Cadmium chloride-induced DNA and lysosomal damage in a hepatoma cell line.
Cadmium is a toxic metal and no uniform mechanism of toxicity has so far been proposed. The aim of this study was to investigate the biochemical effects of cadmium chloride in a rat hepatoma cell line (HTC cells) and the cellular events mediating DNA damage. HTC cells were exposed to various concentrations of cadmium chloride for 5 and 8 h and lysosomal damage was assessed with the neutral red assay (NR) and fluorescence microscopy. Mitochondrial integrity was assessed from ATP levels and DNA damage determined with the single cell gel electrophoresis/comet assay. The formation of reactive oxygen species (ROS) was also determined under the same experimental conditions with the dichlorofluorescein assay. Cytotoxicity was assessed with the LDH leakage assay and the levels of glutathione were measured and correlated with the other effects. The results indicate that lysosomal damage occurs at a lower concentration of cadmium chloride (20 microM) than DNA damage (500 microM) in HTC cells. The latter effect was accompanied by an increase of reactive oxygen species without any significant LDH leakage whereas lysosomal damage was significant as determined by the neutral red assay and confirmed with fluorescence microscopy. The effect of CdCl2 on mitochondria and glutathione levels were observed at concentrations or incubation times higher than the ones required to induce lysosomal damage. The data suggest that DNA damage may be due to the formation of reactive oxygen species. It is possible that cadmium induced lysosomal damage is an earlier event than DNA damage and can mediate other cellular events that lead to cell death.
Fotakis
George
G
Department of Pharmacy, Franklin Wilkins Building, 150 Stamford Street, King's College London, London SE1 8WA, UK.
Cemeli
Eduardo
E
Anderson
Diana
D
Timbrell
John A
JA
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D019256:Cadmium Chloride / Q000633:toxicity*; D002105:Cadmium Poisoning / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D020552:Comet Assay; D004249:DNA Damage / Q000187:drug effects*; D004273:DNA, Neoplasm / Q000187:drug effects*; D005978:Glutathione / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008114:Liver Neoplasms, Experimental / Q000378:metabolism* / Q000648:ultrastructure; D008247:Lysosomes / Q000187:drug effects* / Q000648:ultrastructure; D009363:Neoplasm Proteins / Q000378:metabolism; D009499:Neutral Red; D051381:Rats; D017382:Reactive Oxygen Species
D004273:DNA, Neoplasm; D009363:Neoplasm Proteins; D017382:Reactive Oxygen Species; D009499:Neutral Red; D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase; D005978:Glutathione; D019256:Cadmium Chloride