Pipoxolan suppresses the inflammatory factors of NF-κB, AP-1, and STATs, but activates the antioxidative factor Nrf2 in LPS-stimulated RAW 264.7 murine macrophage cells

Although pipoxolan (PIPO) is a smooth muscle relaxant, its anti-inflammatory capability has not been studied. Therefore, we investigated the anti-inflammatory molecular mechanisms of PIPO in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages. In this study, we used the MTT assay to evaluate the cytotoxicity, applied the enzyme-linked immunosorbent assay to determine the inflammatory cytokines, and performed Western blotting to assess protein expression. The results showed that PIPO significantly inhibited cytokine production, including nitric oxide, prostaglandin E₂ , tumor necrosis factor-α, and interleukin-6. PIPO also suppressed the pro-inflammatory mediator expression with inducible nitric oxide synthase and cyclooxygenase-2. Moreover, PIPO prohibited the multiple inflammatory transcription factor pathways, including inhibitor kappa B/nuclear factor of the κ light chain enhancer of B cells (NF-κB), mitogen-activated protein kinase/activator protein-1 (AP-1), Janus kinase/signal transducer and activator of transcription (STAT), and Toll-like Receptor 4 (TLR4)/serine/threonine kinase (Akt). Besides, PIPO effectively activated the nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 antioxidative pathway. Collectively, PIPO may attenuate the inflammatory effects via influencing the LPS/TLR4 receptor binding; suppress the expression of anti-inflammatory transcription factors NF-κB, AP-1, and STAT; and activating the antioxidative transcription factor Nrf2 in LPS-stimulated mouse RAW 264.7 cells.