Elsevier

Brain and Development

Volume 24, Issue 8, December 2002, Pages 770-775
Brain and Development

Original article
Oxidative stress and disturbed glutamate transport in spinal muscular atrophy

https://doi.org/10.1016/S0387-7604(02)00103-1Get rights and content

Abstract

Spinal muscular atrophy (SMA) is a hereditary motor neuron disease, and three clinical subtypes of autosomal recessive SMA, including Werdnig Hoffmann disease (type 1), have been shown to be induced by deletion within the same genes. In order to clarify the pathogenesis of motor neuron degeneration in SMA, we immunohistochemically examine the expressions of oxidative stress-related materials (oxidative products) and glutamate transporters, which can prevent glutamate neurotoxicity, in five autopsy cases of SMA type 1. Age-matched controls did not show any deposition of oxidative products in the brain. In contrast, the abnormal deposition of 4-hydroxy-2-nonenal-modified protein, a product of membrane lipid oxidation, was observed in the spinal motor neurons in three cases, although the motor neurons did not show an increase of nitrotyrosine, which was observed in adult-onset amyotrophic lateral sclerosis. In addition, the nuclei of neurons and glial cells in the precentral gyrus, thalamus or cerebellar cortex were immunoreactive for 8-hydroxy-2′-deoxyguanosine in two cases, which was one of the most commonly used markers for oxidative DNA damage. Regarding glial glutamate transporters, three of five cases of SMA type 1 showed a reduction in immunoreactivity for excitatory amino acid transporter-1 (GLAST) in the ventrolateral nucleus of the thalamus, in which there was neither neuronal loss nor gliosis in routine histochemistry. One case, having mechanical ventilation, demonstrated a reduced expression of another glial glutamate transporter (GLT-1) throughout the central nervous system. These data suggest that oxidative stress and disturbed glutamate transport can partly be involved in the motor neuron devastation and/or latent thalamic degeneration in SMA type 1.

Introduction

Spinal muscular atrophy (SMA) is a hereditary neurodegenerative disease, showing progressive muscular weakness and atrophy, due to the degeneration of motor neurons in the spinal cord and brainstem [1], [2]. The gene responsible for three clinically and genetically distinct forms of autosomal recessive SMA consisting of Werdnig Hoffmann disease (type 1), intermediate form (type 2) and Kugelberg–Welander disease, is mapped to chromosome 5q13. The survival motor neuron gene (smn) is deleted in more than 90% of the SMA patients, in addition to the deletion of another neuronal apoptosis inhibitory protein (naip) located near the smn gene [3], [4], [5]. Since both SMN and NAIP proteins have anti-apoptotic activity, an apoptotic scenario has been investigated in the motor neuron degeneration in SMA. However, the occurrence of apoptotic cell death was extremely rare in the spinal and brainstem motor neurons in autopsy materials [6], [7]. Recently, oxidative stress and disturbed glutamate transport have been shown to be involved in adult-onset amyotrophic lateral sclerosis (ALS), which is also characterized by progressive degeneration of motor neurons in the spinal cord and brainstem [8], [9]. Excessive nitration of tyrosine by peroxynitrite and subsequent increase of nitrotyrosine have been observed in the spinal cord in ALS [8]. Glutamate transport is critical for preventing excessive extracellular accumulation of neurotoxic chemicals. Since variant mRNA transcript for the main glial glutamate transporter GLT-1 has been detected in brain tissue of 60% of patients of sporadic ALS, disturbed glutamate transport has been postulated in the motor neuron degeneration [9]. Neither oxidative stress nor glutamate transport have been investigated in child-onset SMA. Here, we immunohistologically examine the deposition of oxidative products and the expression of glutamate transporters in the brain in autopsy cases of SMA type 1 to clarify involvement of both pathologic processes in neurodegeneration in SMA.

Section snippets

Materials and methods

The clinical subjects comprised five patients with clinicopathologically confirmed Werdnig Hoffmann disease (SMA type 1), which showed onset before the age of 6 months (Table 1), and eight control subjects (Table 2). In all subjects, the brains were fixed in a buffered formalin solution for 10–14 days. Each formalin-fixed brain was cut coronally and was embedded in paraffin. Then, 10 μm thick sections in the areas of interest were subjected to hematoxylin and eosin (HE), Klüver–Barrera (KB),

Results

All subjects, including the patients and controls, failed to show an abnormal deposition of nitrotyrosine and AGE in the brain regions tested including the motor neurons. The immunoreactivity for 4HNE was increased in the motor neurons in the hypoglossal nucleus and spinal ventral horn in three of five cases of SMA type 1 aged over 1 years (Table 1 and Fig. 1A), whereas 4HNE deposition was not observed in controls. Controls under 40 years of age did not show nuclei immunoreactive for 8OHdG. In

Discussion

Unlike ALS [8], the nitrotyrosine deposition could not pathologically be detected in autopsy cases of SMA type 1. It may partly be because the onset is intrauterine and the disease develops more rapidly in SMA type 1 than ALS. In addition, there was no increased deposition of AGE, which is a product of protein glycation and oxidation (the Maillard reaction) and is increased in the hippocampus in Alzheimer's disease [10]. In contrast, all of the cases over 1 year in age showed the 4HNE

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