Stein Eye Institute and Department of Ophthalmology, David Geffen School of Medicine at UCLA, 100 Stein Plaza, Los Angeles, CA, 90095, USA.
Department of Microbiology Immunology and Molecular Genetics, Los Angeles, CA, USA.
Stem Cell Res Ther. 2017 Oct 2;8(1):217. doi: 10.1186/s13287-017-0652-9.
Dysfunction of the retinal pigment epithelium (RPE) is implicated in numerous forms of retinal degeneration. The readily accessible environment of the eye makes it particularly suitable for the transplantation of RPE cells, which can now be derived from autologous induced pluripotent stem cells (iPSCs), to treat retinal degeneration. For RPE transplantation to become feasible in the clinic, patient-specific somatic cells should be reprogrammed to iPSCs without the introduction of reprogramming genes into the genome of the host cell, and then subsequently differentiated into RPE cells that are well characterized for safety and functionality prior to transplantation.
We have reprogrammed human dermal fibroblasts to iPSCs using nonintegrating RNA, and differentiated the iPSCs toward an RPE fate (iPSC-RPE), under Good Manufacturing Practice (GMP)-compatible conditions.
Using highly sensitive assays for cell polarity, structure, organelle trafficking, and function, we found that iPSC-RPE cells in culture exhibited key characteristics of native RPE. Importantly, we demonstrate for the first time with any stem cell-derived RPE cell that live cells are able to support dynamic organelle transport. This highly sensitive test is critical for RPE cells intended for transplantation, since defects in intracellular motility have been shown to promote RPE pathogenesis akin to that found in macular degeneration. To test their capabilities for in-vivo transplantation, we injected the iPSC-RPE cells into the subretinal space of a mouse model of retinal degeneration, and demonstrated that the transplanted cells are capable of rescuing lost RPE function.
This report documents the successful generation, under GMP-compatible conditions, of human iPSC-RPE cells that possess specific characteristics of healthy RPE. The report adds to a growing literature on the utility of human iPSC-RPE cells for cell culture investigations on pathogenicity and for therapeutic transplantation, by corroborating findings of others, and providing important new information on essential RPE cell biological properties.
视网膜色素上皮（RPE）功能障碍与多种形式的视网膜变性有关。眼睛这个易于接近的环境非常适合移植 RPE 细胞，而现在可以从自体诱导多能干细胞（iPSC）中获得这些细胞，以治疗视网膜变性。为了使 RPE 移植在临床上可行，应该对患者特有的体细胞进行重编程，使其成为 iPSC，而不会将重编程基因引入宿主细胞的基因组中，然后再将其分化为 RPE 细胞，这些细胞在移植前需要对安全性和功能进行充分的表征。
我们使用非整合 RNA 将人皮肤成纤维细胞重编程为 iPSC，并在符合良好生产规范（GMP）的条件下将 iPSC 向 RPE 命运分化（iPSC-RPE）。
使用细胞极性、结构、细胞器运输和功能的高度敏感检测方法，我们发现培养中的 iPSC-RPE 细胞表现出了天然 RPE 的关键特征。重要的是，我们首次用任何干细胞衍生的 RPE 细胞证明，活细胞能够支持动态细胞器运输。这种高度敏感的测试对于用于移植的 RPE 细胞至关重要，因为已经表明，细胞内运动缺陷会促进类似于黄斑变性的 RPE 发病机制。为了测试它们在体内移植中的能力，我们将 iPSC-RPE 细胞注射到视网膜变性的小鼠模型的视网膜下腔，并证明移植的细胞能够挽救丧失的 RPE 功能。
本报告记录了在符合 GMP 条件下成功生成具有健康 RPE 特定特征的人 iPSC-RPE 细胞。该报告通过证实其他研究的发现，并提供有关 RPE 细胞生物学特性的重要新信息，为人类 iPSC-RPE 细胞在致病性的细胞培养研究和治疗性移植方面的应用增加了越来越多的文献。
