Sporadic Motor Neuron Disease Research Articles

Does lead damage motor neurons?

It has been suggested that lead (Pb) may damage motor neurons selectively after retrograde axonal transport from the neuromuscular junction. This has reinforced suspicions that Pb could be a cause of some sporadic motor neuron diseases. However, the long-term effect of Pb on spinal motor neurons is not known. To investigate this, we injected mice in the right hindlimb with 0.1 ml of 2.5% Pb nitrate. Nine months later we measured the sizes and counted the numbers of motor neuron cell bodies in the anterior horns of the L4 spinal cord. There was no significant difference in sizes or numbers of motor neuron cell bodies between the left and right sides of the spinal cord in either group or between groups. These data do not therefore support the hypothesis that Pb transported from muscle damages spinal motor neurons.

CNS tissue Cu/Zn superoxide dismutase (SOD1) mutations in motor neurone disease (MND).

DNA extracted from CNS tissue of 79 cases of motor neurone disease (MND) was screened by single strand conformation analysis (SSCA) and heteroduplex analysis (HA) for mutations in the Cu/Zn superoxide dismutase (SOD1) gene. The aims were to determine whether somatic mutations of SOD1 may underlie some cases of MND and to characterize the genetic abnormalities by sequencing, for subsequent correlation with the molecular pathological phenotype. In 3 cases a point mutation was found in exon 4: E100G in one familial case, and I113T in two cases (one familial, one sporadic). Two cases had previously undescribed mutations in the 3' untranslated region (3'UTR) of SOD1 and one case had a single base substitution in the intronic sequence upstream from exon 2. None of these patients had a positive family history. Non-CNS tissue was available for 3 out of the 6 cases in whom changes were found. In all 3 the same changes were consistently found in both CNS and non-CNS tissue, excluding the presence of somatic mutations in SOD1. We investigated many MND blood samples and normal controls for the presence of the 3'UTR deletions. We found the 4 bp deletion in 1/90 sporadic MND patients and 1/209 non-MND controls. If the 3'UTR deletions are pathogenic, they would have to operate via a loss of the function mechanism, and further work is necessary to define their significance.

British motor neuron disease twin study.

OBJECTIVES: To investigate the cause of sporadic motor neuron disease (MND) by twin study, so allowing (1) estimation of the genetic contribution, and (2) collection of matched pairs for a...

Immunocytochemical study of the distribution of the free radical scavenging enzymes CU/ZN superoxide dismutase (SOD1); MN superoxide dismutase (MN SOD) and catalase in the normal human spinal cord and in motor neuron disease

Interest in free radical-mediated mechanisms of motor neuron injury has arisen following the discovery of point mutations in the CU/ZN superoxide dismutase (SOD1) gene in a proportion of cases of familial motor neuron disease (MND). Evidence is emerging which indicates that oxidative stress may contribute to the pathophysiology of sporadic MND. Free radical scavenging enzymes form a major component of the anti-oxidant defense system. The aim of this study was to compare the distribution and density of immunoreactivity to CU/ZN SOD, Mn SOD and catalase in the spinal cord of sporadic MND cases ( n = 10) compared to normal controls ( n = 8). There was abundant expression of CU/ZN SOD, Mn SOD and catalase in spinal motor neurons, suggesting important roles for these enzymes in neuroprotective pathways in this cell group. In MND cases, there was no evidence in surviving motor neurons of a consistent alteration in the protein expression of any of these enzymes. There was evidence of increased expression of these enzymes in glial cells present in the ventral and intermediate grey matter and degenerating descending motor pathways of the spinal cord in MND cases. The changes observed were more marked in the cervical compared to lumbar spinal segments. Further investigation is required to determine whether these findings represent a compensatory response to a pathophysiological process involving oxidative stress.

Ultrastructural localization of heparan sulfate‐like immunoreactivity in spinal spheroids of motor neuron disease

It has recently been reported that heparan sulfate (HS)‐like immunoreactivity exists in spheroids of the spinal cord in motor neuron disease (MND). However, the association of the HS‐like immunoreactivity with neurofilaments, which are the main structure in spinal spheroids of MND, remains to be clarified. The aim of the present study is to elucidate the ultrastructural localization of HS‐like immunoreactivity. The spinal cords from five autopsy cases of sporadic MND and two control cases were immunocytochemically investigated with an antibody against axonal neurofilament proteins and several antibodies against glycosaminoglycans including HS, dermatan sulfate, keratan sulfate, and HS‐proteoglycan. Ultrastructural localization of the immunoreactivity was also studied by using pre‐embedding immunoperoxidase and immunogold methods. On the basis of the findings of this study, only HepSS‐1‐recognized HS among several glycosaminoglycans is clearly detected in the spinal spheroids in MND, and it exists in close association with accumulated neurofilaments. Because of its capacity to bind various proteins, HS may act as a trigger or an accelerator for the accumulation of neurofilaments and may play important roles in the formation of spheroids in MND.

Immunocytochemical co-localization of the proteasome in ubiquitinated structures in neurodegenerative diseases and the elderly.

To determine at the tissue level whether the proteasome (Ps), a unique nonlysosomal protease, is involved in the metabolism of ubiquitinated proteins, we examined for the first time the immunocytochemical localizations of both Ps and ubiquitin (Ub) in sections of various abnormal structures that are known to be ubiquitinated in various neurodegenerative diseases and in the elderly. Concomitant increases of Ps and Ub were observed at the sites of most dystrophic neurites in Alzheimer disease (AD) and parkinsonism-dementia complex on Guam (PDC) and in Lewy bodies in Parkinson's disease and diffuse Lewy body disease, but not in neurofibrillary tangles in AD or PDC, in filamentous inclusions within anterior horn cells in sporadic motor neuron disease, or in eosinophilic granules in the olivary nucleus of the elderly. These results at the tissue level indicated that Ps is involved in the metabolism of some, but not all, ubiquitinated proteins and structures in various neurodegenerative disorders. This suggests that the involvement of Ps in the metabolism of ubiquitinated structures differs in different cases and at different stages of disease. These results and our previous immunocytochemical studies of lysosomal cathepsin proteases suggest that both nonlysosomal and lysosomal systems are involved in the metabolism of various ubiquitinated proteins and that their involvements differ in different structures and at different stages of degeneration of the structures.

The relationship of spinal muscular atrophy to motor neuron disease: Investigation of SMN and NAIP gene deletions in sporadic and familial ALS

Amyotrophic lateral sclerosis (ALS) is found in a familial form in around 5–10% of cases. Of these familial cases around 20% are associated with mutations of SOD-1. The genetic basis of the disease in the remaining familial cases, and genetic risk factors in sporadic cases, are unknown. Recently, the common forms of spinal muscular atrophy (SMA) have been associated with mutations of the SMN and NAIP genes on chromosome 5, in the region q11.2–13.3. Some patients with both familial and sporadic motor neuron disease show only lower motor neuron signs, in common with SMA patients, and families containing individuals with phenotypes of both childhood SMA and adult motor neuron disease have been reported. We therefore examined the SMA locus as a candidate for ALS, in 54 patients with sporadic motor neuron disease, and 10 single-generation familial patients (with no evidence of SOD-1 mutations), and in a single patient with Brown-Vialetto-Van Laere syndrome. No mutations of the SMN or NAIP genes were detected. The difficulties of classification of lower motor neuron presentations of motor neuron diseases are discussed. The demonstration that mutations diagnostic of SMA are not found in ALS patients helps distinguish these conditions. © 1997 Elsevier Science B.V.

Motor neuron disease with pyramidal tract dysfunction involves the cortical generators of the early somatosensory evoked potential to tibial nerve stimulation.

We evaluated somatosensory evoked potentials (SEPs) to tibial nerve stimulation in 39 patients with sporadic motor neuron disease using multiple scalp derivations (earlobe reference). SEPs were altered in 22 of 29 amyotrophic lateral sclerosis (ALS) patients, whereas they were unaffected in 10 progressive muscular atrophy (PMA) patients. The main changes involved the amplitude and the field distribution of the early P40 and N37 cortical potentials with different modalities varying from a selective loss of the P40 potential (33% of tested sides) to absence of all early cortical SEPs (22% of tested sides). The later components following N50 were generally spared. The commonly used Cz-Fz montage was inadequate for detecting these alterations. Central afferent conduction was slightly affected. The selective loss of cortical SEPs and their close correlation with clinicoelectrophysiologic evidence of central motor system involvement strongly support a cortical origin of the SEP alterations in ALS. We suggest that neuronal loss in the somatosensory cortex may selectively affect the generator sites of the cortical SEPs to lower limb stimulation.

Absence of serological evidence for foamy virus infection in patients with amyotrophic lateral sclerosis.

Foamy virus (FV) infection has been implicated in the pathogenesis of sporadic motor neuron disease (MND) by means of serological assays. To confirm these results we tested serum and cerebrospinal fluid (CSF) samples from 23 cases of clinically verified non-familial MND and 11 cases of suspected non-familial MND for the presence of FV infection as determined by Western blot (WB) and indirect immunofluorescence assay (IFA). Using the same tests we also screened sera from 87 healthy chimpanzees for the presence of FV antibodies. None of the human samples in question tested positive. However, the testing revealed that 84 of 87 chimpanzees (96.6%) were seropositive for FV, indicating that combined WB and IFA are suitable methods for the serodiagnosis of FV infection. Given these results an association of FV infection and sporadic MND is highly improbable. Furthermore a suggested therapeutic trial with anti-retroviral drugs appears unjustified.

Oxidative damage and motor neurone disease difficulties in the measurement of protein carbonyls in human brain tissue.

It has been suggested in the literature that elevated oxidative protein damage, measured as protein carbonyls, is present in the nervous system of patients with sporadic motor neurone disease (MND). However, the actual reported levels of brain protein carbonyls vary over a wide range. We show here that this is probably due to the use of the different protocols for the carbonyl assay; results differ depending on when the dinitrophenylhydrazine reagent is added and at what stage in the procedure protein is assayed for the calculation of carbonyls on a unit protein basis. Using a range of different procedures, we were unable to confirm reports of elevated protein carbonyls in motor cortex from brains of patients with MND. We also measured thiobarbituric acid-reactive material in the brain samples using an HPLC-based TBA test in the presence of butylated hydroxytoluene. In general, there was no significant elevation of TBARS in MND motor cortex. However, four patients showed values higher than any of the control patients (both 'normal' control and 'disease control'). There was no correlation of TBARS with protein carbonyl values. We suggest that oxidative damage in motor cortex in sporadic MND, if it occurs, may be confined to a small group of patients and may affect different molecular targets in each patient.

Motor neuron uptake of low dose inorganic mercury

In animals, inorganic mercury can bypass the blood brain barrier and enter motor neurons. We sought to determine the lowest injected dose of mercury that could be detected in mouse motor neurons. Mice were injected intraperitoneally with mercuric chloride in doses from 0.05 to 2 μg/g body weight and studied between 5 days and 18 months after injection. After formalin fixation, 7 μm sections of cerebrum, cerebellum, brain stem, spinal cord and kidney were stained with silver nitrate autometallography. Five days after injection, mercury granules were detected at doses from 0.2 μg/g upwards in the cell bodies of spinal and brain stem motor neurons, more granules being seen at the higher doses. Mercury granules were also seen in 5% of posterior root ganglion neurons. At doses from 0.05 μg/g upwards mercury was detected 5 days later in renal tubule cells. Mercury was still present in motor neurons 6–11 months after injection, but by this time mercury had been cleared from the kidneys. Low doses of inorganic mercury are therefore selectively taken up and retained by motor neurons, making this neurotoxin a good candidate for a cause of sporadic motor neuron disease.

Oxidative damage to protein in sporadic motor neuron disease spinal cord

The recent discovery that defects in the gene encoding copper-zinc superoxide dismutase (SOD1) are associated with some cases of familial motor neuron disease has heightened interest in the possibility that free radical mechanisms may contribute to selective motor neuron injury. Sporadic and familial motor neuron diseases are clinically and pathologically very similar and may share common pathophysiological mechanisms. Thus the role of free radical mechanisms as a contributory factor to motor neuron injury in the common sporadic form of motor neuron disease requires urgent exploration, particularly as this may provide an avenue for therapy aimed at retarding pathological progression. We investigated oxidative damage to proteins in the lumbar spinal cord by quantifying the protein carbonyl level from 19 patients with sporadic motor neuron disease, 8 neurologically normal control subjects, and 11 neurological disease control subjects, most of whom had slowly progressive neurodegenerative disease. In sporadic motor neuron disease the mean protein carbonyl level in the spinal cord was increased by 119% (p < 0.02) compared to normal control subjects and by 88% (p < 0.04) compared to the neurological disease control subjects. These data contribute to the emerging evidence that oxidative damage may play a contributory role in the neuronal death in sporadic motor neuron disease. This mechanism may be particularly important in a subset of patients with motor neuron disease.

Ciliary neurotrophic factor receptor expression in spinal cord and motor cortex in amyotrophic lateral sclerosis

Ciliary neurotrophic factor (CNTF) is known to rescue motor neurones in animal models of injury and neurodegeneration and we have recently described regional changes of CNTF protein levels in spinal cord from patients with sporadic motor neurone disease of the amyotrophic lateral sclerosis (ALS) type. However, information is lacking about the CNTF receptor in this condition. We have therefore studied mRNA levels for the α subunit of the CNTF receptor (CNTFRα) and for CNTF itself in postmortem spinal cord and cerebral cortex in patients with sporadic ALS and matched controls. We report that in the spinal cord of ALS patients there is a marked increase in the hybridisation signal for CNTFRα subunit mRNA overlying motor neurones. In contrast, very little mRNA for CNTFR α subunit was found in the motor cortex and no differences were seen between ALS and controls. We were unable to detect any hybridisation signal for CNTF mRNA. Our findings provide evidence for regional differences in CNTF receptor expression in upper- and lower-motor neurones in ALS.

Motor neuron disease. Studies of the corticospinal excitation of single motor neurons by magnetic brain stimulation.

The responses to magnetic brain stimulation of 57 motor units (MUs) in the first dorsal interosseous muscle of 33 patients with sporadic motor neuron disease (MND), and 44 MUs of 15 healthy control subjects have been compared. Thresholds for MU excitation both in the resting state and during tonic activity have been measured and the modulation of MU discharge by series of magnetic stimuli has been determined from peristimulus time histograms (PSTHs). Abnormalities in the MND MUs have been correlated with the physical signs of impaired hand muscle function. Motor units from hands showing pure lower motor neuron (LMN) signs fired at increased frequency when recruited, had normal or reduced thresholds for activation by magnetic scalp stimuli and had normal or large primary excitatory peaks with normal inter-subpeak intervals on PSTHs. Occasionally muscles with pure LMN signs also had fasciculations that could be driven by cortical stimuli. In contrast, MUs from hands showing pure upper motor neuron (UMN) signs fired at reduced frequency when recruited, had raised thresholds for excitation by cortical stimuli, had prolonged primary peaks with wide inter-subpeak intervals and had large secondary peaks on PSTHs. The rise times and amplitudes of the underlying post-synaptic potentials released by cortical stimuli in spinal motor neurons have been deduced from measurements of the peaks in PSTHs. This analysis suggests that in some MUs from muscles showing pure LMN signs, stimuli evoke larger than normal excitatory post-synaptic potentials (EPSPs), probably resulting from abnormalities in central motor fibres. It is proposed that a major cause of the changes found in MND MUs from hands showing either pure UMN or pure LMN signs may be an abnormality in the motor cortex leading to faulty initiation of the descending corticospinal volley from magnetic stimuli.

Risk estimates for developing motor neurone disease in first‐degree relatives

Patients with motor neurone disease resident in the three counties of South Glamorgan, Mid Glamorgan and Gwent (total population 1,394,400) from 1/11/91 to 31/12/92, were ascertained through general practitioners and neurology department records: 62 out of 71 (51 male, 20 female) index cases ascertained agreed to be visited. These individuals were clinically reviewed, had a detailed family history taken and, after discussion, had a blood sample taken for testing for the X-linked bulbospinal muscular atrophy mutation. The period prevalence of motor neurone disease was 5.02/100,000 with a mean age of onset of 59.4 (S.D. 12.9) years. One 66-year-old male was found to have the X-linked bulbospinal muscular atrophy mutation. A single, definite first-degree relative pair was identified, of whom only one was included in the data. The index cases had a total of 131 offspring and 173 siblings. The risk estimates, by current age, for these first-degree relatives of patients with apparently sporadic motor neurone disease decreased from 1 in 85 at age 20 years to 1 in 1683 at age 80 years. These estimates require confirmation in an independent sample, as they are based on a single "event".

Iron, selenium and glutathione peroxidase activity are elevated in sporadic motor neuron disease

Oxygen free radical damage is strongly implicated in the pathogenesis of familial motor neuron disease (MND) associated with mutation of the Cu Zn superoxide dismutase gene, and may be relevant in sporadic MND. Selenium (Se) and iron (Fe) have important roles in free radical metabolism. Using neutron activation analysis we have demonstrated significant elevation of Se and Fe in lumbar spinal cord in MND cases (38) compared to controls (22). Analysis of enzymes involved in free radical scavenging showed a significant and specific increase in the activity of the selenoprotein enzyme glutathione peroxidase in MND spinal cord.

Sequences specific for enterovirus detected in spinal cord from patients with motor neurone disease.

To investigate the association of enteroviruses with motor neurone disease, also known as amyotrophic lateral sclerosis. Analysis by enterovirus polymerase chain reaction of wax embedded material from spinal cords taken at necropsy from subjects with motor neurone disease and from age and sex matched controls. Specimens were collected in the west of Scotland and in London between 1982 and 1992. Sequences specific for a non-poliovirus type enterovirus were detected in spinal cord tissue from subjects with motor neurone disease. Amplification of a 414 base RNA target sequence in the conserved enterovirus 5' untranslated region from wax embedded tissue sections was successful in tissue from eight of 11 cases of sporadic motor neurone disease, one of two cases of familial motor neurone disease, and the one case of poliomyelitis, but not in the six matched controls or one case of antecedent poliomyelitis. In addition, sequences were detected in spinal cords from one monkey infected with wild type poliovirus and one monkey infected with polio vaccine. Comparison of sequences from cases of motor neurone disease with sequences of corresponding regions of the 5' untranslated regions of known picornaviruses showed them to be tightly grouped within the enterovirus genus closely related to coxsackievirus type B but not to polioviruses. Sequences derived from different parts of the spinal cord of the same subjects were identical, but sequences differed between individual subjects. Conserved enteroviral sequences closely related to coxsackie B virus sequences were detectable in spinal cords from subjects with sporadic motor neurone disease and from one subject with possible familial motor neurone disease.

Heparin sulfate-like immunoreactivity in the spinal cord in motor neuron disease.

The spinal cords from eight autopsy cases of sporadic motor neuron disease (MND) and two control cases were immunohistochemically examined using antibodies directed to neurofilament proteins (anti-Nf) and to heparan sulfate (HepSS-1). Variable numbers of spheroids were observed in the anterior horns in the MND cases. In one case of MND, one third to half of the remaining anterior horn cells contained conglomerate inclusions in their perikarya. These pathological structures were not encountered in the control cases. The immunohistochemical study revealed that both anti-Nf and HepSS-1 intensely labelled all spheroids and conglomerate inclusions in the MND cases. The colocalization of heparan sulfate with neurofilamentous accumulation suggests that heparan sulfate is required for the aggregation of neurofilaments, resulting in the formation of spheroids and conglomerate inclusions in MND.

Utility of Scottish morbidity and mortality data for epidemiological studies of motor neuron disease.

To determine the accuracy of (1) hospital discharge data and (2) death certificates, coded as motor neuron disease (MND). Comparison of data from The Scottish Motor Neuron Disease Register (SMNDR) with routinely collected Scottish Hospital In-Patient Statistics (SHIPS) and death certificate coding. Scotland UK. 1) 379 adults (> 15 years) discharged for the first time from a Scottish hospital in 1989-90 and (2) 281 deaths in the same period assigned to the International Classification of Diseases (ICD)-9, category 335 (MND). The sensitivity and positive predictive value of a diagnosis of MND as retrieved by (1) the Information and Statistics Division of the Common Services Agency for the Scottish Health Service for morbidity data and (2) the Registrar General's office for mortality data, using the SMNDR as the 'gold standard'. (1) Thirty per cent of adult patients identified as having MND by SHIPS did not have this disease and 23% of patients with MND did not appear on SHIPS. The sensitivity of a diagnosis of MND, as retrieved by SHIPS, was 84% and the positive predictive value was 70% overall. Miscoding of patients with pseudobulbar palsy caused by cerebrovascular disease was the major source of false positive error. The incidence of adult onset sporadic MND was over estimated by SHIPS by a factor of 1.6. (2) Mortality data were more accurate, with a false negative rate of 6% and a positive predictive value of 90%. Coded hospital discharge data are an inaccurate record of a diagnosis of MND and cannot, in their present form, be used as a reliable measure of disease incidence in Scotland. Greater care is required in the preparation of discharge summaries and coding if these data are to be useful for health care planning and epidemiological research. SHIPS is, however, an important source of information to achieve a complete sample of patients with MND. There is also a problematic false positive rate for mortality data but this source more closely approximates true incidence.

Eosinophilic cytoplasmic inclusions in sporadic equine motor neuron disease: an electron microscopic study.

Equine motor neuron disease (EMND) is a sporadic, progressive neurodegenerative disorder that has been identified recently in horses of different breeds in North America. The cause is unknown. Pathologic changes which occur in spinal and certain brain stem motor neurons include chromatolysis, swelling, neurofilamentous accumulation, and development of eosinophilic cytoplasmic inclusions. Punctate eosinophilic inclusions, the type usually encountered in degenerating neurons, resembled Bunina bodies at the light microscopic level, but differed in their ultrastructural composition. These and less common but larger juxtanuclear inclusions appeared to be aggregated vesicular residues of membranous organelle degradation. The third kind of eosinophilic inclusion was a marginated derivative of endoplasmic reticulum and consisted of large membrane-bound accumulations of finely granular material. It was concluded that, although the distribution and nature of the lesions in EMND appeared similar to those of human motor neuron disease, none of the equine eosinophilic inclusions duplicated the ultrastructure of Bunina bodies.