EFFECTS OF CYTOKINES ON MYCOBACTERIAL PHAGOSOME MATURATION

One of the major mechanisms permitting intracellular pathogens to para sitize macrophages is their ability to alter maturation of the phagoso me or affect its physical integrity. These processes are opposed by th e host innate and adaptive immune defenses, and in many instances mono nuclear phagocytes can be stimulated with appropriate cytokines to res trict the growth of the microorganisms within the phagosomal compartme nt, Very little is known about the effects that cytokines have on phag osome maturation, Here we have used green fluorescent protein (GFP)-la beled mycobacteria and a fixable acidotropic probe, LysoTracker Red DN D-99, to monitor maturation of the mycobacterial phagosome, The macrop hage compartments that stained with the LysoTracker probe were examine d first. This dye was found to colocalize preferentially with the late endosomal and lysosomal markers rab7 and Lamp1, and with a fluid phas e marker chased into the late endosomal compartments. In contrast, Lys oTracker showed only a minor overlap with the early endosomal marker r ab5. Pathogenic mycobacteria are believed to reside in nonacidified va cuoles sequestered away from late endosomal compartments as a part of their intracellular survival strategy. We examined the status of mycob acterial phagosomes in macrophages from IL-10 knockout mice, in quiesc ent cells, and in mononuclear phagocytes stimulated with the macrophag e-activating cytokine IFN-gamma. When macrophages were derived from th e bone marrow of transgenic IL-10 mice lacking this major deactivating cytokine, colocalization of GFP-fluorescing mycobacteria with the Lys oTracker staining appeared enhanced, suggestive of increased acidifica tion of the mycobacterial phagosome relative to macrophages from norma l mice, When bone marrow-derived macrophages from normal mice or a J77 4 murine macrophage cell line were stimulated with IFN-gamma and LPS, this resulted in increased colocalization of mycobacteria and LysoTrac ker, but no statistically significant enhancement was observed in IL-1 0 transgenic animals, These studies are consistent with the interpreta tion that proinflammatory and antiinflammatory cytokines affect matura tion of mycobacterial phagosomes, Although multiple mechanisms are lik ely to be at work, we propose the existence of a direct link between c ytokine effects on the host cell and phagosome maturation in the macro phage.