Berberine Alleviates Intestinal Inflammation by Disrupting Pathological Macrophage–Epithelial Crosstalk in Macrophage–Organoid Co-Culture Model

Berberine (BBR), a benzylisoquinoline alkaloid isolated from Chinese herb <i>Coptis chinensis</i>, has been widely used clinically to treat intestinal infectious diseases. Recently, it has been found to have multiple pharmacological effects, including anti-inflammatory activity and immune effects in inflammatory bowel disease (IBD). However, its exact targets remain to be elucidated. In this study, we used a mouse intestinal organoid-macrophage co-culture model to investigate the anti-inflammatory effects and immune effects of BBR. Our findings demonstrated that lipopolysaccharide (LPS) induced more robust inflammatory responses and epithelium damage in the co-culture system compared to the organoid alone. BBR effectively attenuated inflammation and restored epithelial barrier integrity by suppressing M1 macrophage polarisation and infiltration, alongside upregulating the expression and organisation of tight junction protein zonula occludens-1 (ZO-1). RNA sequencing and proteomic analysis revealed that BBR disrupted organoid-macrophage interaction by inhibiting chemokine (e.g., C-X-C motif chemokine ligand 1 (CXCL1) and macrophage migration inhibitory factor (MIF)) release from epithelial cells, thereby reducing macrophage recruitment. Collectively, our study establishes the organoid-macrophage co-culture system as a more physiologically relevant model for studying epithelial-immune interactions and elucidates the multi-target mechanism of BBR, which concurrently modulates epithelial cells, macrophages, and their crosstalk. These findings lay the foundation for further exploration of the therapeutic potential of BBR in inflammatory bowel disease and the development of targeted therapies that regulate cell interactions.