Chloride channels in endothelial cells

项目摘要 内皮细胞调节各种各样的血管功能，包括调节器官的收缩能力 血流量和全身血压。几乎所有心血管疾病的一个特征就是功能不全。 内皮细胞，但涉及的病理机制尚不清楚。表达了几种阳离子通道 在内皮细胞中调节动脉收缩能力，包括小(SK)和中(IK)电导钙 (Ca2+)激活的钾通道和瞬时受体电位通道，包括TRPV4。相比之下， 内皮细胞中阴离子通道的生理功能还知之甚少。ECS表达钙离子TMEM16A- 激活的氯离子通道，但该阴离子通道是否调节动脉收缩尚不清楚。类似 内皮细胞TMEM16A通道在血管功能障碍中的病理参与尚不清楚 这发生在高血压期间。使用广泛的方法，包括他莫昔芬诱导的内皮细胞 细胞特异性基因敲除小鼠，我们提供了血管扩张剂刺激激活TMEM16A通道的证据 血管内皮细胞诱导血管扩张。初步数据还表明，内皮细胞TMEM16A通道 在高血压期间会出现功能障碍。在这项提案中，我们将调查三个具体目标。目标1将 探讨生理性血管扩张剂激活内皮细胞TMEM16A通道的信号机制 以诱导血管松弛。AIM 2将研究TMEM16A通道激活的机制 内皮细胞可引起血管扩张。目标3将研究高血压与 内皮细胞中TMEM16A通道的病理变化抑制了这些蛋白的血管扩张。 使用的方法将包括分子生物学和生化技术、超分辨 免疫荧光显微镜、高速双倒置选择性平面照明显微镜(DiSPIM)、 双光子共聚焦显微镜，膜片钳和传统电生理学，动脉肌电图和 无线电遥测。该项目将提供重要的生理和病理方面的新信息。 内皮细胞中TMEM16A通道对血管的调节作用。

Project Summary Endothelial cells regulate a wide variety of vascular functions, including contractility, which modulates organ blood flow and systemic blood pressure. A hallmark of virtually all cardiovascular diseases is dysfunctional endothelial cells, but pathological mechanisms involved are uncertain. Several cation channels are expressed in ECs that modulate arterial contractility, including small- (SK) and intermediate (IK)-conductance calcium (Ca2+)-activated potassium channels and transient receptor potential channels, including TRPV4. In contrast, physiological functions of anion channels in ECs are poorly understood. ECs express TMEM16A, a Ca2+- activated Cl- channel, but whether this anion channel regulates arterial contractility is unclear. Similarly uncertain is the pathological involvement of endothelial cell TMEM16A channels in the vascular dysfunction that occurs during hypertension. Using a broad range of approaches, including tamoxifen-inducible endothelial cell-specific knockout mice, we provide evidence that vasodilator stimuli activate TMEM16A channels in endothelial cells to induce vasodilation. Preliminary data also suggest that endothelial cell TMEM16A channels are dysfunctional during hypertension. In this proposal, we will investigate three specific aims. Aim 1 will investigate the signaling mechanisms by which physiological vasodilators activate TMEM16A channels in ECs to induce vasorelaxation. Aim 2 will examine the mechanisms by which the activation of TMEM16A channels in endothelial cells elicits vasodilation. Aim 3 will study the hypothesis that hypertension is associated with pathological alterations in TMEM16A channels in endothelial cells which inhibits vasodilation by these proteins. Methods used will include molecular biology and biochemistry techniques, super-resolution immunofluorescence microscopy, high-speed dual-inverted selective plane illumination microscopy (diSPIM), two-photon confocal microscopy, patch-clamp and conventional electrophysiology, arterial myography and radiotelemetry. This project will provide significant novel information regarding physiological and pathological vasoregulation by TMEM16A channels in endothelial cells.