A Protocol for Stepwise Differentiation of Induced Pluripotent Stem Cells into Retinal Pigment Epithelium

2021 ◽  
Author(s):  
Miho Tagawa ◽  
Hanako Ohashi Ikeda ◽  
Masayuki Hata ◽  
Yumi Inoue ◽  
Sachiko Iwai ◽  
...  
Keyword(s):  
Stem Cells ◽  
Retinal Pigment Epithelium ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Induced Pluripotent

scholarly journals Spheroid transplantable and functional retinal pigment epithelium derived from non-colony dissociated human induced pluripotent stem cells

2020 ◽  
Author(s):  
Xiaoling Guo ◽  
Deliang Zhu ◽  
Ruiling Lian ◽  
Qiaolang Zeng ◽  
Sanjana Mathew ◽  
...  
Keyword(s):  
Stem Cells ◽  
Retinal Pigment Epithelium ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Great Promise ◽  
Rpe Cells ◽  
Cell Spheroids ◽  
Induced Pluripotent

Abstract Background: Retinal pigment epithelium (RPE) cells derived from human induced pluripotent stem cells (hiPSCs) exhibit great promise in treating retinal degenerative diseases. Here, we would explore the feasibility of non-colony dissociated hiPSCs to differentiate into functional RPE cells (hiPSC-RPE), and offer an alternative transplantation method based on cell spheroids.Methods: hiPSC-RPE cells were identified using reverse transcription-polymerase chain reaction (RT-PCR), immunofluorescence assay, Western blotting, and flow cytometry assay. The functions of hiPSC-RPE cells in vitro and in vivo were assessed by fluorescein leakage test, transepithelial electrical resistance (TEER) assay, atomic force microscopy observation, POS phagocytosis assay, frozen tissue sections, live/dead assay, SA-β-Gal staining, and immunocytochemistry.Results: 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%). hiPSC-RPE cells displayed RPE-like characteristics including barrier function, phagocytic activity, and polarized membrane. The cells derived from hiPSC-RPE spheroids positively expressed Nestin and exhibited reduced SA-β-Gal staining. hiPSC-RPE cell spheroids could form monolayer on decellularized corneal matrixes (DCM). After one month of subretinal transplantation, hiPSC-RPE cell spheroids could survive and maintain segmental sheet growth in sodium iodate (NaIO3) induced RPE-degenerated chinchilla rabbits. Conclusion: This study suggested that non-colony dissociated hiPSCs were effectively differentiated into functional RPE cells, and hiPSC-RPE cell spheroids maintained segmental sheet growth in the subretinal of RPE degenerate chinchilla rabbits in vivo, which may lay the foundation for cell spheroid transplantation as an alternative method for RPE degenerative disease therapy in the future.

scholarly journals Differentiation/Purification Protocol for Retinal Pigment Epithelium from Mouse Induced Pluripotent Stem Cells as a Research Tool

PLoS ONE ◽  
2016 ◽  
Vol 11 (7) ◽  
pp. e0158282 ◽  
Author(s):  
Yuko Iwasaki ◽  
Sunao Sugita ◽  
Michiko Mandai ◽  
Shigenobu Yonemura ◽  
Akishi Onishi ◽  
...  
Keyword(s):  
Stem Cells ◽  
Retinal Pigment Epithelium ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Research Tool ◽  
Induced Pluripotent ◽  
Purification Protocol
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