RPE Energy Metabolism and Cell Phenotype

RPE 能量代谢和细胞表型

• 批准号: 10218680
• 负责人: Douglas E. Vollrath
• 金额: $ 5.2万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10218680
• 关键词: Address; Affect; Age; Age related macular degeneration; Aging; Atrophic; Cell Aging; Cells; Characteristics; Choroid; DNA Damage; DNA biosynthesis; Development; Disease; Diurnal Rhythm; Electron Transport; Energy Metabolism; Epithelial; Epithelium; Equilibrium; Eye; FRAP1 gene; Food Energy; Foundations; Generations; Genes; Genetic Transcription; Glucose; Glycolysis; Goals; HGF gene; Homeostasis; Human; Hypertrophy; Individual; Knowledge; Light; Link; Malignant Neoplasms; Mediating; Mesenchymal; Metabolic; Metabolism; Mitochondria; Mitochondrial DNA; Mitotic; Modeling; Morphology; Mus; Mutation; NADH; Nerve; Nonexudative age-related macular degeneration; Oxidative Phosphorylation; Oxygen; Pathway interactions; Phagocytes; Phagocytosis; Pharmacology; Phenotype; Photoreceptors; Production; Proteins; Respiration; Retina; Retinal Diseases; Risk Factors; Series; Severity of illness; Shapes; Sirolimus; Structure of retinal pigment epithelium; Time; Tissues; Variant; Vision; Work; Yeasts; aerobic glycolysis; alternative oxidase; cell growth; cost; experimental study; glucose metabolism; human model; in vivo; insight; macula; metabolomics; mitochondrial DNA mutation; mouse model; postnatal; response; retinal damage; stem cell differentiation; success; therapy development; transdifferentiation

The retinal pigment epithelium (RPE) is a highly differentiated, post-mitotic cell layer that performs a host of functions critical to retinal homeostasis. To discharge its many functions, the RPE requires ample energy. Studies of cultured RPE cells show that they can derive energy from glucose by either aerobic glycolysis or oxidative phosphorylation (OXPHOS), depending upon culture conditions. However, the balance between these two primary modes of RPE glucose metabolism in vivo is unknown, and it is unclear whether alterations in this balance occur under normal and/or disease conditions. Abundant evidence links changes in cellular energy metabolism with alterations in cell phenotype in a variety of fields including cancer, development, stem cell differentiation and aging. In the outer retina, mutations in mitochondrial DNA that compromise OXPHOS cause macular retinopathy. Moreover, disproportionate damage to mitochondrial DNA has been documented in the RPE of individuals with age-related macular degeneration (AMD), suggesting a causal link. We previously showed that postnatal loss of mitochondrial DNA and OXPHOS capacity in the murine RPE in vivo has surprising effects on cell phenotype, causing activation of cell growth pathways, increased glycolytic flux, and loss of epithelial functions and integrity. Our findings demonstrate that enforced changes in cellular energy metabolism in vivo can drive dedifferentiation and transdifferentiation of the RPE, and support a causal connection between diminished RPE OXPHOS capacity and AMD. However, our results raise new questions about how particular aspects of altered cellular energy metabolism read out as changes in cell phenotype. Can increased aerobic glycolysis alone, in the presence of intact OXPHOS, activate cell growth pathways, cause dedifferentiation/transdifferentiation and loss of epithelial integrity? Are features of the RPE glycolytic phenotype reversible through rebalancing of metabolism? What aspects of the altered phenotype of OXPHOS- deficient RPE result from lack of ATP production via OXPHOS versus loss of electron transport to oxygen? Does diurnal variation in energy metabolism affect the capacity of the RPE to phagocytize outer segment tips? We propose an ensemble of experiments to address these questions. Specifically we will modulate RPE aerobic glycolysis in vivo in the context of intact OXPHOS, restore respiration without ATP generation to OXPHOS-deficient RPE in vivo, and probe the relationship between RPE energy metabolism and diurnal phagocytic capacity. Through detailed characterization and quantification of the alterations in RPE cell phenotype caused by these in vivo metabolic changes, we will uncover mechanistic connections between energy metabolism and RPE cell phenotype. Because we will work in vivo, the impact of RPE metabolic modulation on photoreceptors will be apparent. Thus, success of this project will not only provide foundational knowledge of in vivo RPE metabolism and its relationship to cell phenotype, it will also inform studies of abnormal RPE metabolism in human retinal disease.

视网膜色素上皮（RPE）是一种高度分化的有丝分裂后细胞层，其执行许多功能， 对视网膜的自我平衡至关重要为了履行其许多功能，RPE需要充足的能量。 对培养的RPE细胞的研究表明，它们可以通过有氧糖酵解或 氧化磷酸化（OXPHOS），这取决于培养条件。然而，平衡 体内RPE葡萄糖代谢的这两种主要模式尚不清楚， 这种平衡发生在正常和/或疾病条件下。大量证据表明， 能量代谢与细胞表型的改变，包括癌症、发育、干细胞、 细胞分化和衰老。在视网膜外层，线粒体DNA突变损害OXPHOS 引起黄斑视网膜病变。此外，线粒体DNA的不成比例的损伤已被记录在 与年龄相关性黄斑变性（AMD）的个体的RPE，表明因果关系。我们之前 表明，出生后小鼠RPE中线粒体DNA和OXPHOS能力的丧失， 对细胞表型的令人惊讶的影响，引起细胞生长途径的激活，增加糖酵解通量，以及 上皮功能和完整性的丧失。我们的研究结果表明，细胞能量的强制变化 体内代谢可以驱动RPE的去分化和转分化，并支持一种因果关系。 减少RPE OXPHOS能力和AMD之间的联系。然而，我们的研究结果提出了新的问题， 关于细胞能量代谢改变的特定方面如何解读为细胞表型的变化。可以 在完整OXPHOS存在下，单独增加有氧糖酵解激活细胞生长途径， 去分化/转分化和上皮完整性丧失？是RPE糖酵解的特征 表型通过代谢再平衡可逆？OXPHOS表型改变的哪些方面- RPE缺陷是由于缺乏通过OXPHOS产生ATP而不是失去电子传递到氧？ 能量代谢的昼夜变化是否影响RPE吞噬外节尖端的能力？ 我们提出了一套实验来解决这些问题。具体来说，我们将调节RPE 在完整OXPHOS的情况下，体内有氧糖酵解，恢复呼吸而不产生ATP， OXPHOS缺陷的RPE细胞，探讨RPE能量代谢与昼夜节律的关系。 吞噬能力通过对RPE细胞中的改变进行详细的表征和定量， 表型引起的这些体内代谢的变化，我们将揭示机械之间的联系 能量代谢和RPE细胞表型。因为我们会在体内工作，影响RPE的代谢 对光感受器的调节将是明显的。因此，该项目的成功不仅将提供基础 了解体内RPE代谢及其与细胞表型的关系，还将为以下研究提供信息： 人视网膜疾病中的异常RPE代谢。