Fig. 4.
Spp1 expression is required for induction of TH1 and TH17 cells by lung DCs. (A) Effect of Spp1 siRNA on SPP1 mRNA. Human emphysematous lung DCs were transfected with 10 ng of Spp1 or scrambled siRNA, after which SPP1 mRNA was quantitated by PCR. (B) Human lung DCs require Spp1 to differentiate T cells. Similarly treated human lung DCs were cultured with allogeneic human PBMC-derived CD4 T cells in the presence of anti-CD3 antibody (1 μg/ml) for 3 days, followed by intracellular cytokine staining for IL-17A and IFN-γ. Data are representative of at least five independent experiments. (C and D) IL-17A (C) and IFN-γ (D) were quantitated from 3-day cultures of lung APCs isolated from smoke-exposed WT and Spp1−/− mice that were incubated with spleen-derived CD4 T cells in the presence of soluble anti-CD3 antibody (1 μg/ml) (n = 3 independent experiments with triplicate replicates in each experiment).