Regulation of substrate-linked mitochondrial ROS generation

描述(由申请人提供)： 葡萄糖的供应影响细胞的生长、增殖和存活。这些影响中的许多似乎依赖于葡萄糖代谢，特别是底物水平对线粒体通量的控制。胰岛β细胞中的葡萄糖感应通过线粒体氧化磷酸化与胰岛素的分泌偶联，直接影响细胞的能量电荷。包括内皮细胞和胰岛β细胞在内的多种细胞长期暴露在高糖浓度下，导致底物供应与氧化磷酸化的低效偶联，通过将还原当量转移到分子氧的单电子还原来形成超氧自由基。这项应用解决了细胞如何对正能量平衡做出反应的关键问题。使用内皮细胞和胰岛细胞系作为原型细胞谱系，我们将描述细胞和线粒体对超过代谢需求的营养源葡萄糖的可获得性的反应。特别是，我们发现原代内皮细胞在25 mM葡萄糖中孵育诱导c-myc转录，激活核因子-kB致炎通路，下调PI3-K/Akt信号转导。这些途径中的每一个都与线粒体代谢和氧化还原动态平衡有关。具体目标如下：1.为了验证这一假说，葡萄糖刺激的氧化应激依赖于依赖于NRF-1的线粒体基因表达的c-myc反式激活。2.验证核因子-kB转录因子诱导的Bcl-xL通过调节线粒体氧化还原载体之间的平衡来减轻葡萄糖刺激的氧化应激的假说。从这些研究中获得的见解将帮助我们理解高血糖这一信息对于为美国日益增长的糖尿病患者设计有效的器官损伤预防和治疗方案是多么重要。

DESCRIPTION (provided by applicant): Glucose availability affects cell growth, proliferation and survival. Many of these effects appear to depend on glucose metabolism, and in particular, substrate level control of mitochondrial flux. Glucose sensing in pancreatic beta cells is coupled to insulin secretion via mitochondrial oxidative phosphorylation with direct effects on cellular energy charge. Longer term exposure of multiple cell types, including endothelium and pancreatic beta cells, to high glucose concentrations results in inefficient coupling of substrate supply to oxidative phosphorylation, with formation of superoxide radicals by diversion of reducing equivalents to single electron reduction of molecular oxygen. This application addresses key questions about how cells respond to positive energy balance. Using endothelial cells and islet cell lines as prototypic cell lineages, we will characterize cellular and mitochondrial responses to the availability of a nutrient source, glucose, in excess of metabolic requirements. Specifically, we have found that incubation of primary endothelial cells in 25 mM glucose induces c-myc transcription, activation of NF-KB pro-inflammatory pathways and downregulation of PI3-K/Akt signaling. Each of these pathways is relevant to mitochondrial metabolism and redox homeostasis. The following specific aims are proposed: 1. To test the hypothesis glucose-stimulated oxidative stress is dependent on c-myc transactivation of NRF- 1-dependent mitochondrial gene expression. 2. To test the hypothesis that Bcl-xL, induced by NF-kB transcription factors, reduces glucose-stimulated oxidative stress by modulating the equilibrium between mitochondrial redox carriers. Insights gained from these studies will help us understand how hyperglycemia This information will be important for designing effective prevention and treatment for end organ damage for the growing population in the United States with diabetes.