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Arsenic (As) and Copper (Cu) Serve as Cofactors to Produce Hepatotoxicity With Mitochondrial Damage, Double Strand DNA Breaks and Disruption of ATM Signaling
Author(s) -
Chinnasamy Thirunavukkarasu,
Sharma Yogeshwar,
Gupta Priya,
Gupta Sanjeev
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.1026.3
Subject(s) - toxicity , dna damage , comet assay , microbiology and biotechnology , biology , acridine orange , oxidative stress , necrosis , chemistry , biochemistry , apoptosis , dna , organic chemistry , genetics
Despite increasing exposure of populations to As the role of cofactors in organ toxicity, e.g., reactive Cu ions, is poorly defined. Since As and Cu ions may combine to increase oxphos events and processes directing mitochondrial and cellular DNA damage, we studied metal toxicity in cultured adult and fetal human hepatocytes with IC50 concentrations of As in MTT assays. In even nontoxic conditions by itself Cu dose‐dependently worsened As toxicity. This worsening included alterations related to oxphos activity with mitochondrial and nuclear perturbations (JC‐1 dye assays, acridine orange nuclear stain, 8‐OHdG stain), double strand DNA breaks (Comet assay, γH2AX stain), and replicative stress including loss of cycling cells and/or arrest in G2/M (Ki‐67staining, flow cytometry). In addition, verapamil‐sensitive side populations denoting stem cells were depleted by As toxicity. Use of cells stably transduced lentivirally to express hATM promoter construct with tdT reporter showed that Cu exacerbated cellular DNA damage by perturbing ATM signaling pathways. To further establish whether Cu accumulation may also worsen As toxicity in vivo , we studied Atp7b −/− mice that model Wilson's disease (WD) and accumulate Cu in liver due to absence of hepatobiliary Cu excretion, along with their healthy littermates. After As was given i.p. for 10d, liver necrosis increased significantly in Atp7b −/− mice (H&E tissue staining, serum hyaluronic acid levels). Moreover, we noted increased ATM pathway‐dependent DNA strand breaks (Comet assay, γH2AX stain). This As and Cu toxicity was ameliorated in human hepatocytes cultured with an anti‐oxidant, indole‐3 propionic acid (IPA), as shown by MTT, JC‐1 and hATM promoter assays. Similarly, IPA decreased As toxicity in Atp7b −/− mice. Conclusions Cu ions markedly exacerbated As toxicity with mitochondrial and ATM pathway‐regulated DNA damage. These findings will help understand how toxic metals may alter physiological processes and contribute additionally in human diseases, e.g., WD with Cu toxicosis.