Stratified corneal limbal epithelial cells are protected from UVB-induced apoptosis by elevated extracellular K⁺

The goal of this study was to determine whether elevated [K(+)] protects stratified corneal epithelial cells from entering apoptosis following exposure to ambient levels of UVB radiation. Human corneal limbal epithelial (HCLE) cells were stratified to form multilayered constructs in culture. The cells were exposed to UVB doses of 100-250 mJ/cm(2) followed by incubation in medium with 5.5-100 mM K(+). The protective effect of K(+) was determined by measuring the caspase-3 and -8 activity and TUNEL staining of the stratified HCLE constructs. In response to UVB exposure, activation of apoptotic pathways peaked at 24 h. Caspase-8 in stratified cells was activated by exposure to UVB at 100-250 mJ/cm(2), and activity was significantly reduced in response to 50 or 100 mM K(+). Caspase-3 was activated in the stratified cells in response to 100-250 mJ/cm(2) UVB and showed a significant reduction in activity in response to 25, 50 or 100 mM K(+). DNA fragmentation, as indicated by TUNEL staining, was elevated after exposure to 200 mJ/cm(2) UVB, and decreased following incubation with 25-100 mM K(+). These results show that in a culture system that models the intact corneal epithelium, elevated extracellular K(+) can reduce UVB-induced apoptosis which is believed to be initiated by loss of K(+) from cells. This is the basis of damage to the corneal epithelium caused by UVB exposure. Based on these observations it is suggested that the relatively high K(+) concentration in tears (20-25 mM) may play a role in protecting the corneal epithelium from ambient UVB radiation.