Original contribution
Detection of elevated reactive oxygen species level in cultured rat hepatocytes treated with aflatoxin B1

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Abstract

Accumulating evidence demonstrates that oxidative damage is one of the underlying mechanisms to the cytotoxicity and carcinogenicity of AFB1. The main objective of this study is to show that AFB, increases reactive oxygen species (ROS) formation in hepatocytes. The ROS level was detected using a fluorescence probe, 2′,7′-dichlorofluorescin diacetate (DCFH-DA), which could be converted to highly fluorescent dichlorofluorescein (DCF) with the presence of intracellular ROS. It was found that AFB1 exposure significantly enhanced DCF fluorescence formation in cultured rat hepatocytes. A dose-response of AFB1 was also observed within the range of 10 nM to 1000 nM. Catalase (CAT) was able to completely prevent the increase of DCF fluorescence in AFB1-treated cells in a dose-dependent manner (from 500 to 2000 U/ml). Moreover, the significant inhibitory effects of desferrioxamine (DFO) and dimethyl sulfoxide (DMSO) on DCF fluorescence formation were also observed in both control and AFB1-treated hepatocytes. Therefore, results from the present study provide in vitro evidence indicating the generation of ROS in cultured rat hepatocytes caused by AFB1 exposure. It is postulated that the metabolic process of AFB1 by cytochrome P450 might be the possible source of the elevated ROS level in AFB1-treated hepatocytes. The enhanced level of ROS may be responsible for the oxidative damage caused by AFB1, which may ultimately contribute to the cytotoxic and carcinogenic effects of AFB1.

References (39)

  • G. Decad et al.

    Maintenance of cytochrome P-450 and metabolism of aflatoxin B1 in primary hepatocyte cultures

    Biochem. Biophys. Res. Commun.

    (1977)
  • G.S. Jennings et al.

    Metabolism and cytotoxicity of aflatoxin B1 in cultured rat hepatocytes and nonparenchymal cells: Implications for tumorigenesis

    Toxicol. Appl. Pharmacol.

    (1994)
  • J.S. Beckman et al.

    Superoxide dismutase and CAT conjugated to polyethylene glycol increases endothelial enzyme activity and oxidant resistance

    J. Biol. Chem.

    (1988)
  • I. Morel et al.

    An tioxidant and free radical scavenging activities of the iron chelators pyoverdin and hydroxypyrid-4-ones in iron-loaded hepatocyte cultures: Comparison of their mechanism of protection with that of desferrioxamine

    Free Radic. Biol. Med.

    (1992)
  • A.I. Cederbaum et al.

    The effects of dimethylsulfoxide and other hydroxyl radical scavengers on the oxidation of ethanol by rat liver microsomes

    Biochem. Biophys. Res. Commun.

    (1977)
  • P. Villa et al.

    Mechanism of maintenance of liver-specific functions by DMSO in cultured rat hepatocytes

    Exp. Cell Res.

    (1991)
  • J.L. Farber et al.

    Biology of disease: Mechanisms of cell injury by activated oxygen species

    Lab. Invest.

    (1990)
  • J.P. Kehrer

    Free radicals as mediators of tissue injury and disease

    Crit. Rev. Toxicol.

    (1993)
  • B.N. Ames et al.

    Oxidants, antioxidants, and the degenerative diseases of aging

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