Pomegranate peel polyphenols inhibits inflammation in LPS-induced RAW264.7 macrophages via the suppression of TLR4/NF-κB pathway activation

Backgrounds: Inflammatory response mediated by activated immune cells is a vital process in host defense system while responding to various stresses. Our previous studies have indicated that pomegranate peel polyphenols (PPPs) and their main components punicalagin (PC) and ellagic acid (EA) decreased pro-inflammatory cytokines and inflammatory mediators by regulating the mitogen-activated protein kinases (MAPKs) pathway, but whether these tested polyphenols play an important role in NF-κB signaling pathway, another crucial pathway of inflammation, remains unclear. Objective: In this study, we analyzed the anti-inflammatory effect of these polyphenols via TLR4-NF-κB pathway in lipopolysaccharide (LPS)-induced RAW264.7 macrophages. Methods: Different concentrations of PPPs, PC, and EA were pre-incubated with RAW264.7 macrophages and then stimulated with LPS (1 μg/mL), and the effects of reactive oxygen species and TLR4 were investigated. Moreover, NF-κB p65 nuclear translocation and phosphorylation, and degradation of IκB were measured by Western blot. Furthermore, the influence of pro-inflammatory cytokines was detected by enzyme-linked immunosorbent assay (ELISA). Results: Our data showed that PPPs, PC, and EA inhibited LPS-induced intracellular ROS production and suppressed the mRNA and protein expression levels of TLR4 in a dose-dependent manner. Moreover, the anti-inflammatory mechanism was involved in blocking LPS-induced phosphorylation, degradation of IκB, and nuclear translocation of p65. Additionally, PPPs and PC exhibited a stronger anti-inflammatory effect than that of EA. Conclusion: The results indicated that PPPs possess potent anti-inflammatory effect, and PC was the main effective component in PPPs, which provided new insights into the utilization of PPPs to prevent inflammation-associated disorders.