Bacterial challenge stimulates formation of arachidonic acid metabolites by human keratinocytes and neutrophils in vitro

Although the interactions of bacteria with keratinocytes induce the synthesis of various mediators, the capability of epithelial cells to form arachidonic acid mediators has not been studied, and therefore the first part of this study was initiated. The complex mixture of epithelium-derived mediators suggests that chemoattraction is not their only effect on neutrophils and that they may also affect neutrophil mediator synthesis. The effect of epithelium-derived mediators on neutrophil eicosanoide synthesis was evaluated in the second part of this study. We incubated human keratinocytes with human-pathogenic bacteria for 2 It and harvested the supernatants after 4, 6, 10, and IS h of culture. Subsequently, the supernatants were coincubated for 5 min with human neutrophils with or without arachidonic acid. The formation of the arachidonic acid metabolites prostaglandin E-2 (PGE(2)), leukotriene B-4 (LTB4), 12-hydroxyeicosatetraenoic acid (12-HETE), and 15-HETE in keratinocytes and neutrophils was measured by reverse-phase high-pressure liquid chromatography. We demonstrated for the first time that keratinocytes produced significant amounts of LTB4 and 12-HETE 4 to 6 h after bacterial challenge. Upon stimulation with epithelial supernatants, neutrophils produced significant amounts of PGE(2), LTB4,12-HETE, and 15-HETE throughout the observation period of 18 h, with a maximum synthesis by supernatants harvested 4 to 10 h after bacterial infection. The results of the study suggest that arachidonic acid mediator formation by epithelial cells following bacterial challenge may act as an early inflammatory signal for the initiation of the immune response. The epithelial supernatants were capable of inducing the formation of arachidonic acid mediators by neutrophils, which may have further regulatory effects on the immune response.