Exploring the potential pharmacological mechanism of Huiyang Zhitong plaster in treating knee osteoarthritis through network pharmacology, molecular docking, and experimental validation

Abstract

Knee osteoarthritis (KOA) is a common degenerative disease of the joints, and the Huiyang Zhitong Plaster (HYZTP) formulation exhibits distinct therapeutic advantages in the clinical treatment of KOA. However, the underlying pharmacological mechanisms of HYZTP remain unclear. This study aimed to explore the mechanism underlying the therapeutic effect of HYZTP in the treatment of KOA. Methods: The active components and targets of HYZTP were identified from the TCMSP database, and the KOA-related genes were retrieved from the GeneCards database, Therapeutic Target Database (TDD) database, and Online Mendelian Inheritance in Man (OMIM) database. The potential targets related to both HYZTP and KOA were identified using the online Venny tool. Protein-protein interaction (PPI) network analysis was performed using the online STRING data analysis tool, and the network was optimized using Cytoscape. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were additionally performed using the Oebiotech cloud platform. The interactions between the core targets and active components of HYZTP were investigated by molecular docking, and animal experiments were conducted for investigating the mechanism of action of HYZTP in the treatment of KOA. Results: A total of 138 active components of HYZTP, which are associated with its therapeutic effect in the treatment of KOA, were identified by integrating the results of database search. Of the 129 targets, 39 key genes were found to be significantly enriched in the TNF, IL-17, HIF-1, and Toll-like receptor (TLR) signaling pathways. The results of molecular docking revealed that the key components of HYZTP, namely, beta-sitosterol and stigmasterol, exhibited a favorable binding potential for the targets in the TNF signaling pathway, including TNF-α, IL-1β, and AKT1. Further histopathological analysis revealed that HYZTP repaired cartilage damage in a rabbit model of KOA. The results of western blotting revealed that the expression levels of TNF-α, RIP1, CASP3, and BAX proteins were downregulated in the knee joint cartilage of the HYZTP group, compared to those of the model group. Conclusion: HYZTP likely hinders the progression of KOA by inhibiting the TNF signaling pathway and suppressing cellular apoptosis. The study provides novel insights into the underlying pharmacological mechanism of HYZTP in the treatment of KOA, providing a foundation for further research into the clinical applications of HYZTP.