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Y. Kumagai, M. S. Kurokawa, H. Ueno, M. Kayama, K. Tsubota, N. Nakatsuji, S. Nito, S. Ueno, N. Suzuki; Induction of Corneal Epithelium-Like Cells From Cynomorgus Monkey Embrionic Stem Cells and Their Experimental Transplantation to Damaged Cornea. Invest. Ophthalmol. Vis. Sci. 2007;48(13):477.
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We have reported the successful transplantation of corneal epithelium-like cells derived from mouse embryonic stem (ES) cells onto damaged mouse cornea. The aim of this study was to induce differentiation into corneal epithelium-like cells of cynomolgus monkey (CM) ES cells and we have tested whether the CMES derived corneal epithelium-like cells were applicable for the experimental transplantation to damaged cornea of mice.
CMES cells were cultivated on typeIV collagen-coated dishes for 4 to 8 days to induce differentiation into corneal epithelium-like cells. The differentiation was evaluated by RT-PCR and immunofluorescence staining. Mouse corneas were scraped and corneal epithelial cells were peeled off to make the damaged cornea. The corneal epithelium-like cells were transplanted to the injured cornea, and the recipient mice were sacrificed at the various time points. Reconstitution of the corneal epithelium was evaluated by immunofluorescence staining and subsequent confocal laser-microscopy.
The cells cultured on typeIV collagen showed cobble stone like appearance. They expressed mRNA of pax6, e-cadherin and keratin3 in vitro. Confocal analysis confirmed the protein expressions of Pax6, E-cadherin and Keratin3, suggesting differentiation to corneal epithelium-like cells of CMES cells. When the corneal epithelium-like cells of CMES origin were transplanted onto injured cornea, they adhered to the corneal lesion, leading to the formation of monolayer of corneal epithelium-like cells. The overlaid cells were stained with anti-human nuclei antibody which crossreacted with nuclei of CM cells but not those of mouse cells. They retained the expressions of E-cadherin and Keratin3 in situ.
We have succeeded in the induction of corneal epithelium-like cells from CMES cells, and the cells were successfully transplanted onto the injured mouse cornea. This is the first demonstration that corneal epithelium-like cells differentiated from nonhuman primate ES cells were applicable for transplantation to an animal model of corneal injury.
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