Mechanisms of Intracellular NAMPT-regulated GSNOR in Vessel Wall

细胞内 NAMPT 调节血管壁 GSNOR 的机制

基本信息

批准号：
8660371
负责人：
Adam Carl Straub
金额：
$ 23.7万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-06-15 至 2016-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8660371
关键词：
Anabolism Arteries Attenuated Blood Pressure Blood Vessels Cardiovascular Diseases Cell Signaling Process Cells Coculture Techniques Communication Connexin 43 Connexins Coupling Cysteine Electrons Elements Endothelial Cells Endothelium Enzymes Equilibrium Excision Figs - dietary Functional disorder Future Gap Junctions Grant Hypertension In Vitro Lead Measures Mediating Modeling Modification Molecular Mus Niacinamide Nicotinamide adenine dinucleotide Nitric Oxide Oxidoreductase Pathway interactions Peptides Permeability Pilot Projects Play Post-Translational Protein Processing Process Proteins Proteomics Recombinants Regulation Resistance Role S-Nitrosoglutathione Side Smooth Muscle Smooth Muscle Myocytes Sulfhydryl Compounds Testing Time Vascular Endothelial Cell Vascular Smooth Muscle Vascular resistance Vasoconstrictor Agents Vasodilator Agents Work abstracting base blood pressure regulation cell type constriction human NOS3 protein hypertension treatment inhibitor/antagonist interest protein expression protein protein interaction response trafficking vasoconstriction

项目摘要

Abstract The regulation of resistance arterial tone involves communication between vascular smooth muscle and endothelium, which is tightly controlled by an intricate, but yet to be fully defined, cell signaling processes. Recently, we made the discovery that S-nitrosylation/denitrosylation, the addition or removal of a nitric oxide group from a cysteine-thiol side chain, serves as an important post-translational modification on connexin 43 gap junction (GJ) proteins, and that this modification is associated with control of resistance arterial tone. Regulation of connexin 43 nitrosylation appeared to be predominant at the myoendothelial junction (MEJ), the point where endothelial cells and smooth muscle cells make contact in resistance arteries. At the MEJ, endothelial nitric oxide synthase (eNOS), and the denitrosylase S-nitrosoglutathione reductase (GSNOR), work in concert to modulate the permeability of GJs. The mechanisms regulating eNOS activity have been well characterized, however the molecular mechanisms regulating GSNOR activity remain poorly understood. To identify enriched proteins at the MEJ capable of regulating GSNOR activity, we recently performed an in vitro MEJ proteomic screen. From this analysis, we found enriched expression of nicotinamide phoshoribosyltransferase (NAMPT), a rate-limiting enzyme in the nicotinamide adenine dinucleotide (NAD) biosynthesis pathway. The localized protein expression of intracellular NAMPT at the MEJ suggested to us that it is critical for the regulation of NAD levels, which are known to modulate GSNOR activity and thus might control heterocellular communication in the vessel wall. In our pilot studies, we explored key elements of this concept by showing that intracellular NAMPT can regulate GSNOR activity and resistance arterial tone. Based on these observations we formulated the central hypothesis that vascular resistance and thus, systemic blood pressure control is mediated through a localized NAMPT-regulated GSNOR mechanism. We will test this hypothesis using three specific aims: AIM 1 will test whether NAMPT regulates GSNOR activity and heterocellular communication in vitro, AIM 2 will determine if NAMPT is critical in the regulation of resistance arterial tone, AIM 3 will elucidate how cell-type specific modulation of NAMPT expression in endothelium or smooth muscle modifies the responses to vasoconstrictors or vasodilators in resistance arteries. Our results will impact our understanding of these enzymes in blood pressure control and provide a framework to determine whether dysfunctions in the expression and/or activity of NAMPT and GSNOR contribute to cardiovascular diseases including hypertension.

抽象的动脉阻力张力的调节涉及血管平滑肌和血管之间的通讯内皮细胞受到复杂但尚未完全定义的细胞信号传导过程的严格控制。最近，我们发现S-亚硝基化/去亚硝基化，添加或去除一氧化氮来自半胱氨酸-硫醇侧链的基团，作为连接蛋白 43 上重要的翻译后修饰间隙连接（GJ）蛋白，并且这种修饰与阻力动脉张力的控制有关。连接蛋白 43 亚硝基化的调节似乎在肌内皮连接处 (MEJ) 占主导地位，肌内皮连接处是阻力动脉中内皮细胞和平滑肌细胞接触的点。在MEJ，内皮一氧化氮合酶 (eNOS) 和脱亚硝基酶 S-亚硝基谷胱甘肽还原酶 (GSNOR) 发挥作用共同调节 GJ 的渗透性。调节 eNOS 活性的机制已得到很好的证实然而，调节 GSNOR 活性的分子机制仍知之甚少。到为了鉴定 MEJ 中能够调节 GSNOR 活性的富集蛋白质，我们最近进行了一项体外实验 MEJ 蛋白质组筛选。从该分析中，我们发现烟酰胺的富集表达磷酸核糖基转移酶 (NAMPT)，烟酰胺腺嘌呤二核苷酸 (NAD) 中的限速酶生物合成途径。 MEJ 细胞内 NAMPT 的局部蛋白表达向我们提示它对于 NAD 水平的调节至关重要，已知 NAD 水平可以调节 GSNOR 活性，因此可能控制血管壁中的异细胞通讯。在我们的试点研究中，我们探索了这一点的关键要素通过证明细胞内 NAMPT 可以调节 GSNOR 活性和阻力动脉张力来提出这一概念。基于根据这些观察结果，我们制定了中心假设，即血管阻力以及全身血液压力控制是通过局部 NAMPT 调节的 GSNOR 机制介导的。我们将测试这个使用三个特定目标的假设：AIM 1 将测试 NAMPT 是否调节 GSNOR 活性以及体外异细胞通讯，AIM 2 将确定 NAMPT 在耐药性调节中是否至关重要动脉张力，AIM 3 将阐明内皮或细胞中 NAMPT 表达的细胞类型特异性调节如何平滑肌改变阻力动脉中对血管收缩剂或血管扩张剂的反应。我们的成果将影响我们对这些血压控制酶的理解，并提供一个框架确定 NAMPT 和 GSNOR 的表达和/或活性功能障碍是否导致心血管疾病包括高血压。