Investigation of the Underlying Mechanism of Zhibai Dihuang Pill in Treating Osteoporosis by using Network Pharmacology and Molecular Dynamics Simulation

Background With society aging, the rising prevalence of osteoporosis (OP) has enormous social and economic implications. At present, the Zhibai Dihuang pill has been clinically applied in OP treatment and shown significant efficacy, but its underlying mechanism remains unclear. Aims This study was designed to explore the mechanism of the Zhibai Dihuang pill treating OP. Methods In this study, the active ingredients and corresponding targets in the Zhibai Dihuang pill were searched using the TCMSP platform. Based on the mRNA expression data of OP patients in the GEO database, differential expression analysis was conducted by bioinformatics means. By using the differentially expressed genes (DEGs), protein-protein interaction (PPI) network was constructed, and random walk with restart (RWR) analysis based on seed genes intersected from DEGs and drug target genes was conducted. On this basis, the drug-active ingredient-gene interaction network was built. The topological property of the network (degree) was statistically analyzed to find the key therapeutic target AKT1 for the treatment of OP. Molecular docking between AKT1 and the active ingredients was conducted, and according to the affinity score, diosgenin was determined as the key small molecule of the Zhibai Dihuang pill in OP treatment. Based on molecular dynamics simulation and cellular thermal shift assay validation, it was found that diosgenin had a good binding ability with AKT1. Results Cell experiments showed that diosgenin could affect the expression of bone markers and the mineralization of extracellular matrix in mouse osteoblasts by inhibiting the phosphorylation of AKT1, thus achieving the effect of OP treatment. Conclusion Based on network pharmacology, this study clarified the key small molecule compounds in the Zhibai Dihuang pill and their action targets and preliminarily analyzed the molecular mechanism of the Zhibai Dihuang pill treating OP, providing a theoretical basis for the clinical use of the Zhibai Dihuang pill.