Mechanism of Coronary Endothelium-mediated Vasodilation

冠状动脉内皮介导的血管舒张机制

• 批准号: 6326391
• 负责人: PinLan Li
• 金额: $ 28.19万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-01-01 至 2005-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6326391
• 关键词: ADP ribosylation; CD38 molecule; adenosine diphosphate; animal tissue; biological signal transduction; calcium channel; calcium flux; coronary artery; enzyme activity; fluorescence spectrometry; nitric oxide; nucleoside monophosphate; receptor binding; receptor expression; ribose; sarcoplasmic reticulum; tissue /cell culture; vascular endothelium; vasodilation; video microscopy; voltage /patch clamp

DESCRIPTION (Verbatim from the application): Recent studies from our laboratory and by others have indicated that cyclic adenosine diphosphate-ribose (cADPR) is produced and hydrolyzed in coronary arterial smooth muscle cells, and that this nucleotide serves as a second messenger to stimulate Ca24 release from the sarcoplasmic reticulum (SR). This proposal will examine the hypothesis that endogenous cADPR-induced Ca2+ release contributes to the control of [Ca2+], in vascular smooth muscle cells from small coronary arteries and that NO decreases intracellular cADPR production and consequently lowers [Ca2+], thereby resulting in vasodilation. We will first examine the role of cADPR in the regulation of ryanodine receptor/Ca2+ release channel activity using SR Ca2+ channel reconstitution and lipid bilayer clamp techniques. Then, we will further determine the role of cADPR-mediated activation of ryanodine receptors in the control of intracellular [Ca2+] in coronary arterial smooth muscle cells and the contribution of this signaling pathway to the inhibitory effect of NO on [Ca2+]i using single cell fluorescence microscopic spectrometry. Since cADPR has been reported to act through Ca2+-induced Ca2+ release (CICR), we will address whether the cADPR-mediated effects of NO are associated with inhibition of CICR or ryanodine receptors. We will also examine the effects of NO on the production or metabolism of cADPR by reverse phase-HPLC analysis of ADP-ribosylcyclase and cADPR hydrolase activities and to explore the mechanisms by which NO modulates these enzyme activities in coronary arterial smooth muscle. The involvement of cGMP and nitrosylation-mediated dimerization of ADP-ribosylcyclase in the effect of NO will be determined. Using videomicroscopy of isolated pressurized small coronary arteries, we will determine the role of cADPR, CICR and ryanodine receptors in the development of coronary tension and in mediating the vasodilator response to NO. Finally, we will explore the precise mechanisms by which cADPR binds or activates ryanodine receptors and produces Ca2+ release from the SR using radioligand binding or ADP ribosylation assays. These studies will define a new signaling mechanism regulating [Ca2+]i and vasomotor response in coronary resistance arteries and increase our understanding of the cellular mechanism mediating the vasodilator effect of NO.

描述（申请中的逐字记录）：我们实验室的最新研究 和其他人指出，环腺苷二磷酸核糖（cADPR） 在冠状动脉平滑肌细胞中产生并水解，并且 该核苷酸作为第二信使刺激Ca24从细胞中释放 肌浆网（SR）。该提案将检验以下假设： 内源性 cADPR 诱导的 Ca2+ 释放有助于控制 [Ca2+]， 来自小冠状动脉的血管平滑肌细胞，NO 减少 细胞内 cADPR 的产生，从而降低 [Ca2+]，从而 从而导致血管舒张。我们将首先研究 cADPR 在 使用 SR Ca2+ 调节兰尼碱受体/Ca2+ 释放通道活性 通道重建和脂质双层钳技术。那么，我们将 进一步确定 cADPR 介导的兰尼碱受体激活的作用 控制冠状动脉平滑肌细胞的细胞内 [Ca2+] 以及该信号通路对NO抑制作用的贡献 使用单细胞荧光显微光谱法测定[Ca2+]i。自从 cADPR 据报道通过 Ca2+ 诱导 Ca2+ 释放 (CICR) 起作用，我们将 解决 NO 的 cADPR 介导的作用是否与抑制相关 CICR 或兰尼定受体。我们还将研究 NO 对 通过反相 HPLC 分析 cADPR 的产生或代谢 ADP-核糖基环化酶和cADPR水解酶活性并探讨其机制 NO 通过其调节冠状动脉平滑肌中的这些酶活性 肌肉。 cGMP 和亚硝基化介导的二聚化的参与 ADP-核糖基环化酶对NO的影响将被测定。使用 孤立的加压小冠状动脉的视频显微镜检查，我们将 确定 cADPR、CICR 和兰尼定受体在发育中的作用 冠状动脉张力并介导血管舒张剂对 NO 的反应。最后，我们 将探索 cADPR 结合或激活兰尼碱的精确机制 受体并使用放射性配体结合从 SR 中释放 Ca2+ 或 ADP 核糖基化测定。这些研究将定义新的信号机制 调节冠状动脉阻力动脉中的 [Ca2+]i 和血管舒缩反应 增加我们对介导血管舒张剂的细胞机制的理解 NO的影响。