Nuclear factor κB (NF-κB) activation by hydrogen peroxide in human epidermal keratinocytes and the restorative effect of interleukin-10

Abstract The heterodimeric form of nuclear factor κB (NF-κB), NF-κB1/RelA, is one of the pluripotential transcription factors that activates various genes encoding cytokines and cell adhesion molecules. To clarify the involvement of radical oxygen species in the NF-κB activation pathway in keratinocytes, we examined the effect of hydrogen peroxide (H 2 O 2 ) on the activation of NF-κB in cultured normal human epidermal keratinocytes. After the treatment of keratinocytes with 300 μM H 2 O 2 , a translocation of NF-κB from the cytoplasm to nucleus was observed in an immunofluorescence study using anti-human NF-κB1 and anti-human RelA antibodies. Specific DNA binding was observed with the nuclear extract prepared from the H 2 O 2 -treated keratinocytes by the electrophoretic mobility shift assay. The presence of N -acetyl- l -cysteine or pyrrolidine dithiocarbamate during H 2 O 2 treatment prevented the nuclear localization of NF-κB. The involvement of radical oxygen species in the NF-κB activation pathway was suggested. Pretreatment of keratinocytes with 10 ng/ml of recombinant human interleukin-10 (IL-10) for 24 h suppressed the nuclear translocation of NF-κB induced by H 2 O 2 . IL-10, which increases in ultraviolet-irradiated skin and suppresses delayed type hypersensitivity in vivo, may play an inhibitory role in cutaneous inflammation by inhibiting the NF-κB activation pathway in keratinocytes.