Critical role of the JNK-p53-GADD45α apoptotic cascade in mediating oxidative cytotoxicity in hippocampal neurons.
OBJECTIVE
Glutamate-induced oxidative stress plays a critical role in the induction of neuronal cell death in a number of disease states. We sought to determine the role of the c-Jun NH(2) -terminal kinase (JNK)-p53-growth arrest and DNA damage-inducible gene (GADD) 45α apoptotic cascade in mediating glutamate-induced oxidative cytotoxicity in hippocampal neuronal cells.


METHODS
HT22 cells, a mouse hippocampal neuronal cell line, were treated with glutamate to induce oxidative stress in vitro. Kainic acid-induced oxidative damage to the hippocampus in rats was used as an in vivo model. The signalling molecules along the JNK-p53-GADD45α cascade were probed with various means to determine their contributions to oxidative neurotoxicity.


RESULTS
Treatment of HT22 cells with glutamate increased the mRNA and protein levels of GADD45α, and these increases were suppressed by p53 knock-down. Knock-down of either p53 or GADD45α also prevented glutamate-induced cell death. Glutamate-induced p53 activation was preceded by accumulation of reactive oxygen species, and co-treatment with N-acetyl-cysteine prevented glutamate-induced p53 activation and GADD45α expression. Knock-down of MKK4 or JNK, or the presence of SP600125 (a JNK inhibitor), each inhibited glutamate-induced p53 activation and GADD45α expression. In addition, we also confirmed the involvement of GADD45α in mediating kainic acid-induced hippocampal oxidative neurotoxicity in vivo.


CONCLUSIONS
AND IMPLICATIONS Activation of the JNK-p53-GADD45α cascade played a critical role in mediating oxidative cytotoxicity in hippocampal neurons. Pharmacological inhibition of this signalling cascade may provide an effective strategy for neuroprotection.
Department of Pharmacology, Toxicology and Therapeutics, School of Medicine, University of Kansas Medical Center, Kansas City, KS 66160, USA.
Hye Joung
HJ
Choi
Ki Sung
KS
Kang
Masayuki
M
Fukui
Bao Ting
BT
Zhu
D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D006624:Hippocampus / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D007608:Kainic Acid / Q000494:pharmacology; D048670:MAP Kinase Kinase 4 / Q000378:metabolism*; D051379:Mice; D008856:Microscopy, Fluorescence; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D018384:Oxidative Stress; D016133:Polymerase Chain Reaction; D034741:RNA, Small Interfering; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D000097642:GADD45 Proteins
D018797:Cell Cycle Proteins; D007608:Kainic Acid; D048670:MAP Kinase Kinase 4; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D016159:Tumor Suppressor Protein p53; C499072:Gadd45a protein, mouse; D000097642:GADD45 Proteins