Purpose: The purpose of the present study is to determine the role of K+ channel activity as an early event in UV-induced corneal epithelial cell apoptosis.
Method: Both cell-attached and nystatin-perforated patch-clamping were performed to record K+ channel activity in rabbit corneal epithelial (RCE) and primary cultured rabbit corneal epithelial (PRCE) cells exposed to UV irradiation. On exposure of corneal epithelial cells or intact corneas to UV-C irradiation or treatment of corneal epithelial cells with etoposide, cell apoptosis was determined by DNA fragmentation, ethidium bromide-acridine orange nuclear stain and TdT-mediated dUTP nick-end labeling (TUNEL).
Results: In the present study, UV-irradiation-induced corneal epithelial cell apoptosis through activation of a K+ channel in the cell membrane was an early event in response to UV irradiation. UV-C irradiation (42 microJ/cm(2)) activated robust K+ channel activity in RCE and PRCE cells at both the single-channel and whole-cell levels, when measured with the cell-attached and nystatin-perforated patch clamps, respectively. Suppression of UV-irradiation-induced K+ channel activity with the specific K+ channel blocker 4-aminopurydine (4-AP) prevented UV-irradiation-induced apoptosis in the RCE and PRCE cells, loss of the superficial layer of corneal epithelium, and apoptosis in the basal layer corneal epithelium. However, suppression of K+ channel activity did not protect RCE and PRCE cells from etoposide, a topoisomerase II inhibitor, which induced cell death by bypassing the membrane. Furthermore, application of valinomycin, a K+ ionophore, to mimic the effect of mass activation of the K+ channel in RCE and PRCE cells caused cell apoptosis.
Conclusions: The results indicate that UV irradiation induces superactivity of K+ channels in the membrane is an early event mediating signaling transduction and resulting in corneal epithelial cell death in response to UV irradiation.
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