Abstract
Hepatotoxicity of the antidiabetic drug metformin has been reported, but the underlying mechanisms remain unclear. We here investigated the effect of metformin in immune-mediated liver damage. While not hepatotoxic alone, metformin (200 mg/kg) aggravated concanavalin A (Con A, 12 mg/kg)-induced hepatitis, an experimental model of T cell-mediated liver injury, in both relatively resistant BALB/c and highly susceptible C57Bl/6 mice. Metformin + Con A-treated mice had elevated serum levels of pro-inflammatory cytokines TNF-α and IFN-γ, accompanied by a massive mononuclear cell infiltration in the liver. This was associated with the higher numbers of CD4+ T cells producing TNF-α, IFN-γ and IL-17, CD4+ T cells expressing chemokine receptor CXCR3 and activation marker CD27, CD4+CD62L−CCR7− and CD8+CD62L−CCR7− effector memory cells, IFN-γ producing NK cells, IL-4 and IL-17 producing NKT cells and IL-12 producing macrophages/dendritic cells. The percentage of CD4+CXCR3+Tbet+IL-10+ and CD4+CD69+CD25− regulatory T cells was reduced. Metformin stimulated inducible nitric oxide synthase (iNOS) expression in the liver and spleen, and genetic deletion of iNOS attenuated the hepatotoxicity of metformin. Metformin increased the autophagic light chain 3 conversion and mRNA expression of important autophagy-inducing (beclin-1, Atg5 and GABARAP) and pro-apoptotic (p21, p27, Puma, Noxa, Bax, Bad, Bak1, Bim and Apaf1), but not anti-apoptotic molecules (Bcl-xL, survivin and XIAP), which correlated with the apoptotic caspase-3/PARP cleavage in the liver. The autophagy inhibitor chloroquine (20 mg/kg) prevented liver injury and apoptotic changes induced by metformin. Therefore, metformin aggravates immune-mediated hepatitis by promoting autophagy and activation of immune cells, affecting effector, as well as liver-specific regulatory T cells and iNOS expression.
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Acknowledgments
This study was supported by Grants ON175069, ON175103 and III41025 from the Ministry of Education, Science and Technology, Republic of Serbia, and MP01/12 from The Faculty of Medical Sciences University of Kragujevac, Serbia. The authors thank Mr. Milan Milojevic for technical support, Dr. Mirjana Sumarac-Dumanovic (Clinic for Endocrinology, Diabetes and Diseases of Metabolism, School of Medicine, University of Belgrade) for useful discussion and Dr. Zeljka Radulovic (Galenika a.d., Belgrade) for providing metformin.
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Volarevic, V., Misirkic, M., Vucicevic, L. et al. Metformin aggravates immune-mediated liver injury in mice. Arch Toxicol 89, 437–450 (2015). https://doi.org/10.1007/s00204-014-1263-1
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DOI: https://doi.org/10.1007/s00204-014-1263-1