In view of the current speculation regarding the possible role of reactive oxygen species (ROS) in apoptosis, both under physiological conditions and in response to chemicals that promote their intracellular formation, the present investigation was undertaken to examine whether DNA fragmentation during oxidative stress results from endonuclease activity (apoptosis) or from direct attack by ROS. We report that the incubation of HepG2 cells (a human-derived hepatoma cell line) with the copper(II) complex of 1,10-phenanthroline, CuII(OP)2, results in internucleosomal DNA fragmentation, which is widely recognized as being a hallmark of apoptosis. DNA fragmentation did not occur at low temperature, but activity was restored by the addition of ascorbic acid. It is proposed that DNA fragmentation results from the direct attack of hydroxyl radicals upon DNA. Hydroxyl radicals are produced from oxygen by the redox-cycling of CuII(OP)2, which is supported by metabolic processes at normal temperature. At low temperature ascorbic acid provides an artificial cellular reducing environment, thereby restoring hydroxyl radical formation. These findings were confirmed by the detection of internucleosomal DNA fragmentation following the exposure of isolated chromatin to a biomimetic CuII(OP)2 redox-cycling system. We conclude that DNA laddering, the widely employed hallmark of apoptosis, is not unique to endonuclease activity and may also result from direct attack upon DNA by the hydroxyl radical.
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July 1996
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Research Article|
July 01 1996
Research communication copper-1,10-phenanthroline induces internucleosomal DNA fragmentation in HepG2 cells, resulting from direct oxidation by the hydroxyl radical
Shui Ying TSANG;
Shui Ying TSANG
*Boyd Orr Research Centre§ at the Rowett Research Institute, Division of Biochemical Sciences, Greenburn Road, Bucksburn, Aberdeen AB2 9SB, Scotland, U.K.
†Boyd Orr Research Centre§ at the University of Aberdeen, Department of Chemistry, Meston Walk, Old Aberdeen AB9 2UE, Scotland, U.K.
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Shuk Ching TAM;
Shuk Ching TAM
†Boyd Orr Research Centre§ at the University of Aberdeen, Department of Chemistry, Meston Walk, Old Aberdeen AB9 2UE, Scotland, U.K.
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Ian BREMNER;
Ian BREMNER
*Boyd Orr Research Centre§ at the Rowett Research Institute, Division of Biochemical Sciences, Greenburn Road, Bucksburn, Aberdeen AB2 9SB, Scotland, U.K.
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Mark J. BURKITT
Mark J. BURKITT
‡
*Boyd Orr Research Centre§ at the Rowett Research Institute, Division of Biochemical Sciences, Greenburn Road, Bucksburn, Aberdeen AB2 9SB, Scotland, U.K.
‡To whom correspondence should be addressed.
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Publisher: Portland Press Ltd
Received:
May 03 1996
Accepted:
May 13 1996
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London © 1996
1996
Biochem J (1996) 317 (1): 13–16.
Article history
Received:
May 03 1996
Accepted:
May 13 1996
Citation
Shui Ying TSANG, Shuk Ching TAM, Ian BREMNER, Mark J. BURKITT; Research communication copper-1,10-phenanthroline induces internucleosomal DNA fragmentation in HepG2 cells, resulting from direct oxidation by the hydroxyl radical. Biochem J 1 July 1996; 317 (1): 13–16. doi: https://doi.org/10.1042/bj3170013
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