The modulation of IL-6 and TNF-alpha release by nitric oxide following stimulation of J774 cells with LPS and IFN-gamma.

Abstract

Lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma) induced nitric oxide synthase (NOS) activity, tumour necrosis factor-alpha (TNF-alpha), interleukin (IL)-6 and prostaglandin (PG)E2 release in J774 cells, a murine macrophage cell line. The role of endogenous NO in modulating TNF-alpha and IL-6 release was investigated using N-iminoethyl-L-ornithine (L-NIO), a specific inhibitor of NOS. L-NIO (10-1000 microM) produced a concentration-dependent potentiation of LPS and IFN-gamma induced IL-6 release. Time-course studies demonstrated a significant potentiation of IL-6 release at 12 h with a maximum effect at 48 h. By contrast to its effects on IL-6, L-NIO significantly attenuated TNF-alpha release, and at 48 h reduced PGE2 release. The NO-donor S-nitroso-N-acetyl-penicillamine (SNAP,300 microM), significantly inhibited LPS and IFN-gamma induced IL-6 release, but potentiated TNF-alpha release. In addition, SNAP prevented the potentiation of IL-6 and the inhibition of TNF-alpha release by L-NIO. Stimulation of J774 cells with a combination of LPS and IFN-gamma for 24 h or 48 h reduced cell viability which was prevented by L-NIO. Furthermore, SNAP also reduced cell viability determined after 24 h incubation. These results indicate that NO can differentially modulate LPS and IFN-gamma-induced cytokine release from J774 cells, up-regulating TNF-alpha but down-regulating IL-6, and that NO is cytotoxic to these cells.