Abstract
Purpose :
To assess the effects of Wnt acitvation in a in vitro model of partial limbal stem cell deficiency.
Methods :
To mimic LSCD from a chemical burn, one of the most frequent etiologies of LSCD worldwide, an in vitro alkali burn model was developed using single limbal epithelial cells. The model was established by exposing cultured LSCs to various concentrations of sodium hydroxide. After model validation, Wnt activation was attained in the damaged LSCs by treating the cells with lithium chloride (LiCl), a widely used GSK-3β inhibitor, and a small-molecule Wnt mimic, respectively. The cell phenotype (% of cytokeratin 12, 14, p63 bright cells, vimentin) and proliferation were assessed.
Results :
A progressive loss of their stem/progenitor phenotype was observed: the percentage of p63bright cells (putative LSCs marker) and cytokeratin (K)14+ cells (undifferentiation marker) decreased while the percentage of K12+ (differentiation marker) increased. After 18 hours of treatment by Wnt activators, LSCs proliferation was increased. The LSC phenotype was recovered in the Wnt mimic and the LiCl groups, while the untreated cells lost their phenotype and did not proliferate. In the Wnt mimic group, the percentage of p63bright cells was significantly higher in LiCl-treated cells (2.27 fold change increase, p<0.05) and Wnt mimic-treated cells (4.12 fold change increase, p<0.05) while the percentage of K12+ cells was significantly lower.
Conclusions :
These findings suggest that the local Wnt activation may rescue LSCs upon alkaline injury.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.