Inhibition of NF-κB, iNOS mRNA, COX2 mRNA, and COX catalytic activity by phenyl-N-tert-butylnitrone (PBN)

Previously, the spin trapping agent phenyl-N-tert-butylnitrone (PBN) has been shown to decrease the level of nitric oxide synthase mRNA in vivo. This inhibition is suggested to be an underlying mechanism for PBN's wide variety of pharmacological actions in animal models. However, the determination of PBN's cellular pharmacological activities has not been carried out, but is necessary for the understanding of the effects in vivo. Since the known pharmacological effects of PEN are primarily anti-inflammatory in nature, in this study we determined the inhibitory activities of PEN against two inflammatory factors: inducible nitric oxide synthase (iNOS) and inducible cyclooxygenase (COX2). We show here that PEN decreases steady state COX2 mRNA level and COX2 catalytic activity in macrophage cell culture at supra-pharmacological concentrations. While PEN decreases iNOS mRNA, it does not inhibit iNOS catalytic activity, which is consistent with previous in vivo studies. We also studied nuclear factor kappa B (NF-kappa B), a transcription factor that can rapidly activate the expression of genes involved in inflammatory, immune and acute phase responses. The binding of NF-kappa B to iNOS gene has been shown to be critical for iNOS gene expression, and the promoter region of COX2 gene contains NF-kappa B consensus sequence. We show that PEN inhibits lipopolysaccharide-mediated increase of NF-kappa B DNA binding activity with a lower concentration than that for the non-steroidal anti-inflammatory drug (NSAID), salicylate. Furthermore, we show that PEN inhibits COX2 catalytic activity, suggesting that PEN has an NSAID-like function. (C) 1998 Elsevier Science B.V. All rights reserved.