Proteomic identification of hippocampal proteins vulnerable to oxidative stress in excitotoxin-induced acute neuronal injury

Ayako Furukawa, Yoshiyuki Kawamoto, Yoichi Chiba, Shiro Takei, Sanae Hasegawa-Ishii, Noriko Kawamura, Keisuke Yoshikawa, Masanori Hosokawa, Shinji Oikawa, Masashi Kato, Atsuyoshi Shimada

Research output: Contribution to journalArticle

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Abstract

Excitotoxicity is involved in seizure-induced acute neuronal death, hypoxic-ischemic encephalopathy, and chronic neurodegenerative conditions such as Alzheimer's disease. Although oxidative stress has been implicated in excitotoxicity, the target proteins of oxidative damage during the course of excitotoxic cell death are still unclear. In the present study, we performed 2D-oxyblot analysis and mass spectrometric amino acid sequencing to identify proteins that were vulnerable to oxidative damage in the rat hippocampus during kainic acid (KA)-induced status epilepticus. We first investigated the time course in which oxidative protein damage occurred using immunohistochemistry. Carbonylated proteins, a manifestation of protein oxidation, were detected in hippocampal neurons as early as 3 h after KA administration. Immunoreactivity for 8-hydroxy-2'-deoxyguanosine (8-OHdG) was also elevated at the same time point. The increase in oxidative damage to proteins and DNA occurred concomitantly with the early morphological changes in KA-treated rat hippocampus, i.e., changes in chromatin distribution and swelling of rough endoplasmic reticulum and mitochondria, which preceded the appearance of morphological features of neuronal death such as pyknotic nuclei and hypereosinophilic cytoplasm. Proteomic analysis revealed that several hippocampal proteins were consistently carbonylated at this time point, including heat shock 70 kDa protein 4, valosin-containing protein, mitochondrial inner membrane protein (mitofilin), α-internexin, and tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein (14-3-3 protein). We propose that oxidative damage to these proteins may be one of the upstream events in the molecular pathway leading to excitotoxic cell death in KA-treated rat hippocampus, and these proteins may be targets of therapeutic intervention for seizure-induced neuronal death.

Original languageEnglish (US)
Pages (from-to)706-714
Number of pages9
JournalNeurobiology of Disease
Volume43
Issue number3
DOIs
StatePublished - Sep 1 2011

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Neurotoxins
Proteomics
Oxidative Stress
Wounds and Injuries
Kainic Acid
Proteins
14-3-3 Proteins
Hippocampus
Seizures
Cell Death
Tryptophan Hydroxylase
Brain Hypoxia-Ischemia
HSP70 Heat-Shock Proteins
Status Epilepticus
Rough Endoplasmic Reticulum
Protein Sequence Analysis
Tyrosine 3-Monooxygenase
Chromatin
Alzheimer Disease
Mitochondria

All Science Journal Classification (ASJC) codes

  • Neurology

Cite this

Furukawa, A., Kawamoto, Y., Chiba, Y., Takei, S., Hasegawa-Ishii, S., Kawamura, N., ... Shimada, A. (2011). Proteomic identification of hippocampal proteins vulnerable to oxidative stress in excitotoxin-induced acute neuronal injury. Neurobiology of Disease, 43(3), 706-714. https://doi.org/10.1016/j.nbd.2011.05.024
Furukawa, Ayako ; Kawamoto, Yoshiyuki ; Chiba, Yoichi ; Takei, Shiro ; Hasegawa-Ishii, Sanae ; Kawamura, Noriko ; Yoshikawa, Keisuke ; Hosokawa, Masanori ; Oikawa, Shinji ; Kato, Masashi ; Shimada, Atsuyoshi. / Proteomic identification of hippocampal proteins vulnerable to oxidative stress in excitotoxin-induced acute neuronal injury. In: Neurobiology of Disease. 2011 ; Vol. 43, No. 3. pp. 706-714.
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Furukawa, A, Kawamoto, Y, Chiba, Y, Takei, S, Hasegawa-Ishii, S, Kawamura, N, Yoshikawa, K, Hosokawa, M, Oikawa, S, Kato, M & Shimada, A 2011, 'Proteomic identification of hippocampal proteins vulnerable to oxidative stress in excitotoxin-induced acute neuronal injury', Neurobiology of Disease, vol. 43, no. 3, pp. 706-714. https://doi.org/10.1016/j.nbd.2011.05.024

Proteomic identification of hippocampal proteins vulnerable to oxidative stress in excitotoxin-induced acute neuronal injury. / Furukawa, Ayako; Kawamoto, Yoshiyuki; Chiba, Yoichi; Takei, Shiro; Hasegawa-Ishii, Sanae; Kawamura, Noriko; Yoshikawa, Keisuke; Hosokawa, Masanori; Oikawa, Shinji; Kato, Masashi; Shimada, Atsuyoshi.

In: Neurobiology of Disease, Vol. 43, No. 3, 01.09.2011, p. 706-714.

Research output: Contribution to journalArticle

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T1 - Proteomic identification of hippocampal proteins vulnerable to oxidative stress in excitotoxin-induced acute neuronal injury

AU - Furukawa, Ayako

AU - Kawamoto, Yoshiyuki

AU - Chiba, Yoichi

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AU - Hasegawa-Ishii, Sanae

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AU - Yoshikawa, Keisuke

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AU - Oikawa, Shinji

AU - Kato, Masashi

AU - Shimada, Atsuyoshi

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N2 - Excitotoxicity is involved in seizure-induced acute neuronal death, hypoxic-ischemic encephalopathy, and chronic neurodegenerative conditions such as Alzheimer's disease. Although oxidative stress has been implicated in excitotoxicity, the target proteins of oxidative damage during the course of excitotoxic cell death are still unclear. In the present study, we performed 2D-oxyblot analysis and mass spectrometric amino acid sequencing to identify proteins that were vulnerable to oxidative damage in the rat hippocampus during kainic acid (KA)-induced status epilepticus. We first investigated the time course in which oxidative protein damage occurred using immunohistochemistry. Carbonylated proteins, a manifestation of protein oxidation, were detected in hippocampal neurons as early as 3 h after KA administration. Immunoreactivity for 8-hydroxy-2'-deoxyguanosine (8-OHdG) was also elevated at the same time point. The increase in oxidative damage to proteins and DNA occurred concomitantly with the early morphological changes in KA-treated rat hippocampus, i.e., changes in chromatin distribution and swelling of rough endoplasmic reticulum and mitochondria, which preceded the appearance of morphological features of neuronal death such as pyknotic nuclei and hypereosinophilic cytoplasm. Proteomic analysis revealed that several hippocampal proteins were consistently carbonylated at this time point, including heat shock 70 kDa protein 4, valosin-containing protein, mitochondrial inner membrane protein (mitofilin), α-internexin, and tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein (14-3-3 protein). We propose that oxidative damage to these proteins may be one of the upstream events in the molecular pathway leading to excitotoxic cell death in KA-treated rat hippocampus, and these proteins may be targets of therapeutic intervention for seizure-induced neuronal death.

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