Rubiadin, as a key metabolite of the Bushen Huoxue formula, promotes apoptosis of endometrial stromal cells and improves intrauterine adhesions by activating the AMPK/p53/p21 pathway

Background The Bushen Huoxue formula (BSHX) is a classic traditional Chinese medicine prescription for treating gynecological disorders. However, its specific metabolites and mechanisms against IUA remain to be elucidated. Aim This study aimed to identify the key metabolite of BSHX and to investigate its therapeutic effects and underlying mechanism against IUA, both in vitro and in vivo . Methods Network pharmacology and molecular dynamics simulation were utilized to screen the metabolites of BSHX. In vitro , cells were treated with different concentrations of Rubiadin with or without the AMPK inhibitor Dorsomorphin. Cell viability, apoptosis, migration, invasion, and pathway activity were assessed. In vivo , the IUA rat model was treated with Rubiadin and Dorsomorphin, and histopathology, fibrosis, inflammation, pregnancy outcomes, and associated signaling pathways were evaluated. Results In IUA clinical tissues, the AMPK/p53/p21 pathway was significantly inhibited. Rubiadin dose-dependently activated the AMPK/p53/p21 pathway, promoted endometrial stromal cell apoptosis, and inhibited cell migration, invasion, and collagen synthesis in vitro . These effects were reversed by Dorsomorphin. In IUA rats, Rubiadin treatment activated the AMPK/p53/p21 pathway, reduced endometrial fibrosis and inflammation, and significantly improved pregnancy rates. The AMPK inhibitor attenuated these therapeutic benefits. Conclusion Rubiadin, a key metabolite of the Bushen Huoxue formula, ameliorates intrauterine adhesions by activating the AMPK/p53/p21 pathway, which promotes the apoptosis of endometrial stromal cells and restores endometrial function. This study provides a pharmacological basis for using Rubiadin as a potential therapeutic agent for IUA.