Iron toxicity in organotypic cultures of hippocampal slices: role of reactive oxygen species.
Free iron has been assumed to potentiate oxygen toxicity by generating reactive oxygen species (ROS) via the iron-catalyzed Haber-Weiss reaction, leading to oxidative stress. ROS-mediated iron cytotoxicity may trigger apoptotic cell death. In the present study, we used iron treatment of organotypic cultures of hippocampal slices to study potential mechanisms involved in iron-induced neuronal damage. Exposure of mature hippocampal slices to ferrous sulfate resulted in concentration- and time-dependent cell death. After iron treatment, markers of ROS formation and lipid peroxidation, i.e. intensity of dichlorofluorescein (DCF) fluorescence and levels of thiobarbiturate reactive substances (TBARS), were significantly increased. Levels of cytochrome c were increased while levels of pro-caspase-9 and pro-caspase-3 were decreased in cytosolic fractions of iron-treated hippocampal slice cultures. Treatment of cultured slices with a synthetic catalytic ROS scavenger, EUK-134, provided between 50 and 70% protection against various parameters of cell damage and markers of oxidative stress. In addition, inhibition of caspase-3 activity by Ac-DEVDcho partially protected cells from iron toxicity. The combination of EUK-134 and Ac-DEVDcho resulted in an almost complete blockade of iron-induced damage. These results indicate that iron elicits cellular damage predominantly by oxidative stress, and that ROS-mediated iron toxicity may involve cytochrome c- and caspase-3-dependent apoptotic pathways.
Liu
Ruolan
R
Neuroscience Program, University of Southern California, Los Angeles, California, USA.
Liu
Wei
W
Doctrow
Susan R
SR
Baudry
Michel
M
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D020169:Caspases / Q000378:metabolism; D004396:Coloring Agents / Q000493:pharmacokinetics; D003574:Cytochrome c Group / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005296:Ferrous Compounds / Q000633:toxicity; D006624:Hippocampus / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D066298:In Vitro Techniques; D007501:Iron / Q000633:toxicity*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D015227:Lipid Peroxidation / Q000187:drug effects; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D009942:Organometallic Compounds / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects; D011419:Propidium / Q000493:pharmacokinetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012459:Salicylates / Q000494:pharmacology
D004396:Coloring Agents; D003574:Cytochrome c Group; C433928:EUK-134; D005296:Ferrous Compounds; D009942:Organometallic Compounds; D017382:Reactive Oxygen Species; D012459:Salicylates; D011419:Propidium; C020748:ferrous sulfate; D007501:Iron; D007770:L-Lactate Dehydrogenase; D020169:Caspases