Anti-inflammatory function of 4-tert-butylphenyl salicylate through down-regulation of the NF-kappa B pathway

Abstract

The salicylic acid derivative 4-tert-butylphenyl salicylate (4-TBPS) possesses anti-inflammatory activity. We demonstrated this and elucidated the mechanisms involved by using the lipopolysaccharide-stimulated Raw 264.7 mouse macrophage model. The 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, western blot, enzyme-linked immunosorbent assay, and reverse transcriptase-polymerase chain reaction were performed to explore 4-TBPS anti-inflammatory activity. We found that 4-TBPS decreased nitric oxide production without cytotoxic effects on macrophages and reduced expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2 in a dose-dependent manner. Additionally, mRNA expressions of iNOS and COX-2 significantly reduced, with concentrations between 1 and 15 μg/ml. Furthermore, 4-TBPS significantly inhibited the production of pro-inflammatory cytokines including tumor necrosis factor-α (TNF-α), interleukin- (IL)-1β, and IL-6. Moreover, mRNA gene expression of TNF-α, IL-1β, and IL-6 was attenuated in a dose-dependent manner. 4-TBPS potently inhibited translocation of nuclear factor-κB (NF-κB) into the nucleus by degrading IκB kinase (IκBα) following its phosphorylation, thereby causing NF-κB to remain inactive. Collectively, our data indicate that 4-TBPS significantly (p < 0.01) targets the inflammatory response of macrophages via inhibition of iNOS, COX-2, TNF-α, IL-1β, and IL-6 through downregulation of the NF-κB pathway. This indicates that 4-TBPS may have therapeutic potential in inflammatory disorders.