FIG 1. RV activates inflammasome in vivo in immature mice.

A. Eight week-old (adult) or 6 day-old (immature) C57BL/6 mice were inoculated with sham or RV. Lung mRNA and protein expression were measured 1 day later. (N=4, mean±SEM, *different from sham, †different from adult RV, p<0.05, one-way ANOVA with Tukey’s multiple comparisons test.) B. Six day-old immature C57BL/6 mice were inoculated with sham or RV. Lung mRNA and protein expression were measured 1, 2, 3, 4 or 7 days later. (N=4–7, mean±SEM, *different from sham, p<0.05, one-way ANOVA with Tukey’s multiple comparisons test. C. Six day-old immature C57BL/6 mice were inoculated with sham or RV. Lung mRNA was measured 1 day after infection. (N=6, mean±SEM, *different from sham, p<0.05, unpaired t test. D. One day after infection, whole lungs were homogenized within the lysis buffer and subjected to Western blot. Anti-mouse-IL-1β recognizes pro-IL-1β and its bioactive form IL-1β. Anti-mouse-caspase-1 detects both caspase-1 and its cleaved form, caspase-1 p12. E. Group mean relative expression levels were normalized to β-actin. (N=6, mean±SEM, *different from sham, p<0.05, one-way ANOVA.) F and G. Lung IL-1β+ cells in RV-infected six-day-old mice. IL-1β+ cells were identified 1 d after infection. IL-1β+ cells were analyzed as a percentage of live cells (left panel) and F4/80+ and CD11b+ cells were analyzed as a percentage of CD45+IL-1β+ cells (right panel) (n = 4, mean±SEM, *different from WT sham, p<0.05, unpaired t-test). H. Lungs were stained for IL-1β (green) NLRP3 (green), F4/80 (red) and nuclei (DAPI, black; bar, 50 μm). I. and J. Clodronate- or PBS-containing liposomes were delivered to mice intranasally 24 hours before sham or RV infection. One day after infection, lungs were harvested for mRNA and Western blot (N=6, mean±SEM, *different from sham, p<0.05, one-way ANOVA).