Engraftment and Proliferation of Thermoreversible-Gelation-Polymer-Encapsulated Human Corneal Limbal-Stem-Cells on Ocular Surface of a Cadaver Cornea.

Purpose: Corneal limbal stem cell (LSC) transplantation has been reported as a potential approach to treat the damaged corneal epithelium. Scaffolds such as human amniotic membrane (hAM) are commonly employed for the culture and as a carrier during transplantation. However, they carry the risk of biological contamination and donor to donor variability. To overcome these disadvantages, we herein report the capabilities of a synthetic thermoreversible gelation polymer (TGP) scaffold to serve as an encapsulation support during LSC transplantation and to enable engraftment for corneal regeneration.

Methods: Sixteen discarded human corneas were used to isolate the corneal epithelium which was cultured in TGP and hAM. The cell proliferation and characteristics between TGP and hAM culture methods were evaluated by microscopic observation, 3H Thymidine incorporation assay, immunoperoxidase and immunofluorescence staining.

Results: The 3H Thymidine assay's results showed that TGP allowed human-donor cornea-derived LSCs to proliferate well , compared to hAM and the cells encapsulated in TGP and transplanted onto a human cadaver donor cornea denuded of its epithelium, migrated on the ocular surface, and proliferated to form a continuous layer in 25 days. Immunoperoxidase and Immunofluorescence staining of TGP-cultured cells were positive for LSC markers (p63, ABCG2, Connexin 43 and Integrin ), proving that the TGP helps to preserve the limbal cells' stemness.

Conclusion: TGP is found to be a multipurpose scaffold for (i) culture, (ii) encapsulation, and transplantation (iii), enabling engraftment of LSCs in this study, with potentials to extend its application in cell-based therapies in several regenerative medicine approaches.