一氧化氮的形成将与膜电位和细胞硬度有关。工作假说是，内皮细胞膜电位差的大小决定了细胞的变形能力，因此决定了NO的释放和血管平滑肌的张力。通过使用阻滞剂和激动剂，我们将从功能上识别内皮膜电位差异背后的不同离子通道，重点是醛固酮调节的上皮性钠通道。该项目的目的是确定引起细胞变形性变化的膜机制的关键因素，并将它们与内皮的生理功能联系起来。

There is accumulating evidence that blood vessels are targets for aldosterone. Recent data indicate that, under certain circumstances, the steroid renders endothelia stiff and vulnerable. Although the clinical relevance is obvious and aldosterone antagonists are successfully used for treatment, the mechanism of aldosterone action in endothelial cells is unknown. The electrical plasma membrane potential difference, dominated by the activities of various ion channels, could play a key role in the regulation of endothelial stiffness and in the control of the cell's deformability. Aldosterone leads to the expression of epithelial sodium channels in the apical membrane of endothelial cells which is supposed to depolarize the cells. To test the role of the membrane potential and the underlying ion channel activities we designed a project to manipulate and measure the membrane potential difference of endothelial cells applying fluorescence methods. In parallel, cell stiffness and deformability (by using the atomic force microscope as a nanosensor) will be quantified. Finally, nitric oxide formation will be related to membrane potential and cell stiffness. The working hypothesis is that the magnitude of the endothelial cell membrane potential difference determines the cell's deformability and, as a consequence, determines NO release and vascular smooth muscle tone. By using blockers and agonists we will functionally identify the different ion channels underlying the endothelial membrane potential difference with emphasis on aldosteroneregulated epithelial sodium channels. The aim of this project is to identify the key players of the membrane-born mechanisms that cause the changes in cellular deformability and to relate them to the physiological functions of the endothelium.