SLC4A11 Mitochondrial Uncoupling and ROS Production in Corneal Endothelium

SLC4A11 角膜内皮线粒体解偶联和 ROS 产生

• 批准号: 10393579
• 负责人: Joseph Aurelio Bonanno
• 金额: $ 41.2万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10393579
• 关键词: Address; Aging; Amino Acids; Ammonia; Antioxidants; Apical; Apoptosis; Autophagocytosis; Back; Bicarbonates; Blindness; Buffers; Carcinoma; Catabolism; Cell Density; Cell Line; Cell membrane; Cells; Citric Acid Cycle; Colon Carcinoma; Consumption; Cornea; Corneal Endothelium; Corneal dystrophy; Corneal edema; Data; Defect; Down-Regulation; Electron Transport; Endothelial Cells; Endothelium; Feeds; Fuchs' Endothelial Dystrophy; Functional disorder; Genes; Glutamates; Glutamine; Goals; Human; Hydration status; In Vitro; Inflammation; Inherited; Knock-out; Knockout Mice; Laboratories; Lactate Transporter; Lead; Malignant Epithelial Cell; Measurement; Measures; Medical; Membrane Potentials; Membrane Proteins; Membrane Transport Proteins; Metabolism; Mitochondria; Mitochondrial Membrane Protein; Modeling; Mus; Mutation; NADP; Na(+)-K(+)-Exchanging ATPase; Oryctolagus cuniculus; Pathology; Phenotype; Physiology; Production; Property; Proteins; Pump; Reactive Oxygen Species; Reduced Glutathione; Role; Source; Stimulus; Taurine; Testing; Tissues; Transfection; Transplantation; Trauma; UCP2 protein; Variant; Vision; Work; basolateral membrane; carbonate dehydratase; cell type; in vivo; insight; loss of function; mitochondrial dysfunction; mitochondrial membrane; mouse model; novel; prevent; therapy development; transcriptome sequencing

“SLC4A11 Mitochondrial Uncoupling and ROS Production in Corneal Endothelium” ABSTRACT Defects in the gene SLC4A11 cause Congenital Hereditary Endothelial Dystrophy and some forms of Fuchs Dystrophy. The goal of this study is to understand the role of this membrane transporter in normal corneal endothelial metabolism and how SLC4A11 deficiency leads to Corneal Dystrophy. The corneal endothelial “pump” maintains corneal hydration and transparency. When the “pump” fails due to trauma, inflammation, ageing, or dystrophy, corneal edema ensues, transparency is lost, and vision is significantly degraded. The usual therapy is transplantation, which is not without significant compromises and complications. A hallmark of Corneal Endothelial Dystrophies is mitochondrial dysfunction. Our laboratory has shown that SLC4A11 is an NH3 dependent electrogenic H+ transporter. We have found that glutamine is actively metabolized by the endothelium producing NH3 and enhancing ATP formation. Slc4a11 knock out shows significant corneal edema, lactate accumulation, altered mitochondrial physiology, and ROS. These data have led to the overarching hypothesis that corneal endothelium actively metabolize glutamine and that the absence of SLC4A11 alters glutamine metabolism, leading to mitochondrial dysfunction, ROS, and eventual apoptosis. Preliminary data indicate that SLC4A11 is both a plasma membrane and a mitochondrial membrane protein, leading to the novel hypothesis that SLC4A11 is a mitochondrial uncoupler. Using multiple in vitro & in vivo complementary approaches these hypotheses will be tested in three aims. Aim 1 will determine how glutamine metabolism is facilitated in Corneal Endothelial mitochondria. The hypothesis is that Slc4a11 is an NH3 sensitive mitochondrial uncoupler that works in conjunction with Uncoupling Protein-2 and the potential mitochondrial buffer taurine to facilitate Glutamine catabolism. Aim 2 will examine the source of ROS and ROS as a stimulus to Apoptosis in Slc4a11 KO. Our hypothesis is that apoptosis is accelerated by ROS, which is generated by the interaction of NH3 with an energized electron transport chain and reduced by SLC4A11 uncoupling. Aim 3 will identify the cause of corneal edema in Slc4a11 KO Mice. We will test the hypothesis that loss of Slc4a11 secondarily induces downregulation of key proteins that facilitate lactate transport. Completion of this study will establish the role of SLC4A11 and glutamine in endothelial metabolism; provide new insight for mechanisms that facilitate glutamine metabolism yet alleviate NH3 induced ROS production that will be transferable to a wide array of glutamine metabolizing tissues; and provide insight for development of therapies for endothelial dystrophies.

“SLC4A11 线粒体解偶联和角膜内皮细胞中 ROS 的产生” 抽象的 SLC4A11 基因缺陷导致先天性遗传性内皮营养不良和某些形式的 这项研究的目的是了解这种膜转运蛋白在正常角膜中的作用。 内皮代谢以及 SLC4A11 缺乏如何导致角膜营养不良。 “泵”保持角膜水合作用和透明度 当“泵”因创伤、炎症而失效时， 老化或营养不良，角膜水肿，透明度丧失，视力明显下降。 治疗方法是移植，这并非没有严重的损害和并发症。 内皮营养不良是线粒体功能障碍。我们的实验室已证明 SLC4A11 是一种 NH3。 我们发现谷氨酰胺被内皮细胞活跃地代谢。 产生 NH3 并增强 ATP 形成 Slc4a11 敲除显示显着的角膜水肿、乳酸。 积累、线粒体生理学改变和活性氧这些数据得出了总体假设。 角膜内皮活跃地代谢谷氨酰胺，并且 SLC4A11 的缺失会改变谷氨酰胺 代谢，导致线粒体功能障碍、ROS 和最终细胞凋亡。 SLC4A11 既是质膜蛋白又是线粒体膜蛋白，从而提出了新的假设 SLC4A11 是一种线粒体解偶联剂，采用多种体外和体内互补方法。 假设将在三个目标中进行测试，目标 1 将确定如何促进角膜中的谷氨酰胺代谢。 假设 Slc4a11 是一种 NH3 敏感的线粒体解偶联剂。 与解偶联蛋白 2 和潜在的线粒体缓冲牛磺酸一起促进谷氨酰胺 目标 2 将检查 ROS 的来源以及 ROS 作为 Slc4a11 KO 细胞凋亡的刺激物。 假设是 ROS 会加速细胞凋亡，ROS 是由 NH3 与 通电电子传输链并通过 SLC4A11 解偶联减少目标 3 将确定角膜的原因。 Slc4a11 KO 小鼠的水肿 我们将检验 Slc4a11 缺失继发诱导下调的假设。 促进乳酸转运的关键蛋白质的研究的完成将确定 SLC4A11 和的作用。 内皮代谢中的谷氨酰胺；为促进谷氨酰胺代谢的机制提供新的见解 但可以减轻 NH3 诱导的 ROS 产生，这种产生可转移到广泛的谷氨酰胺代谢中 组织；并为内皮营养不良疗法的开发提供见解。