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Kana Orihara, Chiho Yabuta, Mitsuyoshi Azuma, Noriyuki Azuma; Differentiation of human iPSC-derived optic vesicles into numerous types of retinal cells. Invest. Ophthalmol. Vis. Sci. 2018;59(9):572. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
The integration of neuronal cells in mature human retina produces retinal function. These cells differentiate from retinal stem cells. Retinal ganglion cells (RGCs) differentiate early. Horizontal, amacrine, and photoreceptor cells follow. Finally, differentiation of bipolar cells yields an intact retina. Tanaka et al1 successfully produced functional RGCs differentiated from human induced pluripotent stem cell (iPSC)-derived optic vesicles (OV). These RGCs are useful for studying the physiology and pathophysiology of retina. Human iPSC-derived OV probably also differentiate into other cell types; however, this has not been established. Therefore, the purpose of the present experiments was to determine the types of neuronal cells produced by differentiation of human iPSC-derived OV.
Differentiation from human iPSC was induced following a previously reported method.1 Dissociated iPSCs were cultured in V-bottomed, 96-well plates on Day0, and aggregated embryoid bodies (EBs) were then cultured on adhesive substrates starting on D27. OV extruded from EBs were mechanically divided on D28, 29, 30, 31, and 32. Stains for retinal cell markers were: neurofilament-H (NF-H) and Brn3 for RGCs, calbindin for horizontal cells, syntaxin1 for amacrine cells, recoverin for photoreceptor cells, and PKCα for bipolar cells. To confirm biological differentiation, OV at D31 were cultured for 24 hrs with ciliary neurotrophic factor (CNTF), a known promoter of neurite elongation. RGCs with elongated neurites were counted after immuno labeling with antibodies for NF-H and Brn3.
On days 1 and 2 after culturing EBs onto adhesive plates, RGCs, horizontal cells, and amacrine cells appeared in OV. Photoreceptor and bipolar cells appeared at D30. CNTF significantly increased the number of RGCs with elongated neurites.
Numerous differentiated retinal cells were produced by induction of human iPSC-derived OV. Maturation of the RGCs into elongated neurites was promoted by CTNF. These findings suggest that cultured iPSC-derived OV would be a useful system for studying organogenesis. In addition, this system may facilitate clinical studies by evaluating initial drug efficacy and safety in vitro.1 SCIENTIFIC REPORTS. 2015; 5: 8344 : DOI: 10.1038/srep08344
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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