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Kai Liao, Shanyi Li, Shibo Tang, Jiansu Chen; Biomimetic culture promotes the differentiation of iPS cells into corneal endothelial-like cells. Invest. Ophthalmol. Vis. Sci. 2019;60(9):2167.
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© ARVO (1962-2015); The Authors (2016-present)
Corneal endothelial cells (CECs) are critical to maintain the transparency of cornea. However, the donors of CECs for transplantation are limited. Therefore, we developed a biomimetic culture method to promote the differentiation of induced pluripotent stem (iPS) cells into CEC-Like cells for the treatment of corneal endothelial diseases.
In this study, iPS cells were induced to differentiate into CECs using both microenvironment and embryonic development as a guide. The procedure of iPS cells differentiation to CECs includes two-stage differentiation method. (1) iPS cells are induced to differentiate into neural crest cells by using previously neural crest differentiation medium including TGFβ inhibitor and GSK-3β inhibitor. (2) In order to promote neural crest cells differentiation into CEC cells, we use a biomimetic environment by bovine CEC extracellular matrix (B-ECM) and bovine aqueous humor as well as other factors including GSK-3β inhibitor, retinoic acid (RA), bFGF and TGF-β2.
TGFβ inhibitor and GSK-3β inhibitor are key factors for iPS cells induction into neural crest cells, which display positive expressions of P75 and Nestin by immunofluorescence staining, positive expressions of SOX9 and SOX10 by qPCR. (Figure 1). Both bovine aqueous humor and B-ECM are important factors for further differentiation neural crest cells to CEC-like cells. After the second stage of differentiation, most of the cells show CEC-like cells with polygonal shape. In addition, immunofluorescence staining exhibit CEC-like cells express N-Cadherin, ZO-1 and Na+-K+-ATPase markers, and weakly express AQP-1 and Nestin (Figure 2).
This study demonstrates that iPS cells are able to differentiate into CEC-like cells by using a two-step method. Our findings suggest that iPS cells may be a promising resource for the treatment of corneal endothelial diseases in the future.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
Figure 1 iPS cells were differentiated into neural crest cells.
Figure 2 Neural crest cells were differentiated into corneal endothelial-like cells.
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