Serum anti-GM2 and anti-GalNAc-GD1a ganglioside IgG antibodies are biomarkers for immune-mediated polyneuropathies in cats.

Recent work identified anti-GM2 and anti-GalNAc-GD1a IgG ganglioside antibodies as biomarkers in dogs clinically diagnosed with acute canine polyradiculoneuritis, in turn considered a canine equivalent of Guillain-Barré syndrome. This study aims to investigate the serum prevalence of similar antibodies in cats clinically diagnosed with immune-mediated polyneuropathies. The sera from 41 cats clinically diagnosed with immune-mediated polyneuropathies (IPN), 9 cats with other neurological or neuromuscular disorders (ONM) and 46 neurologically normal cats (CTRL) were examined for the presence of IgG antibodies against glycolipids GM1, GM2, GD1a, GD1b, GalNAc-GD1a, GA1, SGPG, LM1, galactocerebroside and sulphatide. A total of 29/41 IPN-cats had either anti-GM2 or anti-GalNAc-GD1a IgG antibodies, with 24/29 cats having both. Direct comparison of anti-GM2 (sensitivity: 70.7%; specificity: 78.2%) and anti-GalNAc-GD1a (sensitivity: 70.7%; specificity: 70.9%) antibodies narrowly showed anti-GM2 IgG antibodies to be the better marker for identifying IPN-cats when compared to the combined ONM and CTRL groups (P = .049). Anti-GA1 and/or anti-sulphatide IgG antibodies were ubiquitously present across all sample groups, whereas antibodies against GM1, GD1a, GD1b, SGPG, LM1 and galactocerebroside were overall only rarely observed. Anti-GM2 and anti-GalNAc-GD1a IgG antibodies may serve as serum biomarkers for immune-mediated polyneuropathies in cats, as previously observed in dogs and humans.

Guillain-Barré Syndrome outbreak associated with Zika virus infection in French Polynesia: a case-control study.

BACKGROUND: Between October, 2013, and April, 2014, French Polynesia experienced the largest Zika virus outbreak ever described at that time. During the same period, an increase in Guillain-Barré syndrome was reported, suggesting a possible association between Zika virus and Guillain-Barré syndrome. We aimed to assess the role of Zika virus and dengue virus infection in developing Guillain-Barré syndrome. METHODS: In this case-control study, cases were patients with Guillain-Barré syndrome diagnosed at the Centre Hospitalier de Polynésie Française (Papeete, Tahiti, French Polynesia) during the outbreak period. Controls were age-matched, sex-matched, and residence-matched patients who presented at the hospital with a non-febrile illness (control group 1; n=98) and age-matched patients with acute Zika virus disease and no neurological symptoms (control group 2; n=70). Virological investigations included RT-PCR for Zika virus, and both microsphere immunofluorescent and seroneutralisation assays for Zika virus and dengue virus. Anti-glycolipid reactivity was studied in patients with Guillain-Barré syndrome using both ELISA and combinatorial microarrays. FINDINGS: 42 patients were diagnosed with Guillain-Barré syndrome during the study period. 41 (98%) patients with Guillain-Barré syndrome had anti-Zika virus IgM or IgG, and all (100%) had neutralising antibodies against Zika virus compared with 54 (56%) of 98 in control group 1 (p<0.0001). 39 (93%) patients with Guillain-Barré syndrome had Zika virus IgM and 37 (88%) had experienced a transient illness in a median of 6 days (IQR 4-10) before the onset of neurological symptoms, suggesting recent Zika virus infection. Patients with Guillain-Barré syndrome had electrophysiological findings compatible with acute motor axonal neuropathy (AMAN) type, and had rapid evolution of disease (median duration of the installation and plateau phases was 6 [IQR 4-9] and 4 days [3-10], respectively). 12 (29%) patients required respiratory assistance. No patients died. Anti-glycolipid antibody activity was found in 13 (31%) patients, and notably against GA1 in eight (19%) patients, by ELISA and 19 (46%) of 41 by glycoarray at admission. The typical AMAN-associated anti-ganglioside antibodies were rarely present. Past dengue virus history did not differ significantly between patients with Guillain-Barré syndrome and those in the two control groups (95%, 89%, and 83%, respectively). INTERPRETATION: This is the first study providing evidence for Zika virus infection causing Guillain-Barré syndrome. Because Zika virus is spreading rapidly across the Americas, at risk countries need to prepare for adequate intensive care beds capacity to manage patients with Guillain-Barré syndrome. FUNDING: Labex Integrative Biology of Emerging Infectious Diseases, EU 7th framework program PREDEMICS. and Wellcome Trust.

Inhibition of complement in Guillain-Barré syndrome: the ICA-GBS study.

The outcome of Guillain-Barré syndrome (GBS) remains unchanged since plasma exchange and intravenous immunoglobulin (IVIg) were introduced over 20 years ago. Pathogenesis studies on GBS have identified the terminal component of complement cascade as a key disease mediator and therapeutic target. We report the first use of terminal complement pathway inhibition with eculizumab in humans with GBS. In a randomised, double-blind, placebo-controlled trial, 28 subjects eligible on the basis of GBS disability grade of at least 3 were screened, of whom 8 (29%) were randomised. Five received eculizumab for 4 weeks, alongside standard IVIg treatment. The safety outcomes, monitored via adverse events capture, showed eculizumab to be well-tolerated and safe when administered in conjunction with IVIg. Primary and secondary efficacy outcomes in the form of GBS disability scores (GBS DS), MRC sum scores, Rasch overall disability scores, and overall neuropathy limitation scores are reported descriptively. For the primary efficacy outcome at 4 weeks after recruitment, two of two placebo- and two of five eculizumab-treated subjects had improved by one or more grades on the GBS DS. Although the small sample size precludes a statistically meaningful analysis, these pilot data indicate further studies on complement inhibition in GBS are warranted.

Anti-ganglioside antibodies are removed from circulation in mice by neuronal endocytosis.

SEE VAN DOORN AND JACOBS DOI101093/BRAIN/AWW078 FOR A SCIENTIFIC COMMENTARY ON THIS ARTICLE : In axonal forms of Guillain-Barré syndrome, anti-ganglioside antibodies bind gangliosides on nerve surfaces, thereby causing injury through complement activation and immune cell recruitment. Why some nerve regions are more vulnerable than others is unknown. One reason may be that neuronal membranes with high endocytic activity, including nerve terminals involved in neurotransmitter recycling, are able to endocytose anti-ganglioside antibodies from the cell surface so rapidly that antibody-mediated injury is attenuated. Herein we investigated whether endocytic clearance of anti-ganglioside antibodies by nerve terminals might also be of sufficient magnitude to deplete circulating antibody levels. Remarkably, systemically delivered anti-ganglioside antibody in mice was so avidly cleared from the circulation by endocytosis at ganglioside-expressing plasma membranes that it was rapidly rendered undetectable in serum. A major component of the clearance occurred at motor nerve terminals of neuromuscular junctions, from where anti-ganglioside antibody was retrogradely transported to the motor neuron cell body in the spinal cord, recycled to the plasma membrane, and secreted into the surrounding spinal cord. Uptake at the neuromuscular junction represents a major unexpected pathway by which pathogenic anti-ganglioside antibodies, and potentially other ganglioside binding proteins, are cleared from the systemic circulation and also covertly delivered to the central nervous system.

Swab pooling enables rapid expansion of high-throughput capacity for SARS-CoV-2 community testing.

BACKGROUND: The challenges of rapid upscaling of testing capacity were a major lesson from the COVID-19 pandemic response. The need for process adjustments in high-throughput testing laboratories made sample pooling a challenging option to implement. OBJECTIVE: This study aimed to evaluate whether pooling samples at source (swab pooling) was as effective as qRT-PCR testing of individuals in identifying cases of SARS-CoV-2 in real-world community testing conditions using the same high-throughput pipeline. METHODS: Two cohorts of 10 (Pool10: 1,030 participants and 103 pools) and 6 (Pool6: 1,284 participants and 214 pools) samples per pool were tested for concordance, sensitivity, specificity, and Ct value differences with individual testing as reference. RESULTS: Swab pooling allowed unmodified application of an existing high-throughput SARS-Cov-2 testing pipeline with only marginal loss of accuracy. For Pool10, concordance was 98.1% (95% Confidence interval: 93.3-99.8%), sensitivity was 95.7% (85.5-99.5%), and specificity was 100.0% (93.6-100.0%). For Pool6, concordance was 97.2% (94.0-99.0%), sensitivity was 97.5% (93.7-99.3%), and specificity was 96.4% (87.7-99.6%). Differences of outcomes measure between pool size were not significant. Most positive individual samples, which were not detected in pools, had very low viral concentration. If only individual samples with a viral concentration > 400 copies/ml (i.e. Ct value < 30) were considered positive, the overall sensitivity of pooling increased to 99.5%. CONCLUSION: The study demonstrated high sensitivity and specificity by swab pooling and the immediate capability of high-throughput laboratories to implement this method making it an option in planning of rapid upscaling of laboratory capacity for future pandemics.

Guillain-Barré Syndrome Following Zika Virus Infection Is Associated With a Diverse Spectrum of Peripheral Nerve Reactive Antibodies.

BACKGROUND AND OBJECTIVES: Recent outbreaks of Zika virus (ZIKV) in South and Central America have highlighted significant neurologic side effects. Concurrence with the inflammatory neuropathy Guillain-Barré syndrome (GBS) is observed in 1:4,000 ZIKV cases. Whether the neurologic symptoms of ZIKV infection are immune mediated is unclear. We used rodent and human live cellular models to screen for anti-peripheral nerve reactive IgG and IgM autoantibodies in the sera of patients with ZIKV with and without GBS. METHODS: In this study, 52 patients with ZIKV-GBS were compared with 134 ZIKV-infected patients without GBS and 91 non-ZIKV controls. Positive sera were taken forward for target identification by immunoprecipitation and mass spectrometry, and candidate antigens were validated by ELISA and cell-based assays. Autoantibody reactions against glycolipid antigens were also screened on an array. RESULTS: Overall, IgG antibody reactivities to rat Schwann cells (SCs) (6.5%) and myelinated cocultures (9.6%) were significantly higher, albeit infrequent, in the ZIKV-GBS group compared with all controls. IgM antibody immunoreactivity to dorsal root ganglia neurones (32.3%) and SCs (19.4%) was more frequently observed in the ZIKV-GBS group compared with other controls, whereas IgM reactivity to cocultures was as common in ZIKV and non-ZIKV sera. Strong axonal-binding ZIKV-GBS serum IgG antibodies from 1 patient were confirmed to react with neurofascin 155 and 186. Serum from a ZIKV-infected patient without GBS displayed strong myelin-binding and putative antilipid antigen reaction characteristics. There was, however, no significant association of ZIKV-GBS with any known antiglycolipid antibodies. DISCUSSION: Autoantibody responses in ZIKV-GBS target heterogeneous peripheral nerve antigens suggesting heterogeneity of the humoral immune response despite a common prodromal infection.

Detection of Autoantibodies Using Combinatorial Glycolipid Microarrays.

Glycolipids cluster in plasma membranes to form heterogenous patches. Whereas lectins and antibodies have been conventionally viewed as binding a single oligosaccharide head group, and assayed accordingly, it is increasingly evident that cis-interactions between glycan headgroups may form unique molecular shapes that either enhance or attenuate binding of antibodies and other proteins. Herein we describe a method for assaying antibody binding to heteromeric glycolipid complexes that allows rapid, simple, inexpensive and high-throughput assessment of binding events, focusing on autoantibodies present in human serum.

Anti-GD1a antibodies activate complement and calpain to injure distal motor nodes of Ranvier in mice.

The motor axonal variant of Guillain-Barré syndrome is associated with anti-GD1a immunoglobulin antibodies, which are believed to be the pathogenic factor. In previous studies we have demonstrated the motor terminal to be a vulnerable site. Here we show both in vivo and ex vivo, that nodes of Ranvier in intramuscular motor nerve bundles are also targeted by anti-GD1a antibody in a gradient-dependent manner, with greatest vulnerability at distal nodes. Complement deposition is associated with prominent nodal injury as monitored with electrophysiological recordings and fluorescence microscopy. Complete loss of nodal protein staining, including voltage-gated sodium channels and ankyrin G, occurs and is completely protected by both complement and calpain inhibition, although the latter provides no protection against electrophysiological dysfunction. In ex vivo motor and sensory nerve trunk preparations, antibody deposits are only observed in experimentally desheathed nerves, which are thereby rendered susceptible to complement-dependent morphological disruption, nodal protein loss and reduced electrical activity of the axon. These studies provide a detailed mechanism by which loss of axonal conduction can occur in a distal dominant pattern as observed in a proportion of patients with motor axonal Guillain-Barré syndrome, and also provide an explanation for the occurrence of rapid recovery from complete paralysis and electrophysiological in-excitability. The study also identifies therapeutic approaches in which nodal architecture can be preserved.

Guillain-Barré syndrome during the Zika virus outbreak in Northeast Brazil: An observational cohort study.

OBJECTIVE: To determine the clinical phenotype of Guillain-Barré syndrome (GBS) after Zika virus (ZIKV) infection, the anti-glycolipid antibody signature, and the role of other circulating arthropod-borne viruses, we describe a cohort of GBS patients identified during ZIKV and chikungunya virus (CHIKV) outbreaks in Northeast Brazil. METHODS: We prospectively recruited GBS patients from a regional neurology center in Northeast Brazil between December 2014 and February 2017. Serum and CSF were tested for ZIKV, CHIKV, and dengue virus (DENV), by RT-PCR and antibodies, and serum was tested for GBS-associated antibodies to glycolipids. RESULTS: Seventy-one patients were identified. Forty-eight (68%) had laboratory evidence of a recent arbovirus infection; 25 (52%) ZIKV, 8 (17%) CHIKV, 1 (2%) DENV, and 14 (29%) ZIKV and CHIKV. Most patients with a recent arbovirus infection had motor and sensory symptoms (72%), a demyelinating electrophysiological subtype (67%) and a facial palsy (58%). Patients with a recent infection with ZIKV and CHIKV had a longer hospital admission and more frequent mechanical ventilation compared to the other patients. No specific anti-glycolipid antibody signature was identified in association with arbovirus infection, although significant antibody titres to GM1, GalC, LM1, and GalNAc-GD1a were found infrequently. CONCLUSION: A large proportion of cases had laboratory evidence of a recent infection with ZIKV or CHIKV, and recent infection with both viruses was found in almost one third of patients. Most patients with a recent arbovirus infection had a sensorimotor, demyelinating GBS. We did not find a specific anti-glycolipid antibody signature in association with arbovirus-related GBS.

Guillain-Barré Syndrome Outbreak in Peru 2019 Associated With Campylobacter jejuni Infection.

OBJECTIVE: To identify the clinical phenotypes and infectious triggers in the 2019 Peruvian Guillain-Barré syndrome (GBS) outbreak. METHODS: We prospectively collected clinical and neurophysiologic data of patients with GBS admitted to a tertiary hospital in Lima, Peru, between May and August 2019. Molecular, immunologic, and microbiological methods were used to identify causative infectious agents. Sera from 41 controls were compared with cases for antibodies to Campylobacter jejuni and gangliosides. Genomic analysis was performed on 4 C jejuni isolates. RESULTS: The 49 included patients had a median age of 44 years (interquartile range [IQR] 30-54 years), and 28 (57%) were male. Thirty-two (65%) had symptoms of a preceding infection: 24 (49%) diarrhea and 13 (27%) upper respiratory tract infection. The median time between infectious to neurologic symptoms was 3 days (IQR 2-9 days). Eighty percent had a pure motor form of GBS, 21 (43%) had the axonal electrophysiologic subtype, and 18% the demyelinating subtype. Evidence of recent C jejuni infection was found in 28/43 (65%). No evidence of recent arbovirus infection was found. Twenty-three cases vs 11 controls (OR 3.3, confidence interval [CI] 95% 1.2-9.2, p < 0.01) had IgM and/or IgA antibodies against C jejuni. Anti-GM1:phosphatidylserine and/or anti-GT1a:GM1 heteromeric complex antibodies were strongly positive in cases (92.9% sensitivity and 68.3% specificity). Genomic analysis showed that the C jejuni strains were closely related and had the Asn51 polymorphism at cstII gene. CONCLUSIONS: Our study indicates that the 2019 Peruvian GBS outbreak was associated with C jejuni infection and that the C jejuni strains linked to GBS circulate widely in different parts of the world.

Eculizumab prevents anti-ganglioside antibody-mediated neuropathy in a murine model.

Anti-GQ1b ganglioside antibodies are the serological hallmark of the Miller Fisher syndrome (MFS) variant of the paralytic neuropathy, Guillain-Barré syndrome, and are believed to be the principal pathogenic mediators of the disease. In support of this, we previously showed in an in vitro mouse model of MFS that anti-GQ1b antibodies were able to bind and disrupt presynaptic motor nerve terminals at the neuromuscular junction (NMJ) as one of their target sites, thereby causing muscle paralysis. This injury only occurred through activation of complement, culminating in the formation and deposition of membrane attack complex (MAC, C5b-9) in nerve membranes. Since this step is crucial to the neuropathic process and an important convergence point for antibody and complement mediated membrane injury in general, it forms an attractive pharmacotherapeutic target. Here, we assessed the efficacy of the humanized monoclonal antibody eculizumab, which blocks the formation of human C5a and C5b-9, in preventing the immune-mediated motor neuropathy exemplified in this model. Eculizumab completely prevented electrophysiological and structural lesions at anti-GQ1b antibody pre-incubated NMJs in vitro when using normal human serum (NHS) as a complement source. In a novel in vivo mouse model of MFS generated through intraperitoneal injection of anti-GQ1b antibody and NHS, mice developed respiratory paralysis due to transmission block at diaphragm NMJs, resulting from anti-GQ1b antibody binding and complement activation. Intravenous injection of eculizumab effectively prevented respiratory paralysis and associated functional and morphological hallmarks of terminal motor neuropathy. We show that eculizumab protects against complement-mediated damage in murine MFS, providing the rationale for undertaking clinical trials in this disease and other antibody-mediated neuropathies in which complement activation is believed to be involved.

The role of complement and complement regulators in mediating motor nerve terminal injury in murine models of Guillain-Barré syndrome.

Recent research into the Guillain-Barré syndromes (GBS) has focused on anti-ganglioside antibodies that correlate with specific clinical phenotypes. Our increasing understanding of the role of antibodies in mediating GBS has naturally focused our attention on complement involvement in the pathological procession. We have studied the axonal and glial components of the murine motor nerve terminal as a model site of antibody and complement mediated injury. Such studies are providing us with clear information on the molecular components underlying our clinicopathological model for GBS and have lead us to the testing of emerging complement therapeutics that are potentially suitable for human use.

C5 inhibitor rEV576 protects against neural injury in an in vitro mouse model of Miller Fisher syndrome.

Guillain-Barré syndrome and its clinical variants, including the anti-GQ1b ganglioside-mediated Miller Fisher syndrome (MFS), comprise the world's leading cause of acute neuromuscular paralysis. Presently, no specific drug therapies exist. The complement cascade, which is activated in these patients, forms an attractive drug target. In this study, we tested whether the complement C5-inhibiting recombinant protein, rEV576, was able to prevent neural injury in a previously developed in vitro mouse model for MFS. Mouse hemidiaphragm preparations were treated with anti-GQ1b antibody and normal human serum as a source of complement with added rEV576 or control protein. Immunohistology in control tissue showed deposition of C3c and membrane attack complex at neuromuscular junctions (NMJs), along with terminal motor axonal neurofilament degradation as well as ethidium homodimer-2 staining showing perisynaptic Schwann cell (pSC) injury. Electrophysiological and functional analyses showed block of synaptic transmission at the NMJ after an initial period of a dramatically high level of asynchronous acetylcholine release. In tissue treated with rEV576, all these indicators of motor neuronal damage were absent, except for the presence of C3c, indicating effective inhibition of C5. These results demonstrate that rEV576 effectively prevents development of neuronal and pSC damage in experimental murine neuropathy.

Differential binding patterns of anti-sulfatide antibodies to glial membranes.

Sulfatide is a major glycosphingolipid in myelin and a target for autoantibodies in autoimmune neuropathies. However neuropathy disease models have not been widely established, in part because currently available monoclonal antibodies to sulfatide may not represent the diversity of anti-sulfatide antibody binding patterns found in neuropathy patients. We sought to address this issue by generating and characterising a panel of new anti-sulfatide monoclonal antibodies. These antibodies have sulfatide reactivity distinct from existing antibodies in assays and in binding to peripheral nerve tissues and can be used to provide insights into the pathophysiological roles of anti-sulfatide antibodies in demyelinating neuropathies.

Autoimmune Neurological Conditions Associated With Zika Virus Infection.

Zika virus (ZIKV) is an emerging flavivirus rapidly spreading throughout the tropical Americas. Aedes mosquitoes is the principal way of transmission of the virus to humans. ZIKV can be spread by transplacental, perinatal, and body fluids. ZIKV infection is often asymptomatic and those with symptoms present minor illness after 3 to 12 days of incubation, characterized by a mild and self-limiting disease with low-grade fever, conjunctivitis, widespread pruritic maculopapular rash, arthralgia and myalgia. ZIKV has been linked to a number of central and peripheral nervous system injuries such as Guillain-Barré syndrome (GBS), transverse myelitis (TM), meningoencephalitis, ophthalmological manifestations, and other neurological complications. Nevertheless, mechanisms of host-pathogen neuro-immune interactions remain incompletely elucidated. This review provides a critical discussion about the possible mechanisms underlying the development of autoimmune neurological conditions associated with Zika virus infection.

Overexpression of GD1a ganglioside sensitizes motor nerve terminals to anti-GD1a antibody-mediated injury in a model of acute motor axonal neuropathy.

Anti-GD1a ganglioside antibodies (Abs) are the serological hallmark of the acute motor axonal form of the post-infectious paralysis, Guillain-Barre syndrome. Development of a disease model in mice has been impeded by the weak immunogenicity of gangliosides and the apparent resistance of GD1a-containing neural membranes to anti-GD1a antibody-mediated injury. Here we used mice with altered ganglioside biosynthesis to generate such a model at motor nerve terminals. First, we bypassed immunological tolerance by immunizing GD1a-deficient, beta-1,4-N-acetylgalactosaminyl transferase knock-out mice with GD1a ganglioside-mimicking antigens from Campylobacter jejuni and generated high-titer anti-GD1a antisera and complement fixing monoclonal Abs (mAbs). Next, we exposed ex vivo nerve-muscle preparations from GD1a-overexpressing, GD3 synthase knock-out mice to the anti-GD1a mAbs in the presence of a source of complement and investigated morphological and electrophysiological damage. Dense antibody and complement deposits were observed only over presynaptic motor axons, accompanied by severe ultrastructural damage and electrophysiological blockade of motor nerve terminal function. Perisynaptic Schwann cells and postsynaptic membranes were unaffected. In contrast, normal mice were not only unresponsive to immunization with GD1a but also resistant to neural injury during anti-GD1a Ab exposure, demonstrating the central role of membrane antigen density in modulating both immune tolerance to GD1a and axonal susceptibility to anti-GD1a Abmediated injury. Identical paralyzing effects were observed when testing mouse and human anti-GD1a-positive sera. These data indicate that anti-GD1a Abs arise via molecular mimicry and are likely to be clinically relevant in injuring peripheral nerve axonal membranes containing sufficiently high levels of GD1a.

Microarray screening of Guillain-Barré syndrome sera for antibodies to glycolipid complexes.

OBJECTIVE: To characterize the patterns of autoantibodies to glycolipid complexes in a large cohort of Guillain-Barré syndrome (GBS) and control samples collected in Bangladesh using a newly developed microarray technique. METHODS: Twelve commonly studied glycolipids and lipids, plus their 66 possible heteromeric complexes, totaling 78 antigens, were applied to polyvinylidene fluoride-coated slides using a microarray printer. Arrays were probed with 266 GBS and 579 control sera (2 µL per serum, diluted 1/50) and bound immunoglobulin G detected with secondary antibody. Scanned arrays were subjected to statistical analyses. RESULTS: Measuring antibodies to single targets was 9% less sensitive than to heteromeric complex targets (49.2% vs 58.3%) without significantly affecting specificity (83.9%-85.0%). The optimal screening protocol for GBS sera comprised a panel of 10 glycolipids (4 single glycolipids GM1, GA1, GD1a, GQ1b, and their 6 heteromeric complexes), resulting in an overall assay sensitivity of 64.3% and specificity of 77.1%. Notable heteromeric targets were GM1:GD1a, GM1:GQ1b, and GA1:GD1a, in which exclusive binding to the complex was observed. CONCLUSIONS: Rationalizing the screening protocol to capture the enormous diversity of glycolipid complexes can be achieved by miniaturizing the screening platform to a microarray platform, and applying simple bioinformatics to determine optimal sensitivity and specificity of the targets. Glycolipid complexes are an important category of glycolipid antigens in autoimmune neuropathy cases that require specific analytical and bioinformatics methods for optimal detection.

Anti-disialoside antibodies kill perisynaptic Schwann cells and damage motor nerve terminals via membrane attack complex in a murine model of neuropathy.

Anti-disialoside antibodies (Abs) that bind NeuAc(alpha2-8) NeuAc epitopes on GQ1b and related gangliosides are found in human autoimmune neuropathy sera and are considered to be pathogenic. In a model system in mice, one mechanism by which anti-disialoside Abs have been demonstrated to induce paralysis is through a complement dependent blocking effect on transmitter release at the neuromuscular junction, similar to the effects of alpha-latrotoxin. Although direct targeting of presynaptic neuronal membranes occurs in this model, concomitant injury to perisynaptic Schwann cells (pSC) could indirectly contribute to this paralytic effect by influencing nerve terminal function and survival. To examine this possibility and the specific complement components that might mediate these effects, we exposed neuromuscular junctions in vivo and in vitro to an anti-disialoside Ab in conjunction with intact and selectively deficient complement sources. Using immuno-electron microscopy, we observed Ab deposits equally distributed on both neuronal and pSC membranes, and ultrastructural evidence of injury at both sites. Presynaptic neuronal injury was demonstrated functionally with microelectrode recordings and histologically as neurofilament loss. As hypothesized, concomitant pSC injury occurred, as indicated by abnormal uptake of ethidium dimer into pSC nuclei. The pSC and nerve terminal damage indicators correlated well with deposition of the pore-forming terminal complement component, membrane attack complex (MAC) in pSC and nerve terminal membranes. Furthermore, both neuronal and pSC injury were exacerbated in tissues from mice lacking the inhibitory complement regulator, CD59, where MAC formation is increased. These data demonstrate that both presynaptic neuronal membranes and pSCs are targets for anti-disialoside Abs, and that the injury to both sites is mediated by MAC and further regulated by CD59. This is the first demonstration that complement mediated pSC injury occurs in a model of autoimmune neuropathy and provides a rationale for investigating the possibility of pSC injury in equivalent conditions in man.

Improving the detection of IgM antibodies against glycolipids complexes of GM1 and Galactocerebroside in Multifocal Motor Neuropathy using glycoarray and ELISA assays.

Antibodies against complexes of GM1:GalC are detected in multifocal motor neuropathy. Previous studies used different techniques, explaining disparities in the results. Antibodies against GM1 and GM1:GalC with different proportions of GalC were measured with both glycoarray and ELISA in 20 multifocal motor neuropathies, and 45 controls. The 1:5 ratio and the 1:1 ratio of GM1:GalC (weight ratio) were respectively the most effective for glycoarray and for ELISA. Testing for anti-GM1:GalC antibodies increased the sensitivity from 40% with anti-GM1 antibodies to 65% with array and 60% with ELISA without loss in specificity (above 91%). Anti-GM1:GalC antibodies are effective biological tools to diagnose multifocal motor neuropathy.