Epithelial regeneration after limbus-to-limbus debridement. Expression of alpha-enolase in stem and transient amplifying cells.

PURPOSE

To examine the expression of the glycolytic enzyme alpha-enolase after limbus-to-limbus epithelial debridement in the rabbit.

METHODS

Corneas were debrided, leaving limbal epithelium intact, and were allowed to heal from 2 days to 8 weeks. Immunofluorescence microscopy was used to observe the expression of alpha-enolase. To quantitate changes in alpha-enolase levels 2 days to 4 weeks after wounding, epithelium was harvested, homogenized, and assayed using anti-alpha-enolase in immunoslot blots.

RESULTS

Expression of alpha-enolase appeared to increase in the limbus and the central cornea during epithelial migration (2-day time point) with intense labeling of all basal cells. These levels were maintained until wound closure (1 week). By 2 weeks, expression in the limbal basal cells decreased to levels present in unwounded corneas. Expression in the corneal epithelium decreased after 2 weeks, progressing from central cornea to the periphery. At 4 weeks, antibody binding decreased concomitantly with a change in the shape of the basal cells from flattened or ovoid to columnar. At 8 weeks, expression of alpha-enolase was similar to that in control corneas. Immunoslot blot data indicated that alpha-enolase made up 0.28% of the total soluble protein in unwounded corneal epithelium and 0.73%, 1.22%, 0.96%, and 0.49% at 2 days, 1 week, 2 weeks, and 4 weeks after debridement, respectively.

CONCLUSIONS

These data indicate that expression of alpha-enolase is elevated during corneal epithelial migration initiating from the stem (limbal basal) cell population and that expression is linked to active migration. Furthermore, it appears that limbal basal cells are metabolically active during the period of epithelial sheet movement, whereas peripheral corneal basal cells remain activated as long as 4 weeks after wounding.