Investigating the role of mitochondrial dynamics in retinal pigment epithelium

研究线粒体动力学在视网膜色素上皮细胞中的作用

• 批准号: 10468097
• 负责人: Nan Wu Hultgren
• 金额: $ 7.23万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2023-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10468097
• 关键词: Affect; Age related macular degeneration; Area; Biogenesis; Biological; Biological Assay; Biology; Blindness; Blood Vessels; Cell Communication; Cell Line; Cell Respiration; Cells; Cellular Metabolic Process; Cellular biology; Cessation of life; Choroideremia; Complement; Consumption; Data; Data Analyses; Defect; Disease; Electron Microscopy; Energy-Generating Resources; Ensure; Etiology; Event; Experimental Designs; Eye diseases; Functional disorder; Future; Genome Stability; Genus Hippocampus; Glycolysis; Goals; Health; Homeostasis; Human; Image; Impairment; In Vitro; Ingestion; Inherited; Knowledge; Link; Measures; Membrane Lipids; Metabolic; Metabolism; Methods; Microscopy; Microtubules; Mission; Mitochondria; Modeling; Molecular; Morphology; Motor; Mus; NADH; Neurons; Organelles; Oxidative Phosphorylation; Pathogenesis; Pathway interactions; Patients; Phagocytosis; Phagosomes; Photoreceptors; Positioning Attribute; Potential Energy; Primary Lesion; Process; Production; Proteins; Public Health; Research Personnel; Resolution; Retina; Retinal Degeneration; Retinal Diseases; Role; Scientist; Signal Pathway; Site; Solid; Speed; Structure of retinal pigment epithelium; Synapses; System; Techniques; Testing; Time; Training; Vision; Vision research; Work; career; career development; cell motility; disease mechanisms study; experience; experimental study; in vivo; induced pluripotent stem cell; inherited retinal degeneration; insight; live cell imaging; loss of function; mitochondrial genome; neuron loss; new therapeutic target; novel; prenylation; prevent; response; skills; temporal measurement; trafficking; training opportunity; vision science

Project Summary The retinal pigment epithelium (RPE) is often the initial site of pathogenesis in many retinal degenerative diseases. The proposed study aims to understand how mitochondrial dynamics responds to and affects RPE function and health. The successful completion of the project will provide additional insights into disease mechanisms. The RPE performs many important functions to ensure photoreceptor homeostasis, including daily phagocytosis of photoreceptor outer segment. Phagocytosis is a metabolically challenging process. The formation, transport and degradation of large numbers of phagosomes are energy intensive for the RPE, as each RPE cell interacts with up to 200 photoreceptor cells. On the other hand, the products of phagosome degradation could represent an energy source. We therefore speculate that, RPE mitochondria have to respond and adapt to different daily metabolic events. In many degeneration diseases, impaired mitochondrial functions have been observed, and this is often associated with defects in mitochondrial dynamics. However, little is known about mitochondrial dynamics in RPE. In this study, we propose to use super-resolution high-speed live imaging and electron microscopy to study the dynamics of RPE mitochondria, including morphology (fission/fusion), distribution and motility. To complement these studies, we will also conduct metabolic analysis such as Seahorse assay and Fluorescent Lifetime Microcopy to measure mitochondrial functions. We will focus on answering three questions: 1) How does RPE phagocytosis affect mitochondrial dynamics? 2) What regulatory machineries drive mitochondrial dynamics? 3) How are RPE mitochondria dynamics affected in the inherited retinal degeneration, choroideremia? To ensure relevance to human health and disease, we will compare and contrast our observations in mice with those in human RPE cell lines, as well as patient-derived iPSC-RPE cultures. By imaging with very high spatial and temporal resolutions, the study will provide novel insight into RPE mitochondrial dynamics, which in turn will allow us to uncover new disease mechanism and identify novel therapeutic targets. The scope and potential public health impact of the proposed study fits well with the missions of NEI to help prevent and treat eye diseases. The proposed project also provides invaluable training opportunity for the applicant towards her long- term career goal to become an independent investigator in molecular and cellular vision research. The applicant will gain knowledge of the vision system, specifically the interaction between photoreceptors and RPE. The applicant will also receive training on various cutting-edge microscopy techniques, as well as patient-derived iPSC as a model to study disease mechanism in vitro. Together with her prior experience in vascular biology, the training proposed here will set the applicant apart in a uniquely qualified position to study molecular mechanisms and cell-cell interactions involved in health and diseases of the vision system.

项目摘要 视网膜色素上皮（RPE）通常是许多视网膜中的发病机理的初始部位 退化性疾病。拟议的研究旨在了解线粒体动力如何响应和 影响RPE功能和健康。该项目的成功完成将提供更多的见解 疾病机制。 RPE执行许多重要功能以确保感光体稳态， 包括光感受器外部段的每日吞噬作用。吞噬作用是一种代谢挑战 过程。大量吞噬体的形成，运输和降解是能量密集型的 当每个RPE细胞与多达200个感光细胞相互作用时，RPE。另一方面， 吞噬体降解可以代表能源。因此，我们推测，RPE线粒体 必须做出反应并适应不同的每日代谢事件。在许多变性疾病中，受损 已经观察到线粒体功能，这通常与线粒体中的缺陷有关 动力学。但是，对RPE中的线粒体动力学知之甚少。在这项研究中，我们建议使用 超分辨率的高速实时成像和电子显微镜研究RPE线粒体的动力学， 包括形态（裂变/融合），分布和运动。为了补充这些研究，我们也将 进行代谢分析，例如海马测定法和荧光寿命微拷贝以测量 线粒体功能。我们将专注于回答三个问题：1）RPE吞噬作用如何影响 线粒体动力学？ 2）哪些监管机械驱动线粒体动力学？ 3）RPE如何 线粒体动力学受到遗传性视网膜变性，脉络膜血症的影响？确保与 人类健康和疾病，我们将在小鼠中比较和对比我们的观察结果与人类RPE中的观察结果 细胞系以及患者来源的IPSC-RPE培养物。通过具有很高的空间和时间成像 决议，该研究将提供对RPE线粒体动力学的新见解，这反过来又使我们能够 发现新的疾病机制并确定新的治疗靶标。范围和潜在的公共卫生 拟议的研究的影响非常符合NEI的任务，以帮助预防和治疗眼病。 拟议的项目还为申请人提供了宝贵的培训机会 任期职业目标是成为分子和细胞视觉研究领域的独立研究者。这 申请人将了解视力系统，特别是感光体之间的相互作用 RPE。申请人还将接受各种尖端显微镜技术的培训，以及 患者衍生的IPSC作为体外研究疾病机制的模型。以及她先前的经历 血管生物学，这里提出的培训将使申请人处于独特的资格研究中 与视力系统的健康和疾病有关的分子机制和细胞 - 细胞相互作用。