Figure 7.

Cognate interactions are essential for PSA-mediated generation of IL-10 in vitro and for protection in the TNBS colitis model. (A) WT CD4+ T cells co-cultured with PDCs from WT and MHCII−/− mice were compared. Co-cultures with WT PDCs had significantly higher levels of IL-10 than those with MHCII-deficient PDCs. (B) Adoptive transfer of PSA-pretreated BMPDCs from MHCII−/− mice (1.5 × 10⁵/dose, 2 doses) did not confer significant protection to WT mice. (C) Bone marrow–derived PDCs (SH+PDCA-1+CD11c+; gating used for evaluation) included a significantly higher frequency of ICOSL+SH+ cells when treated with PSA (60 μg/ml) than when treated with other TLR2 ligands [Pam3CSK4 (PAM), 0.1 μg/ml; FSL-1, 0.1 μg/ml; and lipomannan (LM-MS), 10 ng/ml] or left untreated (medium control). Bar graph shows average of 5 independent experiments. (D, E) Co-cultures of WT CD4+ T cells with PDCs from ICOSL−/− mice (D) or CD86−/− mice (E) produced significantly less IL-10 than co-cultures of WT CD4+ cells with WT PDCs. (F) Enhanced ability of PDCs from WT mice to stimulate IL-10 production more strongly in WT CD4+ T cells than in CD4+ T cells from ICOS−/− or CD28−/− mice. Data are the average of 2 independent experiments. (G) Adoptive transfer of PSA-pretreated WT PDCs did not confer protection in ICOS−/− recipients. (H) Anti-CD86 IgG and anti-ICOSL IgG blocked PSA-mediated protection and significantly reduced protection from that in IgG isotype control–treated mice. (I) WT mice—but not ICOSL−/− mice—were significantly protected after oral PSA treatment. Data shown in A, C, and D–F were analyzed by unpaired Student's t-test. *, **, and *** denote p<0.05, p<0.01, and p<0.001, respectively. Clinical scores shown in B, G, H, and I were assessed for statistical significance by two-tailed nonparametric Mann-Whitney test. *p<0.05; **p<0.01; ns, not significant. Each dot represents one mouse. Error bars indicate SEM values. See also Figure S5.