Possible implication of arachidonic acid metabolism in the decrease of chemiluminescence production after exposure of murine peritoneal macrophages to diethylnitrosamine and tumour promoter, 12-O-tetradecanoylphorbol-13-acetate

Abstract

The effect of the carcinogen diethylnitrosamine (DEN) on prostaglandins (PGs), leukotrienes (LTs) and reactive oxygen intermediates production by murine peritoneal macrophages was assessed. In vitro exposure to DEN (0.8, 1.6 and 8 mM) resulted in a dose-dependent stimulation of the PGs and LTs generation by macrophages. DEN-exposed peritoneal macrophages demonstrated enhanced production of arachidonic acid (AA) metabolites following stimulation with 12-O-tetradecanoylphorbol 13-acetate (TPA) as compared to macrophages stimulated with TPA alone. Studies of [3H]AA release from glycerolipids of prelabelled macrophages and of the distribution of AA metabolites between intra and extracellular compartments indicated that DEN induced de novo synthesis of AA metabolites. The stimulation of AA metabolism by DEN was decreased by H-7 and staurosporine, protein kinase C (PKC) inhibitors, and so could be dependent on PKC activation. The generation of PGs by macrophages after DEN exposure was also inhibited by indomethacin (cyclo-oxygenase inhibitor). DEN at high concentrations (1.6-16 mM) inhibited chemiluminescence production by peritoneal macrophages in a dose-dependent manner, triggered by tumour promoter TPA; lower concentrations (0.8 and 1.2 mM) increased this reactive oxygen intermediates dependent chemiluminescence production induced by TPA. The role of AA metabolism in the alteration of chemiluminescence production by murine peritoneal macrophages treated in vitro with DEN and triggered by TPA has been evaluated by using AA metabolism inhibitors. The stimulation of chemiluminescence by TPA was inhibited by the addition of phospholipase A2 (PLA2) inhibitor, 4-p-BPB; this metabolic inhibitor did not affect the decrease of chemiluminescence production induced by DEN. The cyclo-oxygenase (CO) inhibitor, indomethacin, reversed the inhibition of TPA-induced chemiluminescence caused by DEN. These results suggest that AA and/or a lipoxygenase product can potentiate the reactive oxygen intermediates production by macrophages stimulated by TPA. The CO pathway could be involved in the inhibition by DEN of the reactive oxygen intermediates generating enzyme system. It is suggested that this inhibition could be related to AA metabolites issued from the CO pathway or to DEN oxygenated metabolites issued from the co-oxidation of the DEN by the PGs endoperoxide synthase. These results also raise the problem of macrophage dysfunction by chemical carcinogens and the implication of the CO pathway in this process.