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Biomedical subjects

V Askanas

Publications and source records attributed to V Askanas.

At least 55 records · Page 3Linked to original sources

Human muscle macrophages express beta-amyloid precursor and prion proteins and their mRNAs.

Well-characterized antibodies against beta-amyloid precursor protein (beta APP) and prion protein (PrP), and specific cRNA probes, were used to localize beta APP and PrP and their mRNAs in human muscle macrophages. Macrophages present in muscle biopsies of 51 patients with various neuromuscular disorders showed accumulation of beta APP and PrP, and strongly expressed beta APP and PrP mRNAs. These were present in all muscle macrophages unrelated to their localization within the muscle tissue or diagnosis. Our study provides the first demonstration that human muscle resident macrophages synthesize and accumulate beta APP and PrP. We suggest that those proteins play a role in biology of muscle macrophages, including their participation in inflammatory and immune responses.

Adolescent↗

New advances in the understanding of sporadic inclusion-body myositis and hereditary inclusion-body myopathies.

This review emphasizes new advances in seeking the pathogenic mechanisms of sporadic inclusion-body myositis and hereditary inclusion-body myopathy syndromes. Clinical and pathologic similarities and differences between sporadic and hereditary forms are described. Hypotheses are presented regarding the possible causes and consequences of abnormally accumulated intramyofiber beta-amyloid precursor protein (beta APP) (including beta-amyloid protein and C- and N-terminal epitopes), hyperphosphorylated tau, alpha 1-antichymotrypsin, apolipoprotein E, prion protein, ubiquitin, nicotinic acetylcholine receptor and its 43-kD associated protein, fibroblast growth factor, and transforming growth factor-beta. Also increased are beta APP mRNA and prion protein mRNA. Striking similarities between the pathology of muscle specimens from sporadic inclusion-body myositis and samples from the brains of patients with Alzheimer's disease in regard to Congo red positivity and accumulations of several proteins are discussed. Because most of the proteins that pathologically accumulate throughout the abnormal muscle fibers also accumulate focally at normal human neuromuscular junctions, the possible "junctionalization" of nonjunctional nuclei as a pathogenic mechanism in the muscle fiber is discussed.

Diagnosis, Differential↗

Alpha 1-antichymotrypsin is strongly immunolocalized at normal human and rat neuromuscular junctions.

alpha 1-Antichymotrypsin (alpha 1-ACT) is an early-stage acute-phase plasma protein and a serpin that preferentially inactivates chymotrypsin, cathepsin G, and chymase. Using immunofluorescence with four rabbit polyclonal and two monoclonal specific antibodies against human alpha 1-ACT, we have localized alpha 1-ACT at human and rat neuromuscular junctions (NMJs). Strong alpha 1-ACT immunoreactivity (IR) was present at all NMJs identified by bound alpha-bungarotoxin (alpha-BT). alpha 1-ACT immunoreactivity typically extended slightly deeper into the muscle fiber than alpha-BT, and it closely co-localized with immunoreactivities of post-synaptic desmin, beta-amyloid precursor protein, and dystrophin at the same double- or triple-labeled NMJs. Topography of alpha 1-ACT-IR was the same at human and rat NMJs. The muscle non-junctional sarcolemma was either not immunoreactive or was only very slightly so. When the primary antibody was omitted, absorbed, or replaced by a non-immune serum, there was no immunostaining. Thus, alpha 1-ACT is a novel component of the NMJ. Although its role in the postsynaptic domain of the NMJ is unknown, it might be involved in the interaction between the presynaptic and postsynaptic components and/or inhibit excessive or unwanted serine proteases that may exist in the region of the NMJ.

Animals↗

Expression of beta-amyloid precursor protein gene is developmentally regulated in human muscle fibers in vivo and in vitro.

Regenerating muscle fibers in 25 human muscle biopsies, obtained from patients with a variety of neuromuscular diseases, manifested increased mRNA for the beta-amyloid precursor protein (beta APP) that contains the Kunitz-type protease inhibitor (KPI) motif, whereas adult human muscle fibers were negative in their vast non-extrajunctional region. Aneurally cultured normal human muscle fibers also expressed strong KPI-beta APP mRNA signal, which became significantly down-regulated during muscle differentiation. Our study demonstrates that in human muscle KPI-beta APP mRNA is developmentally regulated, and it suggests that beta APP may play a role in human muscle development.

Adolescent↗

Idiopathic inflammatory myopathies: inclusion-body myositis, polymyositis, and dermatomyositis.

In this review, the main emphasis is on new advances concerning sporadic inclusion-body myositis and hereditary inclusion-body myopathy. Polymyositis and dermatomyositis are reviewed briefly. Hypotheses are presented regarding the possible cause and significance of abnormally accumulated beta-amyloid protein, two other epitopes of beta-amyloid precursor protein, hyperphosphorylated tau, alpha 1-antichymotrypsin, ubiquitin, and prion protein in sporadic inclusion-body myositis and hereditary inclusion-body myopathy. Because most of those proteins are also accumulated at the neuromuscular junction, "junctionalization" of other muscle fiber nuclei is a possibility. Attention is given to the fact that vacuolated muscle fibers in hereditary inclusion-body myopathy may represent early changes because they are virtually free of congophilic amyloid deposit but, like sporadic inclusion-body myositis, contain large accumulations of beta-amyloid protein and prion.

Aged↗

Abnormal accumulation of prion protein mRNA in muscle fibers of patients with sporadic inclusion-body myositis and hereditary inclusion-body myopathy.

Sporadic inclusion-body myositis is the most common progressive muscle disease of older patients. The muscle biopsy demonstrates mononuclear cell inflammation and vacuolated muscle fibers containing paired helical filaments and 6 to 10-nm fibrils, both resembling those of Alzheimer brain, and Congo-red positivity. Hereditary inclusion-body myopathy designates patients cytopathologically similar but without inflammation. In both muscle diseases, prion, and several proteins characteristic of Alzheimer brain--eg, beta-amyloid protein and hyperphosphorylated tau (which normally are expressed mainly in neurons), and apolipoprotein E--are abnormally accumulated in vacuolated muscle fibers, by unknown mechanisms. We now demonstrate in both muscle diseases that prion mRNA is strongly expressed in the vacuolated muscle fibers, which suggests that their accumulated prion protein results, at least partly, from increased gene expression. This, to our knowledge, is the first demonstration of abnormally increased prion mRNA in human disease. Another novel finding is the increased prion mRNA in human muscle macrophages, and both increased prion protein and prion mRNA in regenerating muscle fibers. The latter indicates that prion may play a role in human muscle development.

Adult↗

Twisted tubulofilaments of inclusion body myositis muscle resemble paired helical filaments of Alzheimer brain and contain hyperphosphorylated tau.

We immunostained muscle biopsies of 8 patients with sporadic inclusion body myositis (S-IBM), 7 patients with autosomal recessive hereditary inclusion body myopathy (H-IBM) (both diseases being characterized by similar muscle fiber vacuoles containing inclusions), and 11 normal and disease controls. We used the following well-characterized antibodies against tau protein: Tau-1, Alz-50, and anti-paired helical filament (PHF) antiserum. By light microscopy, in all S-IBM muscle biopsies virtually all vacuoles immunoreactive for ubiquitin and beta-amyloid protein also contained inclusions immunoreactive with Alz-50 and anti-PHF antiserum. With tau-1 antibody, strong immunoreactivity in the vacuoles was obtained only after dephosphorylation of muscle sections. By electronmicroscopy, all three antibodies immunodecorated exclusively cytoplasmic twisted tubulofilaments (TTFs). In H-IBM, virtually all ubiquitin and beta-amyloid-positive muscle fiber vacuoles contained inclusions immunoreactive with anti-PHF antiserum, but in only 40% of those fibers were the inclusions immunoreactive with Alz-50. In six H-IBM patients there were no tau-1 immunoreactive inclusions in any of their vacuolated muscle fibers; in one patient, 24% of the vacuolated fibers had tau-1 immunoreactivity. By demonstrating that hyperphosphorylated tau, which is characteristic of Alzheimer brain PHFs, is a component of S-IBM-muscle TTFs (which are also ultrastructurally similar to PHFs), our study: 1) provides the first demonstration of abnormally accumulated tau in nonneural tissue and 2) suggests that the cytopathogenesis in Alzheimer brain and S-IBM muscle may share some similar mechanisms. Whether the difference in tau immunoreactivity between S-IBM and most of the H-IBM patients reflects a difference in genetically determined transcriptional or posttranslational modifications of tau protein or other factors remains to be determined.

Adult↗

Prion protein is abnormally accumulated in inclusion-body myositis.

In muscle biopsies of 8 sporadic inclusion-body myositis (S-IBM) and 4 hereditary inclusion-body myopathy (H-IBM) patients, vacuolated muscle fibers contained within their vacuoles strongly immunoreactive inclusions with 2 polyclonal and 1 monoclonal antibodies against prion protein (PrP). By light-microscopy, PrP deposits co-localized with beta-amyloid protein (A beta) and ubiquitin (Ub). By immuno-electronmicroscopy, both PrP and A beta were present on amorphous material and on 6-10 nm amyloid-like fibrils; and PrP and Ub co-localized on cytoplasmic twisted tubulofilaments (TTFs) and on amorphous material. Our study provides the first demonstration of abnormally accumulated PrP in pathological tissue other than brain, and it suggests that PrP may play a role in the pathogenesis of IBM.

Adult↗

Prion protein is strongly immunolocalized at the postsynaptic domain of human normal neuromuscular junctions.

Using three well-characterized polyclonal and monoclonal antibodies against prion protein (PrP), we demonstrated a strong concentration of PrP at human neuromuscular junctions (NMJs). Applying double and triple fluorescence-labeling, we found that PrP immunoreactivity exactly co-localized with alpha-bungarotoxin (alpha-BT) identified acetylcholine receptors, as well as with the high junctional concentrations of beta-amyloid precursor protein, beta-amyloid protein, desmin, ubiquitin and dystrophin. Therefore, PrP was considered to be located on the postsynaptic muscle membrane. At all NMJs identified by bound alpha-BT, strong PrP immunoreactivity was obtained with all PrP antibodies. This appears to be the first demonstration of PrP concentrated at human NMJs.

Antibodies, Monoclonal↗

beta-Amyloid precursor epitopes in muscle fibers of inclusion body myositis.

Sporadic inclusion body myositis (IBM) and hereditary inclusion body myopathy (hIBM) are severe and progressive muscle diseases, characterized pathologically by vacuolated muscle fibers that contain 15- to 21-nm cytoplasmic tubulofilaments (CTFs). Those vacuolated muscle fibers also contain abnormally accumulated ubiquitin and beta-amyloid protein (A beta), and they contain amyloid in beta-pleated sheets as indicated by Congo red and crystal violet positivity. Using several well-characterized antibodies, we have now demonstrated that, in addition to A beta, two other epitopes, N-terminal and C-terminal, of the beta-amyloid precursor protein (beta PP) are abnormally accumulated in IBM vacuolated muscle fibers and similarly in hIBM. At the light microscopy level, immunoreactivities of N- and C-epitopes of beta PP closely colocalized with A beta and ubiquitin immunoreactivities. However, by immunogold electronmicroscopy, even though N-, C-, and A beta epitopes of beta PP and ubiquitin colocalized at the amorphous and dense floccular structures, only A beta was localized to the 6- to 10-nm amyloid-like fibrils and only ubiquitin was localized to CTFs. beta PP immunoreactive structures were often in proximity to CTFs, but CTFs themselves never contained beta PP immunoreactivities. The fact that A beta but not C- or N-terminal epitopes of beta PP localized to the 6- to 10-nm amyloid-like fibrils suggests that free A beta might be generated during beta PP processing and, after aggregation, may be responsible for the amyloid present within IBM muscle fibers. Our study demonstrates that three epitopes of beta PP accumulate abnormally in diseased human muscle, and therefore this phenomenon is not unique to Alzheimer's disease, Down's syndrome brain, and Dutch-type cerebrovascular amyloidosis.

Adolescent↗

Tropomodulin is highly concentrated at the postsynaptic domain of human and rat neuromuscular junctions.

Tropomodulin is expressed in skeletal muscle and recent studies suggest that tropomodulin is associated with synaptic membranes. Therefore, we have examined neuromuscular junctions by immunofluorescence analysis for tropomodulin localization. Anti-tropomodulin antibodies generated against either the whole protein or a 15-amino acid peptide fragment label human neuromuscular junctions as demonstrated by colocalization with alpha-bungarotoxin label. Tropomodulin labeling also colocalizes with desmin and beta-amyloid proteins which are concentrated in the postsynaptic region. Comparable results are found with immunostaining of neuromuscular junctions in rat diaphragm muscle. These immunofluorescence results suggest that tropomodulin is an integral component of the cytoskeletal lattice associated with the postsynaptic region of neuromuscular junctions.

Animals↗

Strong immunoreactivity of alpha 1-antichymotrypsin co-localizes with beta-amyloid protein and ubiquitin in vacuolated muscle fibers of inclusion-body myositis.

In 10 of 10 inclusion-body myositis (IBM) patients, including 1 hereditary case, vacuolated muscle fibers contained large or small cytoplasmic inclusions immunoreactive for alpha 1-antichymotrypsin (alpha 1-ACT). All IBM muscle biopsies had characteristic cytoplasmic tubulo-filaments by electron microscopy. None of 17 control muscle biopsies contained the alpha 1-ACT immunoreactive inclusions characteristic of IBM. In vacuolated muscle fibers, alpha 1-ACT immunoreactive inclusions colocalized with beta-amyloid protein and ubiquitin immunoreactivities. Our study provides the first demonstration of alpha 1-ACT accumulations in abnormal human muscle, and it suggest that, as in Alzheimer's disease and Down's syndrome, alpha 1-ACT may be involved in the pathogenesis of IBM.

Adolescent↗

New organotypic model to culture the entire fetal rat spinal cord.

We have developed a system for culturing separately the entire longitudinally-cut ventral and dorsal parts of the whole spinal cord of 12-14 day rat embryos, either on polylysine-collagen mixture or co-cultured with human muscle. At the time of explanation and throughout the period of culturing (up to 21 days) choline acetyltransferase (ChAT) activity in dorsal macroexplants (D-MEs) was negligible (2% of that in the ventral macroexplants (V-MEs)). During culturing neuronal survival was evaluated by measurements of ChAT and enolase. The densities and lengths of neuronal processes outgrowing from the MEs were measured in living and fixed cultures. In the latter, immunocytochemical and cytochemical stainings, allowing visualization of neurites, were evaluated by computerized video image analysis. The ChAT activity in the MEs cultured under our two experimental conditions differed only during the first few days of culturing, being significantly higher in the V-MEs co-cultured with muscle as compared to those cultured on a polylysine-collagen mixture. After 3 weeks of culturing, ChAT activity was not statistically different. However, the co-cultured muscle fibers definitely influenced: (a) neurite outgrowth, (b) the expression of neurofilament proteins and (c) the ability to maintain cultured MEs long-term. This new system allows study of motor neurons and the influence on them of putative growth, survival and maturation factors in a tissue culture milieu that more closely resembles in vivo conditions than other culture systems. Our established parameters can now serve as a basis for such studies.

Acetylcholinesterase↗

beta-Amyloid precursor protein mRNA is increased in inclusion-body myositis muscle.

Vacuolated muscle fibers in muscle biopsies of 8 out of 8 inclusion body myositis (IBM) patients, including 2 hereditary patients, manifested increased mRNA for the beta-amyloid precursor protein (beta APP) that contains Kunitz-type protease inhibitor motif. In affected fibers, increased beta APP-mRNA correspond to abnormally accumulated beta APP immunoreactivity (including beta-amyloid protein epitope). In normal human muscle fibers increased beta APP-mRNA was present only at the neuromuscular junctions. Our study (a) suggests that abnormally accumulated beta APP in IBM vacuolated fibers results, at least partly, from increased beta APP generation, and (b) provides the first demonstration of up-regulated beta APP-mRNA in pathologic human tissue other than brain of Alzheimer's disease and Down's syndrome.

Adolescent↗

New advances in inclusion-body myositis.

The major new advances in seeking the pathogenic mechanisms of sporadic inclusion-body myositis and hereditary inclusion-body myopathy are discussed. Hypotheses are presented regarding the possible causes and significance of amyloid deposits in sporadic inclusion-body myositis and the roles of abnormally accumulated ubiquitin, beta-amyloid protein, beta-amyloid precursor protein, alpha 1-antichymotrypsin, hyperphosphorylated tau, and prion protein in the vacuolated muscle fibers in both sporadic inclusion-body myositis and hereditary inclusion-body myopathy. Because hereditary inclusion-body myopathy is virtually free of Congophilic amyloid deposits but, like sporadic inclusion-body myositis, contains large accumulations of beta-amyloid protein, it is possible that the lesions in hereditary inclusion-body myopathy may represent "early" changes. There are striking similarities between the pathology of inclusion-body myositis muscle and brains affected by Alzheimer's disease in regard to accumulation of ubiquitin, beta-amyloid protein and its precursor protein, alpha 1-antichymotrypsin, and hyperphosphorylated tau.

Amyloid↗