RPE Energy Metabolism and Cell Phenotype

RPE 能量代谢和细胞表型

• 批准号: 10260148
• 负责人: Douglas E. Vollrath
• 金额: $ 16.73万
• 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/10260148
• 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) 是高度分化的有丝分裂后细胞层，执行许多功能 对视网膜稳态至关重要的功能。为了发挥其多种功能，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糖酵解从葡萄糖中获取能量。 氧化磷酸化 (OXPHOS)，取决于培养条件。然而，之间的平衡 RPE 体内葡萄糖代谢的这两种主要模式尚不清楚，也不清楚是否会发生改变 这种平衡发生在正常和/或疾病条件下。大量证据表明细胞的变化有关 能量代谢与癌症、发育、干细胞等多个领域的细胞表型改变 细胞分化和衰老。在外层视网膜中，线粒体 DNA 突变会损害 OXPHOS 引起黄斑视网膜病变。此外，线粒体 DNA 受到不成比例的损伤已被记录在案。 年龄相关性黄斑变性 (AMD) 患者的 RPE，表明存在因果关系。我们之前 研究表明，出生后小鼠 RPE 体内线粒体 DNA 和 OXPHOS 能力的丧失 对细胞表型产生令人惊讶的影响，导致细胞生长途径的激活，糖酵解通量的增加，以及 上皮功能和完整性丧失。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，并支持因果关系 RPE OXPHOS 容量减少与 AMD 之间的联系。然而，我们的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。能 在完整的 OXPHOS 存在下，单独增加有氧糖酵解，激活细胞生长途径，导致 去分化/转分化和上皮完整性丧失？ RPE 糖酵解的特点 通过新陈代谢的重新平衡可以逆转表型吗？ OXPHOS- 改变表型的哪些方面 RPE 缺陷是由于 OXPHOS 产生的 ATP 不足以及电子向氧气传输的损失造成的？ 能量代谢的昼夜变化是否会影响 RPE 吞噬外节尖端的能力？ 我们提出了一系列实验来解决这些问题。具体来说，我们将调节 RPE 在完整的 OXPHOS 背景下进行体内有氧糖酵解，在不产生 ATP 的情况下恢复呼吸 体内OXPHOS缺陷的RPE，并探讨RPE能量代谢与昼夜节律的关系 吞噬能力。通过对 RPE 细胞变化的详细表征和量化 通过这些体内代谢变化引起的表型，我们将揭示之间的机制联系 能量代谢和 RPE 细胞表型。因为我们将在体内工作，RPE代谢的影响 对光感受器的调节将是显而易见的。因此，该项目的成功不仅为 了解体内 RPE 代谢及其与细胞表型的关系，它还将为以下研究提供信息 人类视网膜疾病中 RPE 代谢异常。