MACROPHAGE ACTIVATING FACTOR(S) SECRETED BY MITOGEN-STIMULATED GOLDFISH KIDNEY LEUKOCYTES SYNERGIZE WITH BACTERIAL LIPOPOLYSACCHARIDE TO INDUCE NITRIC-OXIDE PRODUCTION IN TELEOST MACROPHAGES

Recent studies in our laboratory demonstrated that fish macrophages pr oduce nitric oxide. To elucidate the mechanisms which regulate nitric oxide production in teleosts, we examined whether macrophage activatin g factors (MAFs) secreted by mitogen stimulated leukocytes, induced ni tric oxide production in a long-term cultured macrophage cell line and in primary cultures of kidney macrophages from the goldfish. The resu lts indicate that both primary and long term cultured goldfish macroph ages produce nitric oxide in response to MAF or bacterial lipopolysacc haride (LPS), and co-stimulation with both factors results in a synerg istic induction of nitric oxide production. MAF that induced nitric ox ide production were present in leukocyte supernatants as early as 24 h after addition of mitogens to cell cultures. The production of MAF wa s dependent upon the incubation temperature, presence of serum in the culture medium and duration of incubation: maximal MAF activity was de tected in 72-96 h supernatants raised in media with serum at 30 degree s C. MAF-induced nitric oxide production by long term cultured macroph ages was inhibited by 1000 mu M N-G-monomethyl-L-arginine or amino-gua nidine, indicating an L-arginine-dependent metabolic pathway for the p roduction of the reactive nitrogen intermediates in teleosts. The bioc hemical events of cytokine induced nitric oxide production by teleost macrophages appear to be similar to those of mammalian macrophages.