Adriamycin-enhanced membrane lipid peroxidation in isolated rat nuclei

Isolated rat liver nuclei enzymatically activated Adriamycin to the electron spin resonance-detectable semiquinone free radical in the presence of reduced nicotinamide adenine dinucleotide phosphate (NADPH). This process resulted in the enhancement of oxyradical-mediated peroxidation of the nuclear envelope membrane unsaturated phospholipids, measured as malonaldehyde equivalents by the thiobarbituric acid method. Peroxidation required the inclusion of NADPH and catalytically active protein, presumably NADPH:cytochrome P-450 reductase, and was enhanced more than 5-fold by Adriamycin. It was observed that Adriamycin-stimulated nuclear membrane peroxidation was diminished by superoxide dismutase, reduced glutathione, the hydroxyl radical scavenger, 1,3-dimethylurea, and by the metal ion chelator, diethylenetriaminepentaacetic acid, indicating that multiple species of reactive oxygen and trace amounts of metal ions (iron) were required in the peroxidation reaction. The generation of superoxide and hydroxyl radicals was confirmed by 5,5-dimethyl-1-pyrroline-N-oxide spin trap electron spin resonance spectroscopy. Calf thymus DNA added to incubations containing nuclei and NADPH caused a pronounced concentration-dependent inhibition of Adriamycin-stimulated lipid peroxidation. It was found that nuclei incubated with Adriamycin (300 microM) accumulated 128 nmol of the drug per mg of nuclear protein within 1 h, apparently because the Adriamycin was internalized and bound to the DNA and nuclear protein. These results suggest that some of the toxic effects of Adriamycin observed in the nucleus could result, directly or indirectly, through the peroxidation of the unsaturated lipids of the nuclear membrane.