Rhona McGonigal

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.

Anti‐GM2 ganglioside antibodies are a biomarker for acute canine polyradiculoneuritis

Acute canine polyradiculoneuritis (ACP) is considered to be the canine equivalent of the human peripheral nerve disorder Guillain‐Barré syndrome (GBS); an aetiological relationship, however, remains to be demonstrated. In GBS, anti‐glycolipid antibodies (Abs) are considered as important disease mediators. To address the possibility of common Ab biomarkers, the sera of 25 ACP dogs, 19 non‐neurological, and 15 epileptic control dogs were screened for IgG Abs to 10 glycolipids and their 1 : 1 heteromeric complexes using combinatorial glycoarrays. Anti‐GM2 ganglioside Abs were detected in 14/25 ACP dogs, and anti‐GA1 Abs in one further dog. All controls except for one were negative for anti‐glycolipid Abs. In this cohort of cases and controls, the glycoarray screen reached a diagnostic sensitivity of 60% and a specificity of 97%; a lower sensitivity (32%) was reported using a conventional glycolipid ELISA. To address the possible pathogenic role for anti‐GM2 Abs in ACP, we identified GM2 in canine sciatic nerve by both mass spectrometry and thin layer chromatography overlay. In immunohistological studies, GM2 was localized predominantly to the abaxonal Schwann cell membrane. The presence of anti‐GM2 Abs in ACP suggests that it may share a similar pathophysiology with GBS, for which it could thus be considered a naturally occurring animal model.

Neuronal expression of GalNAc transferase is sufficient to prevent the age-related neurodegenerative phenotype of complex ganglioside-deficient mice

Gangliosides are widely expressed sialylated glycosphingolipids with multifunctional properties in different cell types and organs. In the nervous system, they are highly enriched in both glial and neuronal membranes. Mice lacking complex gangliosides attributable to targeted ablation of the B4galnt1 gene that encodes β-1,4-N-acetylegalactosaminyltransferase 1 (GalNAc–transferase; GalNAcT−/−) develop normally before exhibiting an age-dependent neurodegenerative phenotype characterized by marked behavioral abnormalities, central and peripheral axonal degeneration, reduced myelin volume, and loss of axo-glial junction integrity. The cell biological substrates underlying this neurodegeneration and the relative contribution of either glial or neuronal gangliosides to the process are unknown. To address this, we generated neuron-specific and glial-specific GalNAcT rescue mice crossed on the global GalNAcT−/− background [GalNAcT−/−-Tg(neuronal) and GalNAcT−/−-Tg(glial)] and analyzed their behavioral, morphological, and electrophysiological phenotype. Complex gangliosides, as assessed by thin-layer chromatography, mass spectrometry, GalNAcT enzyme activity, and anti-ganglioside antibody (AgAb) immunohistology, were restored in both neuronal and glial GalNAcT rescue mice. Behaviorally, GalNAcT−/−-Tg(neuronal) retained a normal “wild-type” (WT) phenotype throughout life, whereas GalNAcT−/−-Tg(glial) resembled GalNAcT−/− mice, exhibiting progressive tremor, weakness, and ataxia with aging. Quantitative electron microscopy demonstrated that GalNAcT−/− and GalNAcT−/−-Tg(glial) nerves had significantly increased rates of axon degeneration and reduced myelin volume, whereas GalNAcT−/−-Tg(neuronal) and WT appeared normal. The increased invasion of the paranode with juxtaparanodal Kv1.1, characteristically seen in GalNAcT−/− and attributed to a breakdown of the axo-glial junction, was normalized in GalNAcT−/−-Tg(neuronal) but remained present in GalNAcT−/−-Tg(glial) mice. These results indicate that neuronal rather than glial gangliosides are critical to the age-related maintenance of nervous system integrity.

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

See van Doorn and Jacobs (doi:10.1093/brain/aww078) for a scientific commentary on this article.   How does anti-ganglioside autoantibody clearance from the circulation affect their detection and neurotoxicity? Cunningham et al. describe an uptake pathway for these autoantibodies at motor nerve terminals and their delivery into the brain parenchyma. This highlights the limitations of serum antibody measurements and suggests a possible entry mechanism for their central effects.

C1q-targeted inhibition of the classical complement pathway prevents injury in a novel mouse model of acute motor axonal neuropathy

IntroductionGuillain-Barré syndrome (GBS) is an autoimmune disease that results in acute paralysis through inflammatory attack on peripheral nerves, and currently has limited, non-specific treatment options. The pathogenesis of the acute motor axonal neuropathy (AMAN) variant is mediated by complement-fixing anti-ganglioside antibodies that directly bind and injure the axon at sites of vulnerability such as nodes of Ranvier and nerve terminals. Consequently, the complement cascade is an attractive target to reduce disease severity. Recently, C5 complement component inhibitors that block the formation of the membrane attack complex and subsequent downstream injury have been shown to be efficacious in an in vivo anti-GQ1b antibody-mediated mouse model of the GBS variant Miller Fisher syndrome (MFS). However, since gangliosides are widely expressed in neurons and glial cells, injury in this model was not targeted exclusively to the axon and there are currently no pure mouse models for AMAN. Additionally, C5 inhibition does not prevent the production of early complement fragments such as C3a and C3b that can be deleterious via their known role in immune cell and macrophage recruitment to sites of neuronal damage.Results and ConclusionsIn this study, we first developed a new in vivo transgenic mouse model of AMAN using mice that express complex gangliosides exclusively in neurons, thereby enabling specific targeting of axons with anti-ganglioside antibodies. Secondly, we have evaluated the efficacy of a novel anti-C1q antibody (M1) that blocks initiation of the classical complement cascade, in both the newly developed anti-GM1 antibody-mediated AMAN model and our established MFS model in vivo. Anti-C1q monoclonal antibody treatment attenuated complement cascade activation and deposition, reduced immune cell recruitment and axonal injury, in both mouse models of GBS, along with improvement in respiratory function. These results demonstrate that neutralising C1q function attenuates injury with a consequent neuroprotective effect in acute GBS models and promises to be a useful new target for human therapy.

Peroxisomal dysfunctions cause lysosomal storage and axonal Kv1 channel redistribution in peripheral neuropathy

Impairment of peripheral nerve function is frequent in neurometabolic diseases, but mechanistically not well understood. Here, we report a novel disease mechanism and the finding that glial lipid metabolism is critical for axon function, independent of myelin itself. Surprisingly, nerves of Schwann cell-specific Pex5 mutant mice were unaltered regarding axon numbers, axonal calibers, and myelin sheath thickness by electron microscopy. In search for a molecular mechanism, we revealed enhanced abundance and internodal expression of axonal membrane proteins normally restricted to juxtaparanodal lipid-rafts. Gangliosides were altered and enriched within an expanded lysosomal compartment of paranodal loops. We revealed the same pathological features in a mouse model of human Adrenomyeloneuropathy, preceding disease-onset by one year. Thus, peroxisomal dysfunction causes secondary failure of local lysosomes, thereby impairing the turnover of gangliosides in myelin. This reveals a new aspect of axon-glia interactions, with Schwann cell lipid metabolism regulating the anchorage of juxtaparanodal Kv1-channels. DOI: http://dx.doi.org/10.7554/eLife.23332.001

Reduced axonal diameter of peripheral nerve fibers in a mouse model of Rett syndrome

Rett syndrome (RTT) is a neurological disorder characterized by motor and cognitive impairment, autonomic dysfunction and a loss of purposeful hand skills. In the majority of cases, typical RTT is caused by de novo mutations in the X-linked gene, MECP2. Alterations in the structure and function of neurons within the central nervous system of RTT patients and Mecp2-null mouse models are well established. In contrast, few studies have investigated the effects of MeCP2-deficiency on peripheral nerves. In this study, we conducted detailed morphometric as well as functional analysis of the sciatic nerves of symptomatic adult female Mecp2+/- mice. We observed a significant reduction in the mean diameter of myelinated nerve fibers in Mecp2+/- mice. In myelinated fibers, mitochondrial densities per unit area of axoplasm were significantly altered in Mecp2+/- mice. However, conduction properties of the sciatic nerve of Mecp2 knockout mice were not different from control. These subtle changes in myelinated peripheral nerve fibers in heterozygous Mecp2 knockout mice could potentially explain some RTT phenotypes.

Heparanase attenuates axon degeneration following sciatic nerve transection

Axon degeneration underlies many nervous system diseases; therefore understanding the regulatory signalling pathways is fundamental to identifying potential therapeutics. Previously, we demonstrated heparan sulphates (HS) as a potentially new target for promoting CNS repair. HS modulate cell signalling by both acting as cofactors in the formation of ligand-receptor complexes and in sequestering ligands in the extracellular matrix. The enzyme heparanase (Hpse) negatively regulates these processes by cleaving HS and releasing the attached proteins, thereby attenuating their ligand-receptor interaction. To explore a comparative role for HS in PNS axon injury/repair we data mined published microarrays from distal sciatic nerve injury. We identified Hpse as a previously unexplored candidate, being up-regulated following injury. We confirmed these results and demonstrated inhibition of Hpse led to an acceleration of axonal degeneration, accompanied by an increase in β-catenin. Inhibition of β-catenin and the addition of Heparinase I both attenuated axonal degeneration. Furthermore the inhibition of Hpse positively regulates transcription of genes associated with peripheral neuropathies and Schwann cell de-differentiation. Thus, we propose Hpse participates in the regulation of the Schwann cell injury response and axo-glia support, in part via the regulation of Schwann cell de-differentiation and is a potential therapeutic that warrants further investigation.

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.