DIRECT ACTIVATION OF MURINE PERITONEAL-MACROPHAGES FOR NITRIC-OXIDE PRODUCTION AND TUMOR-CELL KILLING BY INTERFERON-GAMMA

Interferon-gamma (IFN-gamma) is known to prime macrophages for tumor cell lysis and nitric oxide (NO) production as measured by enhanced sensitivity to lipopolysaccharide (LPS). In the present study, the ability of IFN-gamma to directly activate peritoneal macrophages from C57BL/6 and Balb/c mice for tumor cytotoxicity and NO production was evaluated. Macrophage-mediated tumor cell killing was measured by an 18 h Cr-51 release assay using P815 mastocytoma cells as targets. Concurrent NO production was measured as nitrite in the supernatants of macrophage cultures. Incubation of macrophages with IFN-gamma resulted in activation of macrophages for tumor cell lysis. IFN-gamma alone also activated macrophages for NO production under identical conditions. Addition of LPS along with IFN-gamma resulted in synergism in the activation of macrophages for both cytolysis and NO production. LPS contamination of the IFN-gamma preparation was absent as evidenced by the following criteria: (1) the IFN-gamma preparation as well as the reagents used were shown to be free of LPS contamination based on LAL endotoxin tests (sensitivity 25 pg/ml), (2) the ability of IFN-gamma to activate macrophages was not abrogated by prior treatment of the cytokine with polymyxin B, whereas the effect of LPS was inhibited (70-100%) under similar conditions, (3) pretreatment of the IFN-gamma preparation with a specific endotoxin neutralizing protein did not abrogate the ability of IFN-gamma to induce macrophage activation, and (4) heat treatment of solutions containing IFN-gamma alone or IFN-gamma + LPS abolished only the effect of IFN-gamma, not that of LPS. A comparison of the IFN-gamma responsiveness in the peritoneal macrophages from C57BL/6 and Balb/c macrophages revealed that C57BL/6 macrophages are more responsive to IFN-gamma. These results demonstrate that IFN-gamma alone is capable of stimulating macrophages for tumor cell killing and NO production and that the magnitude of IFN-gamma responsiveness varies with the strain of mice studied.