Network toxicology, molecular docking technology, and experimental verification revealed the mechanism of cantharidin-induced testicular injury in mice

Abstract

As a protein kinase inhibitor, cantharidin (CTD) exhibits antitumor activities. However, CTD is highly toxic, thereby limiting clinical applications. Moreover, relatively few studies have investigated CTD-induced reproductive toxicity, thus the underlying mechanism remains unclear. In this study, the toxic effects of CTD on mouse testis were confirmed in vivo and the potential mechanism was predicted by network toxicology (NT) and molecular docking technology. Proteins involved in the signaling pathways and core targets were verified. The results showed that different concentrations of CTD induced weight loss increased the testicular coefficient, and caused obvious pathological damage to testicular cells. The NT results showed that the main targets of CTD-induced testicular injury (TI) included AKT1, Caspase 3, Bcl-2, and Bax. The results of pathway enrichment analysis showed that CTD-induced TI was closely related to apoptosis and the PI3K/AKT and HIF-1 signaling pathways. Molecular docking methods confirmed high affinity between CTD and key targets. Western blot analysis showed that CTD inhibited expression of PI3K, AKT, and the anti-apoptotic protein Bcl-2, while promoting expression of the pro-apoptotic proteins Bax and Caspase 3. These results suggest that CTD-induced TI involves multiple targets and pathways, and the underlying mechanism was associated with inhibition of the apoptosis-related PI3K/AKT signaling pathway.