Purpose: We previously reported the successful transplantation of corneal epithelium-like cells derived from mouse embryonic stem (ES) cells onto injured mouse cornea. Here, we tested whether nonhuman primate ES cells have ability to differentiate into corneal epithelial cells and whether monkey ES cell-derived corneal epithelium-like cells were applicable for the experimental transplantation to damaged cornea.
Methods: Monkey ES cells were cultivated on type IV collagen-coated dishes for various days to induce differentiation into corneal epithelium-like cells. The differentiation was evaluated by reverse transcription-polymerase chain reaction and immunostaining. The corneal epithelium-like cells were transplanted to the injured mouse cornea. Reconstitution of the corneal epithelium was evaluated by immunostaining.
Results: The cells cultured on type IV collagen showed cobblestone-like appearance resembling epithelial cells. They expressed messenger RNA of pax6, p63, E-cadherin, CD44, proliferating cell nuclear antigen, keratin 3, and keratin 12. Protein expressions of pax6, keratin 3/12, p63, proliferating cell nuclear antigen, E-cadherin, and CD44 were confirmed by immunostaining. When the corneal epithelium-like cells were transplanted, they adhered to the corneal stroma, leading to formation of multiple cell layers. The grafted cells were stained with anti-human nuclear protein antibody, which cross-reacted with nuclei of monkey cells but not with those of mouse cells. They retained the expressions of keratin 3/12, E-cadherin, and CD44.
Conclusions: We induced corneal epithelium-like cells from monkey ES cells with moderate efficiency. The cells were successfully transplanted onto the injured mouse cornea. This is the first demonstration that nonhuman primate ES cells were induced to differentiate into corneal epithelium-like cells, which were applicable for transplantation to an animal model of corneal injury.