Skein-like Inclusions Research Articles

Characterizations of heterotopic neurons in the spinal cord of amyotrophic lateral sclerosis patients.

This report concerns a comparative immunocytochemical, ultrastructural and morphometric investigation on heterotopic neurons in the white matter of the spinal cords of 19 patients with amyotrophic lateral sclerosis (ALS) and 18 age-matched neurologically normal individuals. The study revealed that the heterotopic neurons were scattered in the white matter, often adjacent to gray matter, that they immunoreacted with the antibody to synaptophysin, and that there were synaptic apparatuses on the surface of their somata and their neuronal processes. Bunina bodies and ubiquitin-positive inclusions such as Lewy body-like inclusions and skein-like inclusions, characteristic of anterior horn neurons of ALS, were present in the cytoplasm of the patients' heterotopic neurons in the anterior or lateral column of the white matter. These findings suggest that heterotopic neurons in the anterior or lateral column have the characteristics of alpha motor neurons. The average number of heterotopic neurons observed in ALS patients was generally less than in normal subjects. This reduction was correlated with the severity of neuronal loss. The heterotopic neurons in ALS were less susceptible to the degenerative process as compared with spinal cord anterior horn cells. We assume that in this disease the heterotopic neurons may be degenerated and their number diminished after or concomitantly with the depletion of anterior horn neurons.

Neuropathology of sporadic amyotrophic lateral sclerosis of long duration

We performed post-mortem examinations of three patients with progressive neurogenic amyotrophy of long duration. One patient had been clinically diagnosed as having sporadic amyotrophic lateral sclerosis (ALS) and two had been diagnosed with progressive spinal muscular atrophy (PSMA). The disease durations were 10, 17 and 20 years, respectively, and all of the patients died of respiratory failure with no artificial respiratory support. In all of the patients, both the upper and lower motor neuron systems were affected; degeneration of the former was definite, but was milder than that usually encountered in sporadic ALS patients, and the histopathology of the latter was identical to that of sporadic ALS. Light microscopy revealed Bunina bodies, which are characteristic of sporadic ALS, in the remaining anterior horn cells of each patient. In addition, ubiquitin-positive skein-like inclusions were also identified, immunohistochemically, in the remaining anterior horn cells of each patient. Neuron counts indicated that the number of neurons was preserved in Clarke's column in these patients, but was significantly reduced in the intermediolateral nucleus, compared with control subjects. Based on these findings, we think that these three patients, with long disease durations, were affected by essentially the same underlying disease process as that of sporadic, classical ALS. Moreover, we question the neuropathological occurrence of sporadic ALS without involvement of the upper motor neuron system, namely, pure PSMA or lower motor neuron disease. © 1997 Elsevier Science B.V. All rights reserved.

Golgi apparatus and intraneuronal inclusions of anterior horn cells in amyotrophic lateral sclerosis: an immunohistochemical study

We applied immunohistochemical techniques to study alterations of the Golgi apparatus, and to determine possible relationships between this complex and the intraneuronal inclusions of lower motor neurons of patients with the sporadic form of amyotrophic lateral sclerosis (ALS). Spinal cords from normal individuals served as controls. Monoclonal antibodies to the Golgi zone and to Golgi beta-COP protein were used. Immunoreactivity with these antibodies was seen in frozen sections of paraformaldehyde-fixed spinal cord tissue, but not in formalin-fixed, paraffin-embedded specimens. The immunoreaction products were seen as granular structures, diffusely distributed in neurons and glial cells. Although immunostaining of some ALS neurons was reduced, fragmentation of the Golgi apparatus was not evident with either antibody. Bunina bodies and skein-like inclusions, characteristically found in spinal anterior horn cells of ALS patients, were not stained by these antibodies to epitopes of the Golgi apparatus, nor were Lewy body-like hyaline inclusions, present in some cases of sporadic ALS and reported to be characteristic to familial ALS with posterior column degeneration. These results suggest that components of the Golgi apparatus are not directly incorporated into intraneuronal inclusions. However, the possibility that abnormal proteinaceous material of the Golgi apparatus may be involved in their genesis cannot be ruled out.

Immunohistochemical study on superoxide dismutases in spinal cords from autopsied patients with amyotrophic lateral sclerosis.

A role mutations in the superoxide dismutase (SOD)-1 gene in the pathogenesis of amyotrophic lateral sclerosis (ALS) has been discussed. To investigate immunohistochemical alterations of SOD in the spinal cord affected with the disease, we examined 3 patients with SOD1 mutation-associated family with ALS, 20 patients with sporadic ALS and 10 control individuals. Lewy body-like hyaline inclusions (LBHIs) were seen in the anterior horn cells of all the familial patients and 10 of the 20 sporadic patients, while skein-like inclusions (SIs) and Bunina bodies (BBs) were present in the 20 sporadic patients but not in the familial patients. The primary antibodies used for immunostaining were rabbit antisera raised against human SOD1 and SOD2. The anti-SOD1 antibody reacted strongly with all LBHIs of each familial patient and with some LBHIs of each sporadic patient. The cytoplasm of morphologically intact and degenerated spinal cord neurons as well as spheroids seen in the cases examined was only weakly stained by the antibody to SOD1 or not at all. The reactive astrocytes displayed weak to moderate staining for SOD1. The anti-SOD2 antibody strongly immunolabeled the reactive astrocytes and microglia. LBHIs of both familial and sporadic ALS were negatively stained for SOD2. Spinal cord neurons and spheroids of each case exhibited no significant SOD2 immunoreactivity. Neither antibodies reacted with SIs nor BBs. These results indicate that SOD1 may be involved in the formative process of LBHIs especially in familial ALS but not always in that of SIs or BBs, and imply that SOD2 may have no connection with any of these ALS-related abnormal structures.

Ultrastructural abnormalities with inclusions in Onuf's nucleus in motor neuron disease (amyotrophic lateral sclerosis).

This study describes an ultrastructural examination focused on motor neurons in Onuf's nucleus in the spinal cord of four control patients without neurological disease (45-70 years) and six motor neuron disease (MND) patients (38-79 years; duration 8 months-19 years) who showed no somato-vesical dysfunction. Prompted by recent studies suggesting some sphincteric motor neurons may succumb to MND, this study sought to determine whether the wider population of neurons in Onuf's nucleus display ultrastructural cytopathology which is normally undetectable in histological preparations. Spinal cords were removed 3-20 h after death, and 1 mm slices of cord rapidly fixed in modified Karnovsky medium were processed for both light- and electronmicroscopy. 'Control motor neurons' had intact neuronal and nuclear membranes. Nissl bodies chiefly comprised ordered structures of alternate lamellae of rough endoplasmic reticulum and arrays of polyribosomes. The Golgi complexes consisted of multilamellated curvilinear stacks of ER. No intraneuronal filamentous or Bunina body inclusions were observed, but occasional axonal spheroids were seen in the neuropil. In MND, histological evidence of sparing in Onuf's nucleus was associated with abnormal ultrastructure of the motor neurons. Some sphincteric neurons were atrophic, whereas in the others, Nissl bodies were reduced in number, showed loss of structural organization or comprised polyribosomal aggregates. Golgi complexes had disrupted lamellated organization or consisted solely of distended ER. Intraneuronal filamentous Lewy-body or skein-like inclusions and Bunina bodies were identified in Onuf's nucleus of three subjects (duration of MND 8 months-2 years). The results of the present study indicate that Onuf's nucleus is vulnerable in MND, and preservation of sphincter function with qualitative histological evidence of 'sparing' does not necessarily imply a corresponding lack of ultrastructural cytopathology in this nucleus.

Formula omitted] superoxide dismutase-like immunoreactivity in Lewy body-like inclusions of sporadic amyotrophic lateral sclerosis

Cu Zn superoxide dismutase (SOD)-like immunoreactivity (LI) was found within Lewy body-like inclusions (LBIs) in the spinal cords of patients with sporadic amyotrophic lateral sclerosis (ALS) by using an antibody to human Cu ZnSOD . LBIs were detected in the anterior horn cells in 10 of 20 patients with sporadic ALS. In each of these patients, 7 to 60% of LBIs showed Cu ZnSOD-LI . No Cu ZnSOD-LI was detected in intact neurons and glia in the 20 ALS patients, as well as in these cells in 10 normal control individuals. The skein-like inclusions and Bunina bodies, which were found in all of the 20 ALS cases, showed no Cu ZnSOD-LI . Thus, Cu ZnSOD appears to play roles in the production and/or degradation process of LBIs.

Ubiquitin and neurofilament expression in anterior horn cells in amyotrophic lateral sclerosis: possible clues to the pathogenesis.

Cytoskeletal abnormalities are a prominent pathological feature of anterior horn cells in amyotrophic lateral sclerosis (ALS), and are thought to be involved in the process of motor neuron death. Skein-like filamentous inclusions have been detected by immunocytochemical staining for ubiquitin, a stress protein involved in targeting abnormal proteins for proteolysis. So far, identification of the target protein has been elusive. We have studied the ultrastructural localization of ubiquitin and neurofilaments by post-embedding immunogold staining. In skein-like arrays, strong ubiquitin labelling was concentrated on abnormally formed 15-20 nm filaments; neurofilament labelling was localized on 10 nm filaments adjacent or in continuity with the abnormal filaments. In addition, Bunina bodies were a major site of ubiquitin accumulation. Our results suggest that ubiquitinated filaments in skein-like inclusions might originate from abnormally aggregated neurofilament proteins, which are no longer recognized by antibodies to neurofilament epitopes. Furthermore, the presence of ubiquitin in Bunina bodies suggests that, in addition to its protective role, ubiquitin might be directly implicated in the mechanism of programmed neuronal death in ALS.

Immunocytochemical and ultrastructural studies of the motor cortex in amyotrophic lateral sclerosis.

This report concerns an immunocytochemical and ultrastructural study of the motor cortices of 11 patients with amyotrophic lateral sclerosis (ALS). Specimens from 12 normal individuals served as controls. Antibodies against phosphorylated neurofilament (PNF; 200 kDa), ubiquitin, glial fibrillary acidic protein (GFAP) and phosphorylated tau protein were used. The pyramidal cells of layer III of all ALS patients were stained, with varying intensities, by the antibody to PNF. By contrast, Betz cells reacted less frequently with this antibody. Staining for GFAP was noted in numerous astrocytes in layer III and at the transition between white matter and motor cortex of most patients. Ubiquitin-positive inclusions were only occasionally seen in Betz cell and pyramidal cell of layer V. These observations indicate that alterations of the motor cortex occur first in the pyramidal cells of layer III rather than in Betz cells. Pyramidal cells and Betz cells were not stained by the antibody to phosphorylated tau protein. In controls, pyramidal cells and Betz cells were less frequently stained with the anti-neurofilament antibody than those from ALS patients. Immunoreactivity of GFAP in layer III and at the junction of white matter and motor cortex was observed in only one patient. Ultrastructural examination revealed that the Betz cells of some ALS patients had Bunina bodies (BB), Lewy body-like inclusions (LBI) and skein-like inclusions (SI), as well as bundles of filaments that were thicker than neurofilaments; some of these filaments appeared to be constricted. The incidence of these inclusions was lower than that seen in anterior horn neurons.(ABSTRACT TRUNCATED AT 250 WORDS)

On the possibility of autolysosomal processing of skein-like inclusions: Electron microscopic observation in a case of amyotrophic lateral sclerosis

Hyaline-like inclusions (HLIs) and skein-like inclusions (SLIs) were recently observed in the lower motor neurons in amyotrophic lateral sclerosis (ALS). HLIs mainly consist of randomly arranged neurofilaments and abnormally large (about 15 nm) fibrils, while SLIs comprise bundles of the abnormally large fibrils alone. Although the abnormal fibrils have been reported to be ubiquitinated in both types of inclusions, their final catabolic destination in the neurons is unknown. Electron microscopic observation of the lower motor neurons in an ALS case revealed several neurons containing SLIs in addition to ones containing HLIs. Bundles of SLIs, which were located close to HLIs or were observed by themselves without an HLI, were frequently associated with flattened cisterns; some were wrapped, some were completely sequestrated, and some were demarcated together with a portion of moderately electron dense cytoplasm by cisterns or double-membraned units. These features resemble reported early stages in the formation of autolysosomes, and thus indicate that at least some of the bundles of SLIs or parts of them in the lower motor neurons in ALS undergo degradation through the autolysosomal pathway.

A fine structural study of Onuf's nucleus in sporadic amyotrophic lateral sclerosis

Onuf's nucleus of the second sacral spinal cord level was examined in twelve patients with sporadic amyotrophic lateral sclerosis (ALS). Under light-microscopic examination, the neurons of Onuf's nucleus were well-preserved in contrast with the severe neuronal depletion of the anterior horn cells. Ultrastructurally, however, substantial pathological changes such as Bunina bodies (BB), randomly arranged filaments associated with fine granules corresponding to Lewy body-like hyaline inclusions (LBHI) and bundles of tubular filaments corresponding to skein-like inclusions (SI) all of which are characteristic features of ALS, were demonstrated ubiquitously in the Onuf's nucleus in eleven of twelve patients. These changes may be of a reactive nature and represent an initial change of neuronal degeneration, since no definite cell destruction was observed in the nucleus. The changes in the Onuf's nucleus correlated with the changes observed in the anterior horns, although to a lesser degree. The more BB, LBHI and SI were observed in the anterior horns of patients with ALS, the more these inclusions were observed in Onuf's nucleus. Thus, we conclude that Onuf's nucleus is also affected by pathological changes in ALS.

Hyaline and Skein‐like Inclusions in Amyotrophic Lateral Sclerosis

Hyaline (HI) and skein‐like inclusions (SLI) are characteristic of familial and sporadic amyotrophic lateral sclerosis (ALS). The HIs are usually round eosinophilic structures, some of which consist of cores and halos and are called Lewy body‐like HIs. The HIs, composed of various amounts of granule‐associated thick filaments and neurofilaments, have varied appearances and may be heterogeneous. The SLI is basically a thready structure of a bundle of thick filaments. The threads may be present singly or in the form of networks or aggregates with each other. Occasionally, aggregates of threads may form single larger inclusions that resemble HIs. These features indicate that the HI and the SLI may be closely related. Other than their intense ubiquitination, the molecular nature of HI and SLI is still unknown. The investigation of the genesis of these inclusions may provide important clues for elucidating the pathomechanism of ALS.

Ultrastructural study of Bunina bodies in the anterior horn neurons of patients with amyotrophic lateral sclerosis

We ultrastructurally investigated Bunina bodies (BB) in the anterior horn neurons of 20 amyotrophic lateral sclerosis patients with BB. As for novel findings, filaments thicker than neurofilaments were not uncommonly observed inside the BB. They were occasionally observed around the periphery of the BB. Some of them were composed of bundles of filaments which appeared constricted at 40–50 nm intervals and were 20–25 nm in maximum width. Others consisted of bundles of unconstricted filaments measuring about 20–25 nm in diameter. The BB occasionally contained bundles of filaments of about 20 nm in diameter that closely resembled those found in ubiquitin-positive skein-like inclusions. It seems that some molecular disturbances such as ubiquitin play a role in the formation of the constricted and unconstricted filaments. Probably cytoskeletal or non-cytoskeletal proteins in anterior horn cells are damaged and accumulate to form aggregation of the filaments associated with BB.

Ubiquitin-positive inclusion in anterior horn cells in subgroups of motor neuron diseases: A comparative study of adult-onset amyotrophic lateral sclerosis, juvenile amyotrophic lateral sclerosis and werdnig-hoffmann disease

This report concerns the expression of ubiquitin in anterior horn cells of various subgroups of adult and infantile motor neuron disease (MNDs); immunohistochemical techniques were employed. Ubiquitin-positive skein-like inclusions (SLIs) were found in all cases of adult-onset amyotrophic lateral sclerosis (ALS), including 16 cases with sporadic ALS, two cases of familial ALS with posterior column degeneration and Lewy body-like hyaline incusions (LBHIs), two sporadic ALS cases with LBHIs, and three cases of sporadic ALS with dementia. SLIs were not found in anterior horn cells of 5 cases with Werdnig-Hoffmann disease (WHD). However, granular ubiquitin-positive deposits were seen in ballooned neurons of WHD patients. No ubiquitinated materials were found in the perikarya of two sporadic juvenile ALS patients with basophilic inclusions (BIs), but granular ubiquitin-immunoreactive deposits were occasionally observed in the BIs. These results suggest that ubiquitin-positive SLIs are characteristic features of various forms of adult-onset ALS and that aggregated ubiquitinated granules are characteristic of ballooned neurons of WHD. Ubiquitinated structures and their distribution patterns may reflect degenerative processes of anterior horn cells, and may be useful for classifying subgroups of motor neuron diseases.

Oculomotor nuclear pathology in amyotrophic lateral sclerosis

We examined the oculomotor and/or trochlear nuclei of 27 amyotrophic lateral sclerosis (ALS) patients and 10 controls by histological and immunohistological methods. Their neurons were relatively well preserved. In 7 of 22 sporadic ALS patients (including 3/3 ALS with ophthalmoplegia) and in 4 of 5 ALS patients with dementia, some morphological changes similar to those in anterior horns (Bunina bodies, ubiquitin-positive skein-like inclusions, Lewy body-like inclusions, conglomerate inclusions and spheroids) were rarely, but clearly seen. These changes were not observed in controls. Our results suggest that the oculomotor and trochlear nuclei in ALS patients are slightly affected in a manner similar to that in the anterior horns, but the degree is less than that necessary for development of ophthalmoplegia in the majority of ALS patients.

Lewy body-like inclusions in Onuf's nucleus from two cases of sporadic amyotrophic lateral sclerosis

Lewy body-like inclusions in Onuf's neurons from two sporadic cases with amyotrophic lateral sclerosis (ALS) were reported. These inclusions in Onuf's neurons as well as those found in the anterior horn cells were immunostained with an anti-ubiquitin antibody. Neuropathological examination of these two cases revealed neuronal loss and associated gliosis in the anterior horn of the spinal cord and hypoglossal nuclei, and degeneration of the corticospinal tract. In addition to Lewy body-like inclusions, ubiquitinated skein-like inclusions, Bunina bodies, or both were observed in the cytoplasm of the remaining neurons in the anterior horn of the spinal cord, and, to a lesser degree, in Onuf's nucleus. Spheroids and cord-like thickening of cell processes were also found in the anterior horn of the spinal cord. Histometrical study of Onuf's nucleus revealed atrophy and loss of Onuf's neurons from Case 1 with a long clinical course. Similar cases of motor neuron disease with or without tract degeneration have been reported, but the presence of Lewy body-like inclusions in Onuf's nucleus is reported here for the first time. It is suggested that Onuf's nucleus is more or less involved in the degenerative process characteristic of ALS.

Ultrastructural study of skein-like inclusions in anterior horn neurons of patients with motor neuron disease

We investigated the ultrastructure of skein-like inclusions (SI) in 11 patients with motor neuron disease. SI mainly consisted of bundles of filaments. Each filament was approximately 15–25 nm in diameter and had a tubular profile on transverse sections. SI occasionally appeared to contain electron-dense material similar to Bunina bodies (BB), and in clusters of SI, electron-dense material closely resembling BB with vesicles were scattered among the bundles of filaments or were directly attached to the filaments, suggesting a close relationship between SI and BB. We also infer that SI might be the precursor of Lewy body-like hyaline inclusions (LBHI), on the basis of the frequent presence of in-between structures, the coexistence of SI and LBHI, the detection of tubular profiles (component of SI) in LBHI, the far greater frequency with which SI were observed compared with LBHI, and the occasional connection between ubiquitin-positive SI and LBHI observed in preceding immunocytochemical studies.

Skein-like inclusions in the anterior horn cells in motor neuron disease

Skein-like inclusions (SLIs) in the anterior horn cells of patients with motor neuron diseases, including familial amyotrophic lateral sclerosis with posterior column degeneration, sporadic lower motor neuron disease and classical amyotrophic lateral sclerosis, were investigated morphologically with hematoxylin and eosin preparations, immunostaining for ubiquitin and immunoelectron microscopy. The SLIs were thready linear or tubular structures which immunostained with antiubiquitin antibodies. They were detected on hematoxylin and eosin preparations as eosinophilic thread-like structures often surrounded by pale areas. SLIs were occasionally present as networks of threads or tubules. Sometimes, they were aggregated and formed larger pale inclusions. Ultrastructurally, the SLIs were bundles of filaments which appeared thicker than neurofilaments. The SLIs tended to have central hollow spaces which were devoid of filaments. When the SLIs were clustered, fuzzy thick filaments were randomly and loosely arranged among the individual SLIs. The SLIs were histologically and ultrastructurally distinct from other inclusions such as Bunina bodies and hyaline inclusions. This unique morphology of SLIs may provide a novel perspective on the degenerative processes of the anterior horn cells in MND.

Fine structure of cells infected with human cytomegalovirus after treatment with 9-(1,3-dihydroxy-2-propoxymethyl)guanine.

Infection with human cytomegalovirus (CMV) is characterized by cytological changes which are readily visualized by electron microscopy using ultrathin sections of infected cells. Treatment of such cells with 9-(1,3-dihydroxy-2-propoxymethyl)guanine (DHPG), a potent inhibitor of CMV, is effective when initiated at early or late times after infection and the response to such treatment has been studied by fine structural analysis. Inhibition of viral DNA synthesis by DHPG treatment (50 microM) late in virus infection resulted in a cessation of virus growth accompanied by a lack of development and possible regression in skein-like intranuclear inclusions together with a depletion in cytoplasmic dense bodies. Such changes were accompanied by the appearance of nuclear dense bodies. These were also present when virus growth was reduced (5 microM-DHPG) rather than completely inhibited (50 microM-DHPG) by treatment initiated from the time of infection. The nuclear bodies were predominantly of a reticular type structure after the early treatment but mainly of a homogeneous form when virus growth was interrupted at late times. Their presence appeared to be connected with the ability of infected cells to initiate the synthesis of late proteins and their morphology may relate to the extent of such protein synthesis. Unlike cytoplasmic dense bodies, provisional findings on the characterization of the nuclear bodies suggested that the 69K matrix protein was not present in abundance.