The Potential Proliferative Capacity of the Human Corneal Endothelium

Abstract

Abstract: : Purpose: To describe and characterize mitotic activity in the human corneal endothelium. Previous studies have indicated BrdU incorporation is increased in cell cultured corneal endothelial cells from the periphery versus the center (Bednarz & Engelmann, ARVO 2001, Abstr. 1484). Additionally, telomerase activity has been found in donor tissue only outside an 8 mm trephined button (Whikehart et al, ISER 2002, XV International Congress). We have compared the central and peripheral endothelial cells by evaluating BrdU incorporation in organ cultured donor corneas. Methods: Initially, donor corneas were wounded either from Schwalbe’s line to Schwalbe’s line or in the periphery (2 mm in length from Schwalbe’s line) using an olive tip cannula. Next, 20 ul of BrdU was added into the 20 ml vial of Optisol GS containing the cornea and incubated at 37°C for 7 days. Another set of experiments were performed using 10 ug/ml EGF added to the Optisol GS to induce division of peripheral endothelia. Then, BrdU incorporation was detected using a double antibody labeling procedure. Additionally, we used an ELISA assay to measure levels of TGFb in the Optisol-GS that corneas were stored in because it has been suggested that TGFb suppresses S-phase entry of corneal endothelial cells. Results: If a stripe wound was made, the BrdU incorporation studies inconsistently revealed nuclear staining adjacent to central wounded areas. However, if peripheral wounds were made without the addition of EGF, no BrdU incorporation was detected. Finally, if peripheral wounds were made and EGF was added, BrdU was detected in the peripheral cells of the monolayer. TGFb levels varied directly with temperature (none detected at 4°C and between 50-650 pg/ml detected at 37°C) and with increased time at 37°C (50 pg/ml after 6 days and 650 pg/ml after 17 days). The limited amount of BrdU incorporation may be related to the TGFb produced by endothelia. Conclusions: This data suggests a difference in cell type and activity between the central and peripheral human corneal endothelial cells. Our data together with telomerase activity shown in past studies may point to the existence of progenitor cells, possibly stem cells or transit amplifying cells. TGFb may be the cytokine that suppresses cellular division of corneal endothelial cells.

Keywords: cornea: endothelium • cornea: basic science • wound healing