In vitro proliferative responses and antibody titers specific to human acetylcholine receptor synthetic peptides in patients with myasthenia gravis and relation to HLA class II genes.

To investigate which parts of the acetylcholine receptor are involved in the initiation and development of myasthenia gravis (MG), peptides representing different sequences of the human acetylcholine receptor alpha-subunit were synthesized. These peptides were tested for their ability to stimulate T cells of myasthenic patients and healthy control patients in proliferation assays and to bind to sera antibodies. Three of eight peptides discriminated significantly between the two groups in the proliferation assay, as well as in their ability to bind to serum antibodies. HLA-DR3 and DR5 were associated with proliferative responses to specific AChR peptides in the group of myasthenics. Acetylcholine receptor epitopes that might play a specific role in myasthenia gravis thus were demonstrated.

Recognition of inter-transmembrane regions of acetylcholine receptor alpha subunit by antibodies, T cells and neurotoxins. Implications for membrane-subunit organization.

Three regions of the alpha chain of Torpedo californica acetylcholine receptor (AChR), corresponding to residues alpha 262-276, alpha 388, 408 and alpha 427-437 were synthesized, purified and characterized. The first two peptides have been proposed to occupy inter-transmembrane regions while the third represented the C-terminal segment, proposed by various models to be either extracellular or intracellular. Peptide alpha 388-408 stimulated a good response in the AChR-primed T cells of H-2s haplotype mice, a low response in the H-2q haplotype and no response in the H-2b haplotype. Peptide alpha 427-437 stimulated AChR-primed T cells of the H-2s haplotype, but caused no response in the q and b haplotypes. Peptide alpha 262-276 evoked no in vitro stimulation in any of the s, q or b haplotypes. In antibody binding studies, peptide alpha 388-408 bound antibodies raised against free AChR or against membrane-bound AChR. The other two peptides showed little or no binding activity. Further, peptide alpha 388-408 bound specifically both 125I-labelled bungarotoxin and cobratoxin, while the other two peptides had no binding activity. These results were consistent with only one of the models for subunit organization within the membrane.

Myasthenogenic significance of synthetic alpha-subunit peptide 183-200 of Torpedo californica and human acetylcholine receptor.

Synthetic peptides of the Torpedo californica and human acetylcholine receptor alpha-subunit, Gly183-Asp200, were studied. Both peptides bound alpha-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 alpha 183-200 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 immunopharmacological blockade of the acetylcholine-binding site. Furthermore, this Torpedo peptide was antigenic in the detection of antibody in human myasthenic sera. The human alpha 183-200 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.