Role of Oxidative Stress in Pathogenesis of Fuchs Endothelial Corneal Dystrophy

氧化应激在福克斯内皮性角膜营养不良发病机制中的作用

• 批准号: 10448283
• 负责人: Ula V. Jurkunas
• 金额: $ 39.77万
• 依托单位: SCHEPENS EYE RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10448283
• 关键词: ATM null mice; Affect; Antioxidants; Apoptosis; Apoptotic; Blindness; CYP1B1 gene; Cell Aging; Cell Cycle; Cell Cycle Arrest; Cell Death; Cell Line; Cells; Cornea; Corneal Endothelium; Corneal edema; DNA; DNA Adduction; DNA Adducts; DNA Damage; DNA Repair; DNA Repair Disorder; DNA Repair Gene; Data; Deposition; Development; Disease; Down-Regulation; Elderly; Endothelial Cells; Endothelium; Enzymes; Estrogen Metabolism; Estrogens; Etiology; Extracellular Matrix; Female; Fuchs' Endothelial Dystrophy; Genes; Genetic; Genotype; Goals; Grant; Human; In Vitro; Incidence; Intervention; Investigation; Keratoplasty; Laboratories; Lead; Light; Link; Liquid substance; M Phase Arrest; M cell; Mesenchymal; Mitochondria; Mitochondrial DNA; Mitosis; Mus; NQO1 gene; Outcome; Oxidants; Oxidative Stress; Pathogenesis; Pathogenicity; Patients; Pharmacological Treatment; Phenotype; Physiological; Predisposition; Resistance; Role; Severities; Sex Differences; Specimen; Tissues; UV induced; UVA induced; Ultraviolet Rays; Woman; age related; aqueous; base; cell injury; genotoxicity; human old age (65+); in vivo; irradiation; male; mitochondrial dysfunction; mouse model; non-genetic; novel; oxidative DNA damage; oxidative damage; repaired; response; response to injury; senescence; sex; ultraviolet

Project Summary/Abstract Fuchs Endothelial Cornel Dystrophy (FECD), a common age-related dystrophy, which is more prevalent in women, is of unknown etiology. In FECD, corneal endothelial (CE) cell loss is accompanied by abnormal extracellular matrix (ECM) deposition in the form of guttae. Our laboratory was the first to link oxidative DNA damage and mitochondrial dysfunction in FECD pathogenesis. Specifically, we showed that DNA damage, induced by ultraviolet-A (UVA) light, causes FECD in mice. Moreover, the UVA induced CE cell cycle arrest in G2/M phase and cellular senescence. We identified the novel involvement of CYP1B1, the key estrogen- metabolizing enzyme, in sex-dependent differences in CE susceptibility to UVA; and detected greater mitochondrial DNA (mtDNA) damage in female mice. However, the mechanism of the observed greater susceptibility of female mice to UVA-induced DNA damage is unknown. Building upon our previous findings, we propose to investigate if UV light–induced oxidant-antioxidant imbalance leads to senescence and ECM deposition by causing G2/M cell cycle arrest; and if this imbalance causes translocation of CYP1B1 into mitochondria triggering greater estrogen-induced mtDNA damage in females. Gene array analysis revealed downregulation of DNA repair genes in FECD; therefore, we will determine whether this leads to decreased DNA damage repair during G2/M cell cycle arrest, triggering the cells to undergo either senescence or apoptosis. Our study is significant, as the investigation sex-dependent mechanisms involved in oxidative stress-induced cellular damage will provide new treatment targets for FECD. In order to achieve these aims, we will use our newly developed non-genetic mouse model of FECD based on physiologic outcome of CE susceptibility to UVA along with immortalized human CE cell lines, human aqueous fluid, and ex vivo specimens of genotyped FECD donors. Our Specific Aims are: Aim 1: Investigate the role of UVA irradiation in G2/M cell cycle arrest and induction of cellular senescence and ECM deposition in FECD. This aim is based on the hypothesis that DNA damage leads to G2/M phase arrest and induces cellular senescence, which in turn leads to aberrant ECM deposition in the form of guttae in FECD. Aim 2: Determine whether UVA irradiation activates CYP1B1 and induces estrogen metabolism causing preferentially greater DNA damage in females. This aim is based on the hypothesis that higher incidence and severity of FECD in women occurs due to UVA- induced translocation of CYP1B1 into mitochondria, which triggers formation of reactive estrogen metabolites, causing mtDNA damage. Aim 3: Determine the role of DNA damage response and DNA repair during UVA- induced cell cycle arrest in FECD. This aim is based on the hypothesis that DNA repair deficiency during G2/M cell cycle arrest determines whether cells undergo senescence or apoptotic cell death in FECD.

项目总结/摘要 Fuchs内皮角膜营养不良（FECD）是一种常见的年龄相关性营养不良，在 女性，病因不明。在FECD中，角膜内皮（CE）细胞损失伴随着异常的 细胞外基质（ECM）以滴状物的形式沉积。我们的实验室是第一个将氧化DNA 损伤和线粒体功能障碍。具体来说，我们发现DNA损伤， 由紫外线A（UVA）光诱导，导致小鼠中的FECD。此外，UVA诱导CE细胞周期阻滞， G2/M期与细胞衰老。我们确定了CYP 1B 1的新参与，CYP 1B 1是关键的雌激素， 在CE对UVA敏感性的性别依赖性差异中， 线粒体DNA（mtDNA）损伤。然而，观察到的机制更大 雌性小鼠对UVA诱导的DNA损伤的敏感性尚不清楚。基于我们之前的发现， 我们建议研究紫外线诱导的氧化-抗氧化失衡是否会导致衰老和ECM 沉积引起G2/M细胞周期阻滞;如果这种不平衡导致CYP 1B 1易位到 线粒体触发更大的雌激素诱导的女性线粒体DNA损伤。基因阵列分析显示 FECD中DNA修复基因的下调;因此，我们将确定这是否会导致FECD中DNA修复基因的下调。 在G2/M细胞周期停滞期间DNA损伤修复，触发细胞经历衰老或衰老。 凋亡我们的研究是有意义的，因为研究性别依赖的机制涉及氧化 应激诱导的细胞损伤将为FECD提供新的治疗靶点。为了实现这些目标， 我们将使用我们新开发的基于CE生理结果的FECD非遗传小鼠模型 对UVA的敏感性沿着永生化的人CE细胞系、人水性流体和离体 基因分型FECD供体的样本。我们的具体目标是：目标1：研究UVA照射在 FECD中G2/M细胞周期阻滞、诱导细胞衰老和ECM沉积。这一目标是基于 DNA损伤导致G2/M期阻滞并诱导细胞衰老， 导致FECD中以滴状物形式的异常ECM沉积。目的2：确定UVA照射是否 激活CYP 1B 1并诱导雌激素代谢，导致女性中更大的DNA损伤。 这一目标是基于这样的假设，即女性FECD的发生率和严重程度较高是由于UVA- 诱导CYP 1B 1易位进入线粒体，引发反应性雌激素代谢物的形成， 造成线粒体DNA损伤。目的3：确定UVA照射过程中DNA损伤反应和DNA修复的作用。 诱导FECD中的细胞周期停滞。这一目的是基于这样的假设，即G2/M期间的DNA修复缺陷 细胞周期停滞决定了FECD中细胞是否经历衰老或凋亡性细胞死亡。