Geniposide Alleviates Inflammatory Bowel Disease by Regulating Intestinal Flora and Arginine Metabolism and Inhibiting the NF-κB Pathway Through Targeting Anxa5.

BACKGROUND

Inflammatory bowel disease (IBD), including Crohn's and ulcerative colitis (UC), is a chronic gut inflammation thought to be caused by gut microbial causes and immune system dysfunction. Geniposide (GEN) is a natural compound shown to prevent IBD; however, its mechanism of modifying the gut microbiome, regulating arginine metabolism, and inhibiting the nuclear factor kappa B (NF-κB) pathway by targeting Annexin A5 (Anxa5) remains unexplored.

METHODS

This study examined the regulation of intestinal immunity, gut microbiota, metabolites, and associated activities and pathways, as well as the NF-κB pathway by GEN in a BALB/c mouse model of IBD. The mouse macrophage cell line (RAW264.7) was further utilized to investigate the role of GEN on the macrophage-arginine metabolism and the Anxa5 axis. qRT-PCR, Western blotting, hematoxylin and eosin (H&E), immunohistochemistry (IHC), immunofluorescence (IF), fecal 16S rDNA sequencing, and UHPLC/Q-TOF-MS were used to evaluate the treatment effect of GEN. Drug target sequencing and small interfering RNA (siRNA) were used to establish the target molecule of GEN.

RESULTS

GEN therapy improved colon and spleen tissues, decreased the disease activity index, helped mice maintain their weight, elevated anti-inflammatory cytokines and tight junction proteins, and decreased proinflammatory cytokines. GEN modulated the quantity of dysfunctional metabolites and enhanced the structure and diversity of the gut microbial community. GEN restored the underpopulated genera Enterorhabdus, Rikenella, Anaerotruncus, and Alistipes, regulating arginine metabolism mediated by G-guanidinobutyrate in RAW264.7 cells and the gut mucosa to protect the colon. GEN targeted Anxa5, enhancing its expression in both animal models and RAW264.7 cells. Following Anxa5 knockdown, cyclooxygenase-2 (COX-2) expression increased, along with the activation of NF-κB pathway-related proteins in RAW264.7 cells. GEN reduced the levels of NF-κB pathway-related proteins in both RAW264.7 cells and animal models.

CONCLUSION

GEN mitigated colitis by regulating the intestinal microbiota and arginine metabolism and inhibiting the NF-κB pathway via targeting Anxa5.