Limbal epithelium is more resistant to 5-fluorouracil toxicity than corneal epithelium

There is increasing evidence indicating that the limbal epithelium contains slow-cycling stem cells, whereas both limbal and corneal epithelia contain rapid-cycling transit amplifying cells. 5-Fluorouracil (5-FU), an antimetabolite used in glaucoma filtering procedures, is reported to inhibit rapid-cycling cells. After a 1-day subconjunctival (20 mg/eye) 5-FU injection followed by a 2-day rest, continuous BrdU labeling for 5 additional days (to also label slow-cycling stem cells) revealed that the limbal labeling index was not changed, whereas the corneal labeling index was markedly suppressed (p < 0.001). Following subconjunctival 5-FU treatments, peripheral and central corneal explant cultures showed a significantly higher magnitude of reduction of epithelial outgrowth size than limbal cultures (p < 0.05). Such a resistant nature of limbal epithelium to 5-FU toxicity was further confirmed by two different single-cell clonal growth assays. Taken together, these results indicate that the limbal epithelium has more slow-cycling cells than the corneal epithelium, and these cells selectively harvested by the subconjunctival 5-FU treatment can be used for future studies of their regulatory mechanism.