Evidence suggests that auranofin (AF) exhibits anticancer activity by inhibiting thioredoxin reductase (TrxR). Here, in this study, we have investigated the synergistic effects of AF and morin and their mechanism for the anticancer effects focusing on apoptosis in Hep3B human hepatocellular carcinoma cells. We assessed the anticancer activities by annexin V/PI double staining, caspase, and TrxR activity assay. Morin enhances the inhibitory effects on TrxR activity of AF as well as reducing cell viability. Annexin V/PI double staining revealed that morin/AF cotreatment induced apoptotic cell death. Morin enhances AF-induced mitochondrial membrane potential (ΔΨm) loss and cytochrome c release. Further, morin/AF cotreatment upregulated death receptor DR4/DR5, modulated Bcl-2 family members (upregulation of Bax and downregulation of Bcl-2), and activated caspase-3, -8, and -9. Morin also enhances AF-induced reactive oxygen species (ROS) generation. The anticancer effects results from caspase-dependent apoptosis, which was triggered via extrinsic pathway by upregulating TRAIL receptors (DR4/DR5) and enhanced via intrinsic pathway by modulating Bcl-2 and inhibitor of apoptosis protein family members. These are related to ROS generation. In conclusion, this study provides evidence that morin can enhance the anticancer activity of AF in Hep3B human hepatocellular carcinoma cells, indicating that its combination could be an alternative treatment strategy for the hepatocellular carcinoma.
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