Cellular death mediated by nuclear factor kappa B (NF-κB) translocation in cultured human lens epithelial cells after ultraviolet-B irradiation

Abstract

PURPOSE: To determine the role of nuclear factor kappa B (NF-kappaB) in the death of lens epithelial cells (LECs) after ultraviolet (UV) irradiation. SETTING: Department of Ophthalmology, Ilsan Paik Hospital, Inje University, Korea. METHODS: Cultures of simian virus 40 transfected human LECs (HLE B-3 cells) were were irradiated with a UVB source (312 nm) located 10 cm from the bottom of the slides for 1, 2, 3, or 4 minutes. Cytotoxicity was evaluated using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide method. Translocation of NF-kappaB was examined by immunocytochemistry using anti-NF-kappaB p65 antibody and electrophoretic mobility shift assay (EMSA). Sulfasalazine, a specific NF-kappaB inhibitor, was used to confirm the role of NF-kappaB by pretreating samples for 30 minutes before UV irradiation, after which cytotoxicity and NF-kappaB translocation were evaluated. RESULTS: When HLE B-3 cells were irradiated with UVB, translocation of NF-kappaB was observed with immunocytochemistry. These translocations peaked during EMSA 6 hours after UV irradiation. In HLE B-3 cells pretreated with sulfasalazine, the translocation of NF-kappaB was blocked. Cellular death after UV irradiation was also markedly reduced by sulfasalazine pretreatment. Ultraviolet irradiation can translocate NF-kappaB, and sulfasalazine is a useful blocking agent in this pathway. In this experimental model, sulfasalazine prevented cellular death after UV irradiation.