Figure 10

MB49 harboring BCG activates a MyD88-dependent inflammatory response in macrophages. MB49 cells used for co-culture were previously infected with BCG (MB49 + BCG) for 24 h and washed to remove free BCG in the supernatant, previously to the addition of C57BL/6 WT or MyD88^−/− macrophages in the co-culture. Co-cultured cells were maintained together for 24 h (A–E) or 48 h (F), when the samples were collected for analysis. MB49 or macrophages were also infected separated (not in co-culture) and used as controls. Measurement of TNF-α (A), IL-6 (B), IL-1β (C) and IL-10 (D) production in the supernatants after 24 h were performed by ELISA. (E) iNOS mRNA expression in co-cultured cells after 24 h was evaluated by qPCR. All qPCR results were relative to β-actin mRNA as a normalizer and C57BL/6 WT BMDM (mock) used as control. (F) NO production was evaluated by the Griess method in supernatants after 48 h of stimulation. Mock treated data are not shown in graphs. The values are representative of at least three independent experiments. *Statistically significant compared to MB49 + BCG, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. ^#Statistically significant data comparing C57BL/6 WT and MyD88^−/− BMDMs, ^#P ≤ 0.05, ^##P ≤ 0.01, ^###P ≤ 0.001, ^####P ≤ 0.0001. ^&Statistically significant data comparing C57BL/6 WT BMDM in co-culture or not, ^&&P < 0.01, ^&&&&P ≤ 0.0001.