Luitgard Weyer

Neurofilament Light and Polyadenylated mRNA Levels Are Decreased in Amyotrophic Lateral Sclerosis Motor Neurons

The presence of large neurofilamentous accumulations in the perikaryon and proximal axon of motor neurons in amyotrophic lateral sclerosis (ALS) suggests that the expression of this abundant cytoskeletal protein may be altered. We performed quantitative in situ hybridization for the low molecular weight neurofilament subunit (NF-L) messenger RNA in six cases of sporadic ALS and six controls. We found a 41% decrease (p < 0.02) in the NF-L mRNA levels in anterior horn cells in ALS, with a 60% decrease (p > 0.01) in alpha motor neurons. This alteration may represent a non-specific response to axonal or neuronal injury or, alternatively, reflect the regenerative activity of residual normal motor neurons. NF-L mRNA levels were consistently low (in the third and fourth quartiles) in spheroid-bearing motor neurons, indicating that the neurofilamentous accumulations observed in ALS are not likely the result of overexpression of the NF-L gene. Total neuronal polyadenylated mRNA levels were also 50% lower (p = 0.02) in anterior horn cells and 48% lower (p > 0.05) in alpha motor neurons in ALS, possibly reflecting a decrease in selected mRNA species in diseased motor neurons.

Cortical degeneration in progressive supranuclear palsy. A comparison with cortical-basal ganglionic degeneration.

We report 3 patients with progressive supranuclear palsy (PSP) who developed limb apraxia, focal dystonia, and arm levitation late in the course of the disease. Neuropathological examination revealed cortical degeneration in addition to the characteristic pathological findings of PSP. Semiquantitative comparative histological and immunohistological studies of the neocortex of these patients as well as 5 cases of classical PSP and 4 cases of cortical-basal ganglionic degeneration (CBGD) revealed a distinctive form of cortical degeneration in PSP. The cortical degeneration was often circumscribed and confined to premotor and motor cortex. It was characterized by neuronal loss and gliosis. Swollen neurons were only rarely observed in neocortex of PSP cases in contrast with CBGD, where they were abundant. Neuronal and glial tau as well as tau immunoreactive threads were seen in both PSP and CBGD, but were more abundant in CBGD. The appearance of tau reactive astrocytes also differed in both disorders; tufted astrocytes were seen exclusively in PSP, while typical annular astrocytic plaques were confined to CBGD. These observations indicate that cortical degeneration occurs in PSP and may be associated with atypical clinical manifestations that lead to diagnostic difficulties.

Copper/zinc superoxide dismutase mRNA levels are increased in sporadic amyotrophic lateral sclerosis motorneurons

Mutations of the Cu/Zn superoxide dismutase (SOD-1) gene were recently implicated in the pathogenesis of familial amyotrophic lateral sclerosis (ALS). We measured SOD-1 mRNA levels in motorneurons of the more common sporadic form of the disease and found a 42% increase in ALS motorneurons (P = 0.058) as compared with controls. These results suggest that oxidative stress may also play a role in the pathogenesis of sporadic ALS.

Characterization of a shared epitope in cortical Lewy body fibrils and Alzheimer paired helical filaments

The straight fibrils of the Lewy body contain an epitope related to phosphorylation of the KSPV motif common to the C termini of the 200- and 170-kDa neurofilament subunits and τ. To further characterize this phosphorylated neurofilament/τ epitope in Lewy bodies and to analyze the constituents of isolated Lewy bodies we used a combined biochemical and immunochemical approach. In formalin-fixed paraffin-embedded tissue cortical Lewy bodies were labelled by monoclonal antibodies directed to phosphorylation-dependent KSPV epitopes in the sequences of neurofilament and phosphorylation-independent epitopes. Immunoblotting of solubilized Lewy body fibrils with the same antibodies which stained Lewy bodies in tissue sections revealed that the immunoreactive Lewy body proteins were phosphorylated neurofilament subunits. An antibody to the 68-kDa neurofilament subunit labelled Lewy bodies and Lewy body protein at 50–68 kDa. We conclude that the shared phosphorylated epitope in Lewy body fibrils and paired helical filaments is related to the common KSPV sequence in neurofilament and τ, and that all three neurofilament subunits are present in the Lewy body. This result indicates that although Lewy bodies and neurofibrillary tangles share epitopes they are comprised of distinct structural subunits.

Detergent‐Insoluble Cortical Lewy Body Fibrils Share Epitopes with Neurofilament and τ

Abstract: Lewy bodies are cytoskeletal inclusions associated with neuronal injury and death in idiopathic Parkinsons disease and other neurodegenerative disorders. The chemical composition of the 8–10‐nm fibrils of the Lewy body is unknown, although they are related to both normal cytoskeletal elements and paired helical filaments of Alzheimer neurofibrillary tangles. From the Lewy body‐rich cerebral cortex of patients with diffuse Lewy body disease we have isolated intact Lewy bodies using a high salt buffer/nonionic detergent gradient centrifugation procedure and extracted the constitutive fibrils with urea and sodium dodecyl sulfate. Urea/detergent‐resistant Lewy body fibrils were solubilized with formic acid and found to contain a single protein band of 68 kDa, which was not found in identically prepared normal brain homogenates. The Lewy body derived‐polypeptide was recognized on immunoblots by a polyclonal antibody that reacted with both the 68‐kDa neurofilament subunit and the microtubule‐associated protein τ. The 68‐kDa Lewy body protein was not labeled by the monoclonal antibody τ‐1 despite prior in vitro enzymatic dephosphorylation. We conclude that the detergent‐insoluble component of the cortical Lewy body fibril shares epitopes with neurofilament and τ and may be a posttranslationally modified derivative of either neurofilament or τ with substantially altered biochemical and immunologic properties.

Deposition of detergent-resistant neurofilaments into Lewy body fibrils.

To assess the contribution of neurofilaments (NF) to the detergent-resistant cortical Lewy body (LB) fibril we extracted LBs from Diffuse LB diseased brains and used monoclonal antibodies to probe Western transfers of solubilized LB-derived protein. Antibodies to epitopes located in the COOH-termini of the 200- and 170-kDa NF subunits (NF-H and NF-M) labelled LB proteins corresponding to full length and partially truncated or variably phosphorylated NF-H and NF-M. LB-derived protein at approximately 70-kDa did not contain epitopes detected by monoclonal antibodies to NF-L, tau or the COOH-termini of the NF-H and NF-M. We conclude that NF-H and NF-M are incorporated as integral insoluble components of the cortical LB fibril.

OXIDATIVE STRESS PLAYS A ROLE IN THE PATHOGENESIS OF FAMILIAL AND SPORADIC AMYOTROPHIC LATERAL SCLEROSIS

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder characterized by the selective death of upper and lower motorneurons. The disease affects 0.6–2.6:100,000 individuals1, with a mean age at onset of approximately 55 and a duration of two to three years on average2. Motorneuron death results in muscle weakness and paralysis with eventual ventilatory failure and death.

Absence of protease-resistant prion protein in dementia characterized by neuronal loss and status spongiosus

Dementia characterized by neuronal loss and status spongiosus (DNLS) is a non-Alzheimer degenerative process which is characterized by Pick-like lobar atrophy with neuronal depletion and gliosis of the cerebral cortex, corpus striatum, medial thalamus, and substantia nigra and the absence of neuronal inclusions. To further investigate the cause and pathogenesis of DNLS, we probed cerebral homogenates from three cases of DNLS for protease-resistant prion protein to determine if DNLS could be a variant of a human prion disease. Limited proteolysis of prion proteins and guanidine thiocyanate treatment of cortical homogenates was used to enrich potential abnormal prion protein immunoreactivity. Although protease-resistant prion protein was detected in a case of sporadic Creutzfeldt-Jakob disease no abnormal prion protein was found in the cases of DNLS. We conclude that DNLS is not a human prion disease and remains an important dementia of uncertain eitology.