Endogenous IFN-β is important for optimal LPS-induced CD40 expression. (A) RAW264.7 or EOC13 cells were treated with medium or LPS (10 ng/mL) in the presence of 10 μg/mL isotype antibody or neutralizing antibodies against IFN-γ, IFN-α, or IFN-β for 4 hours (RAW264.7 cells) or 8 hours (EOC13 cells). RNA was harvested and analyzed by RPA for CD40 and GAPDH mRNA. Representative of 3 experiments. (B) RAW264.7 cells were treated with medium or LPS (10 ng/mL) in the absence or presence of 10 μg/mL isotype antibody or IFN-β–neutralizing antibody for 36 hours. CD40 protein expression was detected by flow cytometry. Samples were analyzed by measuring MFI. Fold change in CD40 MFI value was calculated and shown as the mean ± SD of 3 experiments. (C) Primary microglia from WT and STAT-1α–deficient mice were treated with LPS (10 ng/mL) for 4 hours, and then mRNA was analyzed by RPA for CD40 and GAPDH expression. (D) Primary microglia from WT and STAT-1α–deficient mice were treated with LPS for 36 hours, then cells were subjected to FACS analysis for CD40 protein expression. Fold change in CD40 MFI value was calculated and shown as the mean ± SD of 3 experiments.