Using hiPSCs to develop physiologically-relevant outer retina tissue mimetics

使用 hiPSC 开发生理相关的外视网膜组织模拟物

基本信息

批准号：
10467753
负责人：
Danielle S. Benoit
金额：
$ 52.67万
依托单位：
UNIVERSITY OF ROCHESTER
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-30 至 2025-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10467753
关键词：
3-Dimensional Adhesives Adult Age related macular degeneration Age-Years American Architecture Biochemical Biology Biophysics Blindness Blood Vessels Blood capillaries Blood-Retinal Barrier Bruch&apos s basal membrane structure Caliber Cell Communication Cell Death Cell model Cells Choroidal Neovascularization Clinical Complex Cues Data Degenerative Disorder Deposition Development Diffusion Disease Disease model Drug Screening Drusen Elasticity Embryo Encapsulated Endothelial Cells Engineering Extracellular Matrix Eye Eye diseases Functional disorder Gel Growth Factor Health Human Hydrogels Impairment In Vitro Individual Lead Matrix Metalloproteinases Mediating Membrane Mesenchymal Stem Cells Mesenchyme Microfluidics Modeling Modulus Morphogenesis Morphology Neural Retina Nutrient Outcome Measure Pathologic Pathology Patients Pattern Peptides Perfusion Phagocytosis Phenocopy Photoreceptors Physiological Physiology Pigmentation physiologic function Property Proteomics Retina Retinal Degeneration Retinal Pigments Structure Structure of retinal pigment epithelium Testing Tissue Model Tissues Transplantation Vascular Endothelial Growth Factors Vascular blood supply Vertebrates Vision base cell type cellular engineering density ethylene glycol histological studies in vitro Model in vivo induced pluripotent stem cell microfluidic technology mimetics monolayer multidisciplinary retinal progenitor cell retinogenesis shear stress transcriptomics

项目摘要

The outer blood retina barrier (oBRB) comprises of the retinal pigment epithelium (RPE) cells and underlying fenestrated choriocapillaris (CC) that interfaces with the blood supply. The RPE-CC complex functions synergistically to support photoreceptor cell health that is critical for vision. Consistently, dysfunction of the RPE- CC leads to retinal degeneration in myraid eye diseases, including age-related macular degeneration (AMD), the single biggest cause of irreversible blindness in adults > 50 years of age in the US. However, the lack of in vitro tissue mimetics that faithfully recapitulate the RPE-CC complex has significantly impaired the study of normal and diseased physiology of the oBRB. A major challenge for the development of RPE-CC tissue mimetics is our limited understanding of human retinogenesis. This is especially relevant to the CC layer in which the majority of inferences are drawn from histological studies of embryonic human retina. Human induced pluripotent stem cells (hiPSCs) provide a unique platform to develop in vitro oBRB models. Indeed, several studies have now shown that specific cell types relevant to the RPE-CC complex, including RPE, endothelial cells (ECs) and mesenchymal stem cells (MSCs) can be differentiated from hiPSCs. Furthermore, we have recently developed a primitive RPE-CC tissue mimetic by exploiting the versatility of poly(ethylene glycol)(PEG) hydrogel-based engineered ECM (eECM) and hiPSC-derived target cells to emulate the spatial organization of RPE, ECs, and mesenchyme. The RPE-CC tissue mimetic is able to recapitulate important physiological features of the in vivo RPE-CC complex, such as CC-like fenestrated vasculature, that had previously been elusive in vitro. Although this model provides a framework for physiological RPE-CC development, it currently has several limitations, including unoptimized eECM biochemical and biophysical cues, lack of developmentally-instructed temporal cell- cell cues, and absence of vascular perfusion, resulting in a tissue model that does not fully recapitulate in vivo structure (e.g., well-defined Bruch’s membrane-like ECM and CC spatial angioarchitecture) and function (e.g., nutrient transport and macromolecular diffusion). In this proposal, we hypothesize that better understanding of the eECM requirements (Aim 1), ii) incorporation of temporal developmental cues (Aim 2), and integration of vascular perfusion, (Aim 3) will promote development of modular, spatially relevant, and functional RPE-CC tissue mimetic(s). Ultimately, the development of a physiological and modular human outer retina (RPE-CC) tissue mimetic will have important implications for subsequent disease modeling, drug screening, and transplantation studies.

外血视网膜屏障 (oBRB) 由视网膜色素上皮 (RPE) 细胞和底层细胞组成有孔脉络膜毛细血管 (CC) 与血液供应相连 RPE-CC 复合体发挥功能。协同支持对视力至关重要的感光细胞健康，RPE 功能障碍。 CC 会导致多种眼部疾病中的视网膜变性，包括年龄相关性黄斑变性 (AMD)、这是导致美国 50 岁以上成年人不可逆失明的唯一最大原因。忠实再现 RPE-CC 复合体的体外组织模拟物显着损害了以下研究： oBRB 的正常和患病生理学是 RPE-CC 组织模拟物开发的主要挑战。我们对人类视网膜发生的了解有限，这与 CC 层尤其相关。大多数推论来自人类胚胎视网膜的组织学研究。干细胞 (hiPSC) 提供了开发体外 oBRB 模型的独特平台。现在表明，与 RPE-CC 复合体相关的特定细胞类型，包括 RPE、内皮细胞 (EC) 和此外，我们最近还开发出间充质干细胞（MSC）可以从 hiPSC 中分化出来。通过利用基于聚乙二醇（PEG）水凝胶的多功能性，原始的 RPE-CC 组织模拟物工程化 ECM (eECM) 和 hiPSC 衍生靶细胞以模拟 RPE、EC 和细胞的空间组织 RPE-CC 组织模拟物能够重现体内的重要生理特征。 RPE-CC复合体，例如CC样有孔脉管系统，以前在体外难以捉摸。该模型为生理RPE-CC的发展提供了一个框架，目前它有一些局限性，包括未优化的 eECM 生化和生物物理线索、缺乏发育指导的颞细胞细胞线索和缺乏血管灌注，导致组织模型不能完全重现体内结构（例如，明确的 Bruch 膜状 ECM 和 CC 空间血管结构）和功能（例如，（营养物运输和大分子扩散）。在这项提案中，我们更好地理解了这一点。 eECM 要求（目标 1），ii）纳入时间发展线索（目标 2），以及整合血管灌注（目标 3）将促进模块化、空间相关和功能性 RPE-CC 的开发最终，开发出生理性和模块化的人类外视网膜（RPE-CC）。组织模拟将对后续的疾病建模、药物筛选和研究产生重要影响。移植研究。