Objective To study and analyse the main components of Tongluo Muzu Powder transdermal absorption solution, as well as to investigate its mechanism of action in the treatment of osteoarthritis of the knee.

Methods The transdermal absorption solution of Tongluo Muzu Powder was extracted by Franz vertical diffusion cell, and its components were analyzed by UHPLC-MS/MS. Then TCMSP, Swiss target prediction, Gene Cards and other databases were introduced to predict the possible targets of the effective components of transdermal absorption solution for the prevention and treatment of knee osteoarthritis, and Cytoscape software was used to construct the "drug-component-disease-target" visual network, and then the protein interaction network was obtained through the string database to find the core targets. Finally, AutoDock and PyMOL were used to verify the molecular docking of main active ingredients and targets, and the gene ontology (GO) function analysis of core genes and the enrichment analysis of Kyoto Encyclopedia of genes and genomes (KEGG) pathway were conducted. Primary mouse chondrocytes were extracted, and the predicted targets of network pharmacology were verified by ELISA, Western blot, qPCR, etc.

RESULTS A total of 48 effective components of the transdermal absorption solution of Tongluo Muzu Powder and 88 possible targets for the treatment of knee osteoarthritis were screened out. The results of enrichment analysis showed that it was mainly involved in the process of apoptosis and oxidative stress. Molecular docking suggested that the main active ingredients and targets had good binding ability. The results showed that the lyophilized powder of the transdermal absorption solution of Tongluo Muzu Powder could reduce the concentration of inflammatory factors in the supernatant of chondrocytes after LPS intervention in a dose-dependent manner (P < 0.05, P < 0.01). The protein expression of p-PI3K/PI3K, p-AKT/AKT, MMP13 and p53 was decreased, and the protein expression of collagen Ⅱ was increased P < 0. 05 P < 0. 01. At the same time the mRNA expression of MMP13 and p53 was decreased and the mRNA expression of collagen Ⅱ was increased P < 0. 05, P < 0.01). In addition, it reduced the fluorescence intensity of TUNEL staining in cells (P < 0.01).

CONCLUSION By combining UHPLC-MS/MS technology with network pharmacology, the main active ingredients of Tongluo Muzu Powder transdermal absorption solution are preliminarily understood, and its potential mechanism of action in the treatment of knee osteoarthritis are predicted.