内皮细胞机械感受蛋白/微器官对血管功能与血压的调控

Mechanosensitive proteins and cilia of vascular endothelial cells play a vital role in regulation of endothelial calcium signaling and nitric oxide (NO) synthesis/release induced by mechano-signaling, such as shear stress. In our preliminary studies, we found that members of transient receptor potential (TRP) channel family, TRPP2 and TRPV4 are highly expressed in the cilium, as well as in plasma membrane of vascular endothelial cells and form a calcium-permeable nonselective cation channel. It appears that these functional channel complexes together with cilia are most likely responsible for shear stress induced NO production/release in vascular endothelial cells. Therefore, we hypothesize that synchronization of cilia and TRPP2/TRPV4 channel complex in respond to shear stress is critical for vascular function by affecting intracellular calcium signaling, and that their function is closely related to NO production/release and the sensitivity to vasoactive substances. We will use patch-clamp analysis, laser confocal imaging analysis, molecular biology techniques and vascular function studies to test our hypothesis and will determine the mechanisms by which TRPP2/TRPV4 channel complex and cilia play very important role in regulation of vascular function and blood pressure. Our findings will provide new insight to regulation of vascular function and may also provide potential therapeutic targets for hypertension.

血管内皮细胞机械感受蛋白/微器官在血管内机械信号（剪切力）诱导的血管内皮细胞钙信号和NO合成/释放方面发挥重要的调节作用。我们发现瞬时受体电位(TRP)通道成员TRPP2和TRPV4在血管内皮细胞的微器官-纤毛和细胞膜上高度表达，并形成可通透钙的非选择性阳离子通道异源多聚体和机械感受器，其功能可能与NO的合成/释放密切相关。因此，我们假设：血管内皮细胞的TRPP2/TRPV4功能性通道复合体与纤毛协同感应血管内剪切力的变化，介导剪切力变化引起的细胞内钙瞬变；该通道复合体和纤毛功能调节异常将影响血管内皮细胞细胞内钙和NO合成/释放，并使血管对血管活性物质的敏感性降低从而影响血管功能和血压调控。我们将采用电生理学、激光共聚焦成像、分子生物学和血管功能学等方法验证上述科学假设并阐明其机制。该课题的实施将为血管功能调控机制提出新的理念，并对高血压的发生和发展提出新的观点和潜在的防治靶点。

瞬时受体电位通道家族（Transient receptor potential, TRP）成员TRPP2和TRPV4能够形成异源的通道复合体，但是该复合体在血管内皮中的作用尚未被揭示。本课题采用分子生物学、电生理记录技术、激光共聚焦等实验手段，揭示了TRPP2和TRPV4在血管内皮上形成可通透钙离子和具有机械感受性能的离子通道蛋白。课题首次发现：1. TRPP2/TRPV4复合体具有双重作用，除纤毛外，TRPP2/TRPV4通道复合体为机械感受蛋白，并表现为可通透钙离子的非选择性阳离子通道，并可应对机械刺激而介导钙离子内流；参与层流刺激诱导的内皮细胞钙瞬变和一氧化氮的生成；当TRPP2/TRPV4功能受到抑制时，通过Akt/eNOS途经抑制NO生物合成；2. EGF通过EGFR酪氨酸激酶通路调节调节TRPP2/TRPV4通道复合体，从而可能参与内皮功能的调节；3. 在发生盐敏感高血压中，TRPP2和TRPV4蛋白的表达和活性受到显著抑制，且TRPP2/TRPV4功能性复合体的形成受到显著抑制；其机制是高盐所致的血清中的醛固酮水平异常升高，通过且醛固酮可通过与盐皮质激素受体结合，而抑制TRPP2/TRPV4通道复合体，降低NO的生成，而使血管内皮细胞功能障碍；4. 而NO作为重要的气体小分子，可上调TRPP2/TRPV4通道复合体的活性。我们的研究揭示：TRPP2/TRPV4功能性复合体在调节内皮功能中具有重要作用，以上研究结果为探索血管内皮细胞上的离子通道异常调节参与盐敏感性血压调控提供理论依据。此外，拓展研究了血管内皮上的TRP家族另一成员——TRPM4，揭示TRPM4是钙激活非选择性阳离子通道的编码基因，发现醛固酮和H2O2可上调TRPM4的表达和活性，TRPM4的功能异常参与了高盐饮食导致的早期血管内皮损伤；另外，TRPM4可能参与脂代谢紊乱引起的血管内皮损伤。