DNA damage induced by tumour necrosis factor-alpha in L929 cells is mediated by mitochondrial oxygen radical formation.
Treatment of L929 cells with tumour necrosis factor-alpha (TNF-alpha) plus actinomycin D induced DNA damage (indicated by the appearance of a sub-G1 peak due to extracellular leakage of low molecular weight DNA following DNA fragmentation) before significant cell lysis occurred. The DNA damage occurred in parallel with a decrease of the intracellular total glutathione content and an increase of intracellular reactive oxygen intermediates (ROI), as indicated by increased dihydrorhodamine 123 oxidation. Because the inhibition of mitochondrial respiration suppressed the increase of dihydrorhodamine 123 oxidation and DNA damage as well as the decrease in the total glutathione content, it was suggested that increased mitochondrial formation of ROI was responsible for DNA damage after TNF treatment. Deferoxamine (a ferric iron chelator) and dithiothreitol (a sulfhydryl reagent) both prevented DNA damage and cell killing, indicate that hydroxyl radicals generated from O2- and H2O2 produced by the mitochondria in a process catalysed by iron contributed to DNA damage and that this pathway may be involved in TNF-alpha-induced cytotoxicity. An inhibitor of poly(ADP)-ribose polymerase (3-aminobenzamide), worsened DNA damage, but was protective against cell lysis, suggesting that DNA repair subsequent to injury was more important than DNA damage per se in development of TNF-alpha cytotoxicity.
Shoji
Y
Y
Department of Emergency and Critical Care Medicine, Kansai Medical University, Osaka, Japan.
Uedono
Y
Y
Ishikura
H
H
Takeyama
N
N
Tanaka
T
T
D002453:Cell Cycle / Q000187:drug effects; D002478:Cells, Cultured; D003602:Cytotoxicity, Immunologic; D004249:DNA Damage / Q000502:physiology*; D003609:Dactinomycin / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D010084:Oxidation-Reduction; D017382:Reactive Oxygen Species / Q000378:metabolism*; D011994:Recombinant Proteins / Q000494:pharmacology; D012235:Rhodamines / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D017382:Reactive Oxygen Species; D011994:Recombinant Proteins; D012235:Rhodamines; D014409:Tumor Necrosis Factor-alpha; C058319:dihydrorhodamine 123; D003609:Dactinomycin; D007770:L-Lactate Dehydrogenase; D005978:Glutathione