DCs with defective C3a-C3aR stimulation have impaired function in Ag presentation. (A,B) Effect of C3aRa treatment. C3aRa (50 nM) was added into DC culture medium every other day from the beginning of BM cell culture (from WT or C3^−/− mice) for 6 days. DCs were then used for Ag-presentation assays in vitro and in vivo/ex vivo. (C,D) C3aR^−/− DCs versus WT DCs. DCs were prepared from C3aR^−/− mice and WT control mice, and then used for Ag-presentation assays in vitro and in vivo/ex vivo. (A,C) In vitro assays. Data are shown as mean plus or minus SEM (n = 4 for ELISA; n = 8 for thymidine uptake). Data were analyzed by Student t test. ***P < .001; **P < .003; NS, no significant difference. A representative of 3 independent experiments is shown. (B,D) In vivo/ex vivo assays. Each dot represents a single animal and is shown as mean of 4 replicate wells of the ex vivo culture. Data were analyzed by 2-way ANOVA. A representative of 2 independent experiments is shown.