EXPRESSION OF INDUCIBLE NITRIC-OXIDE SYNTHASE IN RAT PULMONARY CRYPTOCOCCUS-NEOFORMANS GRANULOMAS

Rats, like humans, have extremely effective immune mechanisms for cont rolling pulmonary Cryptococcus neoformans infection. Tbe mechanism(s) responsible for efficient immunity in rat experimental infection is un known. Recently, induction of inducible nitric oxide synthase (iNOS) a nd nitric oxide (NO) have been implicated as an important microbicidal mechanism by which activated macrophages effect cytotoxicity against microbes In this report, we investigated the expression of iNOS in rat pulmonary cryptococcosis. Localization and regulation of NO productio n was studied by immunohistochemistry for iNOS in conjunction with imm unohistochemistry for cell markers, cytokines, and cryptococcal capsul ar polysaccharide. iNOS immunoreactivity was detected in macrophages, neutrophils, vascular endothelium, and respiratory epithelium. Double- immunolabeling studies revealed that the most prominent iNOS immunorea ctivity was localized to epithelioid macrophages (CD11b/c(+)) within g ranulomas; CD4(+) and CD8(+) T cells were numerous around granulomas b ut did not express iNOS. iNOS immunoreactivity was detected itt a sele ctive population of epithelioid macrophages within some granulomas but not others. iNOS(-) granulomas were identical to iNOS(+) granulomas w ith respect to morphology and immunohistochemical profiles Macrophage iNOS immunoreactivity was detected I weeks after infection ia one out of four rats and was strongly expressed in all mts at 2 weeks (in up t o 50% of the granulomas) but declined consider ably by 25 days. iNOS e xpression coincided with granuloma formation and preceded a decrease i n lung fungal burden, suggesting an anticryptococcal role for NO. By d ouble labeling, cytokines that have been shown to promote (interferon- gamma, granulocyte/macrophage colony-stimulating factor) and inhibit ( transforming growth factor-beta) macrophage iNOS expression were detec ted around iNOS(+) granulomas. iNOS immunoreactivity was expressed in selected neutrophils CI and 2 week's) and endothelial cells (1 and 2 w eeks and 25 days) in the inflamed lung. Airway iNOS immunoreactivity w as limited to the luminal border of rare bronchiolar epithelial cells iNOS immunoreactivity was not detected in uninfected rats. The present study provides the first evidence for association of iNOS expression with protective cellular responses to cryptococcal infection in vivo.