Proliferating cell nuclear antigen colocalization with corneal epithelial stem cells and involvement in physiological cell turnover.

Corneal integrity is dependent on a constant turnover of epithelial cells. According to the current hypothesis essential contributors to this process, the so-called stem-cells, are localized in the limbal area of the cornea. However, histological identification of stem cells of the cornea is thus far not possible. In the present study we have used specific antibodies against proliferating cell nuclear antigen (PCNA). The nuclear protein is expressed in cells during replication. Prior to establishment of staining indices the optimal fixation procedure was found to be of major importance for the results and standardized for fixation in 4% formaldehyde. This procedure also gave the best preservation of the morphology. In the corneas of ten New Zealand rabbits the staining index for PCNA was found to be on average 23% (SD +/- 13%) for the basal layer of the limbal epithelium. In the mid-peripheral and central corneal epithelium only occasional staining of PCNA was detected with a 2% staining index (SD +/- 2%). The agglomeration of PCNA-positive cells in the basal layer of the limbal epithelium together with their colocalization with the known location of stem cells strongly suggests that PCNA could serve as a reliable indicator for the proximity of proliferating corneal epithelial stem cells in histological sections, which would be of significant clinical importance.