Corneal surface reconstruction using adult mesenchymal stem cells in experimental limbal stem cell deficiency in rabbits

Purpose: To investigate the ability of mesenchymal stem cells (MSC) to transdifferentiate to corneal epithelial cells in experimental limbal stem cell deficiency in rabbits. Methods: Total limbal stem cell deficiency was produced in 21 right eyes of 21 New Zealand rabbits; 6 eyes served as controls (group 1, G1). After removal of the conjunctival overgrowth, five eyes received amniotic membrane transplantation (AMT; G2). In four eyes, autologous limbal stem cell transplantation from the healthy eye was performed with AMT (G3). In another six eyes, enriched autologous MSC were injected under the amniotic membrane (AM) (G4). Within 280 days, corneoscleral discs were analysed for goblet cells, cytokeratin (CK) 3/12, connexin 43, beta(1)-integrin, CK 19, alpha-enolase, p63 and ATP-binding cassette transporter subtype G-2 (ABCG-2) distribution patterns. Results: Cultivated MSC were positive for CK 3/12 and alpha-enolase, but negative for ABCG-2, p63 and connexin 43. On rabbit corneas, CK 3/12 was expressed in all corneal regions in all groups, but with significantly different intensities. Among all other parameters, expression levels of ABCG-2, beta(1)-integrin and connexin 43 were significantly different between the transplanted groups and the control group. After a mean follow-up time of 172 (47-280) days, goblet cells were rarely present in the central cornea (G1-4). Conclusion: CK 3/12 is not highly specific for differentiated corneal epithelium. Further, goblet cells are not a reliable marker for conjunctivalization in rabbits. Expression of ABCG-2, beta(1)-integrin and connexin 43 after mesenchymal stem cell transplantation may indicate their ability to maintain their stem cell character or to transdifferentiate to epithelial progenitor cells.