Myasthenic Sera Research Articles

Profiling of patient-specific myocytes identifies altered gene expression in the ophthalmoplegic subphenotype of myasthenia gravis

BackgroundWhile extraocular muscles are affected early in myasthenia gravis (MG), but respond to treatment, we observe a high incidence of treatment-resistant ophthalmoplegia (OP-MG) among MG subjects with African genetic ancestry. Previously, using whole exome sequencing, we reported potentially functional variants which associated with OP-MG. The aim of this study was to profile the expression of genes harbouring the OP-MG associated variants using patient-derived subphenotype-specific ‘myocyte’ cultures.MethodsFrom well-characterised MG patients we developed the ‘myocyte’ culture models by transdifferentiating dermal fibroblasts using an adenovirus expressing MyoD. These myocyte cultures were treated with homologous acetylcholine receptor antibody-positive myasthenic sera to induce muscle transcripts in response to an MG stimulus. Gene expression in myocytes derived from OP-MG (n = 10) and control MG subjects (MG without ophthalmoplegia; n = 6) was quantified using a custom qPCR array profiling 93 potentially relevant genes which included the putative OP-MG susceptibility genes and other previously reported genes of interest in MG and experimental autoimmune myasthenia gravis (EAMG).ResultsOP-MG myocytes compared to control MG myocytes showed altered expression of four OP-MG susceptibility genes (PPP6R2, CANX, FAM136A and FAM69A) as well as several MG and EAMG genes (p < 0.05). A correlation matrix of gene pair expression levels revealed that 15% of gene pairs were strongly correlated in OP-MG samples (r > 0.78, p < 0.01), but not in control MG samples. OP-MG susceptibility genes and MG-associated genes accounted for the top three significantly correlated gene pairs (r ≥ 0.98, p < 1 × 10− 6) reflecting crosstalk between OP-MG and myasthenia pathways, which was not evident in control MG cells. The genes with altered expression dynamics between the two subphenotypes included those with a known role in gangliosphingolipid biosynthesis, mitochondrial metabolism and the IGF1-signalling pathway.ConclusionUsing a surrogate cell culture model our findings suggest that muscle gene expression and co-expression differ between OP-MG and control MG individuals. These findings implicate pathways not previously considered in extraocular muscle involvement in myasthenia gravis and will inform future studies.

Heterogeneity of auto-antibodies against nAChR in myasthenic serum and their pathogenic roles in experimental autoimmune myasthenia gravis

Many myasthenia gravis (MG) patients have auto-antibodies against the nicotinic acetylcholine receptor (nAChR), and monoclonal antibodies against the main immunogenic region (MIR) of nAChR can induce experimental autoimmune MG (EAMG). We investigated whether Fab fragment of MIR antibody (Fab35) could block the pathogenicity of polyclonal antibodies. Fab35 partially inhibited nAChR downmodulation, blocked EAMG serum-induced binding of polyclonal antibodies and complement deposition in vitro. Moreover, Fab35 did not ameliorate the EAMG serum-induced EAMG phenotype in rats. These results suggested that the EAMG serum possessed several different pathogenic antibodies that might be sufficient to induce the EAMG phenotype.

Why does acetylcholine exacerbate myasthenia caused by anti‐MuSK antibodies?

According to current knowledge, MuSK is a transmembrane protein with four immunoglobulin-like domains and a cysteine-rich domain extracellularly and an intracellular kinase domain. It is expressed on the surface of non-innervated myotubes and on the postsynaptic membrane of the neuromuscular junction (NMJ). Activation of MuSK is a prerequisite for local concentration of acetylcholine receptor (AChR) molecules via rapsyn. In non-innervated myotubes, MuSK is activated by cytosolic Dok-7 to form primitive AChR clusters. At the innervated NMJ, MuSK is activated by neural agrin through interaction with its co-receptor, the transmembrane lipoprotein receptor-related protein LRP4. At the NMJ, MuSK also participates in anchoring the collagenic tail of AChE, collagen Q (ColQ), in the synaptic space. By interacting with additional postsynaptic proteins that include Dok-7, MuSK emerges as a master organizer of postsynaptic development of the NMJ (Wu et al. 2010). Most patients suffering from autoimmune myasthenia gravis (MG) harbour anti-AChR antibodies (AChR-MG) but 5–15% do not. Clinical interest in MuSK arose when enzyme-linked immunosorbent assay (ELISA) and immunoprecipitation experiments revealed that a high proportion of AChR-seronegative myasthenic sera immunoreacted with MuSK (Hoch et al. 2001). Subsequent studies of MuSK antibody-positive myasthenia (MuSK-MG) showed it differs from AChR-MG in several respects. First, the acetylcholinesterase (AChE) inhibitor pyridostigmine, which is effective in AChR-MG, is ineffective in, or worsens, MuSK-MG. Second, MuSK-MG selectively affects bulbar, cervical and respiratory muscles whereas generalized AChR-MG also affects limb muscles. Third, intercostal muscle NMJs in AChR-MG are depleted of AChR, have low-amplitude miniature endplate potentials (MEPPs) and show signs of complement-mediated lysis of the junctional folds whereas in MuSK-MG, NMJs of intercostal or biceps brachii muscles have normal AChR content, generate normal-amplitude MEPPs and have well-preserved junctional folds that bind little or no complement. The reasons for the differences between AChR-MG and MuSK-MG are now reasonably well understood. First, the pathogenic MuSK immunoglobulin is predominantly of class 4 that does not fix complement. Second, combined results of a number of studies show that passive transfer to mice of immunoglobulin G (IgG) isolated from sera of MuSK-MG patients as well as active immunization of mice with rat MuSK causes muscle weakness, reduces MuSK expression by dispersal of synaptic AChR and decreases the synaptic response to spontaneous or evoked release of ACh. Importantly, the deleterious effects on the NMJ are greatest in cranial, paraspinal and masseter muscles that express lower levels of MuSK and MuSK RNA than the less affected muscles (Punga et al. 2011). Finally, it was proposed that pyridostigmine is generally ineffective or harmful in MuSK-MG because MuSK autoantibodies interfere with binding of ColQ to MuSK. This would reduce AChE in the synaptic space and result in cholinergic overactivity at the NMJ. Examination of intercostal muscle NMJs in human MuSK-MG did not show reduced AChE expression, but NMJs of more severely affected muscles were not examined (Kawakami et al. 2011). In this issue of the Journal of Physiology, Morsch and co-workers (2013) report that in the antibody-induced mouse model of MuSK-MG, relevant doses of pyridostigmine worsen the postsynaptic depletion of AChR and reduce the synaptic response to ACh. In contrast, 3,4-diaminopyridine (3,4-DAP), which augments quantal release by nerve impulse, improves neuromuscular transmission without worsening the loss of AChR from the NMJ. What are the possible reasons for this paradox? A partial explanation would be that pyridostigmine increases the lifetime of ACh in the synaptic space by a few milliseconds, during which ACh continues to bind to multiple AChR molecules as it exits the synaptic space by diffusion. In contrast, 3,4-DAP increases quantal release, but the lifetime of ACh in the synaptic space is less than 0.1 ms, during which ACh binds to AChR; and when ACh is released from the receptor over an exponentially distributed period, it is instantly destroyed by AChE. It is also well recognized that the agrin–LRP4–MuSK–Dok-7 signalling pathway promotes AChR clustering, whereas ACh activates an opposite pathway involving subsynaptic AChRs, inositol trisphosphate (IP3) receptors, calcium and cyclin-dependent kinase-5 to disassemble AChR clusters (Misgeld et al. 2005; Zhu et al. 2011). But is the increased exposure to ACh in the model animals sufficient to enhance the synaptic injury? In the experiments of Morsch and co-workers, treatment with conventional doses of pyridostigmine lasted only 7–9 days whereas in other studies chronic treatment of normal animals with high doses of neostigmine was required to cause disintegration of the synaptic folds with loss of AChR. This strongly implies that the enhanced toxicity of ACh in the model animals as well as in human MuSK-MG is caused by impaired cluster stabilization by MuSK. This, in turn, could be due to antibody-dependent reduced expression of MuSK, or is caused by direct interference of anti-MuSK antibodies with MuSK signalling, or both.

Presynaptic membrane receptor in human brain.

Myasthenia gravis (MG) is an autoimmune disease that results from antibody mediated damage of Acetylcholine receptor (AChR) at the neuromuscular junction. The autoimmune character of MG and pathogenic role of AChR antibodies have been established by several workers i.e., the demonstration of anti-AChR antibodies in about 90% of MG patients. It has been demonstrated that patients with MG also have antibodies against a second protein named presynaptic membrane receptor (PsmR), which is identified by utilizing β-Bgtx, a ligand which binds to PsmR. Using β-Bgtx Sepharose 4B affinity matrix, the PsmR was purified from different regions of human cadaver brain by affinity chromatography. Purified receptor was characterized both by biochemical and immunological procedures. PsmR purified from different regions of the brain shows a specific activity of 0.37±0.01, 0.39±0.02 and 0.43±0.005nM/μg of protein in Parietal lobe, Occipital lobe and Frontal lobe respectively. The affinity purified PsmR from the brain of 87 and 68kd (parietal lobe, occipital lobe and frontal lobe) shows immunoreactivity with myasthenic sera. These findings suggest that PsmR from brain is another antigen against which autoantibodies are developed in Myasthenia gravis patients. Upon treatment with various enzymes we concluded that PsmR from brain is a glycoprotein in which the immunoreactivity resides in the carbohydrate as well as the peptide epitopes. In conclusion the PsmR is another antigen against which autoantibodies are formed in different regions of brain. These can be used as a diagnostic tool for detecting antibodies in the sera or cerebrospinal fluid of MG patients.

Determination of Anti-Acetylcholine Receptor Antibodies in Myasthenic Patients by Use of Time-resolved Fluorescence

Autoantibodies against nicotinic acetylcholine receptor (nAChR) in myasthenia gravis (MG) patients are usually detected by radioimmunoprecipitation assays using extracted acetylcholine receptors labeled irreversibly with 125I-alpha-bungarotoxin (alpha-BuTx). To provide a nonradioactive immunoassay, we established an assay using nAChRs labeled with Eu(3+)-alpha-cobratoxin (alpha-CTx). We derivatized alpha-CTx with a diethylenetriaminepentaacetate moiety and formed a complex with Eu(3+). The complex was purified by HPLC, and the fractions were tested for binding to Torpedo and human nAChRs. The most active fractions were used to label nAChRs for the immunoprecipitation assay, and the bound Eu(3+) was quantified by time-resolved fluorescence. Eu(3+)-labeled alpha-CTx competed with 125I-alpha-BuTx for binding to Torpedo nAChRs and saturated the binding sites of human nAChRs, with a K(d) of 7.2 x 10(-9) mol/L. Results of the immunoassay performed with Eu(3+)-labeled alpha-CTx were similar to those obtained with 125I-alpha-BuTx, with a slightly higher limit of detection [0.3 nmol/L (n = 6) vs approximately 0.1 nmol/L for isotopic assay]. None of 34 negative sera tested (16 healthy controls, 10 patients with nonmyasthenia-related disease, 8 patients seronegative for MG) gave a value >0.3 nmol/L. Of the 35 positive myasthenic sera (with antibody values, previously determined by isotopic assay, of 0.4-1290 nmol/L) compared in the two assays, 32 tested positive with the Eu(3+) assay. Linear regression analysis yielded the equation: y = 1.035x - 0.013 nmol/L; S(y:x) = 0.172 nmol/L; r(2) = 0.977. The new time-resolved fluorescence method for quantification of antibodies to nAChRs in MG patients provides a performance similar to that of the widely used isotopic assay and could be used in laboratories with restricted use of isotopes.

Ryanodine receptor gene expression thymomas.

Ryanodine receptor (RyR) antibodies are present in sera of myasthenics with thymoma, and their titer correlates with morbidity and mortality. We investigated whether skeletal muscle RyR expression in thymic tissues could be the source of immune sensitization to the RyR. Skeletal muscle RyR gene expression was investigated using reverse transcription followed by semiquantitative polymerase chain reaction. Hyperplastic and normal thymuses expressed significant levels of RyR, but RyR gene transcripts was statistically less likely in thymomas than in hyperplastic and normal thymus (P < 0.05). The presence of RyR transcripts in thymomas did not correlate with myasthenic manifestations, thymic pathology, or serum RyR antibodies. We conclude that the skeletal muscle RyR in thymoma is not the inciting antigen for immune sensitization to RyR epitopes in thymoma-associated myasthenia gravis.

Effects of electrical stimulation and tetrodotoxin paralysis on antigenic properties of acetylcholine receptors in rat skeletal muscle

To examine the role of muscle activity in the expression of fetal- and adult-type acetylcholine receptors (AChRs), we studied the effects of muscle stimulation in cell culture and of tetrodotoxin (TTX)-induced paralysis and denervation in adult rat muscles. The AChR content of these muscles was determined using [ 125I]α-bungarotoxin and the proportion of fetal-type receptors was estimated using a radioimmunoprecipitation assay with a myasthenic serum that was highly specific for fetal-type receptors. We found that both stimulated, aneural muscle cells in vitro and inactive muscles in vivo produced predominantly fetal-type AChRs. However the TTX-paralysed muscles had a lower proportion of fetal-type receptors than the denervated muscles. We conclude that neither muscle activity nor innervation alone, but a combination of both, is required for full regulation of AChR antigenicity.

Antibodies to synthetic peptide (125‐148) of the α‐subunit of human nicotinic acetylcholine receptor in sera from patients with myasthenia gravis

We measured the amount of antibodies to a synthetic peptide that corresponds to the alpha-subunit residues Lys125-Thr148 of human acetylcholine receptor (AChR) in myasthenic sera. We detected anti-peptide antibodies in 52% (89/171) of the patients with myasthenia gravis (MG), but none in any of the healthy controls. Anti-peptide antibodies should provide a valuable immunologic parameter for the clinical evaluation of MG, but no apparent correlation was observed between the titers of anti-peptide and anti-AChR antibodies.

Expression of functional mouse muscle acetylcholine receptors in Chinese hamster ovary cells

CHO cells were transfected with cDNAs for all 4 subunits of the mouse muscle acetylcholine receptor (AChR) to obtain a stable cell line (CHO-AR) expressing the AChR on its surface. Immunoprecipitation experiments established that the AChR contained alpha- and beta-subunits assembled with gamma- and/or delta-subunits. In addition, one of the toxin-binding sites of the AChR was blocked by a myasthenic serum that specifically recognizes AChR containing the gamma-subunit. AChR from the CHO-AR cells had the same sedimentation rate, association rate constant for the binding of alpha-bungarotoxin (alpha BTX), and the same metabolic half-life as the AChR in myotubes of the mouse muscle cell line C2C12. Electrophysiological assay of CHO-AR cells by single-channel recording showed the presence of ACh-responsive ion channels with the characteristics of the embryonic AChR (gamma = 40 pS, tau = 5.6 msec). In some patches a smaller conductance channel was also seen that may represent partially assembled receptor. Fluorescence microscopy of fixed, permeabilized cells stained with rhodamine-alpha-BTX demonstrated both perinuclear and diffuse surface staining. The expression of fully assembled, functional mammalian muscle AChR in nonmuscle cells will allow detailed investigation of its properties and interactions with other cellular components.

Properties of embryonic and adult muscle acetylcholine receptors transiently expressed in COS cells

We used transient transfection in COS cells to compare the properties of mouse muscle acetylcholine receptors (AChRs) containing α, β, S, and either γ or ε subunits. γ- and ε-AChRs had identical association rates for binding 125I-a-bungarotoxin, and identical curves for inhibition of toxin binding by d-tubocurarine, but ε-AChRs had a significantly longer half-time of turnover in the membrane than γ-AChRs. A myasthenic serum specific for the embryonic form of the AChR reduced toxin binding to γ-, but not ε-AChRs. The γ-AChRs had channel characteristics of embryonic AChRs, whereas the major class of ε-AChR channels had the characteristics of adult AChRs. Two minor channel classes with smaller conductances were also seen with ε AChR. Thus, some, but not all, of the differences between AChRs at adult endplates and those in the extrasynaptic membrane can be explained by the difference in subunit composition of γ- and ε-AChRs.

Antigenic role of single residues within the main immunogenic region of the nicotinic acetylcholine receptor

The target of most of the autoantibodies against the acetylcholine receptor (AChR) in myasthenic sera is the main immunogenic region (MIR) on the extracellular side of the AChR alpha-subunit. Binding of anti-MIR monoclonal antibodies (mAbs) has been recently localized between residues alpha 67 and alpha 76 of Torpedo californica electric organ (WNPADYGGIK) and human muscle (WNPDDYGGVK) AChR. In order to evaluate the contribution of each residue to the antigenicity of the MIR, we synthesized peptides corresponding to residues alpha 67-76 from Torpedo and human AChRs, together with 13 peptide analogues. Nine of these analogues had one residue of the Torpedo decapeptide replaced by L-alanine, three had a structure which was intermediate between those of the Torpedo and human alpha 67-76 decapeptides, and one had D-alanine in position 73. Binding studies employing six anti-MIR mAbs and all 15 peptides revealed that some residues (Asn68 and Asp71) are indispensable for binding by all mAbs tested, whereas others are important only for binding by some mAbs. Antibody binding was mainly restricted to residues alpha 68-74, the most critical sequence being alpha 68-71. Fish electric organ and human MIR form two distinct groups of strongly overlapping epitopes. Some peptide analogues enhanced mAb binding compared with Torpedo and human peptides, suggesting that the construction of a very antigenic MIR is feasible.

A sensitive rosetting assay for detection of acetylcholine receptor antibodies using BC 3H-1 cells: positive results in ‘antibody-negative’ myasthenia gravis

Antibodies to acetylcholine receptor (AChR) were measured in a group of patients with myasthenia gravis (MG), some of whom had previously been classified as “antibody negative” using the standard anti-AChR radioimmunoassay (RIA). AChR antibodies were measured using the resotting assay, a new detection method which utilizes protein A-coated red blood cells and live BC 3H-1 cells, a murine cell line which expresses muscle nicotinic AChR. The results of the rosetting assay were compared with those obtained in the anti-AChR RIA. 76% of all myasthenic sera tested showed rosetting at titers higher than any of the control sera (from patients with non-myasthenic neurologic disease and normal individuals). Of the myasthenic patients previously classified as ‘antibody negative’ in the RIA using human AChR, 71% demonstrated positive rosetting. There was no correlation between the anti-AChR antibody titer obtained in the rosetting assay and that obtained in the RIA using either human or denervated rat AChR. The results suggest that the rosetting assay may meausure a subpopulation of antibodies that differs from those detected in the RIA.

Identification and molecular cloning of a cytoplasmic antigen utilising selected myasthenic sera

cDNA clones encoding a 250kDa protein (P250) localised to the cortical cytoplasm were isolated by immunological screening of a λgt11 cDNA library with serum from a patient with myasthenia gravis. The identity of the cDNA-encoded protein as P250 was shown by immunoblotting of polyclonal antibodies raised against recombinant fusion protein and the finding of a mRNA species of sufficient size on Northern blots. Nested deletions were constructed from a 2 kilobase antibody positive clone using Bal 31 exonuclease and the sequence of single-stranded DNA determined by the method of Sanger. The insert contained a poly A tail, a polyadenylation signal 15 bases upstream, and an open reading frame coding a 33kDa peptide which was moderately hydrophilic and strongly acidic. The predominant predicted secondary structure was alpha helix and numerous continuous antigenic sites were predicted. Neither protein nor cDNA sequences were present in GENBANK or NBRF databases as from 5/89. The expression of P250 was cell type-specific on protein and mRNA blots, occuring only in adherent cell lines and being inducible by phorbol esters. We hypothesize that P250 is a new member of a super-family of actin cross-linking proteins which includes spectrin and dystrophin; P250 may be involved in some of the variations in the endothelial cytoskeleton and link other cytoskeletal components to transmembrane proteins.

Fab fragments of monoclonal antibodies protect the human acetylcholine receptor against antigenic modulation caused by myasthenic sera

The human cell line TE671 produces large amounts of muscle nicotinic acetylcholine receptor (AChR). TE671 cells were used to determine the specificity of antibodies which can increase the internalization rate of AChR (antigenic modulation) and to test procedures for protecting AChR against this mechanism. The half-life of AChR both in the absence and the presence of anti-AChR antibodies was very similar to that of AChR on human muscle cell cultures. The relative contribution of different anti-AChR antibody fractions to the total antigenic modulation capacity of human myasthenic sera was investigated by competition experiments between Fab fragments of anti-AChR monoclonal antibodies (MoAbs) and intact antibodies (MoAb or myasthenic sera). Fab fragments, which do not induce antigenic modulation, were allowed to shield the corresponding regions of the AChR. Intact antibodies were subsequently added. It was found that protection of the main immunogenic region (MIR), but not of a region on the β-subunit, essentially blocked the modulatory effect of the intact anti-MIR MoAbs, and approximately 80% of that of myasthenic sera. These data suggest that anti-MIR antibodies are mainly responsible for the loss of human AChR via antigenic modulation. Furthermore the observation that Fab fragments of anti-MIR MoAbs can efficiently protect AChR against antigenic modulation may have therapeutic implications.

Changes in tetrodotoxin-resistant action potentials after passive transfer of myasthenia gravis patient sera

Muscle electrical activity has been studied in mice after intraperitoneal injection of sera from myasthenia gravis (MG) patients. Myasthenic serum did not modify the electrical properties of innervated muscle fibres. The resting membrane potential and the action potential parameters remained unchanged. However, tetrodotoxin (TTX)-resistant action potentials of denervated muscles were reduced by myasthenic serum, possibly in association with receptor endocytosis induced by the immunoglobulin. However, a direct effect of MG serum on TTX-resistant sodium channels cannot be ruled out.

Loss of antigenic properties of acetylcholine receptors in rat skeletal muscle after birth

Antibodies in a myasthenic serum were used to follow the changes in antigenic properties of acetylcholine receptors in rat skeletal muscle during development. In binding assays at saturating concentrations of antigen or antibody, the antibodies reacted with extrajunctional receptors of fetal and denervated adult muscle but showed little binding to junctional receptors of adult rat muscle. They did, however, bind to junctional receptors of adult chicken muscle which, unlike rat receptors, do not appear to undergo a change in their channel properties during development. Binding studies with acetylcholine receptors of developing rat muscle carried out at saturating concentrations of antibody showed that the loss of antigenic determinant(s) begins at 1–2 days after birth.

Myasthenogenic significance of synthetic α-subunit peptide 183–200 of Torpedo californica and human acetylcholine receptor

Synthetic peptides of the Torpedo californica and human acetylcholine receptor α-subunit, Gly183–Asp200, were studied. Both peptides bound α-bungarotoxin to a significant extent, and inhibited toxin-binding with Torpedo or human acetylcholine receptor. The human peptide was, however, less potent than the Torpedo peptide. One can assume, therefore, that this sequence participates in cholinergic binding. The Torpedo α183–200 segment was immunogenic in the induction of experimental autoimmune myasthenia gravis in rats, accompanied by elevation of anti-peptide and anti-rat acetylcholine receptor blocking antibody and reduced amplitudes of miniature end-plate potentials. Sera did not accelerate the degradation of rat acetylcholine receptor. Thus, one may interpret this to be the animal model induced by an immuno-pharmacological blockade of the acetylcholine-binding site. Furthermore, this Torpedo peptide was antigenic in the detection of antibody in human myasthenic sera. The human α183–200 segment had neither of these 2 properties, although the corresponding anti-peptide antibody was elicited in immunized rats. It is feasible that the synthetic segment of this human sequence may not be maintained in such a conformation to be immunogenic.

Presynaptic Modification by AntiAcetylcholine Receptor Antibody: Study on Myasthenic Serum and Monoclonal Antibody Produced by Cloned Cells

Presynaptic Modification by AntiAcetylcholine Receptor Antibody: Study on Myasthenic Serum and Monoclonal Antibody Produced by Cloned Cells

Complement-mediated muscle damage produced by myasthenic sera.

Annals of the New York Academy of SciencesVolume 505, Issue 1 p. 180-193 Complement-Mediated Muscle Damage Produced by Myasthenic Sera LISBETH A. CHILDS, LISBETH A. CHILDS Department of Biochemistry University of Bath Bath BA2 7AY, England Recipient of a travel grant from the Wellcome Trust.Search for more papers by this authorROGER HARRISON, ROGER HARRISON Department of Biochemistry University of Bath Bath BA2 7AY, England Recipient of project grants from the Wellcome Trust and the Muscular Dystrophy Group of Great Britain and Northern Ireland.Search for more papers by this authorGEORGE G. LUNT, GEORGE G. LUNT Department of Biochemistry University of Bath Bath BA2 7AY, England Recipient of project grants from the Wellcome Trust and the Muscular Dystrophy Group of Great Britain and Northern Ireland.Search for more papers by this author LISBETH A. CHILDS, LISBETH A. CHILDS Department of Biochemistry University of Bath Bath BA2 7AY, England Recipient of a travel grant from the Wellcome Trust.Search for more papers by this authorROGER HARRISON, ROGER HARRISON Department of Biochemistry University of Bath Bath BA2 7AY, England Recipient of project grants from the Wellcome Trust and the Muscular Dystrophy Group of Great Britain and Northern Ireland.Search for more papers by this authorGEORGE G. LUNT, GEORGE G. LUNT Department of Biochemistry University of Bath Bath BA2 7AY, England Recipient of project grants from the Wellcome Trust and the Muscular Dystrophy Group of Great Britain and Northern Ireland.Search for more papers by this author First published: August 1987 https://doi.org/10.1111/j.1749-6632.1987.tb51291.xCitations: 5 AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat Citing Literature Volume505, Issue1Myasthenia Gravis: Biology and TreatmentAugust 1987Pages 180-193 RelatedInformation

Nicotine induces T-suppressor cells: Modulation by the nicotinic antagonist d-tubocurarine and myasthenic serum

To further unravel the basic immunoregulatory defect present in myasthenia gravis, we undertook to study nicotinic acetylcholine receptor (AchR) activity on human peripheral blood leukocytes. A biphasic suppressive effect of nicotine was observed on lymphocyte proliferative responses to phytohemagglutinin, concanavalin A, and poke-weed mitogen with peaks at 10 −9 and 10 −5 M. Using a coculture approach, T cells preincubated with nicotine for 24 hr showed enhanced suppressor cell activity. This was partially blocked by serum from myasthenic patients and by the nicotinic antagonist d-tubocurarine. These studies suggest that suppressor T lymphocytes bear functional nicotinic AchR, which may be modulated by myasthenic serum, with possible relevance to the pathogenesis of myasthenia gravis.