Calcitonin gene-related peptide immunoreactivity in spinal spheroids in motor neuron disease

Summary The spinal cords from seven autopsy cases of sporadic motor neuron disease (MND) and two controls were immunohistochemically examined using anti-bodies directed to calcitonin gene-related peptide (CGRP) and to neurofilament proteins (Nf). CGRP immunoreactivity was observed in the posterior horns, especially in the laminae I and II, of all the spinal cords examined. In MND cases in addition, a considerable number of spheroids in the anterior horns were labelled with the antibody. In some spheroids, their entire area was homogeneously immunostained, whereas in others the immunoreactivity was confined to a focal area(s) within the profile of the spheroids and between these two forms of staining several variations of the staining patterns were seen. The anti-Nf intensely and homogeneously labelled all spheroids and there was no appreciable difference in the Nf-immunoreactive pattern, between CGRP-posittive and-negative spheroids. It is possible that the accumulation of CGRP in spheroids may result from entrapment of CGRP during the anterograde axonal transport causing loss of CGRP at the neuromuscular junction and producing weakness and atrophy of the muscles.Supported in part by the Grant-in-Aid for Encouragement of Young Scientists (the Ministry of Education, Science and Culture, Japan) and in part by the Amyotrophic Lateral Sclerosis Society (USA)     

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.     

FKBP12 immunoreactivity in the human spinal cord of motor neuron disease patients

We investigated the FKBP12 immunoreactivity in the spinal cord of neurological controls and motor neuron disease (MND) patients. In the neurological controls, the spinal neurons were markedly stained with antihuman FKBP12 (N‐19 and C‐19) antibodies. FKBP12 immunoreactivity was associated with lipofuscin in formalin‐fixed paraffin‐embedded samples. In an electron microscopic view, the 10‐nm colloidal gold particles labeled by the anti‐FKBP12 (N‐19) antibody were present on the lipofuscin of the spinal anterior horn neurons. In the MND cases, atrophic neurons with an abundance of lipofuscin granules in the anterior horns of the spinal cord were mildly stained with the anti‐FKBP12 (N‐19 and C‐19) antibodies. Normal‐appearing neurons, inclusion‐laden neurons and chromatolytic neurons of MND cases were weak or negatively stained with anti‐FKBP12 (N‐19) antibodies. These findings suggest that FKBP12 (N‐19) may decrease in the early stages of degeneration in MND. Complexes of FKBP12 and ligands were reported to have neuroprotective and/or neuroregenerative properties. It is speculated that the decrease in FKBP12 (N‐19) plays some causative role in the development of neurodegeneration in MND. Further investigation of FKBP12 and ligands may help elucidate the pathogenesis of MND.     

运动神经元病血清特异抗原成分的检测

目的检测运动神经元病（ＭＮＤ）病人血清中是否存在运动神经元特异抗原成分，并探索ＭＮＤ潜在的诊断标志物。方法制备５株抗运动神经元单克隆抗体，并证明其对大鼠脊髓前角运动神经元具有高度特异的免疫组织化学反应。应用抗运动神经元单克隆抗体２４Ｂ０－ＭｃＡｂ，用ＥＬＩＳＡ法对２５例运动神经元病病人血清中的特异抗原成分进行检测。根据临床表现将２５例病人分为肌萎缩侧索硬化（ＡＬＳ）、脊肌萎缩症（ＳＭＡ）及进行性球麻痹（ＰＢＰ）３组，再按年龄段分３个亚组（＜２０岁组、２０～３９岁组、＞４０岁组）。结果发现８５％（２２／２５）临床确诊的ＭＮＤ病人存在较高浓度的特异抗原成分，ＭＮＤ病人与正常对照组对２４Ｂ０－ＭｃＡｂ的反应性差异有显著性意义（Ｐ＜０．０５），ＡＬＳ、ＳＭＡ及ＰＢＰ亚型之间差异也有显著性意义（Ｐ＜０．０５），而年龄组之间差异虽有显著性意义，其临床意义尚需进一步研究。性别组之间的差异无显著性意义。结论ＭＮＤ病人血清中存在运动神经元特异抗原成分。用抗运动神经元单克隆抗体以ＥＬＩＳＡ法检测运动神经元特异抗原可以作为诊断ＭＮＤ的辅助检查。     

Decreased synaptophysin immunoreactivity of the anterior horns in motor neuron disease

This study concerns the synaptophysin expression in anterior horn neurons of 15 patients with amyotrophic lateral sclerosis and of 4 patients with lower motor neuron disease, who had no upper motor neuron and corticospinal tract involvement. Immunohistochemical procedures were employed and specimens from 13 patients without neurological disease served as controls. A decrease in synaptophysin expression was observed in the anterior horn neuropil of all motor neuron disease patients and this reduction was correlated with the degree of degeneration or neuronal loss of anterior horn cells. Synaptophysin immunoreactivity was preserved in the proximal dendrites and around the somata of the remaining normal-appearing neurons, but it was reduced around the somata and dendrite, especially the distal portion of the proximal dendrites of small degenerated neurons with central chromatolysis, pigmentary atrophy, or simple neuronal atrophy. These observations suggest that presynaptic terminal loss is not secondary to motor cortical neuronal loss, but that the synaptic alterations in anterior horns occur in the wake of motor neuron degeneration.Supported by a Grant-in-Aid for General Scientific Research (C) from the Japanese Ministry of Education, Science and Culture, and a research grant for New Drug Development in ALS from the Ministry of Health and Welfare in Japan     

运动神经元病162例的节段性运动神经传导测定分析

目的探讨运动神经元病(motor neuron d isease,MND)常规节段运动神经传导和位移技术检测的特点。方法对162例MND患者和60名健康对照进行常规节段运动神经传导测定,同时对部分神经采用位移技术测定,并进行分析比较。结果(1)健康人常规节段运动神经传导测定显示:近端与远端比较,波幅和面积下降程度均小于20%,时限增宽小于15%;(2)在MND患者,常规节段测定共有76个节段(5.57%)波幅下降超过20%,45个节段(3.30%)面积下降超过20%,76个节段(5.57%)时限延长超过15%。仅有4例(2.5%)患者4条神经的4个常规节段(0.29%)达到运动神经部分性传导阻滞标准,但采用位移技术测定时均未达到短节段传导阻滞的诊断标准。结论在大部分MND患者常规节段运动神经传导测定正常,在部分患者也可以出现“传导阻滞样”的电生理表现,但其发生率极低,进一步采用位移技术测定有助于鉴别。     

Oculomotor abnormalities in motor neuron disease

To determine whether there are oculomotor abnormalities in motor neuron disease (MND), electrooculographic recordings were performed prospectively in 16 MND patients and the results compared with age-matched healthy controls. Parameters analysed included random and fixed saccades (latency, velocity and accuracy), smooth pursuit (gain, total harmonic distortion and number of saccadic intrusions) and optokinetic nystagmus (maximal and mean slow component velocity). Increased saccadic latencies and decreased smooth pursuit gain were the main alterations in the MND group. Correlation with clinical variables showed a positive relationship between smooth pursuit saccadic intrusions and the bulbar clinical score and the rate of progression and a lower optokinetic nystagmus maximal velocity in patients with pseudobulbar syndrome. Our results demonstrate the presence of subclinical supranuclear abnormalities in MND, and support the notion that MND is not merely a degeneration of the motor system.     

Sporadic motor neuron disease with severe sensory neuronopathy

We report a 62-year-old man with sporadic motor neuron disease (MND) of 52 months’ duration with progressive sensory disturbance and high cerebrospinal fluid protein content. Neuropathologically, both the upper and lower motor neuron systems were severely affected, and light and electron microscopy revealed Bunina bodies and skein-like inclusions, which are characteristic of amyotrophic lateral sclerosis, in the remaining anterior horn cells. Moreover, there was severe degeneration without inflammatory infiltrates in the spinal posterior columns, spinal ganglia, and peripheral sensory nerves. These findings suggest that this case may be an unusual variant of sporadic MND with severe somatic sensory system involvement. Received: 27 July 1997 / Accepted: 29 October 1997     

平山病误诊运动神经元病二例临床分析

目的探讨平山病诊断要点,提高临床医生对该病认识,降低误诊率。方法报道我院近两年被误诊为运动神经元病的2例平山病病例,结合文献复习就平山病与运动神经元病的临床特点、神经电生理特性以及影像学诊断要点进行总结分析和鉴别。结果 :结合文献报道分析,平山病平均起病年龄16.91±2.47岁,男女比18.6:1,均表现以上肢远端受累为主的肌无力和肌萎缩,其中寒冷麻痹72.3%,手伸展震颤74.3%,肌电图示神经源性异常肌电改变,屈颈位颈椎MR有特征性改变:硬膜外腔见月牙形异常信号或流空血管影。结论 "寒冷麻痹及手平举震颤",是平山病临床表现,也是避免误诊的重要鉴别要点。颈椎MR特征性改变对确诊平山病具有重要价值。     

Electrophysiological diagnosis of motor neuron disease and pure motor neuropathy

Motor neuron disease (MND) is a group of disorders in which there is degeneration of upper and lower motor neurons to a variable degree. Amyotrophic lateral sclerosis is the most frequent form of the disease, presenting with both upper and lower motor neuron involvement. Frequently, especially in the early stages of the disease, only lower motor neuron signs are present. In these conditions, some pure motor neuropathies may resemble MND. The diagnosis is of importance because some of these motor neuropathies are “dysimmune” disorders and may respond to immune therapies. In such diseases the multifocal motor neuropathy with conduction block appears to be the more frequent. In MND and pure motor neuropathies, the electrophysiological examination is the most decisive test. In MND, it is of diagnostic importance. In addition, it is useful in the assessment of disease severity and progression, in the evaluation of therapeutic trials and in the understanding of etiopathogenesis of the disease. In pure motor neuropathies, the presence of conduction block leads to immune treatment with good response in more than 50% of the cases. Received: 20 August 1998 Accepted: 10 October 1998     

运动神经元病患者的记忆功能研究

目的观察运动神经元病(motor neuron disease,MND)患者是否存在记忆功能障碍;并研究患者的病程对其记忆功能的影响。方法对比分析了32例MND患者与60例正常对照的临床记忆量表评分,并比较不同病程的MND患者的记忆功能评分。结果MND患者记忆商、临床记忆量表总等值量表分、指向记忆等值量表分、无意义图形再认等值量表分评分均明显低于对照组(P<0.05),且不同病程的MND患者临床记忆量表各项评分比较无显著性差异(P>0.05)。结论MND患者大多存在记忆障碍,提示MND存在运动区域以外的脑组织受累。MND患者的记忆障碍与病程长短无明显关系。     

New insights in motor neuron disease

In motor neuron disease there is a characteristic pattern of nerve cell loss and degeneration of related pathways. In surviving anterior horn cells several morphologically distinct, but generally non-specific, intracytoplasmic inclusion bodies have been recognized. Recently accumulations of previously unrecognized ubiquitinated material have been described in surviving neurons, which cannot be demonstrated with routine histological methods. These changes appear unique to this disease, and provide a new insight into the underlying pathology that may help understand the pathogenesis of this intriguing disorder. In this article we review the new information on the clinical, toxicological and pathological features of the disease.     

Stiff person syndrome associated with lower motor neuron disease and infiltration of cytotoxic T cells in the spinal cord

We present a 67-year-old non-diabetic male who presented with muscle cramps, paresis, atrophy and fasciculations in the left leg, followed by rapidly progressive muscle stiffness and superimposed spasms which subsequently also affected the right leg and the trunk. GAD65 autoantibodies were elevated in serum and CSF, compatible with systemic and intrathecal synthesis of oligoclonal and high-avidity autoantibodies, and GAD65 specific T cells were clonally expanded in the CSF. The patient did not respond to GABAergic and immunomodulatory treatment or plasma exchange, and died from respiratory failure after 18 months. Autopsy revealed unilateral axonal swelling, chromatolysis and vacuolisation of anterior horn cells of the lower spinal cord, accompanied by microglia proliferation and discrete infiltration of CD8+ cytotoxic T cells. No CD4+ T helper cells, B cells or complement deposition were detected. To our knowledge, this is the first report of stiff person syndrome with lower motor signs restricted to a lower limb, and also the first attempt to characterize the infiltrating T cells. The finding of CD8+ cytotoxic T cells in the absence of B cells in the inflamed area of the spinal cord suggests that the intrathecal synthesis of GAD65 autoantibodies takes place in areas of the CNS not strictly related to the clinically relevant lesions.     

Synaptic loss in anterior horn neurons in lower motor neuron disease

This report concerns an ultrastructural investigation of the synapses of anterior horn neurons in the lumbar spinal cord of four patients with lower motor neuron disease (LMND) who had no upper motor neuron and corticospinal tract involvement. Anterior horn neurons of five normal individuals served as controls. The cell body area and the number of synapses of the normal-appearing neurons of the LMND patients were significantly reduced (P < 0.0001). These findings suggest that synaptic changes of anterior horn neurons could be ascribed to the degeneration of lower motor neurons rather than to the influence of upper motor neuron system degeneration. On the other hand, the lengths of individual synapses (P < 0.0001) and of their active zones (P < 0.05) were significantly increased in the patients. These increases would indicate that synapses on anterior horn neurons of individuals with LMND appear to have the capacity to react to progressive degeneration and loss of other synapses by means of a compensatory response or plasticity that enhances their efficiency. Received: 4 September 1995 / Revised: 3 November 1995 / Accepted: 16 November 1995     

三叉神经-颈反射对运动神经元病球部损害的诊断价值

目的 建立三又神经-颈反射(TCR)的肌电检测方法,探讨其对运动神经元病(MND)球部损害的诊断价值。方法受检测者取仰卧位,头部轻度抬高。刺激一侧眶下神经,于双侧胸锁乳突肌记录。结果 刺激对照者一侧的眶下神经,可于双侧胸锁乳突肌引出正/负波。MND组:7例正常(23.3％),8例未引出(26.7％),11例潜伏期延长(36.7％),4例双侧反射超常不对称(13.3％)。结论TCR能够可靠测定,作为颈-球区病变的一种辅助检查手段,有助于运动神经元病的早期诊断。     

Osteopontin is an alpha motor neuron marker in the mouse spinal cord

Motor neurons (MNs) are designated as alpha/gamma and fast/slow based on their target sites and the types of muscle fibers innervated; however, few molecular markers that distinguish between these subtypes are available. Here we report that osteopontin (OPN) is a selective marker of alpha MNs in the mouse spinal cord. OPN was detected in approximately 70% of postnatal choline acetyltransferase (ChAT)-positive MNs with relatively large somas, but not in those with smaller somas. OPN+/ChAT+ MNs were also positive for NeuN, an alpha MN marker, but were negative for Err3, a gamma MN marker. The size distribution of OPN+/ChAT+ cells was nearly identical to that of NeuN+/ChAT+ alpha MNs. Group Ia proprioceptive terminals immunoreactive for vesicular glutamate transporter-1 were selectively detected on the OPN+/ChAT+ cells. OPN staining was also detected at motor axon terminals at neuromuscular junctions, where the OPN+ terminals were positive or negative for SV2A, a marker distinguishing fast/slow motor endplates. Finally, retrograde labeling following intramuscular injection of fast blue indicated that OPN is expressed in both fast and slow MNs. Collectively, our findings show that OPN is an alpha MN marker present in both the soma and the endplates of alpha MNs in the postnatal mouse spinal cord.     

Functional and morphometric study of the liver in motor neuron disease

Summary In routine liver function tests, 23 of 44 patients with motor neuron disease (MND) had abnormal findings, and there was disturbance of unconjugated bilirubin metabolism in 10 of the 33 patients tested. Liver-biopsy specimens from 10 MND patients were compared by electron microscopic examination with specimens from age-matched controls who had chronic persistent hepatitis. The MND patients had a higher incidence of intramitochondrial inclusions, less abundant mitochondria in a given area of cytoplasm and enlarged mitochondria. Electron-probe X-ray microanalysis of hepatocytic lysosomes found copper in 8 of 13 MND patients, but not in the controls. These findings suggest that the pathogenetic processes in MND may involve not only motor neurons but also hepatic cells.     

Spontaneous lower motor neuron disease in rabbits (Oryctolagus cuniculus)

Summary Spontaneous neurologic disease was observed in 6 to 8-week-old rabbits. Both males and females from several different litters were affected but all were sired by the same male. Clinically, the disease was characterized initially by posterior weakness and incoordination which progressed to tetraplegia within 3–4 weeks. With light microscopy there was neuronal degeneration and loss within the ventral horns of the spinal cord and brain stem and type-II fiber atrophy of skeletal muscles. Ultrastructurally the neuronal degeneration was charactered by accumulations of 100 Å neurofilaments within the perikaryon. These findings are compared to diseases with neurofibrillary accumulation in animals and man.This work was supported in part by NIH grant RR00685-05 and by the Scott-Ritchy Research Fund of Auburn University     

Radiological evidence of subclinical dysphagia in motor neuron disease

Dysphagia in motor neuron disease (MND) may lead to dangerous complications such as cachexia and aspiration pneumonia. Functional evaluation of the oropharyngeal tract is crucial for identifying specific swallowing dysfunctions and planning appropriate rehabilitation. As part of a multidisciplinary study on the treatment of dysphagia in patients with neuromuscular diseases, 23 MND patients with different degrees of dysphagia underwent videoflouroscopy, videopharyngolaryngoscopy and pharyngo-oesophageal manometry. The results of the three instrumental investigations were analysed in order (1) to define the pattern of swallowing in MND patients complaining of dysphagia; (2) to evaluate whether subclinical abnormalities may be detected; and (3) to assess the role of videofluoroscopy, videopharyngolaryngoscopy and manometry in the evaluation of MND patients with deglutition problems. Correlations between the instrumental findings and clinical features (age of the patients, duration and severity of the disease, presence and degree of dysphagia) were also assessed. The results of our study showed that: (1) The oral phase of deglutition was compromised most often, followed by the pharyngeal phase. (2) In all patients without clinical evidence of dysphagia, subclinical videofluoroscopic alterations were present in a pattern similar to that found in the dysphagic group. (3) Videofluoroscopy was the most sensitive technique in identifying oropharyngeal alterations of swallowing. Impairment of the oral phase, abnormal pharyngo-oesophageal motility and incomplete relaxation of the upper oesophageal sphincter were the changes most sensitive in detecting dysphagia. Videofluoroscopy was also capable of detecting preclinical abnormalities in non-dysphagic patients who later developed dysphagia. Practical guidelines for the use of instrumental investigations in the assessment and management of dysphagia in MND patients are proposed. Received: 24 March 1997 Received in revised form: 11 November 1997 Accepted: 18 November 1997