Purchase this article with an account.
Jinkyu Park; Stepwise and robust differentiation methods of functional retinal ganglion cells derived from human pluripotent stem cells. Invest. Ophthalmol. Vis. Sci. 2019;60(9):3807.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
We aimed to establish a robust method for retinal ganglion cell (RGC) differentiation from human pluripotent stem cells (hPSCs) with stepwise manner.
To define the contribution of specific signal pathways to RGC development and optimize the differentiation of hPSCs toward RGCs, we examined RGC differentiation in three stage: (1) eye field progenitors expressing the eye field transcription factors (EFTFs), (2) RGC progenitors expressing MATH5, and (3) RGCs expressing BRN3B and ISLET1. By monitoring the condition that elicited the highest yield of cells expressing stage-specific markers, we determined the optimal concentrations and combinations of signaling pathways required for efficient generation of RGCs from hPSCs.
Precise modulation of signaling pathways, including Wnt, insulin growth factor-1, and fibroblast growth factor, in combination with mechanical isolation of neural rosette cell clusters significantly enriched RX and PAX6 double-positive eye field progenitors from hPSCs by day 12. Furthermore, Notch signal inhibition facilitated differentiation into MATH5-positive progenitors at 90% efficiency by day 40. RGCs differentiated via this method were functional as exemplified by their ability to generate action potentials, express microfilament components on neuronal processes, and exhibit axonal transportation of mitochondria.
This protocol offers highly defined culture conditions for RGC differentiation from hPSCs and in vitro disease model and cell source for transplantation for disease related to RGCs.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
This PDF is available to Subscribers Only