SYNERGIC INHIBITORY ACTIVITY OF AMPHOTERICIN-B AND GAMMA-INTERFERON AGAINST INTRACELLULAR CRYPTOCOCCUS-NEOFORMANS IN MURINE MACROPHAGES

Cryptococcus neoformans is responsible for pulmonary and meningal infections in HIV patients. The lack of effective cellular cooperation caused by the low level of CD4(+) cells, and the resistance of C. neoformans to phagocytosis allows growth and persistence of the yeast in the host. We describe here an in-vitro model of intracellular replication of C. neoformans inside J774-A.1 macrophages, and the determination of the intracellular antifungal activity of amphotericin B and fluconazole alone or in association with IFN-gamma. The maximum inhibitory effect was observed with one MIC of amphotericin B and 100 or 1000 IU/mL of IFN-gamma. amphotericin B alone (at 1 x MIC), or either 1 x or 50 x MIC of fluconazole in normal or IFN-gamma activated macrophages, did not eradicate the ingested yeast. A potential underlying mechanism of the synergy of amphotericin B in IFN-gamma primed macrophages was investigated by measurement of nitrite level and by use of the NO synthase competitive inhibitor, N-G-monomethyl L-arginine (NMMA). One MIC of amphotericin B was able to activate the synthesis of nitrogen reactive intermediates in IFN gamma-primed macrophages. NMMA treated infected macrophages responded less well to IFN-gamma priming, resulting in a moderate inhibition in subsequent amphotericin B exposure.