Total saponins of Panax japonicus alleviate adjuvant-induced arthritis in rats by regulating macrophage polarization and intestinal microbiota.

Background: Heightened glycolysis-driven activation of pro-inflammatory M1 macrophages is a characteristic of rheumatoid arthritis (RA). Dysbiosis of the intestinal microbiota is considered a trigger for the immune response in RA. Panax japonicus is used in ethnomedicine to treat RA, but its molecular mechanisms regarding macrophage polarization and the intestinal microbiota remain to be explored. Methods: In vitro and in vivo models were established using lipopolysaccharide (LPS)-induced RAW 264.7 macrophage cells and adjuvant-induced arthritis (AIA) rats, and treated with total saponins from Panax japonicus (TSPJ). Inflammatory cytokines were quantified by enzyme-linked immunosorbent assay (ELISA). Glucose, lactate, and LPS were measured by biochemical kits. The M1/M2a ratio in RAW 264.7 cells was assessed by flow cytometry. Joint bone erosion was evaluated by micro-CT. Immunohistochemistry and western blotting were used to evaluate glycolytic regulators in rat joints. Colonic contents and mucosa were analyzed by 16S rRNA sequencing. Results: TSPJ inhibited inflammatory response, lactate production, and glucose consumption in M1 macrophages, and restored the M2a/M1 balance. In vivo, TSPJ alleviated joint swelling in AIA rats, reduced serum inflammatory cytokines, lactate, and LPS levels, while increasing anti-inflammatory cytokine levels. The protein expressions of hypoxia-inducible factor-1α (HIF-1α), glucose transporter 1 (GLUT1), hexokina se 2 (HK2), and lactate dehydrogenase A (LDHA) in the ankle joints were down-regulated after TSPJ intervention. Furthermore, TSPJ restored microbiota diversity and community structure, and enriched probiotic abundances like Eubacterium_coprostanoligenes_group and Christensenella. Conclusion: TSPJ alleviates arthritis by inhibiting glycolysis pathway in M1 macrophages, modulating colon microbiota, and promoting the abundance of mucosa-associated probiotics.