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

• 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
• 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' 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

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）细胞和下层组成 与血液供应相结合的脉络膜毛细血管（CC）。 RPE-CC复合功能 协同以支持对视觉至关重要的感光细胞健康。始终如一，RPE的功能障碍 CC导致近视眼疾病的残留变性，包括与年龄相关的黄斑变性（AMD）， 在美国，成年人> 50岁以上的不可逆转失明的最大原因。但是，缺乏 忠实地概括RPE-CC复合物的体外组织模拟物严重损害了研究 OBRB的正常生理和脱离生理学。 RPE-CC组织模拟物的发展的主要挑战 是我们对人类视网膜生成的有限理解。这与CC层特别相关 大多数推论是从胚胎人视网膜的组织学研究中得出的。人类诱导的多能 干细胞（HIPSC）为开发体外OBRB模型提供了独特的平台。确实，一些研究有 现在表明，与RPE-CC复合物相关的特定细胞类型，包括RPE，内皮细胞（EC）和 间充质干细胞（MSC）可以与HIPSC区分开。此外，我们最近开发了 原始的RPE-CC组织模拟图，利用聚（乙二醇）（PEG）基于水凝胶的多功能性 设计的ECM（EECM）和HIPSC衍生的目标细胞模仿RPE，ECS和 间充质。 RPE-CC组织模拟物能够概括体内重要的物理特征 RPE-CC复合物，例如CC样fentrecty的脉管系统，以前在体外难以捉摸。虽然 该模型为物理RPE-CC开发提供了一个框架，目前有几个限制， 包括未优化的EECM生化和生物物理提示，缺乏开发的临时细胞 细胞提示和缺乏血管灌注，导致组织模型未完全概括体内 结构（例如，定义明确的Bruch膜状ECM和CC空间血管结构）和功能（例如， 营养转运和大分子扩散）。在此提案中，我们假设对 EECM要求（AIM 1），ii）纳入临时发展线索（AIM 2）和整合 血管灌注（AIM 3）将促进模块化，空间相关和功能性RPE-CC的发展 组织模仿。最终，生理和模块化人类外视网膜（RPE-CC）的发展 组织模拟物将对随后的疾病建模，药物筛查和 移植研究。