IFN-γ-activated lymphocytes boost nitric oxide production in grass carp monocytes/macrophages

It is well known that IFN-γ is a prime activator of nitric oxide (NO) production by monocytes/macrophages in mammals and fish. In parallel, whether IFN-γ-activated lymphocytes are associated with NO production remains unclear. In this study, grass carp monocytes/macrophages and lymphocytes from head kidney were isolated and effects of recombinant grass carp IFN-γ (rgcIFN-γ) on NO releases by these two cell populations were determined. Results showed that rgcIFN-γ time- and dose-dependently increased NO production by monocytes/macrophages but not lymphocytes, which are consistent with the findings in mammals. Interestingly, rgcIFN-γ displayed a greater stimulation on NO production in the co-cultures of monocytes/macrophages and lymphocytes when compared with that in the culture of monocytes/macrophages alone. Furthermore, the media harvested from rgcIFN-γ-treated lymphocytes were effective in boosting NO release in monocytes/macrophages. These data suggest that secretions from rgcIFN-γ-treated lymphocytes may be involved in the NO release by monocytes/macrophages. To address this hypothesis, effect of rgcIFN-γ on the gene expression of inflammatory cytokines in grass carp lymphocytes was examined, showing that it consistently stimulated the mRNA expression of grass carp TNF-α and IL-1β but not IFN-γ. Furthermore, treatment of rgcIFN-γ combined with recombinant grass carp IL-1β (rgcIL-1β) induced a NO production by monocytes/macrophages, which was significantly higher than those induced by either cytokine alone. It provides the evidence that the cytokines secreted by the activated lymphocytes may facilitate the NO production by monocytes/macrophages. Taken together, our findings point out a new mechanism for the involvement of IFN-γ-activated lymphocytes in the NO production by monocytes/macrophages in fish. This knowledge not only strengthens the role of IFN-γ in immune system but also provides the evidence for the existence of a close relationship between lymphocytes and monocytes/macrophages in fish.