IFNγ expression by NK cells is IL-18-dependent but not required for mounting tissue inflammation during the first 12h of the infection.

<p>(a) Volcano plot of all cytokines differentially expressed in the cecal mucosa of the <i>Il18</i>^<i>-/-</i> mice and littermate shown in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005723#ppat.1005723.g002" target="_blank">Fig 2A</a>. We plotted log₂ (fold change) against -log₁₀ (p-value). NK cell effector cytokines are highlighted in red. (b-d) <i>Il18</i>^-/- mice and littermate controls were Sm-pretreated and infected orally with 5x10⁷ CFU <i>S</i>.Tm for 12h (b) or 18h (c-d). (b) <i>Ifng</i>, <i>Iigp1</i> and <i>Cxcl10</i> transcripts in whole cecum tissue were analyzed by RT-qPCR. Results are presented relative to the expression of <i>Actb</i> (n = 8–9 per group). (c) IFNγ protein concentration in whole cecum tissue lysates (n = 5 per group) as measured by CBA; dashed line = detection limit. (d) Quantification of IFNγ-producing cells by flow cytometric analysis of isolated cecal LP cells (n = 6 per group). (e and f) C57BL/6 mice were infected for 18h with 5x10⁷ CFU <i>S</i>.Tm and cecal LP cells were isolated for staining. Data are shown from one out of three independent experiments. (e) Representative dot plot of IFNγ-expressing cells, pre-gated on single live lymphocytes. (f) FACS-analysis of CD3 and NK1.1 surface marker expression by IFNγ⁺ cell populations. (g) <i>Ifng</i>^-/-, <i>Ifngr1</i>^-/- and littermate controls were Sm-pretreated, infected orally with 5x10⁷ CFU <i>S</i>.Tm for 12h and pathological scores were assessed (n = 6–8 per group). (h) Mesenteric lymph node loads as determined by plating of organs from (left) <i>Ifng</i>^-/- and (right) <i>Il18</i>^-/- mice and their littermate controls. Mice were Sm-pretreated and infected orally with 5x10⁷ CFU <i>S</i>.Tm for 12h or 72h. Statistical analysis was performed using the Mann-Whitney-U test (ns = not significant, * = p<0.05; ** = p<0.01; *** = p<0.001).