A somatically mutated human antiganglioside IgM antibody that induces experimental neuropathy in mice is encoded by the variable region heavy chain gene, V1-18.

Abstract

IgM paraproteins associated with autoimmune peripheral neuropathy and anti-Pr cold agglutinins react with sialic acid epitopes present on disialylated gangliosides including GD1b, GT1b, GQ1b, and GD3. A causal relationship between the paraprotein and the neuropathy has never been proven experimentally. From peripheral blood B cells of an affected patient, we have cloned a human hybridoma secreting an antidisialosyl IgM mAb, termed Ha1, that shows identical structural and functional characteristics to its serum counterpart. Variable region analysis shows Ha1 is encoded by the same V H 1 family heavy chain gene, V1-18, as the only other known anti-Pr antibody sequence and is somatically mutated, suggesting that is arose in vivo in response to antigenic stimulation. In the rodent peripheral nervous system, Ha1 immunolocalizes to dorsal root ganglia, motor nerve terminals, muscle spindles, myelinated axons, and nodes of Ranvier. After intraperitoneal injection of affinity-purified antibody into mice for 10 d, electrophysiological recordings from the phrenic nerve-hemidiaphragm preparation demonstrated impairment of nerve excitability and a reduction in quantal release of neurotransmitter. These data unequivocally establish that an antidisialosyl antibody can exert pathophysiological effects on the peripheral nervous system and strongly support the view that the antibody contributes to the associated human disease. ( J. Clin. Invest. 1996. 97:1155-1164.)

Monoclonal antibodies raised against Guillain-Barré syndrome–associated Campylobacter jejuni lipopolysaccharides react with neuronal gangliosides and paralyze muscle-nerve preparations

“…IgG and IgM mAb's were then purified on HiTrap protein G (Supelco, Bellefonte, Pennsylvania, USA) and recombinant Protein L columns (Actigen, Cambridge, United Kingdom), respectively. mAb's were analyzed for monoclonality, light chain type, and subclass by isoelectric focusing and Western blotting using standard methods and reagents as described previously (16). The concentrations of all mAb's were measured using a radial immunoglobulin diffusion kit (Binding Site, Birmingham, United Kingdom).…”

“…The basic structure of GQ1b and related gangliosides is shown in Figure 1. Mouse sera were tested for IgG and IgM response to gangliosides and LPSs by ELISA (16). Immunolon 2 microtiter plates (Dynatech, Ashford, United Kingdom) were coated with 200 ng ganglioside in methanol or 1 µg LPS in PBS per well.…”

“…However, some in vitro electrophysiological evidence suggests they may be responsible for muscle weakness (15)(16)(17), possibly via their action on the neuromuscular junction (NMJ). Using the mouse phrenic nerve hemidiaphragm preparation as an ex vivo model for motor nerve terminal transmission, we have recently shown that antiGQ1b antibody containing sera, IgG fractions, and a cloned anti-disialosyl IgM antibody from a chronic patient with MFS exerts a complement-dependent, α-latrotoxin-like effect at the NMJ, i.e., a temporary dramatic increase in spontaneous neurotransmitter release, followed by block of evoked release resulting in paralysis (18).…”

Probing the cis interactions of the inhibitory receptor Siglec-7 with α2,8-disialylated ligands on natural killer cells and other leukocytes using glycan-specific antibodies and by analysis of α2,8-sialyltransferase gene expression

“…The anti-GD3 mAb, R24, was a kind gift from Dr. Paul Chapman (Memorial Sloan-Kettering Cancer Center, New York, NY) and was used as a purified IgG. The human Ha1 and Pi1 IgM mAb were described and purified as reported previously [28,29]. Biotinylation of these antibodies was performed using the EZ-link biotinylation kit (Pierce, Cramlington, UK).…”

“…To investigate the potential expression of ␣2,8-linked disialic acid moieties on the surface of NK cells, we used human IgM mAb Ha1 and Pi1, both of which are derived from patients with peripheral neuropathy [28,29]. In previous studies, these antibodies have been shown to react similarly with b-series gangliosides GD1b, GT1b, GQ1b, and GD3 in a manner consistent with their minimal epitope being NeuAc␣2,8- [30,31].…”