Protein S-glutathionylation and vascular dysfunction with aging

项目摘要 血管内皮功能障碍（VED）是血管老化的一个重要特征，是老年人易患的基础。 成人CVD持续的氧化应激是VED的主要驱动力，通过协同失调， 产生血管舒张剂NO和血管收缩剂内皮素-1（ET-1）。但潜在的氧化还原机制重新- 主要不清楚。逆转氧化应激的新方法特别引人注目，因为它们是无效的- 针对寿命极短的ROS的一般抗氧化疗法的缺乏。蛋白S-谷胱甘肽化（PrS- SG），一种稳定但可逆的氧化剂诱导的翻译后修饰主导了分子机制 氧化还原信号。PrS-SG的还原由谷氧还蛋白特异性催化，因此谷氧还蛋白在氧化还原中是关键的。 调节细胞功能。谷氧还蛋白-1（Glutaredoxin-1，Glutaredoxin-1）功能障碍，是去谷胱甘肽化的主要参与者， 与心脏和大脑老化有关它在血管老化中的作用尚不清楚。这个应用程序是建立在一个强大的 前提我们已发表的和初步的研究表明Glrx 1缺陷和PrS-SG诱导促进 内皮功能障碍和衰老相关的代谢综合征，以及在衰老过程中，主动脉Glrx 1是 减少，同时增加内皮PrS-SG。文献中的证据也支持因果关系 Pr-SSG在VED中的作用在于S-谷胱甘肽化可以解偶联eNOS并激活p21 Ras，已知其 降低NO生物利用度并刺激Erk 1/2依赖性ET-1表达。因此，我们假设年龄相关的 Glrx 1下调通过失调eNOS和ET-1的综合氧化还原信号促进VED， 可通过补充内皮细胞中的Grlx 1逆转。这一中心假设是通过追求两个具体的测试 目的：1）探讨Glrx 1/PrS-SG与VED发生、发展的关系。 使用C57 BL 6 J小鼠，一种良好表征的衰老模型，我们将测量主动脉Glrx 1的时间变化， PrS-SG和内皮依赖性血管舒张在年轻，中年和老年小鼠，和纵向 无创性光学相干断层成像在体研究股动脉血流介导的舒张功能变化 法为了检测Glrx 1下调在VED中的因果作用，我们将检测VED是否在 Glrx 1基因敲除小鼠; 2）验证补充血管内皮Glrx 1可逆转VED的新概念。 我们将使用一种新的可诱导内皮特异性Glrx 1转基因小鼠模型， 以及eNOS和ET-1在青年、中年和老年人中的整合信号传导 小鼠R21赠款的积极结果将有助于推进我们对氧化还原信号传导如何介导的理解 衰老对内皮细胞的多方面影响，促进VED，并提供一种新的抗氧化治疗策略， 恢复老年人的血管功能。

PROJECT SUMMARY Vascular endothelial dysfunction (VED) is a key feature of vascular aging underlying the predisposition of old adults to CVD. Sustained oxidative stress is a major driving force for VED by synergically dysregulating the production of vasodilator NO and vasoconstrictor endothelin-1(ET-1). But the underlying redox mechanisms re- main unclear. Novel approaches to reversing oxidative stress are particularly compelling given the ineffective- ness of general antioxidant therapies that targets extremely short-lived ROS. Protein S-glutathionylation (PrS- SG), a stable but reversible oxidant-induced posttranslational modification dominates the molecular mechanisms for redox signaling. Reversal of PrS-SG is catalyzed specifically by glutaredoxin which thereby is critical in redox regulation of cellular function. Dysfunction of glutaredoxin-1 (Glrx-1), a major actor in the de-glutathionylation is implicated in cardiac, and brain aging. Its role in vascular aging is unknown. This application is built upon a strong premise. Our published and preliminary studies indicate that Glrx1 deficiency and PrS-SG induction promote endothelial dysfunction and aging-associated metabolic syndromes and that during aging, aortic Glrx1 is decreased with a concomitant increase in endothelial PrS-SG. Evidence in the literature also supports a causal role of Pr-SSG in VED in that S-glutathionylation can uncouple eNOS and activate p21Ras, which are known to impair NO bioavailability and stimulate Erk1/2 dependent-ET-1 expression. We thus hypothesize that age-related Glrx1 downregulation promotes VED by dysregulating the integrated redox signaling of eNOS and ET-1, which can be reversed by replenishing Grlx1 in endothelium. This central hypothesis is tested by pursuing two specific aims: 1) To understand the relationship between Glrx1/PrS-SG and the onset and progression of VED with aging. Using C57BL6J mice, a well-characterized model of aging we will measure the temporal changes in aortic Glrx1, PrS-SG, and endothelium-dependent vasorelaxation in young, middle-aged and old mice, and longitudinal changes of flow-mediated dilation of femoral artery in vivo using a non-invasive Optical-Coherence-Tomography technique. To test the causal role of Glrx1 downregulation in VED, we will test whether VED is aggravated in Glrx1 knockout mice; 2) To test a new concept that replenishing Glrx1 in endothelium can reverse VED in aging. Using a novel inducible endothelial specific Glrx1 transgenic mouse model, we will determine the impact on VED and the integrated signaling of eNOS and ET-1 of Glrx1 transgene expression in young, middle-aged, and old mice. The positive results in the R21 grant will help advance our understanding how redox signaling mediates the multifaceted effects of aging on ECs, promoting VED, and offer a new antioxidant therapeutic strategy to restore vascular function in older adults.