Moutan cortex ameliorates perimenopausal syndrome by regulating the KISS1R-ERK1/2-GnRH signaling pathway: Evidence from serum pharmacochemistry and network pharmacology.

[ETHNOPHARMACOLOGICAL RELEVANCE] Moutan cortex (known as Mu Danpi in Chinese, or MD) has long been used in traditional Chinese medicine for treating perimenopausal syndrome (MPS) since its first recorded use in the ancient medical classic Shennong Bencao Jing. Hormone replacement therapy remains the conventional treatment for MPS; however, its long-term use is associated with increased risks of breast cancer and endometrial hyperplasia. This safety concern has driven demand for non-hormonal alternatives. Despite the historical efficacy of MD, its molecular mechanism of action, particularly its potential to modulate neuroendocrine pathways without estrogenic side effects, remains unclear.

[AIM OF THE STUDY] To elucidate the molecular mechanisms of MD in the treatment of MPS.

[MATERIALS AND METHODS] The therapeutic effect of MD on MPS was evaluated in bilateral ovariectomy (OVX)-induced perimenopausal rats using body weight, tail temperature, HE staining, and hemorheological tests. The main active components of MD in rat plasma were identified and screened by ultraperformance liquid chromatography-high resolution mass spectrometry, followed by a network pharmacology analysis of the compounds. Component-target-disease and protein-protein interaction networks of MD were constructed using a network pharmacology approach. Subsequently, Gene Ontology (GO) functional enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed. Molecular docking was performed to verify the interactions between the core components and targets of MD, ensuring the reliability of the network pharmacology prediction results. Additionally, ELISA was used to evaluate the effects of MD on GnRH secretion levels both in vivo and in vitro, while Western blot analysis was conducted to validate the expression of key target proteins in the KISS1R-ERK1/2-GnRH signaling pathway.

[RESULTS] MD significantly alleviated MPS symptoms in OVX rats. We identified 9 nine bioactive constituents in rat plasma and predicted 87 potential targets through network pharmacology. GO/and KEGG analyses revealed that these targets were enriched in the GnRH secretion pathway. Molecular docking confirmed that eight components spontaneously bound to key pathway proteins (e.g.., KISS1R, MAP2K1; docking score < -5.0). Furthermore, MD inhibited GnRH secretion in vivo and in vitro and downregulated KISS1R, PKC, p-raf-1, p-MEK1/2, p-ERK1/2, and CaM expression levels in the KISS1R-ERK1/2-GnRH pathway. Notably, the KISS1R inhibitor KP234 abolished the MD's-mediated suppression of GnRH secretion, further validating the significance of this pathway's role in MD'sthe mechanism of MD.

[CONCLUSIONS] MD alleviates MPS by suppressing GnRH secretion via the KISS1R-ERK1/2 pathway, providing mechanistic evidence supporting its use as a non-hormonal therapy. This study integrated serum pharmacochemistry and network pharmacology to elucidate herbal medicine mechanisms.