Hypothermia injury/cold-induced apoptosis--evidence of an increase in chelatable iron causing oxidative injury in spite of low O2-/H2O2 formation.
When incubated at 4 degrees C, cultured rat hepatocytes or liver endothelial cells exhibit pronounced injury and, during earlier rewarming, marked apoptosis. Both processes are mediated by reactive oxygen species, and marked protective effects of iron chelators as well as the protection provided by various other antioxidants suggest that hydroxyl radicals, formed by classical Fenton chemistry, are involved. However, when we measured the Fenton chemistry educt hydrogen peroxide and its precursor, the superoxide anion radical, formation of both had markedly decreased and steady-state levels of hydrogen peroxide did not alter during cold incubation of either liver endothelial cells or hepatocytes. Similarly, there was no evidence of an increase in O2-/H2O2 release contributing to cold-induced apoptosis occurring on rewarming. In contrast to the release/level of O2- and H2O2, cellular homeostasis of the transition metal iron is likely to play a key role during cold incubation of cultured hepatocytes: the hepatocellular pool of chelatable iron, measured on a single-cell level using laser scanning microscopy and the fluorescent indicator phen green, increased from 3.1 +/- 2.3 microM (before cold incubation) to 7.7 +/- 2.4 microM within 90 min after initiation of cold incubation. This increase in the cellular chelatable iron pool was reversible on rewarming after short periods of cold incubation. The cold-induced increase in the hepatocellular chelatable iron pool was confirmed using the calcein method. These data suggest that free radical-mediated hypothermia injury/cold-induced apoptosis is primarily evoked by alterations in the cellular iron homeostasis/a rapid increase in the cellular chelatable iron pool and not by increased formation of O2-/H2O2.
Rauen
U
U
Institut für Physiologische Chemie, Universitätsklinikum, D-45122 Essen, Germany. ursula.rauen@uni-essen.de
Petrat
F
F
Li
T
T
De Groot
H
H
D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D003080:Cold Temperature / Q000009:adverse effects*; D004727:Endothelium / Q000166:cytology / Q000502:physiology; D006861:Hydrogen Peroxide; D007501:Iron / Q000378:metabolism*; D007502:Iron Chelating Agents / Q000494:pharmacology; D008099:Liver / Q000166:cytology / Q000502:physiology*; D008297:Male; D018384:Oxidative Stress / Q000502:physiology*; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism; D017694:Rewarming; D013481:Superoxides
D007502:Iron Chelating Agents; D017382:Reactive Oxygen Species; D013481:Superoxides; D006861:Hydrogen Peroxide; D007501:Iron