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D Beeson

Publications and source records attributed to D Beeson.

At least 37 records · Page 2Linked to original sources
PubMed2000

IgG from "seronegative" myasthenia gravis patients binds to a muscle cell line, TE671, but not to human acetylcholine receptor.

Antibodies to acetylcholine receptor (AChR) are found in 85% of patients with myasthenia gravis (seropositive MG [SPMG]) and are thought to be pathogenic; but in 15% of MG patients, the standard immunoprecipitation test for anti-AChR is negative (seronegative MG [SNMG]). Here, we used a novel approach, fluorescence-activated cell sorting analysis, to measure binding of SPMG and SNMG IgG antibodies to rhabdomyosarcoma cell lines that express human adult (TE671-epsilon) or fetal (TE671-gamma) AChR, and to human embryonic kidney (HEK) fibroblasts that express adult human AChR (HEK-AChR). We found that whereas most SPMG antibodies bind to all three cell lines, IgG from 8 of 15 SNMG sera/plasmas bind to the surface of both TE671 cell lines but not to HEK-AChR cells. These results indicate that SNMG antibodies bind to a muscle surface antigen that is not the AChR, which strongly supports previous studies that suggest that SNMG should be considered a distinct subtype of MG.

Adult↗
PubMed1999

Mapping of autosomal dominant progressive external ophthalmoplegia to a 7-cM critical region on 10q24.

OBJECTIVE: To map the gene responsible for autosomal dominant progressive external opthalmoplegia. BACKGROUND: The pathogenesis of progressive external ophthalmoplegia (PEO) can be associated with multiple deletions of mitochondrial DNA (mtDNA). PEO may show autosomal dominant (adPEO) or autosomal recessive (arPEO) patterns of inheritance, indicating that the genetic defect has a Mendelian basis and most likely involves a nuclear gene encoding a protein that interacts with the mitochondrial genome. adPEO is heterogeneous genetically, and thus far disease loci have been identified on chromosomes 3 and 10. The locus on chromosome 10q23-q25 was assigned by linkage analysis in a single Finnish family. METHODS: Samples from a large Pakistani family with adPEO, in which clinical symptoms are bilateral ptosis, limitations of eye movements, and varying degrees of proximal muscle weakness, were collected. Muscle biopsy and mtDNA rearrangement analysis was used to confirm the diagnosis. Genomewide linkage analysis was set up using a set of 391 microsatellite markers. RESULTS: The muscle biopsy from an affected member showed ragged red fibers, increased succinic dehydrogenase staining, lack of cytochrome oxidase activity, and multiple deletions of mtDNA. The disease locus was mapped to 10q23.31-q25.1 by linkage analysis, and a maximum lod score of 5.72 was obtained with D10S1267. CONCLUSION: By analysis of meiotic recombinations in affected individuals, the critical region was restricted to the 7-cM interval between D10S198 and D10S1795.

Adult↗
PubMed1999

Monoclonal antibodies raised against human acetylcholine receptor bind to all five subunits of the fetal isoform.

The human muscle acetylcholine receptor (AChR) is an oligomeric membrane protein consisting of (alpha1)2,beta,delta,epsilon subunits in the adult form and (alpha 1)2,beta,gamma,delta in the fetal form. The adult AChR is the target for autoantibodies in myasthenia gravis (MG), and antibodies that block the function of fetal AChR can cross the placenta and paralyse the developing baby causing joint contractures. Monoclonal antibodies (mAbs) raised against purified AChR were characterised previously in terms of binding to five regions, three of which appeared to partially overlap, but the subunit localisation of the regions was not clearly established and they were assumed to be mainly on the immunodominant alpha subunits. We have studied binding of the mAbs to AChR subunit extracellular fragments expressed in E. coli, and to AChRs derived from TE671 cells and from fibroblast cell lines expressing human/Torpedo and Torpedo/mouse hybrid receptors. Using a combination of Western blotting and immunoprecipitation experiments, we demonstrate the subunit specificity of each mAb. The results confirm our previous observations but importantly show that only two of the regions are on the alpha subunit, the three others being on the beta, gamma and delta subunits of human AChR. Thus these mAbs should be useful in studies of AChR subunit expression in normal and diseased tissue, and to define further the binding sites of antibodies in MG patients.

Animals↗
PubMed1999

Early-onset myasthenia gravis: a recurring T-cell epitope in the adult-specific acetylcholine receptor epsilon subunit presented by the susceptibility allele HLA-DR52a.

No immunodominant T-cell epitopes have yet been reported in the human acetylcholine receptor (AChR), the target of the pathogenic autoantibodies in myasthenia gravis (MG). We have selected and characterized T cells from MG patients by restimulation in culture with recombinant human AChR to alpha, gamma and epsilon subunits; the gamma and epsilon distinguish the fetal and adult AChR isoforms, respectively. We obtained clones specific for the epsilon, rather than the alpha or gamma, subunit in 3 of the first 4 early-onset MG cases tested. They all responded to peptide epsilon201-219 and to low concentrations of adult but not fetal AChR. Moreover, although using different T-cell receptor genes, they were all restricted to HLA-DR52a (DRB3*0101), a member of the strongly predisposing HLA-A1-B8-DR3 haplotype. This apparently immunodominant epsilon201-219 epitope (plus DR52a) was also recognized by clones from an elderly patient whose MG had recently been provoked by the drug D-penicillamine. In all 4 cases, however, the serum antibodies reacted better with fetal than adult AChR and may thus be end products of determinant spreading initiated by adult AChR-specific T cell responses. Furthermore, as these T cells had a pathogenic Th1 phenotype, with the potential to induce complement-activating antibodies, they should be important targets for selective immunotherapy.

Amino Acid Sequence↗
PubMed1999

Novel functional epsilon-subunit polypeptide generated by a single nucleotide deletion in acetylcholine receptor deficiency congenital myasthenic syndrome.

Acetylcholine receptor (AChR) deficiency is a recessively inherited congenital myasthenic syndrome in which fatigable muscle weakness results from impaired neuromuscular transmission caused by reduced AChR numbers. In mature muscle, AChRs consist of alpha2 betadelta together with the adult-specific epsilon subunit. We have identified a deletion of the first nucleotide in exon 12 of the AChR epsilon-subunit gene (epsilon1267delG) and demonstrate its recessive inheritance segregates with disease in 6 unrelated cases of AChR deficiency. In addition, we found that both healthy and AChR-deficient muscle contain a population of AChR epsilon-subunit mRNA transcripts that retain intron 11. We investigated the possible consequences of combining this mutation with the alternative mRNA species through AChR expression studies in human embryonic kidney cells and Xenopus oocytes. Epsilon1267delG generates a polypeptide that lacks M4 and is not detected in surface AChR, whereas retention of intron 11 in the RNA transcript restores the reading frame, conserves M4, and generates a polypeptide that is incorporated into functional surface AChR, although at a reduced level, consistent with the disease phenotype. Our results indicate that for some AChR deficiency mutations located between M3 and M4, the retention of intron 11 in the epsilon-subunit mRNA transcripts may rescue adult AChR function.

Adult↗
PubMed1999

Association of the APOE epsilon4 allele with disease activity in multiple sclerosis.

OBJECTIVES: Allelic variants of the APOE gene are known to influence the course of many neurological diseases and there is increasing evidence that apolipoprotein E (APOE) is a pivotal component in reinnervation and dendritic remodelling after neuronal injury. Previous studies did not show significant differences in the APOE allele frequencies in multiple sclerosis compared with controls but did not examine for correlation with disease severity. This study explores the relation of APOE genotypes with the disease severity. METHODS: Ninety five patients with multiple sclerosis were studied. Age of onset, type, and activity of the disease were recorded prospectively and genotyping was performed according to standard protocols. RESULTS: APOE allele frequencies of the group as a whole, the relapsing group, or the primary progressive group were not significantly different from those reported from matched historical controls. The epsilon4 allele was found to be more common in patients with a more aggressive type of multiple sclerosis (odds ratio=2.95, p=0.03). CONCLUSIONS: Although APOE does not seem to be implicated in the early pathogenesis of the disease, patients possessing the epsilon4 allele might have a reduced capacity for neuronal remodelling after relapses.

Adult↗
PubMed1999

Mutation of the acetylcholine receptor epsilon-subunit promoter in congenital myasthenic syndrome.

Congenital myasthenic syndrome comprises a heterogeneous group of inherited disorders of neuromuscular transmission. Acetylcholine receptor (AChR) deficiency is the most common form of congenital myasthenic syndrome and in most cases results from mutations within the coding region of the AChR epsilon subunit. However, studies in mice have established that synapse-specific expression of AChR is dependent on a sequence contained within the AChR-subunit promoter regions, termed an N-box. We describe a consanguineous family in which 2 of 7 siblings had clinical and electromyographic features consistent with AChR deficiency. Muscle biopsy demonstrated low AChR numbers, establishing the disorder as postsynaptic. Single-strand conformational polymorphism analysis identified an abnormal conformer in the AChR epsilon-subunit gene promoter of the patients. DNA sequence and restriction endonuclease analysis shows that the disorder cosegregates with recessive inheritance of a single point mutation, a transition (C-->T) in the N-box of the epsilon-subunit promoter. Analysis of an intercostal biopsy from 1 of the patients showed a dramatic reduction in epsilon-subunit mRNA levels compared with disease and normal controls. This is the first evidence in humans that an N-box mutation can lead to disruption of epsilon-subunit transcription, resulting in the loss of adult AChR synthesis and the clinical phenotype of AChR-deficiency congenital myasthenic syndrome.

Adult↗
PubMed1998

Regulation of acetylcholine receptor gene expression in human myasthenia gravis muscles. Evidences for a compensatory mechanism triggered by receptor loss.

Myasthenia gravis (MG) is a neuromuscular disorder mediated by antibodies directed against the acetylcholine receptor (nAChR) resulting in a functional nAChR loss. To analyze the molecular mechanisms involved at the muscular target site, we studied the expression of nAChR subunits in muscle biopsy specimens from MG patients. By using quantitative PCR with an internal standard for each subunit, we found that the levels of beta-, delta-, and epsilon-subunit mRNA coding for the adult nAChR were increased in severely affected MG patients, matching our previous data on the alpha-subunit. Messenger levels were highly variable in MG patients but not in controls, pointing to individual factors involved in the regulation of nAChR genes. The fetal subunit (gamma-chain) transcripts were almost undetectable in the extrajunctional region of MG muscle, suggesting that gene regulation in MG differs from that in the denervation model, in which nAChR gamma-subunit mRNA is reexpressed. Nicotinic AChR loss mediated by monoclonal anti-nAChR antibodies in both the TE671 muscle cell line and cultured normal human myotubes induces a similar increase in beta- alphand delta-subunit mRNA levels, suggesting the existence of a new muscular signaling pathway system coupled to nAChR internalization and independent of muscle electrical activity. These data demonstrate the existence of a compensatory mechanism regulating the expression of the genes coding for the adult nAChR in patients with MG.

Adolescent↗
PubMed1998

A pathogenetic role for the thymoma in myasthenia gravis. Autosensitization of IL-4- producing T cell clones recognizing extracellular acetylcholine receptor epitopes presented by minority class II isotypes.

Myasthenia gravis (MG) is caused by helper T cell-dependent autoantibodies against the muscle acetylcholine receptor (AChR). Thymic epithelial tumors (thymomas) occur in 10% of MG patients, but their autoimmunizing potential is unclear. They express mRNAs encoding AChR alpha and epsilon subunits, and might aberrantly select or sensitize developing thymocytes or recirculating peripheral T cells against AChR epitopes. Alternatively, there could be defective self-tolerance induction in the abundant maturing thymocytes that they usually generate. For the first time, we have isolated and characterized AChR-specific T cell clones from two MG thymomas. They recognize extracellular epitopes (alpha75-90 and alpha149-158) which are processed very efficiently from muscle AChR. Both clones express CD4 and CD8alpha, and have a Th-0 cytokine profile, producing IL-4 as well as IFN-gamma. They are restricted to HLA-DP14 and DR52a; expression of these minority isotypes was strong on professional antigen-presenting cells in the donors' tumors, although it is generally weak in the periphery. The two clones' T cell receptor beta chains are different, but their alpha chain sequences are very similar. These resemblances, and the striking contrasts with T cells previously cloned from non-thymoma patients, show that thymomas generate and actively induce specific T cells rather than merely failing to tolerize them against self antigens.

Amino Acid Sequence↗
PubMed1998

Antibodies affecting ion channel function in acquired neuromyotonia, in seropositive and seronegative myasthenia gravis, and in antibody-mediated arthrogryposis multiplex congenita.

A new autoimmune disease affecting the neuromuscular junction has been defined. Acquired neuromyotonia is associated with antibodies to voltage-gated potassium channels that act, at least in part, by reducing potassium channel function with resulting neuronal hyperactivity. This condition is quite frequently associated with thymoma and, in many cases, antibodies to acetylcholine receptors are present as well as antibodies to VGKC. Improvements in techniques and the availability of cloned DNA and recombinant forms of the AChR subunits have led to new observations concerning the specificity and roles of antibodies in myasthenia gravis. The transfection of a cell line with the epsilon subunit means that we can now accurately compare antibodies reactive with adult and fetal human AChR. This may help to determine the relationship between AChR subunit expression in different tissues and the induction of antibodies that bind specifically to the two forms, as well as to clarify the role of antibodies to fetal or adult AChR in causing ocular muscle symptoms. Serum antibodies from a few mothers with obstetric histories of recurrent arthrogryposis multiplex congenita in their babies specifically inhibit the function of fetal AChR. These observations not only explain the cause of some cases of arthrogryposis multiplex congenita, but also suggest that other fetal-specific antibodies might be responsible for other fetal or neonatal conditions. An animal model has been established to enable us to investigate the role of maternal serum factors in causing such disorders. Seronegative MG has been the subject of many studies from our laboratory over the last ten years. The transience of the effects of SNMG plasmas on AChR function strongly suggests that the plasma antibodies do not bind directly to the AChR, but inhibit function by some indirect mechanism. They do not appear to act via the cAMP-dependent protein kinase pathway, and studies are in progress to investigate the involvement of other second messenger systems.

Adult↗
PubMed1998

Determinant spreading and immune responses to acetylcholine receptors in myasthenia gravis.

In myasthenia gravis (MG), antibodies to the muscle acetylcholine receptor (AChR) cause muscle weakness. Experimental autoimmune myasthenia gravis (EAMG) can be induced by immunisation against purified AChR; the main immunogenic region (MIR) is a conformation-dependent site that includes alpha 67-76. EAMG can also occur after immunisation against extracellular AChR sequences, but this probably involves intramolecular determinant spreading. In MG patients, thymic hyperplasia and germinal centres are found in about 50%, and thymoma in 10-15%. The heterogeneous, high affinity, IgG anti-AChR antibodies appear to be end-products of germinal centre responses, and react mainly with the MIR or a site on fetal AChR; the latter contains a gamma subunit and is mainly expressed on myoid cells in the thymic medulla. T cells cloned against recombinant AChR subunits recognise principally two naturally processed epitopes: epsilon 201-219 derived from adult AChR which is expressed in muscle, and sometimes in thymic epithelium, and alpha 146-160, common to fetal and adult AChR. Since AChR is not normally co-expressed with class II, it is unclear how CD4+ responses to AChR alpha and epsilon subunits are initiated, and how and where these spread to induce antibodies against fetal AChR. Various possibilities, including upregulation of class II on muscle/myoid cells and involvement of CD8+ responses to AChR and other muscle antigens, are discussed.

Animals↗