Figure 3

In vitro activation of the RIG-I/MAVS/IFN-I pathway in dendritic cells does not significantly influence allogeneic T cell activation. (A) Scheme of experimental setup: BM isolated from C57BL/6 WT mice was used to generate BM-derived GM-CSF DCs. GM-SCF DCs were stimulated with 3pRNA with or without additional treatment with anti-IFNaR1. One day later, stimulated DCs were cocultured with allogeneic CD4⁺ or CD8⁺ T cells derived from Balbc/c WT mice. Proliferation and IFN-γ production were analyzed on day 3 (CD8⁺ T cells) or 5 (CD4⁺ T cells) after onset of the mixed lymphocyte reaction (MLR). (B) Representative gating strategy of MLR with CD4⁺ T cells: Analysis of live (live/dead stain negative) CD4⁺ lymphocytes. The gate shows the percentage of proliferated (CFSE negative) and IFN-γ⁺ cells of all CD4⁺ T cells on day 5 after onset of the MLR of GM-CSF DCs generated from WT BM. Representative data from one of four experiments. (C) Percentage of proliferated and IFN-γ⁺ cells of all CD4⁺ T cells on day 5 after onset of the MLR. Pooled data of four independent experiments. (D) Percentage of proliferated and IFN-γ⁺ cells of all CD8⁺ T cells on day 3 after onset of the MLR. Pooled data of four independent experiments. Data were analyzed using two-tailed unpaired t test or ordinary one-way ANOVA for multiple comparisons. Significance was set at p values < 0.05, p < 0.01 and p < 0.001 and was then indicated with asterisks (*^,** and ***). Data are presented as mean ± S.E.M.