Betulinic acid has demonstrated colitis-suppressing properties; however, its poor water solubility limits clinical application. This study investigates the effects of a nanoscale drug delivery system for betulinic acid on dextran sodium sulfate (DSS)-induced colitis and associated anxiety-like behaviors. Betulinic acid-loaded carboxymethyl chitosan/sodium alginate (BA-CS-SA) microspheres were characterized using scanning electron microscopy and dynamic light scattering. A DSS-induced ulcerative colitis mouse model was established, and the therapeutic effects of BA-CS-SA were evaluated based on colon length, weight, histopathological analysis, qPCR, and Western blot. Anxiety-like behaviors were assessed using the open field test and elevated plus maze test. Inflammatory factor levels were measured using enzyme-linked immunosorbent assays. In vitro drug release assays confirmed the effective release of betulinic acid in artificial colonic fluid. BA-CS-SA treatment significantly improved colon length and weight while reducing spleen length compared to free betulinic acid. BA-CS-SA also led to a greater reduction in histological scores and a more pronounced increase in Claudin-1 levels. Furthermore, BA-CS-SA microspheres improved anxiety-like behaviors, as evidenced by increased time spent in the center and open arms. Mechanistically, BA-CS-SA microspheres more effectively suppressed TNF-α and IL-1β expression in colon and cortex tissues compared to free betulinic acid. BA-CS-SA microspheres provide superior protection against DSS-induced colitis and anxiety-like behaviors compared to free betulinic acid, primarily through enhanced anti-inflammatory effects.
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