Oxygen radical-mediated lipid peroxidation and inhibition of Ca2+-ATPase activity of cardiac sarcoplasmic reticulum.
Oxygen radicals have been implicated as important mediators of myocardial ischemic and reperfusion injury. A major product of oxygen radical formation is the highly reactive hydroxyl radical via a biological Fenton reaction. The sarcoplasmic reticulum is one of the major target organelles injured by this process. Using a oxygen radical generating system consisting of dihydroxyfumarate and Fe3+-ADP, we studied lipid peroxidation and Ca2+-ATPase of cardiac sarcoplasmic reticulum. Incubation of sarcoplasmic reticulum with dihydroxyfumarate plus Fe3+-ADP significantly inhibited enzyme activity. Addition of superoxide dismutase, superoxide dismutase plus catalase (15 micrograms/ml) or iron chelator, deferoxamine (1.25-1000 microM) protected Ca2+-ATPase activity. Time course studies showed that this system inhibited enzyme activity in 7.5 to 10 min. Similar exposure of sarcoplasmic reticulum to dihydroxyfumarate plus Fe3+-ADP stimulated malondialdehyde formation. This effect was inhibited by superoxide dismutase, catalase, singlet oxygen, and hydroxyl radical scavengers. EPR spin-trapping with 5,5-dimethyl-1-pyrroline-N-oxide verified production of the hydroxyl radical. The combination of dihydroxyfumarate and Fe3+-ADP resulted in a spectrum of hydroxyl radical spin trap adduct, which was abolished by ethanol, catalase, mannitol, and superoxide dismutase. The results demonstrate the role of oxygen radicals in causing inactivation of Ca2+-ATPase and inhibition of lipid peroxidation of the sarcoplasmic reticulum which could possibly be one of the important mechanisms of oxygen radical-mediated myocardial injury.
Kukreja
R C
RC
Department of Medicine, Medical College of Virginia, Virginia Commonwealth University, Richmond 23298.
Okabe
E
E
Schrier
G M
GM
Hess
M L
ML
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000818:Animals; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors*; D004285:Dogs; D004578:Electron Spin Resonance Spectroscopy; D005290:Ferric Compounds / Q000494:pharmacology; D005650:Fumarates / Q000494:pharmacology; D066298:In Vitro Techniques; D008054:Lipid Peroxides / Q000378:metabolism*; D009206:Myocardium / Q000201:enzymology* / Q000378:metabolism; D010100:Oxygen / Q000494:pharmacology*; D012519:Sarcoplasmic Reticulum / Q000201:enzymology* / Q000378:metabolism
D005290:Ferric Compounds; D005650:Fumarates; D008054:Lipid Peroxides; C005998:dihydroxyfumarate; D000244:Adenosine Diphosphate; D000252:Calcium-Transporting ATPases; D010100:Oxygen