Keratinocyte Superoxide Generation

We have demonstrated using the reduction of cytochrome c, that the keratinocyte cell line H357 generates superoxide at significant rates (8.36 nmol/h/106 cells). The rate of superoxide release decreased as the cells reached confluence. Superoxide production was increased more than twofold following preincubation with IL-1β, or by the addition of the Ca2+ ionophore, Ionomycin. Other stimuli known to activate the NADPH oxidase of phagocytes were ineffective, but the regulatory cytokine IFNγ lowered the rate of release. Inhibitors of lipoxygenase function decreased the rate of superoxide production, whereas inhibitors of cyclo-oxygenase, xanthine oxidase, or NADPH oxidase failed to inhibit. The addition of NADH or NADPH to whole cells increased the rate threefold.