Key Laboratory of Optoelectronic Information and Sensing Technologies of Guangdong Higher Educational Institutes, Jinan University, Guangzhou, China.
Key Laboratory for Regenerative Medicine, Ministry of Education, Jinan University, Guangzhou, China.
Stem Cell Res Ther. 2020 Mar 4;11(1):98. doi: 10.1186/s13287-020-01608-8.
Retinitis pigmentosa (RP) is an inherited retinal disease characterized by progressive loss of photoreceptor cells. This study aim at exploring the effect of retinal pigment epithelium (RPE) derived from human-induced pluripotent stem cell (hiPSC-RPE) on the retina of retinal degeneration 10 (rd10) mice, which are characterized with progressive photoreceptor death.
We generated RPE from hiPSCs by sequential supplementation with retinal-inducing factors and RPE specification signaling factors. The three-dimensional (3D) spheroid culture method was used to obtain optimal injectable hiPSC-RPE cells. Subretinal space transplantation was conducted to deliver hiPSC-RPE cells into the retina of rd10 mice. Neurotrophic factor secretion from transplanted hiPSC-RPE cells was detected by enzyme-linked immunosorbent assay (ELISA). Immunostaining, Western blotting, electroretinography (ERG), and visual behavior testing were performed to determine the effects of hiPSC-RPE on the retinal visual function in rd10 mice.
Our data demonstrated that hiPSC-RPE cells exhibited classic RPE properties and phenotype after the sequential RPE induction from hiPSCs. hiPSC-RPE cells co-cultured with mouse retinal explants or retinal ganglion cells 5 (RGC5) exhibited decreased apoptosis. The viability and functional properties of hiPSC-RPE cells were enhanced by 3D spheroid culture. Transplanted hiPSC-derived RPE cells were identified by immunostaining with human nuclear antigen staining in the retina of rd10 14 days after subretinal space injection. The pigment epithelium-derived factor level was increased significantly. The expression of CD68, microglial activation marker, reduced after transplantation. The light avoidance behavior and ERG visual function in rd10 mice improved by the transplantation of hiPSC-RPE cells.
Our findings suggest that injectable hiPSC-RPE cells after 3D spheroid culture can rescue the structure and function of photoreceptors by sub-retinal transplantation, which lay the foundation for future clinical cell therapy to treat RP and other retinal degeneration diseases.
色素性视网膜炎（RP）是一种遗传性视网膜疾病，其特征是感光细胞逐渐丧失。本研究旨在探讨人诱导多能干细胞（hiPSC）衍生的视网膜色素上皮（RPE）对视网膜变性 10 型（rd10）小鼠视网膜的影响，rd10 小鼠表现出感光细胞进行性死亡。
我们通过顺序补充视网膜诱导因子和 RPE 特化信号因子，从 hiPSC 中生成 RPE。采用三维（3D）球体培养法获得最佳的可注射 hiPSC-RPE 细胞。通过视网膜下腔移植将 hiPSC-RPE 细胞递送至 rd10 小鼠的视网膜中。通过酶联免疫吸附试验（ELISA）检测移植的 hiPSC-RPE 细胞分泌的神经营养因子。通过免疫染色、Western blot、视网膜电图（ERG）和视觉行为测试来确定 hiPSC-RPE 对 rd10 小鼠视网膜视觉功能的影响。
我们的数据表明，hiPSC-RPE 细胞在 hiPSC 连续诱导为 RPE 后表现出典型的 RPE 特性和表型。与小鼠视网膜外植体或视网膜神经节细胞 5（RGC5）共培养的 hiPSC-RPE 细胞凋亡减少。3D 球体培养增强了 hiPSC-RPE 细胞的活力和功能特性。视网膜下腔注射后 14 天，免疫染色显示移植的 hiPSC 衍生的 RPE 细胞在 rd10 鼠的视网膜中被鉴定为人核抗原染色阳性。色素上皮衍生因子水平显著增加。移植后，微胶质激活标志物 CD68 的表达减少。hiPSC-RPE 细胞移植可改善 rd10 小鼠的避光行为和 ERG 视觉功能。
我们的研究结果表明，经过 3D 球体培养的可注射 hiPSC-RPE 细胞可以通过视网膜下移植来挽救光感受器的结构和功能，为未来治疗 RP 和其他视网膜变性疾病的临床细胞治疗奠定了基础。
