Betulinic Acid Reduces Intestinal Inflammation and Enhances Intestinal Tight Junctions by Modulating the PPAR-γ/NF-κB Signaling Pathway in Intestinal Cells and Organoids

Background: Intestinal epithelial barrier (IEB) dysfunction is related to multiple gastrointestinal disorders, notably inflammatory bowel disease (IBD). Betulinic acid (BA), a compound derived from birch bark, has demonstrated potential therapeutic benefits in IBD. Nevertheless, the impact of BA on IEB function has not been fully elucidated. Methods: The current study aimed to explore the potential underlying mechanisms of BA in dextran sodium sulfate (DSS)-induced IBD in mice and co-culture models involving Caco-2/HT29-MTX-E12 cell monolayers or mouse intestinal organoids (IOs) in conjunction with macrophages stimulated by lipopolysaccharide (LPS). Results: In vivo, BA treatment significantly improved body weight and colon length, alleviated disease activity index (DAI) scores, and reduced colonic histopathological injury in IBD mice. In vitro, BA reduced the flux of FITC-dextran; increased the TEER; and decreased the production of IL-6, IL-1β, and TNF-α while increasing IL-10 mRNA levels. Additionally, BA enhanced IEB formation by upregulating ZO-1, occludin (OCLN), and claudin-1 (CLDN1). Molecular docking studies revealed significant docking scores and interactions between BA and PPAR-γ. Moreover, BA significantly upregulated PPAR-γ protein expression, decreased NF-κB and MLC2 phosphorylation, and reduced MLCK protein expression. However, this effect was reversed by GW9662, an effective PPAR-γ antagonist. Conclusions: The findings reveal that BA mitigates IBD by safeguarding the intestinal barrier against dysfunction. This effect may be attributed to its ability to suppress inflammation and enhance the expression of tight junction proteins by modulating the PPAR-γ/NF-κB signaling pathway.

PPAR-γ; betulinic acid; inflammatory bowel disease; intestinal epithelial barrier; intestinal organoids.