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M.E. Farrugia, Specialist Registrar in Neurology, and R.J. Swingler, Consultant Neurologist,Department of Neurology, Ninewells Hospital and Medical School, Dundee The annual incidence of MG varies between 0·25 to 2·00 Myasthenia gravis (MG) is an autoimmune condition of per 100,000 of the population. The frequency is bimodal the neuromuscular junction characterised by fatiguable in the younger population (below the age of 40), being muscle weakness of the limbs, bulbar and ocular muscles.
biased towards female patients; the older population with It is associated with the presence of nicotinic MG (above the age of 60) is represented by male patients acetylcholine receptor antibodies, which act at the postsynaptic plate of the neuromuscular junction,interfering with normal synaptic function. About 15– Simpson was the first to propose that MG might be an 20% of patients may have all the characteristic features autoimmune condition, and it is now clear that MG is without detectable antibodies in the serum, often causing immune-mediated.2 Toyka et al. showed that a circulating some concern about the validity of the diagnosis. Because factor from serum of patients with MG could be injected patients with seronegative myasthenia gravis (SNMG) and passively transferred into mice, reproducing the respond to immunotherapy and have very similar clinical features of the disease in the animal model.3 The mice and pathological features to seropositive disease, it is showed reduced amplitudes of miniature endplate suspected that this condition is also mediated by potentials and reduced numbers of acetylcholine antibodies. Recent studies support this hypothesis, receptors (AChRs) at the neuromuscular junctions. Both demonstrating antibodies to muscle specific receptor B- and T-lymphocyte reactivity play a role in the tyrosine kinase (MuSK) in 70% of seronegative patients.
pathogenesis of MG. Antibodies are directed against the This protein is present at the postsynaptic membrane nicotinic AChR with deposition of immune complexes at where it interacts with other proteins and growth factors the postsynaptic part of the neuromuscular junction. The which maintain the architecture of the neuromuscular antibodies induce accelerated degradation as well as the junction. It is now possible to obtain a serological functional blockade of AChRs of skeletal muscle diagnosis in 95% of patients with myasthenia. However, corresponding to a certain degree to the severity of MG.4 further studies are required to elucidate the clinical Reduction of serum anti-AChR by plasma exchange was characteristics and pathogenesis of MuSK-positive shown to result in a progressive improvement in strength, and when the antibody titres remained depressed theimprovement was sustained.5 Buckley et al. showed how SEROPOSITIVE MYASTHENIA GRAVIS
in MG patients with thymoma the neoplastic tissue may Myasthenia gravis was probably first described by the be exporting mature long-lived T-cells which, in turn, may physiologist Thomas Willis, who in 1672 wrote about ‘a have an important role in supporting antibody woman who temporarily lost her power of speech and became as mute as a fish’. The hallmarks of MG arethose of muscular weakness and fatiguability. It has a Different immunogenetic backgrounds are associated with generalised distribution in about 85% of patients and in the different forms of MG, and this is reflected by the the rest remains localised to the extraocular muscles.
various HLA linkages described. A strong and positive The generalised pattern will tend to affect the limbs in a association exists between seropositive MG (SPMG) with proximal distribution but may also affect the diaphragm antigens B8 and DR3, especially in female patients.7 and the neck extensors.1 The clinical severity of MG has Different HLA associations are reported in patients with been graded functionally and regionally by the Osserman thymic hyperplasia when compared with patients with scale: grade I involves focal disease (restricted to ocular thymoma.8 Further to this, there are different HLA muscles), grade II is generalised disease with mild (IIa) or associations in patients with generalised MG versus those moderate (IIb) sub varieties; grade III is severe generalised presenting with ocular involvement only.9 The HLA disease and grade IV represents a crisis with life- associations in SNMG (discussed in depth below) are threatening impairment of respiration. However, the scale not found to differ significantly to those in SPMG.10 is not dynamic and often fails to detect clinically relevantchanges that may be important in evaluating disability The standard test for antibodies to AChRs is based on and in monitoring the effects of therapy. Electro- immunoprecipitation of 125I-α-Bungarotoxin-labelled physiologically, the findings in MG are characterised by AChR extracted from human muscle cell lines.5 a 15% decrementing amplitude of the evoked muscle Antibodies to the nicotinic AChR antibody are detected response to low frequency repetitive stimulation.1 in approximately 80% of cases with generalised MG J R Coll Physicians Edinb 2002; 32:14–18
(seropositive MG or SPMG). In patients with pure ocular observed that the numbers of the AChR were reduced in symptomatology, the antibodies are detected in 50% of SNMG muscle.17 They also showed that the motor cases.1 Acetylcholinesterase inhibitors have been the endplate potential amplitudes and 125I-α-Bungarotoxin mainstay of treatment since 1934 and patients are binding was reduced in mice injected passively with typically treated with pyridostigmine. However, this has immunoglobulin from SNMG patients, while other mice a short half life and some patients fail to respond, so in were shown to have increased sensitivity of neuromuscular the past two decades immunotherapies have been more transmission to D-tubocurarine without altering 125I-α- widely adopted. Steroid therapy may produce remission Bungarotoxin binding. At the cellular level, SNMG but may also be associated with temporary relapse, so behaves in a similar fashion to SPMG. T-cell responses they are best given in hospital. The long-term are no different to those observed in seropositive patients complications of steroids are well known and it is in their ability to bind to acetylcholine-derived important to aim for alternate day regimens and offer myasthogenic peptides in vitro.18 Self-reactive antibody adequate protection against osteoporosis. Azathioprine repertoires towards thymus antigens are similar in has been shown to produce a ‘steroid-sparing’ effect by patients with SPMG and SNMG, thus suggesting that the inducing remission. Some refractory patients may require two share common immunopathological features.19 These intravenous immunoglobulin treatment or plasma early clinical and pathological observations pointed to exchange. Thymic hyperplasia is observed in 70–80% of indications that SNMG was another immune-mediated patients and is associated with the presence of AChR entity, but the site of action was still to be discovered.
antibody.11 The goal of thymectomy is to induce remissionin MG or an improvement such that immunosuppressive The discovery of antibodies to MuSK
medication can be reduced.1 Approximately 15% of The first clues to the pathogenesis of SNMG were provided myasthenia patients will have a thymoma. Surgical by Blaes et al. This group observed that the antibodies in thymectomy in this group of patients will help to prevent SNMG sera were binding to a distinct antigen expressed spread of tumour. Expert opinion argues against surgery on the muscle derived TE671 cells,20 and this indicated in patients over the age of 45 because the thymus is that the target for SNMG antibodies was different to that in SPMG. This group also found that injection of SNMGsera into mice did not reduce the numbers of AChR, but SERONEGATIVE MYASTHENIA GRAVIS
increased the sensitivity of neuromuscular transmission Clinical features
to D-tubocurarine and reduced the amplitudes of MEPPs.
Approximately 15–20% of patients with symptoms and Therefore, although the numbers were unaltered, the signs of generalised MG will not have detectable antibodies function of the receptor was affected. Sera and non-IgG to the AChR and are therefore classed as SNMG. It is a fractions from SNMG patients reduced AChR function in small, yet distinct and heterogenous group with clinical TE671 cells without affecting AChR number, and the and electrophysiological features which are reduction in acetylcholine-induced currents was partly indistinguishable from those seen in SPMG.12 Patients dependent on intracellular calcium. There was also the respond to the same therapeutic measures, but SNMG suggestion that inhibition of function in SNMG was due may be more difficult and challenging to treat. Physicians to AChR desensitisation secondar y to an intra- may be less confident about the role of potentially toxic cellular signalling mechanism that lead to AChR long-term immunotherapies. Moreover, the thymus is often normal and hence thymectomy is ofteninappropriate in this subgroup of patients.13, 14 Last year investigators at the Max Planck Institute,Germany and the Institute of Molecular Medicine, Oxford Immune mechanisms in SNMG
showed that the optical density values for IgG binding to That SNMG is also immune-mediated was demonstrated human TE671 cell lines corresponded to a MuSK, thus early by the response that these patients showed to indicating that the previously identified cell surface antigen plasma exchange and immunosuppressive treatment.13 on these cell lines might be MuSK.23 Muscle specific Burges et al. also provided support for the hypothesis kinase is a receptor tyrosine kinase selectively expressed that SNMG was immune-mediated when they showed in skeletal muscle and localised at the neuromuscular that passive transfer of serum from SNMG patients into junction (see Figure 1). Phosphorylation of MuSK leads mice caused defects in neuromuscular transmission.15 to recruitment of a phophotyrosine binding domain Mossman et al. demonstrated that immunoglobulins from containing protein that stimulates phophorylation and these patients caused a small but significant loss of clustering of AChRs. Muscle specific kinase is expressed endplate AChR from the muscle diaphragm, but this was at low levels in proliferating myoblasts. In the embryo, it insufficient to explain the decrement in twitch amplitude is expressed specifically in the early myotomes and observed in neuromuscular transmission.16 They developing muscle. Muscle specific kinase is then concluded that these patients had an antibody that bound dramatically downregulated in mature muscle, where it to determinants other than AChR and that also caused remains prominent at the neuromuscular junction.
impairment of neurotransmission. Drachman et al.
Expression of MuSK is induced throughout the adult J R Coll Physicians Edinb 2002; 32:14–18
A representative cartoon of the neuromuscular junction (see text).
MuSK interacts with agrin, a nerve-derived protein, via MASC (Myotube- Associated-Specificity Component). This interactionactivates a clustering process of the AChRs, a process that requires phosphorylation and dimerisation of MuSK. MuSK alsointeracts with Rapsyn at the junction and together they provide a scaffolding effect, which is important in the maintenance of thearchitecture as a whole. Muscle specific kinase will also interact with the dystrophin-utrophin glycoprotein complex at thesame site (not shown here). Neuregulin (or ARIA) is an extracellular signal, concentrated at synaptic sites, that activatessynapse-specific transcription, in turn activating AChR gene expression in cultured muscle cells.
myofibre after denervation, block of electrical activity or in fact, reports confirm the presence of complement physical immobilisation. In humans, MuSK maps to deposition at the motor endplates of SNMG patients.23 chromosome 9q31·3–32 which overlaps with the regionreported to contain the Fukuyama muscular dystrophy MuSK in the maintenance of the architecture of
the neuromuscular junction
To understand the role of MuSK, one must grasp the
Data from Hoch et al.23 indicate that the antibodies to concepts of the stages involved in pre- and postsynaptic MuSK are directed against the extracellular N-terminal differentiation.25 Developing muscle fibres undergo a domain of MuSK. These authors also reported that these complex differentiation program, and signals from the antibodies had an inhibitory effect on agrin-induced muscle regulate differentation of the presynaptic terminal.
clustering of AChRs – an interaction that depends on Two signalling pathways are involved in mediating the N-terminal domain of MuSK. It was postulated that postsynaptic differentiation. The signal for one pathway antibodies bind to MuSK in a manner that prevents its is agrin, a synaptic basal lamina protein which interaction with MASC (Myotube–associated specificity redistributes AChRs to synaptic sites. The signal for the component, a hypothetical agrin-binding component), other pathway is also associated with the synaptic basal thus interfering with the agrin/MuSK/AChR clustering lamina but stimulates expression of the AChR genes in pathway in myotubes with the potential to alter MuSK myofibre nuclei near the synaptic site. Formation of the function at the neuromuscular junction. A limited number neuromuscular junction requires a series of reciprocal of biopsies indicate that antibodies to MuSK not only inductive interactions between the motor neuron and interfere with MuSK function but also alter the numbers the muscle cell that culminate in the precise juxtaposition and distribution of AChRs. The binding of IgG to MuSK of a highly specialised presynaptic nerve terminal with a at the postsynaptic membrane activates complement and, complex postsynaptic endplate on the muscle surface.
J R Coll Physicians Edinb 2002; 32:14–18
This is only possible when the interactions between agrin gravis: passive Transfer from man to mouse. Science 1975; and MuSK come into play.25, 26 Agrin is a nerve-derived protein and is synthesised by motor neurons and Drachman DB, Adams RN, Josifek LF et al. Functional deposited in the extracellular matrix of the activities of autoantibodies to acetylcholine receptorsand the clinical severity of myasthenia gravis. N Engl J Med neuromuscular junction. Muscle specific kinase is a component of the agrin-receptor complex and mice Newsom-Davis J, Pinching AJ, Vincent A et al. Function of lacking agrin or MuSK lack neuromuscular synapses.
circulating antibody to acetylcholine receptor in Muscle specific kinase activation signals cascades that myasthenia gravis: investigation by plasma exchange.
are important in synapse formation, including organisation of the postsynaptic membrane, synapse-specific Buckley C, Douek D, Newsom-Davis J et al. Mature, long- transcription and presynaptic differentiation.26 It appears lived CD4+ and CD8+ T cells are generated by the thymoma that it has a role in the maintenance of the architecture in myasthenia gravis. Ann Neurol 2001; 50(1):64-72.
of the postsynaptic membrane and a scaffolding effect Matej H, Nowakowska B, Kalamarz M et al. HLA antigensand susceptibility to myasthenia gravis. Arch Immunol Ther whereby agrin induces MuSK to activate AChR clustering through a synapse-specific cytoplasmic protein rapsyn.27 Spurkland A, Gilhus NE, Ronningen KS et al. Myasthenia Rapsyn-MuSK interactions are mediated by the gravis patients with thymus hyperplasia and myasthenia ectodomain of MuSK. Rapsyn is necessary not only for gravis patients with thymoma display different HLA its structural role but is involved in MuSK-signaling AChR associations. Tissue Antigens 1991; 37:90-3.
phosphorylation. This requires the ectodomain of MuSK.
Kida K, Hayashi M, Yamada I et al. Heterogeneity in Dimerisation of MuSK, induced by agrin, also has an myasthenia gravis: HLA phenotypes and autoantibody important scaffolding effect for other postsynaptic responses in ocular and generalized types. Ann Neurol proteins, namely the dystrophin/utrophin glycoprotein 10 Evoli A, Batocchi AP, Lo Monaco M et al. Clinical complex. The MuSK ectodomain is not only responsible heterogeneity of seronegative myasthenia gravis.
in mediating ligand binding and receptor dimerisation, but also recruits neuromuscular junction components 11 Matsui M, Wada H, Ohta M et al. Potential role of thymoma and other mediastinal tumours in the pathogenesis ofmyasthenia gravis. J Neuroimmunol 1993; 44:171-6.
WHAT DOES THE FUTURE HOLD?
12 Newsom-Davis J, Willcox N, Schluep M et al.
The discovery of antibodies to MuSK is important in Immunological heterogeneity and cellular mechanisms in helping to define this group of SNMG even further and myasthenia gravis. Ann NY Acad Sci 1987; 505:12-26.
substantially helps in the diagnosis and clinical 13 Birmanns B, Brenner T, Abramsky O et al. Seronegative myasthenia gravis: clinical features, response to therapy management of these patients. Using simple ELISA and synthesis of acetylcholine receptor antibodies in vitro.
techniques it should now be possible to detect AChR antibodies in 95% of patients who have clinical evidence 14 Verma PK, Oger JJ. Seronegative generalized myasthenia of MG. However, many questions remain. Our knowledge gravis: low frequency of thymic pathology. Neurology 1992; of the complex nature of the architecture and electrophysiology of the neuromuscular junction 15 Burges J, Vincent A, Molenaar PC et al. Passive transfer of continues to expand, and we now have clear evidence seronegative myasthenia gravis to mice. Muscle & Nerve that MuSK is a target antigen for immune-mediated myasthenia. However, further laboratory research is 16 Mossman S, Vincent A, Newsom-Davis J. Myasthenia gravis without acetylcholine–receptor antibody: a distinct required to characterise the exact function of this disease entity. Lancet 1986; 1:116-9.
extracellular receptor protein, its interactions with other 17 Drachman DB, de Silva S, Ramsay D et al. Humoral proteins and growth factors and the precise mechanisms pathogenesis of myasthenia gravis. Ann NY Acad Sci 1987; of pathogenesis. In the clinic, the availiability of this test will also allow us to define the clinical and epidemiological 18 Karni A, Zisman E, Katz-Levy Y et al. Reactivity of T cells characteristics of MG more precisely. We will now also from seronegative patients with myasthenia gravis to T be able to define the prognosis of MuSK-positive MG cell epitopes of the human acetylcholine receptor.
more clearly. Finally, it should be possible to design trials for this group of patients, who often prove to be somewhat 19 Sharshar T, Lacroix-Desmazes S, Mouthon L et al. Selective impairment of serum antibody repertoires toward muscle more difficult and challenging to treat.
and thymus antigens in patients with seronegative andseropositive myasthenia gravis. Eur J Immunol 1998;28:2344-54.
REFERENCES
20 Blaes F, Beeson D, Plested P et al. IgG from ‘seronegative’ Drachman DB. Myasthenia gravis. N Engl J Med 1994; myasthenia gravis patients binds to a muscle cell line, TE671, but not to human acetylcholine receptor. Ann Neurol 2000; Simpson JA. Myasthenia gravis: a new hypothesis. Scott 21 Barrett-Jolley R, Byrne N, Vincent A et al. Plasma from Toyka KV, Brachman DB, Pestronk A et al. Myasthenia patients with seronegative myasthenia gravis inhibit nAChR J R Coll Physicians Edinb 2002; 32:14–18
responses in the TE671/RD cell line. Pflugers Arch 1994; expression in embryonic muscle, at the neuromuscular junction, and after injury. Neuron 1995; 15:573-84.
22 Li Z, Forester N, Vincent A. Modulation of acetyl-choline 25 Hopf C, Hoch W. Dimerisation of the muscle-specific receptor function in TE671 (rhabdomyosarcoma) cells by kinase induces tyrosine phophorylation of acetylcholine non-AChR ligands: possible relevance to seronegative receptors and their aggregation on the surface of myasthenia gravis. J Neuroimmunol 1996; 64:179-83.
myotubes. J Biol Chem 1998; 273:6467-73.
23 Hoch W, McConville J, Helms S et al. Autoantibodies to 26 Bowen DC, Park JS, Bodine S et al. Localisation and the receptor tyrosine kinase MuSK in patients with regulation of MuSK at the neuromuscular junction. Dev myasthenia gravis without acetylcholine receptor 27 Apel ED, Glass DJ, Moscoso LM et al. Rapsyn is required 24 Valenzuela DM, Stitt TN, DiStefano PS et al. Receptor for MuSK signaling and recruits synaptic components to a tyrosine kinase specific for the skeletal muscle lineage: MuSK – containing scaffold. Neuron 1997; 18:623-35.
Royal College of Physicians of Edinburgh
Forthcoming Symposia for 2002
All symposia are held at the Royal College of Physicians of Edinburgh unless otherwise stated. Further symposia maybe added at a later date. Full details of speakers and topics to be covered in upcoming symposia are available on theCollege website: www.rcpe.ac.uk Renal Medicine – management issues in 2002 Joint Conference of the Federation of Royal Colleges of Physicians of theUK, York District Hospital Advances in health care of the Older Person Scotland’s Health – Climbing the European League Table? In association with the Public Health Institute of Scotland Paediatric gastroenterology and nutrition Joint with the Royal College of Paediatrics and Child Health Joint with the Royal College of Surgeons of Edinburgh *The date of this symposium has changed. The new date is indicated.
All grades of medical, nursing, scientific staff, professions allied to medicine and trainees in these professions arewelcome to attend. For further information on any of the above, please contact: For an up-to-date list of events, and news of other College activities please contact the College or check ourwebsite: www.rcpe.ac.uk J R Coll Physicians Edinb 2002; 32:14–18

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