Regeneration of corneal endothelial cells following keratoplasty in rats with bullous keratopathy.

PURPOSE

Little is known about the behavior of the endothelial cell (EC) layer following keratoplasty. In vitro experiments suggested that the peripheral endothelium might have a higher regenerative capacity than the central endothelium, and some authors hypothesized that endothelial progenitor cells are present in the limbal area. Thus, we analyzed the corneal endothelial regenerative capacity in vivo in a rat model of bullous keratopathy using either bullous central grafts or bullous peripheral recipient corneas to analyze differences in EC regeneration depending on central versus peripheral cell origin.

METHODS

Bullous keratopathy was induced in Lewis rats with an intracameral injection of benzalkonium chloride (0.05%; BAK). Three days later, the central area of the bullous cornea was excised and used as a bullous graft, transplanted to a healthy, green fluorescent protein (GFP)-transgeneic Lewis receipient rat (group 'bullous graft'). In return, the mentioned rat eye with the bullous keratopathy received a healthy GFP-transgeneic corneal transplant (group 'bullous host'). A subgroup of these animals received a healthy, excentrically trephined including parts of the limbus (group 'bullous host, limbo-keratoplasty'). The grafts were monitored clinically for 7 weeks. Subsequently, the mean EC density was calculated on corneal whole mounts with Alizarin Red S staining. GFP was analyzed with confocal microscopy to determine EC origin. Untreated fellow eyes served as controls.

RESULTS

BAK injection led to a significant decrease in the mean EC density with subsequent bullous keratopathy. In the control eyes, the mean EC density was 3,744 cells/mm² in the center and 2,811 cells/mm² in the periphery. In eyes with bullous keratopathy, only 233 cells/mm² in the center and 622 cells/mm² in the periphery were observed three days after BAK-injection. Bullous transplants in the group 'bullous graft' cleared, and GFP-positive cells were detected on the transplant. In contrast, no GFP-positive ECs were detected on the host cornea in the groups 'bullous host'.

CONCLUSIONS

ECs from the peripheral cornea have the ability to cross the graft border and compensate for the pathologically altered/absent endothelium on the graft. In contrast, ECs derived from the central cornea remain central on the graft and do not replace or regenerate peripheral ECs in our model of bullous keratopathy.