NZ suppresses TLR4/NF-κB signalings and NLRP3 inflammasome activation in LPS-induced RAW264.7 macrophages

Objective The purpose of the present study was to evaluate the potential therapeutic effects of NZ on lipopolysaccharide (LPS)-induced RAW264.7 cells and explore its underlying mechanisms. Methods The effect of NZ on NO generation in LPS-activated macrophage was measured by Griess assay. The concentrations of TNF-alpha, IL-18, IL-1 beta were analyzed with ELISA kits. The LPS-induced production of reactive oxygen species (ROS) was determined by flow cytometry. The protein expressions of TLR4, NF-kappa B and NLRP3 signaling pathway were investigated with Western blot analysis. Reults It was shown that NZ significantly reduced the production of NO and the generation of pro-inflammatory cytokines in LPS-induced RAW264.7 cells. In addition, NZ markedly inhibited the up-regulation of toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88) and the activation of nuclear factor kappa B (NF-kappa B) in LPS-stimulated RAW 264.7 macrophages. Of note, NZ suppressed the expression of the inflammasome component such as NOD-like receptor 3(NLRP3), apoptosis-associated speck-like protein containing CARD(ASC), as well as the levels of cytokines including Interleukin-18(IL-18) and Interleukin-1 beta(IL-1 beta). Conclusion These results indicated that NZ inhibited the generations of NO and pro-inflammatory cytokines by suppressing TLR4/MyD88/NF-kappa B pathway, suggesting that NZ could be an effective candidate for ameliorating LPS-induced inflammatory responses.