Regulation of the Nrf2-mediated Antioxidant Defense In Diabetic Retinopathy

项目摘要 糖尿病视网膜病变是美国工作年龄失明的主要原因，影响超过三分之一。 在约2000万糖尿病患者中。这种疾病的发病机制是由以下几个因素组合而成的 高血糖和胰岛素介导的信号减少，这会影响视网膜神经元、神经胶质细胞和 脉管系统。糖尿病并发症病理生理学的统一理论表明， 导致高血糖引起的组织损伤的途径都与反应性的积累有关 氧物种(ROS)。在糖尿病中，视网膜表现出ROS的产生增加和受损 减少自由基的能力。我的中心假设是糖尿病引起的压力表达 发育和DNA损伤调节反应蛋白1(Redd1)抑制核因子红系-1 2相关因子2(NRF2)相关的视网膜抗氧化反应，导致氧化应激增加和 视网膜病理学。为了支持我的假设，我提出了令人信服的初步数据，证明 糖尿病诱导的氧化应激在Redd1基因敲除小鼠的视网膜中得到缓解。此外，还可以表达 在Redd1基因敲除小鼠视网膜中对Nrf2反应的mRNAs和核Nrf2蛋白的表达增加 在Redd1基因敲除的人视网膜细胞中，培养的MIO-M1细胞的表达和活性增强。我计划这样做 通过追求以下具体目标来检验我的中心假设：1.建立Redd1对Nrf2的影响 糖尿病实验模型的合成。2.描述Redd1对Nrf2降解的影响 糖尿病的实验模型。为了验证我的假设，我将执行一项实验方案，包括 从细胞培养到完整小鼠的模型系统，以及用于分析的尖端技术 信使核糖核酸翻译。这一奖学金还将提供两个关键的培训机会。首先，我会和你一起训练 王穗博士(斯坦福大学)开发操纵视网膜基因表达所需的技能 在活体内。随后，我将接受阿利斯泰尔·巴伯博士(宾夕法尼亚州立医学院)的培训 发展技术专长以评估糖尿病对视网膜病理生理学的影响 相干断层扫描、视网膜电信号和虚拟验光。关于结果，这个项目将 不仅扩展我的技能和分析系统，而且还将确定将 糖尿病代谢环境引起的分子事件对视网膜病变的发展。 确定这类机制具有重要意义，因为它将验证发展的新目标 旨在解决糖尿病视网膜病变分子基础的预防和/或治疗干预措施 和促进健康视力。

Project Summary Diabetic retinopathy is the leading cause of blindness in working age Americans, affecting more than a third of the ~20 million individuals with diabetes. The pathogenesis of this disease is defined by a combination of hyperglycemia and a reduction in insulin mediated signaling, which impacts retinal neurons, glia, and vasculature. A unifying theory for the pathophysiology of diabetic complications suggests that the principle pathways responsible for hyperglycemia-induced tissue damage are all linked to the accumulation of reactive oxygen species (ROS). In diabetes, the retina exhibits an increase in the production of ROS and an impaired capacity to reduce free radicals. My central hypothesis is that diabetes-induced expression of the stress response protein regulated in development and DNA damage 1 (REDD1) inhibits the nuclear factor erythroid- 2-related factor 2 (Nrf2)-related antioxidant response in the retina, leading to increased oxidative stress and retinal pathology. In support of my hypothesis, I present compelling preliminary data demonstrating that diabetes-induced oxidative stress is attenuated in the retina of REDD1 knockout mice. In addition, expression of Nrf2-responsive mRNAs is increased in the retina of REDD1 knockout mice and nuclear Nrf2 protein expression and activity are enhanced in REDD1 knockout human MIO-M1 retinal cells in culture. I plan to test my central hypothesis by pursuing the following specific aims: 1. Establish the impact of REDD1 on Nrf2 synthesis in experimental models of diabetes. 2. Delineate the impact of REDD1 on Nrf2 degradation in experimental models of diabetes. To test my hypothesis, I will pursue an experimental protocol involving model systems ranging from cell culture to intact mice, as well as cutting-edge technologies for analyzing mRNA translation. This fellowship award will also provide two key training opportunities. First, I will train with Dr. Sui Wang (Stanford University) to develop skills necessary to manipulate gene expression in the retina in vivo. Subsequently, I will receive training from Dr. Alistair Barber (Penn State College of Medicine) to develop the technical expertise to assess the impact of diabetes on retinal pathophysiology using optical coherence tomography, electroretinograms, and virtual optomotry. With respect to outcomes, this project will not only expand my skills and systems of analysis, but will also identify novel mechanisms that link the molecular events caused by the diabetic metabolic environment to the development of retinal pathology. Identification of such mechanisms is significant because it will validate new targets for the development of preventive and/or therapeutic interventions aimed at addressing the molecular basis of diabetic retinopathy and promoting healthy vision.