Oxidative stress signalling: a potential mediator of tumour necrosis factor alpha-induced genomic instability in primary vascular endothelial cells.
Studying the potential role of tumour necrosis factor (TNF)alpha in the initiation of genomic instability is necessary to understand whether TNFalpha can serve as a signalling mediator of radiation-induced genomic instability in non-irradiated bystander cells. In this study, we examined whether TNFalpha could initiate processes through oxidative stress signalling that lead to DNA damage and genomic instability in primary vascular endothelium. In these cells, low linear energy transfer (LET) radiation (0.1-2 Gy) induced the secretion of TNFalpha into the culture medium. When added ectopically, TNFalpha at concentrations ranging from 0.1 ng ml(-1) to 10 ng ml(-1) increased (twofold to threefold) intracellular oxidative stress. Next, to examine whether TNFalpha induces genetic damage, cells were treated with TNFalpha for 5 h and analysed immediately using the single cell gel electrophoresis assay or after 3 days, 12 days and 20 days using solid stain chromosomal analysis. Cells exposed to 0.1 Gy, 1 Gy or 2 Gy or treated with 100 microM H2O2 were used as positive controls. The results showed that TNFalpha as low as 0.1 ng ml(-1) could initiate increased DNA damage compared with untreated controls. When examined in the progeny cells after several generations, the chromosomal instability appeared to be carried over even after day 12 and day 20. The increased genetic damage is inhibited in cells that are pre-incubated with the antioxidant enzyme catalase, the antioxidant N-acetyl-L-cysteine or the metal chelator pyrrolidine dithiocarbamate. These results clearly indicate that TNFalpha at concentrations at which no cytotoxicity is observed could induce genetic damage through free radical generation, which could, in turn, lead to the delayed events associated with genomic instability.
Natarajan
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M
Department of Radiation Oncology, University of Texas Health Science Center, San Antonio, TX 78229, USA. Natarajan@uthscsa.edu
Gibbons
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Mohan
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Moore
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Kadhim
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D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002869:Chromosome Aberrations; D004249:DNA Damage / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000187:drug effects / Q000528:radiation effects; D004730:Endothelium, Vascular / Q000166:cytology* / Q000378:metabolism / Q000528:radiation effects; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D042822:Genomic Instability / Q000502:physiology*; D006801:Humans; D018384:Oxidative Stress / Q000187:drug effects / Q000235:genetics*; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000494:pharmacology*
D000975:Antioxidants; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha