Mianserin inhibits Toll-like receptor (TLR) 3, 7, and 9 signaling in murine macrophages. Murine bone marrow-derived macrophages were pre-incubated for 30 minutes with mianserin and then stimulated with (a) 100 ng/mL lipopolysaccharide (LPS), 1 μg/mL resiquimod (R-848), and 500 μM ODN M326 for 6 hours before measuring tumor necrosis factor (TNF) in the supernantants or (b) 20 μg/mL polyinosinic:polycytidylic acid (poly I:C) for 24 hours before measuring RANTES production in the supernantants. Data are shown as a percentage of the TLR ligand-only response at 30 μg/mL mianserin and as a representative graph showing the effect of a range of concentrations of mianserin. (c) Cell viability was measured by 3-[4,5 dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT) assay. **P < 0.01, ***P < 0.001. RANTES, Regulated upon Activation, Normal T-cell Expressed, and Secreted; unstim, unstimulated.