中国病理生理杂志2021，Vol.37Issue(4)：593-610,18.DOI:10.3969/j.issn.1000-4718.2021.04.003
非集落样、分散的人诱导多能干细胞可分化为球型可移植的功能性视网膜色素上皮细胞
Spheroid transplantable and functional retinal pigment epithelium derived from non-colony dissociated human induced pluripotent stem cells
摘要
Abstract
AIM:To explore the feasibility of non-colony dissociated human induced pluripotent stem cells (hiPSCs)to differentiate into functional retinal pigment epithelial(RPE)cells(hiPSC-RPE cells),and offer an alterna-tive transplantation method based on cell spheroids. METHODS:The hiPSC-RPE cells were identified by RT-PCR,im-munofluorescence assay,and flow cytometry. The in vitro and in vivo functions of hiPSC-RPE were assessed by fluorescein leakage test,transepithelial electrical resistance assay,atomic force microscopic observation,photoreceptor outer segment (POS)phagocytosis assay,frozen tissue sections,live/dead cell staining assay,senescence-associated β-galactosidase (SA-β-Gal)staining,and immunocytochemistry. RESULTS:The hiPSC-RPE cells positively expressed biomarkers of RPE cells but not iPSCs,such as CRALBP(97. 4％),EMMPRIN(93. 8％),Oct4(2. 1％),and Sox2(2. 0％). These cells displayed RPE cell-like characteristics including barrier function,phagocytic activity,and polarized membrane. The cells derived from hiPSC-RPE cell spheroids positively expressed nestin and exhibited reduced SA-β-Gal staining(P<0. 01). The hiPSC-RPE cell spheroids were able to form monolayer on decellularized corneal matrices(DCM). After 1 month of subretinal transplantation,the hiPSC-RPE cell spheroids survived and maintained segmental sheet growth in Chinchilla rabbits with sodium iodate-induced RPE degeneration. CONCLUSION:Non-colony dissociated hiPSCs can effectively differentiate into functional RPE cells,and hiPSC-RPE spheroids maintain segmental sheet growth in the sub-retinal space of RPE-degenerative Chinchilla rabbits in vivo,which may offer cell spheroid transplantation as an alternative method for the treatment of RPE degenerative diseases in the future.关键词
人诱导多能干细胞/视网膜色素上皮细胞/细胞分化/细胞球/移植Key words
Human induced pluripotent stem cells/Retinal pigment epithelial cells/Cell differentiation/Cellular spheroids/Transplantation分类
医药卫生引用本文复制引用
郭晓令,朱德良,连瑞玲,曾巧浪,Sanjana MATHEW,唐仕波,陈建苏..非集落样、分散的人诱导多能干细胞可分化为球型可移植的功能性视网膜色素上皮细胞[J].中国病理生理杂志,2021,37(4):593-610,18.基金项目
This work was supported by National Natural Science Foundation of China(No.81701426 （）
No.81371689),Special Funds for Major Science and Technology Projects of Guangdong Province(No.2015B010125007),Medical and Health Re-search Science and Technology Plan Project of Zhejiang Province(No.2018KY523),and Public Welfare Science and Technology Plan Project of Wenzhou City(No.Y20180097). （No.2015B010125007）
