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. 2011 Dec 19;208(13):2705–2716. doi: 10.1084/jem.20110547

Figure 5.

Figure 5.

Defective induction of IFNs in the MLNs of TrifLps2 mice after Y. enterocolitica infection. (A) Real-time PCR analysis of IFN-γ, IFN-β, Il-12p35, and TNF expression in the MLNs taken from control and 48 h after infection of WT and TrifLps2 mice. Averages of samples from three independent experiments (n = 6 each; *, P < 0.0001). Error bars, SEM. NS, not significant. (B) IFN production by MLN cell suspensions harvested 48 h after infection. The protein levels of IFN-β and IFN-γ in the supernatants were measured by ELISA assays after an additional 48 h of incubation in vitro. Combined data from three independent experiments (n = 6 each; *, P < 0.001). Error bars, SEM. (C) Immunofluorescent and FCM analyses of IFN-β– and IFN-γ–expressing cells in the MLN taken from infected WT mice 48 h after infection. IFN-β–expressing cells positive for F4/80 are indicated by the arrowhead. IFN-β–expressing cells positive for CD11c are indicated by the arrow. Bars: (top and middle) 10 µm; (bottom) 5 µm. Representative pictures from infected WT mice (n = 4 from two independent experiments). FCM analysis shows that 76.2 ± 1.1 and 23.3 ± 1.0% of IFN-β–expressing cells are macrophages and myeloid DCs, respectively. Isotype control data (biotin rat IgG with PE streptavidin and APC-hamster IgG within IFN-β–positive cells) used for flow setting is shown (right). In addition, 71.5 ± 2.8% of IFN-γ–expressing cells are NK cells. 4.1 ± 0.3% of IFN-γ–expressing cells are CD3+ NKT cells. Isotype control data (FITC–mouse IgG and APC–hamster IgG within IFN-γ–positive cells) used for flow setting is shown (right). Representative data from infected WT mice (n = 3 each from two independent experiments). (D) Serum IFN-β measured by ELISA 48 h after infection. Combined data from three independent experiments (n = 6 each; *, P < 0.05). Error bars, SEM. (E) Kinetic analysis of cytokine gene expression represents regional (MLN) and systemic (spleen) cytokine responses during Y. enterocolitica infection (real-time PCR: averages of samples from three independent experiments, n = 6 each; *, P < 0.05; error bars, SEM). (F) FCM analysis of splenocytes expressing IFN-β in infected WT mice (n = 3 each from two independent experiments).