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John L. Ubels, Leah R. Koetje, Mark P. Schotanus; Elevated Extracellular K+ Contributes to Protection of Stratified Corneal Epithelial Cells from UVB. Invest. Ophthalmol. Vis. Sci. 2011;52(14):314.
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The overall hypothesis of this study is that the relatively high [K+] in tears promotes the health of the corneal epithelium. We have previously reported that elevated [K+]o reduces apoptosis of human corneal limbal epithelial (HCLE) cells exposed to UVB in monolayer culture (Singleton et al. IOVS 89:140, 2009). We have now investigated effects of K+ on apoptosis and barrier function of stratified corneal epithelial cells after UVB treatment.
Stratified HCLE cells were exposed to 100-250 mJ/cm2 UVB and incubated in culture media containing isosmotic 5.5, 25, 50 or 100 mM K+ for 24-30 hours. This was followed by measurement of activation of caspase-3 and caspase-8 activity or fluorescein permeability of stratified constructs on microporous membranes. Control cells, not exposed to UVB, were incubated in medium with 5.5 mM K+.
Exposure to 100-250 mJ/cm2 UVB caused a 3.7-4 fold increase in caspase-3 activity compared to control 24 hr after exposure. Caspase-3 activation in cells exposed to 100 mJ/cm2 UVB was significantly inhibited by 25 mM K+, while 100 mM K+ inhibited caspase-3 at higher doses of UVB. Caspase-8 was activated 2.7-3.4 fold by UVB at 24 hr. It was inhibited by 50 mM K+ after 100 and 150 mJ/cm2 and by 100 mM K+ after 250 mJ/cm2 UVB. Fluorescein permeability increased 1.4-2 fold compared to control 30 hr after 100-250 mJ/cm2 UVB. K+ at 25-100 mM caused a significant, concentration-dependent reduction in the adverse effect of 150 and 200 mJ/cm2 UVB on permeability, while 100 mM K+ was effective after 100 or 250 mJ/cm2 UVB.
These in vitro data indicate that high [K+]o reduces the activation of apoptotic pathways and protects epithelial barrier function in stratified corneal epithelial cells exposed to UVB at levels relevant to ambient exposure. Since stratified HCLE cells model the intact, multilayered epithelium, the results suggest that the relatively high [K+] in tears may help to protect the cornea from environmental stress.
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