Key proteins of activating cell death can be predicted through a kainic acid-induced excitotoxic stress.

Hsiu-Ling Tsai,Sue-Joan Chang

doi:10.1155/2015/478975

Abstract

Epilepsy is a major neurological disorder characterized by spontaneous seizures accompanied by neurophysiological changes. Repeated seizures can damage the brain as neuronal death occurs. A better understanding of the mechanisms of brain cell death could facilitate the discovery of novel treatments for neurological disorders such as epilepsy. In this study, a model of kainic acid- (KA-) induced neuronal death was established to investigate the early protein markers associated with apoptotic cell death due to excitotoxic damage in the rat cortex. The results indicated that KA induces both apoptotic and necrotic cell death in the cortex. Incubation with high concentrations (5 and 500 μM, >75%) and low concentrations (0.5 pM: 95% and 50 nM: 8%) of KA for 180 min led to necrotic and apoptotic cell death, respectively. Moreover, proteomic analysis using two-dimensional gel electrophoresis and mass spectrometry demonstrated that antiapoptotic proteins, including heat shock protein 70, 3-mercaptopyruvate sulfurtransferase, tubulin-B-5, and pyruvate dehydrogenase E1 component subunit beta, were significantly higher in apoptosis than in necrosis induced by KA. Our findings provide direct evidence that several proteins are associated with apoptotic and necrotic cell death in excitotoxicity model. The results indicate that these proteins can be apoptotic biomarkers from the early stages of cell death.

Highlights

Epilepsy is a major neurological disorder characterized by spontaneous seizures accompanied by motor and/or perceptual dysfunction
A number of studies have reported that apoptosis was the major mode of cell death, while a few studies have suggested the existence of kainic acid- (KA-)induced necrotic cell death, as well as the coexistence of apoptosis and necrosis in the brain [9, 10]
Compared to the group treated with a moderate dose of kainic acid (KA) (0.5 pM) group, reactive oxygen species (ROS) generation was markedly increased at 30, 60, and 180 min in the group treated with high concentrations of KA (5 and 500 μM) (P < 0.01 versus 0.5 pM KA group) (Figure 1(f))

Summary

Introduction

Epilepsy is a major neurological disorder characterized by spontaneous seizures (discharges) accompanied by motor and/or perceptual dysfunction. Researchers have proposed that the underlying pathological mechanisms of epilepsy are excessive activation of excitatory amino acid (EAA) receptors, with accompanying generation of reactive oxygen species (ROS) [1, 2]. A growing number of in vivo and in vitro studies have confirmed that oxidative stress induces cell death in excitotoxic injury [5, 6]. KA is an agonist of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) subtype of EAA receptors and is widely used to study the mechanisms behind neurological disorders by creating models of cell damage. KA has been reported to induce excitotoxic cell death in immature and adult animals by systemic or intraventricular injection and in primary cultures and in vitro neuronal cell lines [7, 8]. A number of studies have reported that apoptosis was the major mode of cell death, while a few studies have suggested the existence of KA-induced necrotic cell death, as well as the coexistence of apoptosis and necrosis in the brain [9, 10]

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