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Biomedical subjects

D Beeson

Publications and source records attributed to D Beeson.

At least 19 recordsLinked to original sources
PubMed2006

Pre- and post-synaptic abnormalities associated with impaired neuromuscular transmission in a group of patients with 'limb-girdle myasthenia'.

The properties of neuromuscular junctions (NMJs) were studied in motor-point biopsy samples from eight patients with congenital myasthenic syndromes affecting primarily proximal limb muscles ['limb-girdle myasthenia' (LGM)]. All had moderate to severe weakness of the proximal muscles, without short-term clinical fatigability but with marked variation in strength over periods of weeks or months, with little or no facial weakness or ptosis and no ophthalmoplegia. Most had a characteristic gait and stance. All patients showed decrement of the compound muscle action potential (CMAP) on repetitive stimulation at 3 Hz, and increased jitter and blocking was detected by SFEMG, confirming the presence of impaired neuromuscular transmission. None of the patients had serum antibodies against acetylcholine receptors (AChRs). Two of the patients had similarly affected siblings. Intracellular recording from isolated nerve-muscle preparations revealed that the quantal content (the number of ACh quanta released per nerve impulse) was only approximately 50% of that in controls. However, the quantal size (amplitude of miniature end-plate currents) and the kinetic properties of synaptic potentials and currents were similar to control values. The area of synaptic contact and extent of post-synaptic folding were approximately 50% of control values. Thus, the quantal content per unit area of synaptic contact was normal. The number of AChRs per NMJ was also reduced to approximately 50% of normal, so the local AChR density was normal. Immunolabelling studies revealed qualitatively normal distributions and abundance of each of 14 proteins normally concentrated at the NMJ, including components of the basal lamina, post-synaptic membrane and post-synaptic cytoskeleton. DNA analysis failed to detect mutations in the genes encoding any of the following proteins: AChR subunits, rapsyn, ColQ, ChAT or muscle-specific kinase. Response of these patients to treatment was varied: few showed long-term improvement with pyridostigmine and some even deteriorated with treatments, while others had intolerable side-effects. Several patients showed improvement with 3,4-diaminopyridine, but this was generally only transient. Ephedrine was helpful in half of the patients. We conclude that impaired neuromuscular transmission in these LGM patients results from structural abnormalities of the NMJ, including reduced size and post-synaptic folding, rather from any abnormality in the immediate events of neuromuscular transmission.

Adolescent↗
PubMed2005

Alpha7-acetylcholine receptor antibodies in two patients with Rasmussen encephalitis.

Rasmussen encephalitis (RE) sera were screened for antibodies to human alpha7 nicotinic acetylcholine receptors (nAChRs) using electrophysiology, calcium imaging, and ligand binding assays. Sera from two of nine patients with RE blocked ACh-induced currents through alpha7 nAChRs and the ACh-induced rise in intracellular free calcium ([Ca2+]i) and inhibited (125)I-alpha-bungarotoxin binding in cells expressing alpha7 nAChRs. Thus, the alpha7 nAChR is a potential target for pathogenic antibodies in patients with RE.

Adolescent↗
PubMed2004

Absence of antibodies to glutamate receptor type 3 (GluR3) in Rasmussen encephalitis.

OBJECTIVE: To determine the prevalence of serum antibodies to the ionotropic glutamate receptor 3 (GluR3) in patients with Rasmussen encephalitis (RE), a severe epileptic disorder, and to compare with serum from control subjects and patients with intractable epilepsy (IE). METHODS: The authors looked for serum immunoglobulin (Ig) G antibodies to GluR3 in 30 patients with RE, including two patients who had plasma exchange and 12 who had been treated with IV Igs with varying results, and 49 patients with IE and 23 healthy individuals, using ELISA with GluR3B peptide, Western blot analysis of recombinant full-length GluR3, immunoprecipitation of [35S]- and [125I]-labeled GluR3 extracellular domains, immunohistochemistry on rat brain sections, and electrophysiology of GluR3 expressed in Xenopus oocytes. RESULTS: Low levels of antibodies to the GluR3B peptide were detected using ELISA in only 4 of the 79 patients with epilepsy (2 with RE and 2 with IE); binding to GluR3B in other sera was shown to be nonspecific. One other patient with IE had antibodies to recombinant GluR3 on Western blot analysis. However, none of the sera tested precipitated either the [35S]- or the [125I]-labeled GluR3 domains; none bound to rat brain sections in a manner similar to rabbit antibodies to GluR3; and none of the nine sera tested affected the electrophysiologic function of GluR3. CONCLUSIONS: GluR3 antibodies were only infrequently found in Rasmussen encephalitis or intractable epilepsy.

Adolescent↗
PubMed2004

Mutation in the AChR ion channel gate underlies a fast channel congenital myasthenic syndrome.

BACKGROUND: Most congenital myasthenic syndromes (CMS) have postsynaptic defects from mutations within the muscle acetylcholine receptor (AChR). Mutations underlying the slow channel syndrome cause a "gain of function" and usually show dominant inheritance, whereas mutations underlying AChR deficiency or the fast channel syndrome cause a "loss of function" and show recessive inheritance. OBJECTIVE: To characterize the disease mechanism underlying an apparently dominantly inherited CMS that responds to IV edrophonium. METHODS: DNA from CMS patients was analyzed for mutations by single-strand conformation polymorphism analysis, DNA sequence analysis, and restriction endonuclease digestion. Functional analysis of mutations was by alpha-bungarotoxin binding studies and by patch clamp analysis of mutant AChR expressed in human embryonic kidney cells. RESULTS: Analysis of muscle biopsies from father and son in an affected kinship showed normal endplate morphology and AChR number but severely reduced miniature endplate potentials. DNA analysis revealed that each harbors a single missense mutation in the AChR alpha-subunit gene, alphaF256L. Expression studies demonstrate this mutation underlies a fast channel phenotype with fewer and shorter ion channel activations. The major effect of alphaF256L, located within the M2 transmembrane domain, is on channel gating, both reducing the opening and increasing the closure rate. CONCLUSIONS: Mutation alphaF256L results in fast channel kinetics. Expression studies suggest a dominant-negative effect within the AChR pentamer, severely compromising receptor function.

Adolescent↗
PubMed2004

Distinct phenotypes of congenital acetylcholine receptor deficiency.

We contrast the phenotypes associated with hereditary acetylcholine receptor deficiency arising from mutations in either the acetylcholine receptor epsilon subunit or the endplate acetylcholine receptor clustering protein rapsyn. Mutational screening was performed by amplification of promoter and coding regions by PCR and direct DNA sequencing. We identified mutations in 37 acetylcholine receptor deficiency patients; 18 had acetylcholine receptor-epsilon mutations, 19 had rapsyn mutations. Mutated acetylcholine receptor-epsilon associated with bulbar symptoms, ptosis and ophthalmoplegia at birth, and generalized weakness. Mutated rapsyn caused either an early onset (rapsyn-EO) or late onset (rapsyn-LO) phenotype. Rapsyn-EO associated with arthrogryposis and life-threatening exacerbations during early childhood. Rapsyn-LO presented with limb weakness in adolescence or adulthood resembling seronegative myasthenia gravis. Awareness of distinct phenotypic features of acetylcholine receptor deficiency resulting from acetylcholine receptor-epsilon or rapsyn mutations should facilitate targeted genetic diagnosis, avoid inappropriate immunological therapy and, in some infants, prompt the rapid introduction of treatment that could be life saving.

4-Aminopyridine↗
PubMed2003

Rapsyn mutations in hereditary myasthenia: distinct early- and late-onset phenotypes.

Rapsyn mutations in 16 unrelated patients with a congenital/hereditary myasthenic syndrome were identified, and a mutation (N88K) common to each of them was found. Two distinct phenotypes were noted: early and late onset. The former is frequently associated with arthrogryposis multiplex congenita and life-threatening crises. The late-onset phenotype developed in adolescence or adulthood and was initially mistaken for seronegative myasthenia gravis. Recognition of this late-onset phenotype should prevent inappropriate immunotherapy.

Adolescent↗
PubMed2003

Properties of the human muscle nicotinic receptor, and of the slow-channel myasthenic syndrome mutant epsilonL221F, inferred from maximum likelihood fits.

The mechanisms that underlie activation of nicotinic receptors are investigated using human recombinant receptors, both wild type and receptors that contain the slow channel myasthenic syndrome mutation, epsilonL221F. The method uses the program HJCFIT, which fits the rate constants in a specified mechanism directly to a sequence of observed open and shut times by maximising the likelihood of the sequence with exact correction for missed events. A mechanism with two different binding sites was used. The rate constants that apply to the diliganded receptor (opening, shutting and total dissociation rates) were estimated robustly, being insensitive to the exact assumptions made during fitting, as expected from simulation studies. They are sufficient to predict the main physiological properties of the receptors. The epsilonL221F mutation causes an approximately 4-fold reduction in dissociation rate from diliganded receptors, and a smaller increase in opening rate and mean open time. These are sufficient to explain the approximately 6-fold slowing of decay of miniature synaptic currents seen in patients. The distinction between the two binding sites was less robust, the estimates of rate constants being dependent to some extent on assumptions, e.g. whether an extra short-lived shut state was included or whether the EC50 was constrained. The results suggest that the two binding sites differ by roughly 10-fold in the affinity of the shut receptor for ACh in the wild type, and that in the epsilonL221F mutation the lower affinity is increased so the sites become more similar.

Acetylcholine↗
PubMed2003

Scenarios for autoimmunization of T and B cells in myasthenia gravis.

We have studied responses in thymoma patients to interferon-alpha and to the acetylcholine receptor (AChR) in early-onset myasthenia gravis (EOMG), seeking clues to autoimmunizing mechanisms. Our new evidence implicates a two-step process: (step 1) professional antigen-presenting cells and thymic epithelial cells prime AChR-specific T cells; then (step 2) thymic myoid cells subsequently provoke germinal center formation in EOMG. Our unifying hypothesis proposes that AChR epitopes expressed by neoplastic or hyperplastic thymic epithelial cells aberrantly prime helper T cells, whether generated locally or infiltrating from the circulation. These helper T cells then induce antibody responses against linear epitopes that cross-react with whole AChR and attack myoid cells in the EOMG thymus. The resulting antigen-antibody complexes and the recruitment of professional antigen-presenting cells increase the exposure of thymic cells to the infiltrates and provoke local germinal center formation and determinant spreading. Both these and the consequently enhanced heterogeneity and pathogenicity of the autoantibodies should be minimized by early thymectomy.

Age of Onset↗
PubMed2002

Recessive inheritance and variable penetrance of slow-channel congenital myasthenic syndromes.

BACKGROUND: Slow-channel congenital myasthenic syndromes (SCCMS) typically show dominant inheritance. They are caused by missense mutations within the subunits of muscle nicotinic acetylcholine receptors (AChR) that result in prolonged ion channel activations. SCCMS mutations within the AChR subunit are located in various functional domains, whereas fully described mutations in AChR non- subunits have, thus far, been located only in the M2 channel-lining domain. The authors identified and characterized two -subunit mutations, located outside M2, that underlie SCCMS in three kinships. In two of the three kinships, the syndrome showed an atypical inheritance pattern. METHODS: These methods included clinical diagnosis, mutation detection, haplotype analysis, and functional expression studies using single-channel recordings of mutant AChR transiently transfected into HEK293 cells. RESULTS: The authors identified two SCCMS mutations in the AChR subunit, L78P and L221F. Both mutations prolonged ACh-induced ion channel activations. L78P is present in a consanguineous family and appears to be pathogenic only when present on both alleles, and L221F shows variable penetrance in one of the two families that were identified harboring this mutation. CONCLUSION: SCCMS mutations may show a recessive inheritance pattern and variable penetrance. A diagnosis of SCCMS should not be ruled out in cases of CMS with an apparent recessive inheritance pattern.

Adolescent↗
PubMed2001

Presynaptic neuronal antigens expressed by a small cell lung carcinoma cell line.

Small cell lung carcinoma (SCLC) is a tumour of neuroendocrine origin often found in association with autoimmune paraneoplastic neurological disorders. We established a SCLC cell line from a woman with Lambert-Eaton myasthenic syndrome (LEMS) who developed antibodies to both the P/Q-type voltage-gated calcium channels (VGCC) and the muscle acetycholine receptor (AChR). We used a range of techniques to establish which neuronal antigens were expressed in her tumour cell line. The results show that many proteins involved in exocytosis are present in the SCLC cells, and that depolarisation-dependent release of [(3)H]-serotonin is linked to calcium influx through P/Q-type VGCCs. In addition, some of the subunits encoding the AChR and both agrin and ARIA, molecules released from the motor nerve during development, were expressed. These results suggest that many potential antigenic targets are present in SCLC, and indicate a surprising 'motor nerve terminal'-like characteristic of this line.

Autoantibodies↗
PubMed2001

Human mitochondrial DNA deletions associated with mutations in the gene encoding Twinkle, a phage T7 gene 4-like protein localized in mitochondria.

The gene products involved in mammalian mitochondrial DNA (mtDNA) maintenance and organization remain largely unknown. We report here a novel mitochondrial protein, Twinkle, with structural similarity to phage T7 gene 4 primase/helicase and other hexameric ring helicases. Twinkle colocalizes with mtDNA in mitochondrial nucleoids. Screening of the gene encoding Twinkle in individuals with autosomal dominant progressive external ophthalmoplegia (adPEO), associated with multiple mtDNA deletions, identified 11 different coding-region mutations co-segregating with the disorder in 12 adPEO pedigrees of various ethnic origins. The mutations cluster in a region of the protein proposed to be involved in subunit interactions. The function of Twinkle is inferred to be critical for lifetime maintenance of human mtDNA integrity.

Amino Acid Sequence↗
PubMed2001

End-plate gamma- and epsilon-subunit mRNA levels in AChR deficiency syndrome due to epsilon-subunit null mutations.

Acetylcholine receptor (AChR) deficiency is the most common of the congenital myasthenic syndromes (CMS). Typically, the number of AChRs, measured by alpha-bungarotoxin binding, is reduced to 10-30% of normal levels, the miniature end-plate potentials are correspondingly reduced, and there are morphological changes at the motor end-plates. The majority of these syndromes are due to either missense or frameshift mutations within the gene encoding the adult-specific epsilon-subunit. These are often null mutations, but some mutant epsilon-subunits can be incorporated, at low levels, into functional AChRs in transfected cell lines. It is not clear, therefore, whether upregulation of the mutant epsilon-subunit mRNA could generate sufficient AChR to support neuromuscular transmission, albeit at a reduced level. Conversely, it might be that the mutant epsilon-subunit transcripts are subject to mRNA surveillance and 'nonsense-mediated' loss, leading to reduced epsilon-subunit mRNA expression. In either case, it is thought that neuromuscular transmission may be provided partly or entirely by incorporation of the foetal-specific gamma-subunit into end-plate AChR. gamma-Subunit mRNA is expressed at low levels in normal human muscle, but might be upregulated in CMS. The study of mRNA levels for AChR subunits should improve our understanding of genotype-phenotype relationships in CMS. Here we have defined homozygous epsilon-subunit mutations in four unrelated families with AChR deficiency and studied the steady-state levels of mRNA for AChR subunits at the motor end-plates by in situ hybridization. Although we demonstrated that each mutation would lead to almost complete absence of surface adult AChR expression, we detected similar robust expression of alpha- and epsilon-subunit mRNAs at end-plates of patient and control muscles, suggesting that mRNA transcripts for the epsilon-subunit are neither upregulated nor degraded preferentially. Interestingly, we were unable to detect any increase in gamma-subunit mRNA expression at CMS end-plates. Transgenic mice lacking the epsilon-subunit die 2-3 months after birth, suggesting that alpha(2)betadelta(2) pentamers cannot sustain neuromuscular transmission. Therefore, we tentatively conclude that the persistent low level expression of the gamma-subunit, which is present in normal human muscles as well as in AChR deficiency syndromes, is sufficient to enable patients with epsilon-subunit null alleles to survive.

Adult↗
PubMed2001

Myasthenia gravis: diagnostic and management dilemmas.

This review focuses on the diagnostic tests that may help to confirm myasthenia in patients without acetylcholine receptor antibodies, including the newly discovered anti-muscle-specific receptor tyrosine kinase antibody and other investigations. We discuss management dilemmas, particularly those that revolve around treatments whose efficacy is questionable or unproved, such as thymectomy. We review the important issue of how treatments are assessed, and examine future treatment trial designs.

Humans↗
PubMed2001

Acetylcholine receptor delta subunit mutations underlie a fast-channel myasthenic syndrome and arthrogryposis multiplex congenita.

Limitation of movement during fetal development may lead to multiple joint contractures in the neonate, termed arthrogryposis multiplex congenita. Neuromuscular disorders are among the many different causes of reduced fetal movement. Many congenital myasthenic syndromes (CMSs) are due to mutations of the adult-specific epsilon subunit of the acetylcholine receptor (AChR), and, thus, functional deficits do not arise until late in gestation. However, an earlier effect on the fetus might be predicted with some defects of other AChR subunits. We studied a child who presented at birth with joint contractures and was subsequently found to have a CMS. Mutational screening revealed heteroallelic mutation within the AChR delta subunit gene, delta 756ins2 and delta E59K. Expression studies demonstrate that delta 756ins2 is a null mutation. By contrast, both fetal and adult AChR containing delta E59K have shorter than normal channel activations that predict fast decay of endplate currents. Thus, delta E59K causes dysfunction of fetal as well as the adult AChR and would explain the presence of joint contractures on the basis of reduced fetal movement. This is the first report of the association of AChR gene mutations with arthrogryposis multiplex congenita. It is probable that mutations that severely disrupt function of fetal AChR will underlie additional cases.

Action Potentials↗
PubMed2001

Dominantly inherited familial myasthenia gravis as a separate genetic entity without involvement of defined candidate gene loci.

Myasthenia gravis (MG) is a sporadic autoimmune disorder affecting neuromuscular transmission. Very rarely autoimmune myasthenia gravis may be inherited within a family. We present here the genetic analysis of a Hungarian family where nine members from two generations are affected by myasthenia gravis. Genetic characterisation of this unique Hungarian family using linkage analysis and mutation screening excludes the involvement of defined candidate gene loci. These findings point to familial MG as a separate genetic entity. Identification of the underlying genetic defect in this family may greatly enhance our understanding of the pathogenesis of myasthenia gravis.

Adolescent↗
PubMed2000

The conformation of the main immunogenic region on the alpha-subunit of muscle acetylcholine receptor is affected by neighboring receptor subunits.

Myasthenia gravis (MG) is caused by autoantibodies to the acetylcholine receptor (AChR). Experiments with fetal (alpha(2)betagammadelta) and adult (alpha(2)betaepsilondelta) AChR and with recombinant subunit dimers showed that some monoclonal antibodies (mAbs) against the main immunogenic region (MIR), located on the alpha-subunit of the AChR, bind better to fetal AChR and to alphagamma subunit dimer than to adult AChR and alphaepsilon dimer and equally to both alphabeta and alphadelta. However, other anti-MIR mAbs prefer adult AChR and alphaepsilon dimer, bind well to alphabeta but weakly to alphadelta. These results suggest that the MIR conformation is affected by the neighboring gamma/epsilon- and delta-subunits and may contribute to understanding the antibody specificities in MG.

Aging↗
PubMed2000

Molecular targets for autoimmune and genetic disorders of neuromuscular transmission.

The neuromuscular junction is the target of a variety of autoimmune, neurotoxic and genetic disorders, most of which result in muscle weakness. Most of the diseases, and many neurotoxins, target the ion channels that are essential for neuromuscular transmission. Myasthenia gravis is an acquired autoimmune disease caused in the majority of patients by antibodies to the acetylcholine receptor, a ligand-gated ion channel. The antibodies lead to loss of acetylcholine receptor, reduced efficiency of neuromuscular transmission and muscle weakness and fatigue. Placental transfer of these antibodies in women with myasthenia can cause fetal or neonatal weakness and occasionally severe deformities. Lambert Eaton myasthenic syndrome and acquired neuromyotonia are caused by antibodies to voltage-gated calcium or potassium channels, respectively. In the rare acquired neuromyotonia, reduced repolarization of the nerve terminal leads to spontaneous and repetitive muscle activity. In each of these disorders, the antibodies are detected by immunoprecipitation of the relevant ion channel labelled with radioactive neurotoxins. Genetic disorders of neuromuscular transmission are due mainly to mutations in the genes for the acetylcholine receptor. These conditions show recessive or dominant inheritance and result in either loss of receptors or altered kinetics of acetylcholine receptor channel properties. Study of these conditions has greatly increased our understanding of synaptic function and of disease aetiology.

Binding Sites↗