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S Fuchs

Publications and source records attributed to S Fuchs.

At least 73 records · Page 4Linked to original sources
PubMed1986

Anti-idiotypic antibodies to anti-acetylcholine receptor antibody: characterization by ELISA and immunoprecipitation assays.

The idiotype network is important both as a means of autoregulation of immune mechanisms and a potential tool for manipulation of abnormal responses. In the autoimmune disease myasthenia gravis the acetylcholine receptor (AChR) is the target of an aberrant immune response. In this study we compare 2 widely used methods of antibody determination--immunoprecipitation radioimmunoassay (IPRA) and enzyme-linked immunoassay (ELISA)--for their ability to detect both anti-AChR antibodies (polyclonal and monoclonal) and anti-idiotypic antibodies raised against polyclonal anti-AChR antibodies. Although the IPRA is considerably more sensitive for the detection of monoclonal anti-AChR antibodies, the 2 methods produce similar results in the detection of anti-idiotypic antibodies to the anti-AChR immune response. The 2 techniques also demonstrated specificity of the reagents for idiotypes associated with the anti-AChR response and absence of effect on an idiotype associated with the control antigen, ovalbumin. The results demonstrate that the idiotypic repertoire of the polyclonal anti-AChR response in C57B1/6 mice is sufficiently restricted that antigen-specific blocking anti-idiotypic antibodies can be raised in rabbits by immunization with anti-AChR antibodies.

Animals↗
PubMed1986

Analysis of ligand binding to the synthetic dodecapeptide 185-196 of the acetylcholine receptor alpha subunit.

A synthetic dodecapeptide corresponding to residues 185-196 of the Torpedo acetylcholine receptor alpha subunit, which contains the adjacent cysteine residues at positions 192 and 193, was recently shown by us to contain the essential elements for alpha-bungarotoxin binding. In the present study, we have used Sepharose-linked peptides for quantitative analysis of the cholinergic binding properties of this and other synthetic peptides. Sepharose-linked peptides corresponding to residues 1-20, 126-143, 143-158, 169-181, 185-196, 193-210, and 394-409 of the alpha subunit of Torpedo acetylcholine receptor, as well as a peptide corresponding to residues 185-196 of the alpha subunit of human acetylcholine receptor, were tested for their toxin-binding capacity. Of these immobilized peptides, only peptide 185-196 of the Torpedo acetylcholine receptor bound toxin significantly, thus verifying that this synthetic peptide contains essential components of the receptor toxin-binding site. Analysis of toxin binding to the peptide yielded a dissociation constant of 3.5 X 10(-5) M. This binding was inhibited by various cholinergic ligands. The inhibition potency obtained was alpha-bungarotoxin greater than Naja naja siamensis toxin greater than d-tubocurarine greater than decamethonium greater than acetylcholine greater than carbamoylcholine. This pharmacological profile resembles that of the nicotinic acetylcholine receptor and therefore suggests that the synthetic dodecapeptide also includes the neurotransmitter binding site. Reduction and carboxymethylation of the cysteine residues on peptide 185-196 inhibit its capacity to bind toxin, demonstrating that an intact disulfide is required for toxin binding. A decrease in toxin binding was also obtained following chemical modification of the tryptophan residue at position 187, thus implying its possible involvement in toxin binding. The failure to detect binding of toxin to the corresponding human sequence 185-196, in which the tryptophan residue is replaced by serine, supports this hypothesis.

Binding Sites↗
PubMed1986

Mapping of the alpha-bungarotoxin binding site within the alpha subunit of the acetylcholine receptor.

Synthetic peptides and their respective antibodies have been used in order to map the alpha-bungarotoxin binding site within the alpha subunit of the acetylcholine receptor. By using antibodies to a synthetic peptide corresponding to residues 169-181 of the alpha subunit, we demonstrate that this sequence is included within the 18-kDa toxin binding fragment previously reported. Furthermore, the 18-kDa fragment was also found to bind a monoclonal antibody (5.5) directed against the cholinergic binding site. Sequential proteolysis of the acetylcholine receptor with trypsin, prior to Staphylococcus aureus V8 protease digestion, resulted in a 15-kDa toxin binding fragment that is included within the 18-kDa fragment but is shorter than it only at its carboxyl terminus. This 15-kDa fragment therefore initiates beyond Asp-152 and terminates in the region of Arg-313/Lys-314. In addition, experiments are reported that indicate that in the intact acetylcholine receptor, Cys-128 and/or Cys-142 are not crosslinked by disulfide bridges with any of the cysteines (at positions 192, 193, and 222) that reside in the 15-kDa toxin binding fragment. Finally, the synthetic dodecapeptide Lys-His-Trp-Val-Tyr-Tyr-Thr-Cys-Cys-Pro-Asp-Thr, which is present in the 15-kDa fragment (corresponding to residues 185-196 of the alpha subunit) was shown to bind alpha-bungarotoxin directly. This binding was completely inhibited by competition with d-tubocurarine.

Amino Acid Sequence↗
PubMed1986

The treatment of passively transferred experimental myasthenia with anti-idiotypic antibodies.

Experimental autoimmune myasthenia gravis (EAMG) was induced in chickens by passive transfer of a monoclonal antibody (mcAb) specific for the cholinergic binding site of the acetylcholine receptor (AChR). Specific anti-idiotypes raised in rabbits against this antisite mcAb (5.5) were demonstrated to prevent the induction of EAMG by a subsequent injection of mcAb 5.5. Also, administration of anti-idiotypes against mcAb 5.5 to chickens in which EAMG has been induced by mcAb 5.5 led to a recovery from myasthenic symptoms. These results suggest that passive transfer of the appropriate anti-idiotypes may have potential in the regulation of myasthenia.

Animals↗
PubMed1985

[Clinical findings in multiple sclerosis].

The mean interval between first and last clinical interview in 170 patients with multiple sclerosis was 8.8 years. The main initial symptoms were sensory disturbances and paresis of the cranial nerves. An uncharacteristic symptomatology was found in 6%, psychiatric symptoms in 6%. In the last investigation psychiatric symptoms were found in 2%, uncharacteristic symptoms were seen in 4%. A normal neurological status was found in 9%. Apart from the clinical well defined typical MS patient, also benign forms of the diseases have been found.

Adolescent↗
PubMed1985

Analysis and modulation of the immune response of mice to acetylcholine receptor by anti-idiotypes.

Anti-idiotypes were raised in mice against three well-characterized anti-acetylcholine receptor (AChR) monoclonal antibodies (mcAbs), as well as against polyclonal mouse anti-AChR antibodies. In binding experiments, the anti-idiotypic antibodies inhibited the binding of AChR only to the immunizing idiotype. However, a less restricted specificity was found in in vivo experiments. Mice producing anti-idiotypes were challenged with AChR and the idiotypic composition of their anti-AChR response was analysed using specific rabbit anti-idiotypic antibodies. It was found that preimmunization with a certain idiotype leads to the preferential suppression of this particular idiotype in the polyclonal response to AChR. However, preimmunization with either polyclonal or monoclonal anti-AChR antibodies resulted in a reduction of the overall anti-Torpedo AChR and anti-muscle AChR titers. This reduction was greater than would be expected from the representation of each of the respective idiotypes in the polyclonal anti-AChR serum, and may imply that in addition to the immunizing idiotype other anti-AChR idiotypes are also suppressed. Our results suggest that anti-idiotypes may have a potential for the modulation of the autoimmune response directed against AChR in myasthenia.

Animals↗
PubMed1985

Antigenic specificity of acetylcholine receptor in developing muscle. Studies with monoclonal antibodies.

Monoclonal antibodies (mcAbs) elicited against the nicotinic acetylcholine receptor (AChR) from Torpedo, were used to follow antigenic changes in AChR during muscle development. Newborn rat muscle and denervated mouse muscle were used as sources of extrajunctional AChR; adult innervated rat and mouse muscle were used as sources of junctional AChR. Most of the mcAbs tested reacted preferably, but not exclusively with extrajunctional AChR (EJR), as compared to junctional AChR (JR). None was found to react with only one of the two forms of AChR. We conclude that the anti-AChR monoclonal antibodies used in this study detect antigenic determinants which are shared by EJR and JR, but which probably undergo structural changes during muscle development.

Animals↗
PubMed1985

Antibodies to synthetic peptides as probes for the binding site on the alpha subunit of the acetylcholine receptor.

Synthetic peptides and their respective antibodies were used in an attempt to localize and identify the ligand-binding site of the nicotinic acetylcholine receptor. Two peptides of the receptor alpha subunit were synthesized, the first corresponding to the NH2-terminal domain (positions 1-20) and the other, to a segment (residues 126-143) that contains the first two cysteine residues. Specific antipeptide antibodies were elicited in rabbits after immunization with the peptides conjugated to bovine serum albumin. The antipeptide antibodies thus obtained cross-reacted with the receptor and bound specifically to its alpha subunit. The antipeptide antibodies were used to test whether the peptide sequences corresponded to the alpha-bungarotoxin (alpha-BTX)-binding site. Staphylococcus aureus V8-protease digestion of the isolated receptor alpha subunit generated several fragments. Antipeptide (1-20) and antipeptide (126-143) both bound a 26-kDa fragment, whereas only antipeptide (126-143) bound a 17-kDa fragment. None of these fragments were found to bind alpha-BTX. On the other hand, alpha-BTX bound to an 18-kDa fragment that did not react with either of the antipeptide antibodies. Moreover, the 26-kDa and 17-kDa fragments were also found to contain the endoglycosidase H-susceptible oligosaccharide chain. Our results indicate that the toxin-binding site lies beyond the first possible V8 protease cleavage site after residues 126-143: i.e., Asp-152. This location is in agreement with the possibility that cysteine residues 192 and/or 193 are in close proximity to or contiguous with the ligand-binding site.

Acetylcholine↗
PubMed1984

Anti-acetylcholine receptor response achieved by immunization with a synthetic peptide from the receptor sequence.

A synthetic peptide corresponding to the first twenty amino acids of the N-terminal region from the alpha-subunit of the Torpedo acetylcholine receptor cross reacts with antibodies to the receptor. A conjugate of this peptide to bovine serum albumin elicits in rabbits an immune response towards the synthetic peptide as well as towards the acetylcholine receptor. Blotting experiments demonstrate that the antipeptide antibodies react exclusively with the alpha-subunit of the acetylcholine receptor. Antibodies against synthetic peptides from various regions of the receptor sequence may provide useful reagents for structural and developmental analysis of the acetylcholine receptor as well as for the regulation of experimental autoimmune myasthenia gravis.

Animals↗
PubMed1984

Induction of acetylcholine receptor-specific suppression. An in vitro model of antigen-specific immunosuppression in myasthenia gravis.

This report describes the in vivo and in vitro induction of murine (AChR)-specific suppressor T cells (Ts) and T cell factors (TsF), and the development of an appropriate assay system for their measurement. The assay described is based on the in vitro Mishell-Dutton culture system. Using this assay, it was shown that the AChR-specific helper cell is an Lyt-2- radiosensitive T cell. Moreover, the proliferating cell measured in the lymphocyte transformation assay was shown to provide AChR-specific T cell help. In vivo induction of Ts cells is achieved by injection of soluble AChR; potent AChR-specific suppression is found in the spleen 1 wk later. In vitro induction of Ts cells involves the primary education of naive splenocytes by culturing them with high concentrations of AChR. Both the in vivo- and in vitro-induced Ts cells were shown to secrete AChR-specific factors that mediate their suppressive effects. The possibility of specifically suppressing the AChR-immune response may be of a particular clinical importance since the AChR is the target autoantigen in the neuromuscular autoimmune disease myasthenia gravis.

Animals↗
PubMed1983

An anti-acetylcholine receptor monoclonal antibody cross-reacts with phosvitin.

Rabbit and mouse anti-Torpedo acetylcholine receptor antibodies cross-reacted partially with the highly phosphorylated protein, phosvitin. We have selected an anti-Torpedo acetylcholine receptor monoclonal antibody which binds specifically to phosvitin; this binding is inhibited by acetylcholine receptor. These findings suggest that a phosphorylated amino acid residue may be a part of the determinant on the acetylcholine receptor recognized by this monoclonal antibody.

Animals↗