Critical role for intracellular cAMP in DC functions and effect of C3a treatment on DC intracellular cAMP level. (A) cAMP levels in 6-day DCs (2 × 106) after forskolin stimulation for 30 minutes at indicated concentrations. Data are shown as mean plus or minus SEM (n = 3). Data were analyzed by one-way ANOVA. (B) Effect of forskolin on cytokine secretion, surface expression of MHC class II, Ag uptake, and Ag presentation in DCs. Five-day DCs were further cultured for 24 hours in the presence or absence of forskolin (10 nM) and then used for Ag uptake and Ag-presentation assays. Six-day DCs were further cultured for 24 hours in the presence or absence of forskolin (10 nM). The cells were then used for surface expression of MHC class II assay, while the supernatants were used for cytokine ELISA. Data are shown as mean plus or minus SEM (n = 4 for cytokine secretion; n = 3 for Ag uptake; n = 2 for MHC II expression; and n = 8 for proliferation). Data were analyzed by Student t test. ***P < .001; **P < .005; *P < .05. (C) cAMP levels in 6-day DCs with or without C3a stimulation for 30 minutes in the presence of forskolin at indicated concentrations. Data are shown as mean plus or minus SEM (n = 3). Data were analyzed by 2-way ANOVA. P values are for comparisons between C3a and medium alone. (D) cAMP levels in 6-day DCs after C3a stimulation at indicated concentrations for 30 minutes in the presence of forskolin (10 nM). Data are shown as mean plus or minus SEM (n = 3). Data were analyzed by one-way ANOVA. (E) cAMP levels in 6-day WT DCs and C3aR−/− DCs after forskolin (10 nM) stimulation for 30 minutes. Data are shown as mean plus or minus SEM (n = 3). Data were analyzed by Student t test.
