Figure 6. S100A8/A9 regulates CD11b expression on neutrophils.

Bone marrow neutrophils were isolated and infected with mCherry-labeled HN878 (MOI of 1) for 3 hours. (A) Mtb uptake by neutrophils was determined in B6 (n = 5) and S100A9KO (n = 5) neutrophils using flow cytometry. (B) MFI of CD11b, Gr-1, and CD18 expression on uninfected (UI) and highly infected (Mtb^hi) neutrophils was determined by flow cytometry. Bone marrow neutrophils were isolated and infected with HN878 (MOI of 1) for 3 hours. (C) Expression of phagocytic markers quantitated via RT-PCR and (D) TNF-α and IL-6 levels as quantitated by Luminex assays. Bone marrow neutrophils were isolated from B6 (n = 5) and S100A9KO (n = 5) mice and infected with HN878 (MOI of 1) for 1 hour. (E) Neutrophil chemotactic activity was assayed against gravity controls (Grav.) (circles) or supernatants from infected epithelial cells (Supe.) (squares). Neutrophils from B6 mice were left untreated or treated with HK Mtb, Mtb CFP, and Mtb CW. (F) CD11b and Gr-1 MFI were assessed using flow cytometry. Bone marrow neutrophils from IKK^fl/fl LysM^Cre mice (n = 5) and IKK^fl/fl (n = 5) mice were infected with Mtb (MOI 1). (G) Mtb uptake and MFI of CD11b and Gr-1 expression on uninfected or infected neutrophils was determined by flow cytometry. Figures depict 1 experiment representative of 2 or combined data from multiple experiments. Data points represent the mean ± SEM of values. (A–G) Student’s t test. *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001; ****P ≤ 0.0001.