Flow-Mediated Dilation of Human Coronary Arterioles

作用于内皮细胞的剪切力会产生血管扩张。这可以说是最具生理学意义的 重要的内皮细胞扩张机制，几乎发生在每一个血管床上。来自我们的最新数据 实验室研究表明，冠心病患者的冠状动脉微动脉发生了血流介导的扩张(FMD)。 然而，疾病通过一种新的机制运作，涉及内皮细胞产生活性氧 物种(ROS)，包括过氧化氢(H2O2)。令人惊讶的是，线粒体呼吸链在 在人类心脏FMD中的必要作用，但目前尚不清楚线粒体是如何参与 在血管内皮细胞表面传递机械剪应力以引起扩张。 我们的总体假设是剪切力通过附着的细胞骨架元件作用于内皮细胞 刺激线粒体释放过氧化氢，一种内皮源性超极化因子(EDHF)。我们会 用三种方式检验这一假设。首先将使用抗霉素A和TNFa来确定药理作用 刺激线粒体ROS释放可引起人冠状动脉小动脉扩张。在单独的研究中，我们 应使用针对线粒体内膜的新型抗氧化剂来确定是否产生过氧化氢 线粒体是口蹄疫所必需的。生物检测系统将确认过氧化氢是否确实是一种 EDHF介导人冠脉循环中的FMD。第二，我们将使用免疫组织化学和特异性 包括siRNA在内的药理学和分子方法来确定内皮细胞骨架 元素在FMD和线粒体ROS的产生中起着必要的作用。第三，我们将检验这一假设 一氧化氮通过对线粒体呼吸的抑制作用降低了人类心脏的FMD。这将是 使用一氧化氮供体，测量线粒体复合体的活性和ROS的产生。这些 实验可能发现一条以前没有描述过的途径，即一氧化氮可以抑制EDHF介导的途径 扩张；即通过阻止线粒体产生ROS。 这些目标共同解决了一种新的内皮依赖性血管扩张机制，涉及 线粒体产生的ROS，到目前为止只在人类心脏中有报道。这些研究应该确定新的联系 在调节生理功能的细胞过程中，包括机械转导、呼吸和氧化还原信号 血管扩张等事件。

Shear stress acting on endothelial cells produces vasodilation. This is arguably the most physiologically important endothelial mechanism of dilation and occurs in virtually every vascular bed. Recent data from our laboratory indicate that flow-mediated dilation (FMD) occurs in coronary arterioles from patients with coronary disease however it operates through a novel mechanism involving endothelial production of reactive oxygen species (ROS) including hydrogen peroxide (H2O2). Surprisingly the mitochondrial respiratory chain plays a necessary role in FMD in the human heart, however it is not known how mitochondria are involved in the transduction of mechanical shear stress on the surface of the endothelium to elicit dilation. Our overall hypothesis is that shear acting on endothelial cells through attached cytoskeletal elements stimulates release from the mitochondria ofH2O2, an endothelial derived hyperpolarizing factor (EDHF). We shall test this hypothesis in three ways. First antimycin A and TNFa will be used to determine if pharmacological stimulation of mitochondrial ROS release can elicit dilation of human coronary arterioles. In separate studies, we shall use novel antioxidants targeted to the mitochondrial inner membrane to determine whether H2O2 generated from within the mitochondria is necessary for FMD. A bioassay system will confirm whether H2O2 is indeed an EDHF mediating FMD in the human coronary circulation. Second we will use immunohistochemistry and specific pharmacological and molecular approaches including siRNA to determine whether endothelial cytoskeletal elements play a necessary role in FMD and mitochondrial ROS generation. Third, we shall test the hypothesis that nitric oxide through its inhibitory effect on mitochondrial respiration reduces FMD in the human heart. This will be done using nitric oxide donors, measuring mitochondrial complex activity and ROS generation. These experiments may identify a pathway not previously described by which nitric oxide can inhibit EDHF-mediated dilation; namely, by blocking mitochondrial production of ROS. Collectively these aims address a novel mechanism of endothelium-dependent vasodilation involving mitochondrial generation of ROS, thus far reported only in human hearts. These studies should identify new links among cell processes including mechanotransduction, respiration, and redox signaling that regulate physiological events such as vasodilation.