Intranuclear localization of apoptosis-inducing factor (AIF) and large scale DNA fragmentation after traumatic brain injury in rats and in neuronal cultures exposed to peroxynitrite.
Programmed cell death occurs after ischemic, excitotoxic, and traumatic brain injury (TBI). Recently, a caspase-independent pathway involving intranuclear translocation of mitochondrial apoptosis-inducing factor (AIF) has been reported in vitro; but whether this occurs after acute brain injury was unknown. To address this question adult rats were sacrificed at various times after TBI. Western blot analysis on subcellular protein fractions demonstrated intranuclear localization of AIF in ipsilateral cortex and hippocampus at 2-72 h. Immunocytochemical analysis showed AIF labeling in neuronal nuclei with DNA fragmentation in the ipsilateral cortex and hippocampus. Immunoelectronmicroscopy verified intranuclear localization of AIF in hippocampal neurons after TBI, primarily in regions of euchromatin. Large-scale DNA fragmentation ( approximately 50 kbp), a signature event in AIF-mediated cell death, was detected in ipsilateral cortex and hippocampi by 6 h. Neuron-enriched cultures exposed to peroxynitrite also demonstrated intranuclear AIF and large-scale DNA fragmentation concurrent with impaired mitochondrial respiration and cell death, events that are inhibited by treatment with a peroxynitrite decomposition catalyst. Intranuclear localization of AIF and large-scale DNA fragmentation occurs after TBI and in neurons under conditions of oxidative/nitrosative stress, providing the first evidence of this alternative mechanism by which programmed cell death may proceed in neurons after brain injury.
Zhang
Xiaopeng
X
Department of Anesthesiology and Critical Care Medicine, Safar Center for Resuscitation Research, University of Pittsburgh, 3434 Fifth Avenue, Pittsburgh, PA 15260, USA.
Chen
Jun
J
Graham
Steven H
SH
Du
Lina
L
Kochanek
Patrick M
PM
Draviam
Romesh
R
Guo
Fengli
F
Nathaniel
Paula D
PD
Szabó
Csaba
C
Watkins
Simon C
SC
Clark
Robert S B
RS
D021581:Active Transport, Cell Nucleus; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D051033:Apoptosis Inducing Factor; D015153:Blotting, Western; D001923:Brain Chemistry; D001930:Brain Injuries / Q000378:metabolism* / Q000473:pathology; D002467:Cell Nucleus / Q000378:metabolism* / Q000648:ultrastructure; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000378:metabolism / Q000473:pathology; D053938:DNA Fragmentation / Q000187:drug effects; D004195:Disease Models, Animal; D018450:Disease Progression; D005420:Flavoproteins / Q000378:metabolism*; D006624:Hippocampus / Q000378:metabolism / Q000473:pathology / Q000648:ultrastructure; D007150:Immunohistochemistry; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D016253:Microscopy, Immunoelectron; D009474:Neurons / Q000378:metabolism* / Q000648:ultrastructure; D030421:Peroxynitrous Acid / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013347:Subcellular Fractions / Q000737:chemistry / Q000378:metabolism
C494831:Aifm1 protein, rat; D051033:Apoptosis Inducing Factor; D005420:Flavoproteins; D008565:Membrane Proteins; D030421:Peroxynitrous Acid