Homocysteine's role in Age-Related Macular Degeneration

同型半胱氨酸在年龄相关性黄斑变性中的作用

• 批准号: 10329922
• 负责人: Amany M Tawfik
• 金额: $ 36.05万
• 依托单位: OAKLAND UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10329922
• 关键词: Age related macular degeneration; Aging; Amino Acids; Biological Assay; Blindness; Blood; Cells; Choroidal Neovascularization; Cystathionine; Data; Development; Dietary Supplementation; Elderly; Endothelial Cells; Endothelium; Enzymes; Evaluation; Exhibits; Experimental Models; Fluorescein Angiography; Folic Acid; Functional disorder; Genetic; Glycolysis; Goals; Homocysteine; Human; Hyperhomocysteinemia; In Vitro; Intestinal Absorption; Joints; Knockout Mice; Lasers; Life Expectancy; Mediating; Medical; Metabolic; Metabolism; Mitochondria; Modality; Modeling; Modification; Molecular; Mus; N-Methyl-D-Aspartate Receptors; N-Methylaspartate; Optical Coherence Tomography; Oxidative Phosphorylation; Pathogenesis; Pathway interactions; Patients; Pharmacology; Phenotype; Photoreceptors; Population; Prevalence; Proteins; Reporting; Respiration; Retina; Risk; Role; SLC2A1 gene; Serum; Signal Pathway; Signal Transduction; Structure of retinal pigment epithelium; Testing; Therapeutic; Tight Junctions; Up-Regulation; Vascular Endothelial Growth Factors; Vision Disorders; Vitamin B 12; Vitamin B6; Vitamins; Wild Type Mouse; bevacizumab; conditional knockout; electric impedance; experimental study; fluorescein isothiocyanate dextran; folic acid supplementation; gene therapy; genetic manipulation; in vivo; inhibitor; intravitreal injection; mouse model; overexpression; preservation; prevent; receptor; retinal damage; therapeutic target

Summary/Abstract Age-related macular degeneration (AMD) is the leading cause of vision loss in among elderly populations. Elevated homocysteine (Hcy), also known as hyperhomocysteinemia (HHcy) has been reported in patients with AMD; thereby suggesting an association between HHcy and the risk of AMD. Recently, we reported retinal changes similar to AMD in a mouse model of HHcy which lacks Cystathionine-β-synthase (cbs+/-) or received intravitreal injections of Hcy. These models showed significant retinal pigment epithelium (RPE) dysfunction and choroidal neovascularization (CNV) However, the lack of understanding the molecular/cellular mechanisms of these changes is a critical barrier in proposing Hcy as a therapeutic target in AMD. Our preliminary data show that HHcy-induced RPE dysfunction is associated with the upregulation of the N-methyl-D-aspartate (NMDAr) and GLUT1 receptors and increased glycolysis. Hence, we hypothesize that HHcy contributes to the pathogenesis of AMD via activation of the NMDAr and GLUT1 signaling pathways that induce the metabolic switch from oxidative phosphorylation to glycolysis. Therefore, elimination of excess Hcy through pharmacological or genetic intervention could be beneficial in the treatment of AMD. To test our hypothesis, we will conduct in vitro experiments, using RPE and choroidal endothelial cells (CEC) and in vivo using cbs+/-, wild type mice receiving intravitreal injection of Hcy and mice lacking the endothelial or RPE NMDAr (NMDAr-/-E or NMDAr-/-R respectively). Our specific aims include: 1: Testing the hypothesis that HHcy induces the metabolic switch from mitochondrial respiration to glycolysis via activation of GLUT1 in RPE cells: We will examine the changes in the retinal expression and localization of GLUT1, mitochondrial respiration, glycolysis and rate-limiting glycolytic enzymes in HHcy models. Moreover, we will determine the effect of GLUT1 inhibition on HHcy-induced RPE dysfunction and CNV. Aim 2: Testing the hypothesis that inhibition of NMDAr preserves RPE function and reduces the development of CNV under HHcy. We will examine the effects of pharmacological inhibition or genetic manipulation of the NMDAr on HHcy-induced RPE dysfunction and CNV. The effect of intravitreal injection of Hcy will be evaluated in NMDAr-/-E or NMDAr-/-R as compared to wild type and cbs+/- mice with or without NAMDAr inhibitors. Parallel in vitro experiments will be performed on RPE and CEC subjected to Hcy with or without NMDAr inhibitors followed by assessment of RPE function and angiogenic potential of CEC. Aim 3: Testing the hypothesis that elimination of excess Hcy by dietary supplementation or genetic/ pharmacological modifications prevents the progression of AMD. Hcy clearance will be enhanced in models of HHcy through two approaches, followed by assessment of RPE function and angiogenic potential of CEC: (a) Enhancing the remethylation pathway of Hcy metabolism using vitamins B6, B12 and folic acid supplementation. (b) Enhancing the transsulforation pathway of Hcy metabolism via CBS overexpression. Successful clearance of excess Hcy holds immense promise in the treatment of AMD.

摘要/摘要 年龄相关性黄斑变性（AMD）是老年人群视力丧失的主要原因。 据报道，以下患者的同型半胱氨酸 (Hcy) 升高，也称为高同型半胱氨酸血症 (HHcy) AMD；从而表明 HHcy 与 AMD 风险之间存在关联。最近，我们报道了视网膜 与缺乏胱硫醚-β-合酶 (cbs+/-) 或接受了 HHcy 小鼠模型中的 AMD 类似的变化 玻璃体内注射Hcy。这些模型显示出显着的视网膜色素上皮（RPE）功能障碍和 脉络膜新生血管（CNV）然而，缺乏对其分子/细胞机制的了解 这些变化是提议 Hcy 作为 AMD 治疗靶点的关键障碍。我们的初步数据显示 HHcy 诱导的 RPE 功能障碍与 N-甲基-D-天冬氨酸 (NMDAr) 的上调有关 和 GLUT1 受体并增加糖酵解。因此，我们假设 HHcy 有助于 AMD 的发病机制是通过激活 NMDAr 和 GLUT1 信号通路来诱导代谢 从氧化磷酸化转变为糖酵解。因此，通过消除多余的 Hcy 药物或基因干预可能有益于 AMD 的治疗。为了检验我们的假设，我们 将使用 RPE 和脉络膜内皮细胞 (CEC) 进行体外实验，并使用 cbs+/-、wild 进行体内实验 接受玻璃体内注射 Hcy 的小鼠和缺乏内皮或 RPE NMDAr 的小鼠（NMDAr-/-E 或 分别为 NMDAr-/-R）。我们的具体目标包括： 1：检验 HHcy 诱导的假设 通过激活 RPE 细胞中的 GLUT1，实现从线粒体呼吸到糖酵解的代谢转变：我们 将检查 GLUT1 的视网膜表达和定位、线粒体呼吸的变化， HHcy 模型中的糖酵解和限速糖酵解酶。此外，我们将确定 GLUT1 的效果 抑制 HHcy 诱导的 RPE 功能障碍和 CNV。目标 2：检验抑制作用的假设 NMDAr 保留 RPE 功能并减少 HHcy 下 CNV 的发展。我们将检查 NMDAr 的药理抑制或基因操作对 HHcy 诱导的 RPE 功能障碍的影响 和CNV。玻璃体内注射 Hcy 的效果将在 NMDAr-/-E 或 NMDAr-/-R 中进行评估，并与 使用或不使用 NAMDAr 抑制剂的野生型和 cbs+/- 小鼠。平行体外实验将在 RPE 和 CEC 在有或没有 NMDAr 抑制剂的情况下接受 Hcy 处理，然后评估 RPE 功能和 CEC 的血管生成潜力。目标 3：检验通过饮食消除过量 Hcy 的假设 补充剂或基因/药理学修饰可预防 AMD 的进展。同型半胱氨酸 将通过两种方法增强 HHcy 模型中的清除率，然后评估 RPE 功能 CEC 的血管生成潜力：(a) 使用维生素增强 Hcy 代谢的再甲基化途径 补充B6、B12和叶酸。 (b) 通过 CBS 增强 Hcy 代谢的转磺途径 过度表达。成功清除过量的 Hcy 为治疗 AMD 带来了巨大的希望。