Network analysis and experimental approach to investigate the potential therapeutic mechanism of Zishen Yutai Pills on premature ovarian insufficiency

As society continues to develop, women are more at risk of gonadotoxic substance exposure. Consequently, the incidence of premature ovarian insufficiency (POI) has increased significantly in the past decades. Hormone replacement therapy (HRT) is recommended as the standard treatment to relieve hypoestrogenic symptoms; however, its potential side effects and contraindications have drawn widespread controversy and concern. As such, the Chinese medicine Zishen Yutai Pill (ZSYTP) commonly used for treating miscarriage and menoxenia, is a highly promising alternative drug candidate against POI, however its therapeutic mechanism has not been completely elucidated. In this study, to systematically analyze the potential targets of ZSYTP on POI, we performed network pharmacology analysis and molecular docking to predict critical target genes. A CTX-induced POI mouse model was employed for further validate the accuracy of our prediction by histopathological observation, immunohistochemical staining and transcriptome sequencing. Besides, to study the difference between ZSYTP and HRT, progynova was also applied in this study. Our results suggested that ZSYTP might have a better therapeutic outcome compared with progynova. Results from RNA-Seq revealed that PI3K-Akt, Hippo, AGE-RAGE, Rap1 signaling pathway and regulation of inflammation, immune and lipid metabolism might mediate the protective mechanism of ZSYTP against POI, which was fairly different from progynova. Collectively, this study indicates that ZSYTP could be a highly promising alternative as a non-HRT-based therapy for POI. Its mechanism involves multiple signaling pathways, alleviating ovarian apoptosis and recovering AMH and FSH level. However, the discrepancy between different research techniques highlight the necessity of further experimental verification from other aspects such as translation and posttranslational modification.