Expression of inducible nitric oxide synthase and nitrotyrosine during the evolution of experimental pulmonary tuberculosis.

Nitric oxide (NO) is a relevant antimycobacterial factor in mouse macrophages. NO is a product of inducible nitric oxide synthase (iNOS). NO toxicity is greatly enhanced by reacting with superoxide to form peroxynitrite that reacts with many biological molecules. Tyrosine is one of the molecules with which NO reacts and the product is nitrotyrosine (NT). The production of peroxynitrite and the nitrosylation of proteins might play a role in bacterial killing and also in mediating host injury. In this study, we used a well-characterized mouse model of pulmonary tuberculosis to examine the local kinetics of expression and cellular distribution of iNOS and NT at the cellular and subcellular level. The histopathological study showed two phases of the disease: early and late. The early phase was characterized by mononuclear inflammation and granuloma formation. During this phase, high percentages of activated macrophages were observed that were immunostained for iNOS and NT. Immuno-electronmicroscopy showed NT immunoreactivity in lysosomes and mycobacterial wall and cytoplasm. The concentration of iNOS mRNA and NO metabolites were also elevated. The late phase was characterized by progressive pneumonia with focal necrosis and a decrease of iNOS mRNA and NO metabolites. The strongest NT immunostained areas were the necrotic tissue. Macrophages became foamy cells with scarce iNOS immunostaining but strong NT immunoreactivity. At the ultrastructural level, these cells showed NT immunolabeling in cytoskeleton, mitochondria, lysosomes and cell membrane. NT was also located in bronchial epithelial cell mitochondria, in cell membranes and cytoplasm of endothelial cells and in actin bundles within smooth muscle cells. These results suggest an important role of NO in mycobacterial killing, particularly during the early phase of the infection. They also suggest an important participation by NO in tissue damage during the late phase of the disease.