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Journal of Pharmacology and Toxicology
  Year: 2012 | Volume: 7 | Issue: 3 | Page No.: 114-127
DOI: 10.3923/jpt.2012.114.127
Cytotoxic Effects of Multiple N-nitrosamines in Human Liver Cell Line Hep2G: Possible Mechanisms of Action
J. Patterson, J. Boateng, L.T. Walker and M. Verghese

Abstract:
N-nitrosamines are considered human carcinogens and have been found in cured meats, seafood, vegetables, apples, beer, drinking water, waste water, tobacco products and rubber products. Limited studies exist on the effects of low dose exposure to multiple N-nitrosamines compounds. The objectives of this study were to investigate the cellular mechanisms of action by which N-nitrosamines exhibit toxicity resulting in liver tumors and other effects. Hep2G human liver cells (ATCC HTB-37) was obtained from the American Type Culture Collection (ATCC, Manassas, VA). For assay, 5x104 Hep2G cells/well were seeded in a 24 well culture plate and incubated at 37°C and 7% CO2 until development of a monolayer. Cells were incubated with a combination of selected N-nitrosamines at selected concentrations (0, 4, 8, 16, 32, 64 mM) for 12 and 24 h. Lactate dehydrogenase (LDH) release (% cytotoxicity), histone-related DNA fragmentation and detoxification enzymes were determined. After 12 and 24 h incubation with N-nitrosamines,% cytotoxicity in Hep2G cells displayed a dose-dependent relationship at concentrations of 4, 8 and 16 mM. Cytotoxicity peaked at 16 mM for both time periods and then decreased with increasing concentration (64 mM) to 19.46 (12 h) and 55.73 (24 h). Overall, levels of glutathione-S-transferase (GST), glutathione peroxidase (GPx), Glutathione Reductase (GR) and Superoxide Dismutase (SOD) were higher with control compared to N-nitrosamines-treated cells. Histone-related DNA fragmentation was highest in cells treated with 8 mM (24 h). Possible mechanisms of action may be due to lower detoxification enzymes and/or an increase in H2O2 production, leading to cell death.
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24 October, 2016
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