Dex suppresses TLR3- and TLR9-mediated inflammatory gene expression. Effect of Dexamethasone (Dex) on (A) Poly (I:C)–mediated (50 μg/mL) and (B) CpG-mediated (2μM) proinflammatory cytokine secretion in control and MGRKO macrophages. Cells were treated with the indicated doses of Dex for 3 hours followed by Poly (I:C) or CpG treatment for 24 hours. Concentrations of IL-6, TNF-α, and IL-12 in the culture media were analyzed by ELISA. Data are shown as mean ± SEM; n = 4; *P < .001 and **P < .005 for control macrophages compared with treatment group with Dex in MGRKO macrophages. Effect of Dex on (C) Poly (I:C)–mediated and (D) CpG-mediated proinflammatory cytokine mRNA transcript synthesis. Cells were pretreated with Dex followed by Poly (I:C) or CpG treatment for 4 hours. IL-6, TNF-α, and IL-12 mRNA was normalized to glyceraldehyde-3-phosphate dehydrogenase mRNA, and expressed relative to that in untreated controls. Each bar in the figure represents the average across 3 independent experiments of 2 independently run experimental replicates. Data are shown as mean ± SEM; *P < .05 and **P < .01 for Dex-treated macrophages compared with treatment group treated with Poly (I:C) or CpG only. Effect of Dex on (E) Poly (I:C)–mediated and (F) CpG-mediated COX-2 expression. Cells were pretreated with Dex followed by Poly (I:C) or CpG treatment for the indicated periods of time. ODNc (2μM) was used as the oligonucleotide control for CpG. Cell lysates were analyzed by Western blot using anti–COX-2 and anti–β-actin antibodies. Representative of 4 to 5 independent experiments. (G) Deletion of GR in macrophages enhanced Poly (I:C)–induced proinflammatory responses in vivo. Control and MGRKO mice were treated with Poly (I:C) for the indicated periods of time (within parentheses). Plasma concentrations of IL-6, TNF-α, and IL-12 were analyzed by ELISA. Data presented as mean ± SEM; n = 8-10. *P < .001 and **P < .005 for MGRKO mice compared with treatment group with Poly (I:C) in control mice.