Effect of Dex on TLR-3– and TLR-9–inducible NFκB activation. (A) Effect of Dex on Poly (I:C)–mediated IκBα degradation. Control macrophages were treated with Dex for 3 hours followed by Poly (I:C) for the indicated periods of time. Cytoplasmic extracts were analyzed by Western blotting using an anti-IκBα and anti–β-actin antibodies. (B) Effect of Dex on CpG-mediated NFκB activation in control macrophages. Cells were treated as described. Cytoplasmic and nuclear extracts were analyzed by Western blotting using anti–phospho-IκBα (P-IκBα), anti–total IκBα (IκBα), and anti–phospho-p65 (P-p65) antibodies. (C) Effect of Dex on CpG-mediated IκBα phosphorylation in MGRKO macrophages. Cytoplasmic extracts were analyzed by Western blotting using anti–phospho-IκBα and anti–β-actin antibodies. (D) Effect of Dex on LPS-mediated IκBα degradation. Macrophages from control and MGRKO mice were treated with the indicated doses of Dex for 3 hours followed by LPS for 30 minutes. Cytoplasmic extracts were analyzed by Western blotting using anti-IκBα and anti–β-actin antibodies. (E) Effect of Dex on LPS-mediated p65 redistribution. Cytoplasmic and nuclear extracts were analyzed by Western blotting using anti-p65 and anti–β-actin antibodies. (F) Effect of Dex on LPS-mediated NFκB DNA-binding activity. Nuclear extracts from control and MGRKO macrophages were prepared and DNA-binding activity to major histocompatibility complex class I H2K–specific oligonucleotide probe were analyzed using EMSA. Representative of 3 to 4 independent experiments.