Involvement of reactive oxygen species in the preservation injury to cultured liver endothelial cells.
We have previously demonstrated an energy-dependent injury to cultured liver endothelial cells during cold incubation in University of Wisconsin (UW) solution. In the present study, we report experimental evidence for the involvement of reactive oxygen species in this injury: LDH release during 48 h of cold incubation in UW solution was decreased from 40-55% under aerobic conditions to less than 20% under hypoxic conditions or by the presence of KCN (1 mM). Similar protection was achieved by the addition of the spin trap 5,5-dimethyl-1-pyrroline N-oxide, the hydroxyl radical scavenger dimethyl sulfoxide, or the flavonoid silibinin to UW solution under aerobic conditions. Preincubating the cells with the iron chelator deferoxamine even decreased the injury to less than 5%. The residual injury (as observed after longer incubation times) under hypoxic conditions or in cells preincubated with deferoxamine was no longer energy dependent. The amount of thiobarbituric acid-reactive substances markedly increased during cold incubation of the cells in UW solution. This increase was not observed in UW solution to which KCN had been added, i.e., under the conditions of energy depletion. These results suggest that an iron-dependent generation of reactive oxygen species with subsequent lipid peroxidation is involved in the pathogenesis of the injury to cultured liver endothelial cells in cold UW solution.
Rauen
U
U
Institut für Physiologische Chemie, Universitätsklinikum, Essen, Germany.
Elling
B
B
Gizewski
E R
ER
Korth
H G
HG
Sustmann
R
R
de Groot
H
H
D000818:Animals; D015687:Cell Hypoxia / Q000502:physiology; D002478:Cells, Cultured; D015925:Cryopreservation; D003451:Cryoprotective Agents / Q000494:pharmacology; D003497:Cyclic N-Oxides / Q000494:pharmacology; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004727:Endothelium / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D004734:Energy Metabolism / Q000187:drug effects / Q000502:physiology*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D008297:Male; D011190:Potassium Cyanide / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000502:physiology*; D013113:Spin Labels
D003451:Cryoprotective Agents; D003497:Cyclic N-Oxides; D017382:Reactive Oxygen Species; D013113:Spin Labels; C017245:5,5-dimethyl-1-pyrroline-1-oxide; D007770:L-Lactate Dehydrogenase; D011190:Potassium Cyanide; D004121:Dimethyl Sulfoxide