Embryonic Stem Cell - Derived Corneal Epithelium - Like Cells In The Treatment Of Corneal Limbal Stem Cells

Purpose: To differentiate human embryonic stem cells(h ESCs) into corneal epithelial-like cells(ESC-CECs),observe the biological character of the differentiated cell and its effect to cure limbal stem cell deficiency, and explore the immunological properties of ESC-CECs and rejection after transplantation.Methods: h ESCs were induced to differentiate by small molecular, and ESC-CECs were identified by immunofluorescence staining and PCR. ESC-CECs and limbal epithelial cells(LECs) were seeded on denuded amniotic membrane for further transplantation. Rabbits and monkeys were used to establish limbal stem cell deficiency model and received ESC-CECs or LECs transplantation. The gene expression profile of ESC-CECs and LECs was obtained by whole genome microarray, and MHC expression was analyzed by flow cytometry. Mixed T lymphocyte proliferation assay and NK cell cytotoxicity assay was used to explore the the immunogenicity of ESC-CECs in vitro with or without interferon-γ. After cell transplantation, the rejection and immune cells in the cornea was observed.Results: ESC-CECs had a similar character and function with LECs in vitro, and were negative for pluripotent markers. The cells formed a stratified layer on denuded amniotic membrane and continuously improved the ocular surface condition with limbal stem cell deficiency after transplantation. In ESC-CECs, a large number of genes related with immune response were down-regulated. The expressions of MHC and co-stimulatory molecules were both low. The ESC-CECs were less responsible for T cell proliferation and NK cell lysis in vitro, and there was less immune cell infiltration after transplantation in vivo compared with LECs. Moreover, the immunological properties were not affected by interferon-γ.Conclusion: The corneal epithelial-like cells can be differentiated from h ESCs by small molecular. ESC-CECs present a similar character and function with LECs both in vivo and in vitro. More importantly, they own a low immunogenicity and are unlikely to initiate immune rejection.