Unimpaired metabolism of pyridine dinucleotides and adenylates in Chinese hamster ovary cells during oxidative stress elicited by cytotoxic doses of copper-putrescine-pyridine.
Copper-putrescine-pyridine (Cu-PuPy) effectively dismutates superoxide but is also known to produce H2O2 in a redox cycle with glutathione. The treatment of Chinese hamster ovary (CHO) cells with 0.2 mM Cu-PuPy reduced clonogenic survival to 10(-3) in 50 min and caused significant oxidation and depletion of glutathione and continuous accumulation of protein-glutathione mixed disulfides. Remarkably, other important functional parameters of cell metabolism were not impaired: adenylate pool size, adenylate energy charge and the redox ratios of NADP(H) and NAD(H) remained constant. Moreover, within 200 min the pool size of NADP(H) increased linearly by a factor of four at the expense of the NAD(H) pool, resulting in an 8-fold increase in the ratio of NADPH to glutathione disulfide. Also, Cu-PuPy led to a dose-dependent, persistent inactivation of glutathione reductase, which could be reversed by copper chelators. In contrast to Cu-PuPy, glucose oxidase-generated H2O2 induced oxidation and loss of pyridine dinucleotides, depletion of the adenylate pool and deterioration of the energy charge. Oxidation and depletion of bulk glutathione were comparable to a Cu-PuPy treatment, but formation of protein-glutathione mixed disulfides was significantly less pronounced and reversible. The data indicate that the critical factor in Cu-PuPy cytotoxicity is not its function as catalyst of glutathione oxidation and H2O2 generation, but essentially its disruption of antioxidative cellular defence by inactivation of glutathione reductase. The data further suggest that Cu-PuPy inhibits ADP-ribosylation. This would explain why pyridine dinucleotide and adenylate pools are unaffected, and may be an essential prerequisite for the observation that cells, albeit sublethally damaged and denuded of their antioxidative defence, may be rescued by extending Cu-PuPy treatment.
Strahlenbiologisches Institut, Ludwig-Maximilians-Universität, München, Germany.
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Nagele
D000227:Adenine Nucleotides / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D005949:Glucose Oxidase / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D005980:Glutathione Reductase / Q000037:antagonists & inhibitors / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism; D009243:NAD / Q000378:metabolism*; D009942:Organometallic Compounds / Q000633:toxicity*; D018384:Oxidative Stress / Q000187:drug effects*; D012545:Schiff Bases / Q000633:toxicity*
D000227:Adenine Nucleotides; D009942:Organometallic Compounds; D012545:Schiff Bases; D009243:NAD; C064004:N,N'-bis(2-pyridylmethylene)-1,4-butanediamine (N,N',N'',N''')-Cu(II)diperchlorate; D006861:Hydrogen Peroxide; D005949:Glucose Oxidase; D005980:Glutathione Reductase; D005978:Glutathione