Figure 5.

S100A8/A9 proteins mediate exacerbated inflammation during Mycobacterium tuberculosis (Mtb) infection. B6 and S100a9^−/− mice were infected with approximately 100 CFU Mtb H37Rv. From Day 9 postinfection, B6 and S100a9^−/− Mtb-infected mice received either isotype (Iso) or IFN-γ neutralizing antibody (α−IFN-γ) (300 μg/mouse every 48 h) and samples for the analyses below were collected on Day 30 postinfection. Formalin-fixed paraffin embedded lung sections were stained with hematoxylin and eosin (A and B) or analyzed by immunofluorescence using antibodies specific for Gr1-1-7/4 (red) and S100A8 (green) (C and D). Average size of inflammatory lesions (B) was quantified in the Mtb-infected lungs using the morphometric tool of the Zeiss Axioplan microscope or number of Gr1⁺ cells per ×20 field counted (C) (×100 original magnification for hematoxylin and eosin images; ×200 original magnification for fluorescent images). Single cell suspensions from lungs of Mtb-infected mice were stained with antibodies specific for CD11b and Gr1 and the mean fluorescent intensity (MFI) of CD11b expression (E) and number of lung neutrophils (Gr1⁺ CD11b⁺) determined by flow cytometry (F). Lung bacterial burden in Mtb-infected mice was determined by plating (G). The data points represent the mean (±SD) of values from 4–6 mice (A–G). *P ≤ 0.05, **P ≤ 0.005, ***P ≤ 0.0005. ns = not significant. One experiment representative of two is shown.