Original articleOxidative stress and disturbed glutamate transport in spinal muscular atrophy
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.
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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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