The study investigates the role of nitric oxide (NO) in activating cyclooxygenase (COX) enzymes, specifically COX-1 and COX-2. NO synthase (NOS) and COX were induced in mouse macrophage cell line RAW264.7 using Escherichia coli lipopolysaccharide (LPS). The production of nitrite (NO2-) and prostaglandin E2 (PGE2), products of NOS and COX, respectively, increased. These effects were blocked by Nω-monomethyl-L-arginine and aminoguanidine, NOS inhibitors. The addition of L-Arg, the precursor for NO synthesis, reversed these inhibitions but not d-Arg. RAW264.7 cells stimulated with LPS in L-Arg-free medium showed reduced NO2- release and lower PGE2 production compared to cells in the presence of L-Arg. In human fetal fibroblasts (HFFs) stimulated with interleukin 1β, exogenous NO increased COX activity and PGE2 production, which was abolished by hemoglobin (Hb), a NO binder. These findings suggest that NO enhances COX activity through a mechanism independent of cGMP, potentially leading to an exacerbated inflammatory response. The study also demonstrates that NO directly interacts with COX to increase its enzymatic activity.The study investigates the role of nitric oxide (NO) in activating cyclooxygenase (COX) enzymes, specifically COX-1 and COX-2. NO synthase (NOS) and COX were induced in mouse macrophage cell line RAW264.7 using Escherichia coli lipopolysaccharide (LPS). The production of nitrite (NO2-) and prostaglandin E2 (PGE2), products of NOS and COX, respectively, increased. These effects were blocked by Nω-monomethyl-L-arginine and aminoguanidine, NOS inhibitors. The addition of L-Arg, the precursor for NO synthesis, reversed these inhibitions but not d-Arg. RAW264.7 cells stimulated with LPS in L-Arg-free medium showed reduced NO2- release and lower PGE2 production compared to cells in the presence of L-Arg. In human fetal fibroblasts (HFFs) stimulated with interleukin 1β, exogenous NO increased COX activity and PGE2 production, which was abolished by hemoglobin (Hb), a NO binder. These findings suggest that NO enhances COX activity through a mechanism independent of cGMP, potentially leading to an exacerbated inflammatory response. The study also demonstrates that NO directly interacts with COX to increase its enzymatic activity.