EtOAc extract of H. attenuatum Choisy inhibits inflammation by suppressing the NF-κB and MAPK pathways and modulating the gut microbiota

Background: Hypericum attenuatum Choisy, a traditional Chinese herb, has been shown to be effective in the treatment of diseases associated with inflammation and has been used to treat rheumatic arthritis in China for centuries. However, the underlying mechanism of its anti-inflammatory effect is poorly understood. Hypothesis/Purpose: In this study, we aimed to investigate the anti-inflammatory mechanisms of EtOAc fractions of H. attenuatum Choisy (Ha-EtOAc) on lipopolysaccharide (LPS)-induced RAW264.7 macrophage inflammation and hypothesized that Ha-EtOAc could attenuate inflammation in the colon. Study design: LPS was utilized to induce RAW264.7 cells inflammation. The anti-inflammatory effect of HaEtOAc in RAW264.7 cells was evaluated by measuring the inhibition ratio of nitric oxide (NO) production. Murine ulcerative colitis (UC) was induced by treatment with 2.5% dextran sulfate sodium (DSS). The basic indexes of the mice, including body weight, food intake and hematochezia, were recorded during mice experiments. Methods: The expression levels of pro-inflammatory cytokines, including TNF-alpha, IL-6 and IL-1 beta, were measured by quantitative real-time PCR and western blot. Additionally, the influences of Ha-EtOAc on the NF-kappa B and MAPK signaling pathways were determined by western blot and immunofluorescence assays. In addition, the impact of Ha-EtOAc on gut microbiota of mice with UC was detected by 16S rDNA sequencing. Results: Ha-EtOAc inhibited the LPS-induced production of NO and decreased the release of TNF-alpha, IL-6 and IL-beta in RAW264.7 cells in a dose-dependent manner. In addition, pretreatment with Ha-EtOAc could suppress the nuclear translocation of p65 and the phosphorylation of Erk1/2, p38 and JNK. Ha-EtOAc treatment ameliorated murine UC, as reflected by a reduced body weight loss, improved colon shortening, alleviated mucosal damage and decreased releases of pro-inflammatory cytokines. Furthermore, Ha-EtOAc could modulate the composition of microbial communities. Conclusion: Our results demonstrated that Ha-EtOAc exhibited anti-inflammatory effects mainly by suppressing the NF-kappa B and MAPK pathways, and Ha-EtOAc treatment may be a potent therapy for the treatment of ulcerative colitis.