Vitamin C, glutathione, and endothelial cell activation

这个项目是基于这样一个假设，即细胞的氧化还原状态 在预防内皮细胞功能障碍方面起着关键作用， 动脉粥样硬化氧化应激（氧化剂和抗氧化剂的不平衡） 氧化剂有利于前者）已被牵连作为一个致病因素 在动脉粥样硬化形成中， (LDL)及其后遗症和刺激单核细胞-内皮细胞 交互.虽然以前的研究已经评估了LDL的影响， 相关的和细胞外的抗氧化剂对这些促动脉粥样硬化，氧化还原- 敏感的过程，很少有人知道细胞内的作用， 抗氧化剂维生素C和谷胱甘肽被积累和合成， 分别以毫莫耳浓度被人体细胞吸收， 在细胞抗氧化防御系统中的控制作用，从而在细胞 完整性和功能在面对氧化剂的挑战。因此 该项目的总体目标是确定细胞维生素C的作用 和谷胱甘肽状态在体外和体内的内皮激活。 体外培养系统将培养人主动脉内皮细胞 （HAEC）和体内系统豚鼠，与人类一样， 合成抗坏血酸。第一个目标是描述和操纵 HAEC的维生素和谷胱甘肽状态。维生素C的作用 对细胞谷胱甘肽状态的负荷，以及相反的影响， 根据维生素C状态操纵细胞谷胱甘肽状态将是 研究了目的2将确定细胞氧化还原状态在HAEC中的作用。 介导的LDL氧化。将通过以下方式探讨基本机制： 测量细胞超氧化物和硫醇的产生，并通过抑制 一氧化氮合成在第三个目标中，我们将确定 HAEC活化中的细胞维生素C和谷胱甘肽状态，即 细胞粘附分子（CAM）和单核细胞的表达 趋化蛋白-1和单核细胞趋化蛋白-1 （MCP-1）和单核细胞粘附。基本机制将是 通过研究氧化还原敏感的细胞核转位， 转录因子NF κ B和一氧化氮的参与。在aim中 4，将研究HAEC研究的体内相关性。的 维生素C和/或谷胱甘肽状态的豚鼠喂食标准或0.3% 含胆固醇的饮食将被操纵， 将测量MCP-1、CAM和氧化LDL的自身抗体， 以及主动脉CAM和MCP-1表达，NfkappaB激活， 单核细胞粘附和动脉粥样硬化病变形成的程度。 这一信息将有助于更好地了解 抗氧化剂通过其改变动脉粥样硬化形成。

This project is based upon the hypothesis that the cellular redox status plays a critical role in preventing endothelial cell dysfunction related to atherogenesis. Oxidative stress (an imbalance of oxidants and anti- oxidants in favor of the former) has been implicated as an etiologic factor in atherogenesis, both through the oxidation of low-density lipoprotein (LDL) and its sequelae and the stimulation of monocyte-endothelial interactions. While previous studies have evaluated the effects of LDL- associated and extracellular anti-oxidants on these pro-atherogenic, redox- sensitive processes, little is known about the role of intracellular antioxidants. Vitamin C and glutathione are accumulated and synthesized, respectively, in millimoral concentrations by human cells, and play a control role in the cellular antioxidant defense system, and thus in cellular integrity and function in the face of an oxidant challenge. Therefore, the overall objective of this project is to identify the role of cellular vitamin C and glutathione status in endothelial activation in vitro and in vivo. The in vitro system will be cultured human aortic endothelial cells (HAEC), and the in vivo system guinea pigs, which, like humans, cannot synthesize ascorbic acid. The first aim is to characterize and manipulate the vitamin and glutathione status of HAEC. The effects of vitamin C loading on cellular glutathione status, and conversely the effects of manipulating cellular glutathione status on vitamin C status will be studied. Aim 2 will identify the role of cellular redox status in HAEC- mediated LDL oxidation. Underlying mechanisms will be explored by measuring cellular superoxide and thiol production, and by inhibiting nitric oxide synthesis. In the third aim we will determine the role of cellular vitamin C and glutathione status in HAEC activation, i.e. expression of cellular adhesion molecules (CAMs) and monocyte chemotactic protein-1 (MCP-1) and monocyte chemotactic protein-1 (MCP-1) and monocyte adhesion. Underlying mechanisms will be explored by studying the nuclear translocation of the redox-sensitive transcription factor NfkappaB and the involvement of nitric oxide. In aim 4, the in vivo relevance of the HAEC studies will be investigated. The vitamin C and/or glutathione status of guinea pigs fed a standard or 0.3% cholesterol-containing diet will be manipulated, and circulating levels of MCP-1, CAMs and autoantibodies to oxidized LDL will be measured, as well as aortic CAM and MCP-1 expression, NfkappaB activation, monocyte adhesion, and the extent of atherosclerotic lesion formation. This information will provide a better understanding of the mechanisms by which antioxidants modify atherogenesis.