ANO inhibited IL-6 production by peritoneal macrophages stimulated with TLR agonists. Peritoneal macrophages (5 × 104/well) were stimulated with LPS (2 μg/mL) (A), Poly I:C (50 μg/mL) (B), Pam3CSK4 (5 μg/mL) (C) and zymosan A (100 μg/mL) (D) in the presence or absence of various concentrations of ANO at 37°C for 24 h. Levels of IL-6 in the culture supernatants were determined by ELISA. Growth of macrophages was assessed by adding 20 μl/well of alamarBlue™ dye for the last 2 to 3 h of the incubation period and expressed as FI values. Values represent the means ± S.D. of triplicate cultures. Results are representative of two separate experiments with similar results. *p < 0.05; **p < 0.01, significantly different when compared with control culture.