Production of hydrogen peroxide in situ in cardiac myocytes during hypoxia-reoxygenation as assessed with cerium.
Free radicals have been implicated in myocardial reperfusion injury. Hydrogen peroxide (H(2)O(2)) is one possible source of reactive oxygen intermediates. We studied the formation and toxicity of H(2)O(2) in isolated myocytes during hypoxia-reoxygenation with the use of cerium. This method involves formation of an electron-dense precipitate when H(2)O(2) reacts with cerium chloride (CeCl(3)). Single myocytes were obtained from rat hearts by collagenase digestion. Isolated myocytes were reoxygenated for 15 min after 30 min of hypoxia. The cells were treated with digitonin to increase the permeability of the plasma membrane, and CeCl(3) was added to detect intracellular H(2)O(2) on electron microscopy. In the nonhypoxia control group, the ultrastructure of cells was well preserved, and no dense deposits were found in myocytes. In the hypoxia-reoxygenation group, precipitates, i.e., Ce-H(2)O(2) reaction products, were found inside and along swollen mitochondria, and cell viability was reduced to 72.3% of control. These results indicate that endogenous H(2)O(2) is generated by mitochondria and that its release into the cytosol may lead to myocyte death under pathological situations such as hypoxia-reoxygenation.
Ueda
Ryuji
R
The Third Department of Internal Medicine, Showa University School of Medicine, 1-5-8, Hatanodai, Shinagawa-ku, Tokyo 142-8666, Japan. ueryu@dh.catv.ne.jp
Konno
Noburu
N
Nakatani
Masaki
M
Katagiri
Takashi
T
D000818:Animals; D015687:Cell Hypoxia; D002563:Cerium; D004577:Electron Probe Microanalysis; D006861:Hydrogen Peroxide / Q000378:metabolism*; D066298:In Vitro Techniques; D008297:Male; D008854:Microscopy, Electron; D015428:Myocardial Reperfusion Injury / Q000378:metabolism / Q000473:pathology; D032383:Myocytes, Cardiac / Q000378:metabolism* / Q000648:ultrastructure*; D051381:Rats; D017208:Rats, Wistar
D002563:Cerium; D006861:Hydrogen Peroxide