Abstract
Aim:
Tetrandrine, an alkaloid with a remarkable pharmacological profile, induces oxidative stress and mitochondrial dysfunction in hepatocytes; however, mitochondria are not the direct target of tetrandrine, which prompts us to elucidate the role of oxidative stress in tetrandrine-induced mitochondrial dysfunction and the sources of oxidative stress.
Methods:
Rat primary hepatocytes were isolated by two-step collagenase perfusion. Mitochondrial function was evaluated by analyzing ATP content, mitochondrial membrane potential (MMP) and the mitochondrial permeability transition. The oxidative stress was evaluated by examining changes in the levels of reactive oxygen species (ROS) and glutathione (GSH).
Results:
ROS scavengers largely attenuated the cytotoxicity induced by tetrandrine in rat hepatocytes, indicating the important role of ROS in the hepatotoxicity of tetrandrine. Of the multiple ROS inhibitors that were tested, only inhibitors of CYP450 (SKF-525A and others) reduced the ROS levels and ameliorated the depletion of GSH. Mitochondrial function assays showed that the mitochondrial permeability transition (MPT) induced by tetrandrine was inhibited by SKF-525A and vitamin C (VC), both of which also rescued the depletion of ATP levels and the mitochondrial membrane potential. Upon inhibiting specific CYP450 isoforms, we observed that the inhibitors of CYP2D, CYP2C, and CYP2E1 attenuated the ATP depletion that occurred following tetrandrine exposure, whereas the inhibitors of CYP2D and CYP2E1 reduced the ROS induced by tetrandrine. Overexpression of CYP2E1 enhanced the tetrandrine-induced cytotoxicity.
Conclusion:
We demonstrated that CYP450 plays an important role in the mitochondrial dysfunction induced by the administration of tetrandrine. ROS generated by CYP450, especially CYP2E1, may contribute to the mitochondrial dysfunction induced by tetrandrine.
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Acknowledgements
This work was supported by National Natural Science Foundation of China (No 30271558); Key projects of National Science and Technology Pillar Program (2012ZX09301001-006 and 2012zx09302003) and the Public Service Platform Project of Shanghai Science and Technology Committee (11DZ2292500).
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Supplementary information
Supplementary information Figure S1
Effect of the inhibitors of ROS sources on ROS production by tetrandrine. (JPG 456 kb)
Supplementary information Figure S2
Tetrandrine-induced mitochondrial dysfunction is sensitive to CsA. (JPG 141 kb)
Supplementary information Figure S3
Effect of the Cyp450 isoform inhibitors on the cytotoxicity and ROS production induced by tetrandrine. (JPG 821 kb)
Supplementary information Figure S4
Protective effect of tetrandrine on oxidative stress induced by various oxidative factors in isolated rat liver mitochondria. (JPG 553 kb)
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Methods of supplementary results (DOC 32 kb)
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Qi, Xm., Miao, Ll., Cai, Y. et al. ROS generated by CYP450, especially CYP2E1, mediate mitochondrial dysfunction induced by tetrandrine in rat hepatocytes. Acta Pharmacol Sin 34, 1229–1236 (2013). https://doi.org/10.1038/aps.2013.62
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DOI: https://doi.org/10.1038/aps.2013.62
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