Novel tools for screening retinal function using improved human retinal organoid models

使用改进的人类视网膜类器官模型筛查视网膜功能的新工具

• 批准号: 10288593
• 负责人: Jordan Michael Renna
• 金额: $ 31.98万
• 依托单位: UNIVERSITY OF AKRON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10288593
• 关键词: 3-Dimensional; 9-cis-retinal; Address; Affect; Biological Assay; Biological Models; Blindness; Calcium; Cell physiology; Cone; Development; Devices; Disease; Disease model; Drug Screening; Electrophysiology (science); Electroretinography; Evaluation; Gene Expression; Generations; Health; Histologic; Human; Human Development; Image; Individual; Light; Measures; Modeling; Organoids; Pharmaceutical Preparations; Pharmacological Treatment; Pharmacotherapy; Photoreceptors; Physiological; Physiology; Protocols documentation; Reproducibility; Research Personnel; Retina; Retinal Cone; Retinal Degeneration; Retinal Diseases; Screening procedure; Signal Transduction; Source; Speed; Supplementation; System; Techniques; Technology; Testing; Therapeutic; Therapeutic Uses; Time; Tissues; Tretinoin; Validation; Work; cost; drug development; experimental study; functional status; human stem cells; improved; interdisciplinary approach; interest; multi-electrode arrays; neurophysiology; new technology; non-invasive system; notch protein; novel; novel therapeutics; patch clamp; screening; sight restoration; small molecule; stem cells; therapy development; tool; transcriptomics; treatment strategy

Project Summary/Abstract Treating retinal degenerative diseases has been hampered by the lack of suitable systems that can evaluate how new treatment strategies affect the function of the human retina. Human stem cell-derived 3- dimensional retinal organoid technologies have been recently developed. Remarkably, human retinal organoids mimic the native tissue's histological organization, cellular composition, and are able to respond to light. These organoids are an ideal model system for investigating novel therapies to treat blinding diseases. However, to fully realize the unprecedented potential of human retinal organoids for the development of treatments for retinal degenerative diseases, we need new technologies that can rapidly measure retinal function under a wide variety of conditions. Current techniques such as patch-clamp electrophysiology, calcium imaging, and multi-electrode array recordings, measure how individual cells function within the circuitry of the retina. However, these techniques are laborious and ineffective at assessing the health, reproducibility, and functional responsivity of the retina as a whole - features that are key to the application of organoid systems to drug screening and validation. The lack of a device and techniques that allow for rapid, non-invasive screening of the functional status of retinal organoids constitutes a major unmet need. In Aim 1 of this proposal, we will develop an electroretinogram (ERG) recording chamber and a recording protocol for real-time assessment of light-evoked retinal organoid physiology by measuring photoreceptor and bipolar cell function. Our findings will be used to establish critical metrics associated with normal organoid light-evoked responsivity. Further, we will evaluate the power of this approach to detect changes in photoreceptor function, to provide evidence of its applicability to the assessment of disease models and therapeutic screening. To take full advantage of this technology for downstream applications it is critical that it be combined with robust organoid models. The variability and low yield of current protocols for retinal organoid generation and the extended time required for functional maturation of organoid photoreceptors hinder their application in drug development and disease evaluation. In Aim 2 of this proposal, we will address this critical gap by developing and evaluating improved protocols for human retinal organoid generation that increase yield and accelerate photoreceptor differentiation. We will then evaluate retinal function in these improved organoids using our ERG platform. Through the combination of these technologies, we will have created the first system for rapid, non- invasive functional screening in human retinal organoids that can be applied to the evaluation of normal, diseased, and drug-treated conditions. Our system has the potential to greatly accelerate the development of novel therapies to reverse vision loss.

项目摘要/摘要 视网膜退行性疾病的治疗一直受到阻碍，因为缺乏合适的系统来 评估新的治疗策略如何影响人类视网膜的功能。人类干细胞来源的3- 立体视网膜有机体技术是最近发展起来的。值得注意的是，人类视网膜有机体 模仿原生组织的组织结构、细胞组成，并能够对光做出反应。这些 有机化合物是研究治疗失明疾病的新疗法的理想模型系统。然而，为了 充分认识到人类视网膜类有机物在开发视网膜治疗方面的前所未有的潜力 退化性疾病，我们需要新的技术来快速测量各种情况下的视网膜功能 条件的问题。 目前的技术，如膜片钳电生理学、钙成像和多电极阵列 记录，测量单个细胞在视网膜回路中的功能。然而，这些技术 在评估视网膜的健康、重复性和功能响应性方面是费力和无效的 整体--有机系统应用于药物筛选和验证的关键特征。匮乏 一种设备和技术，允许快速、非侵入性地筛选视网膜器质的功能状态 构成了一个主要的未得到满足的需求。在这项提议的目标1中，我们将开发一种视网膜电信号(ERG)记录 用于实时评估光诱发视网膜器质生理学的记录方案 测量光感受器和双极细胞功能。我们的发现将被用来建立关键指标 与正常的有机类光诱发反应性有关。此外，我们还将评估这种方法的威力 检测光感受器功能的变化，为其在疾病评估中的适用性提供证据 模型和治疗性筛查。 为了充分利用这项技术在下游应用中的优势，将其与 坚固的有机模型。目前视网膜类器官生成方案的可变性和低产量以及 有机类光感受器功能成熟所需时间延长阻碍了其在药物中的应用 发展和疾病评估。在本提案的目标2中，我们将通过制定 以及评估提高产量和加速人类视网膜类有机物生成的改进方案 光感受器分化。然后，我们将使用我们的ERG来评估这些改进的有机化合物的视网膜功能 站台。 通过这些技术的结合，我们将创建第一个快速、非 人视网膜有机体的侵入性功能筛选，可用于评估正常， 疾病，和药物治疗的条件。我们的系统有潜力极大地加速 逆转视力丧失的新疗法。