Distribution Of GM1 Research Articles

Cell density-dependent membrane distribution of ganglioside GM3 in melanoma cells.

Monosialoganglioside GM3 is the simplest ganglioside involved in various cellular signaling. Cell surface distribution of GM3 is thought to be crucial for the function of GM3, but little is known about the cell surface GM3 distribution. It was shown that anti-GM3 monoclonal antibody binds toGM3 in sparse but not in confluent melanoma cells. Our model membrane study evidenced that monoclonal anti-GM3 antibodies showed stronger binding when GM3 was in less fluid membrane environment. Studies using fluorescent GM3 analogs suggested that GM3 was clustered in less fluid membrane. Moreover, fluorescent lifetime measurement showed that cell surface of high density melanoma cells is more fluid than that of low density cells. Lipidomics and fatty acid supplementation experiment suggested that monounsaturated fatty acid-containing phosphatidylcholine contributed to the cell density-dependent membrane fluidity. Our results indicate that anti-GM3 antibody senses GM3 clustering and the number and/or size of GM3 cluster differ between sparse and confluent melanoma cells.

Prospective Evaluation of Aspergillus fumigatus-Specific IgG in Patients With Cystic Fibrosis.

BackgroundIn Cystic Fibrosis (CF), the airways are often colonized by opportunistic fungi. The most frequently detected mold is Aspergillus fumigatus (Af). Af diseases are associated with significant morbidity and mortality. The most common clinical picture caused by Af is allergic bronchopulmonary aspergillosis (ABPA), triggered by an immunological reaction against Af. Af bronchitis and invasive aspergillosis rarely occur in CF as a result of spore colonization and germination. Since pulmonary mycoses and exacerbations by other pathogens overlap in clinical, radiological, and immunological characteristics, diagnosis still remains a challenge. The search for reliable, widely available biomarkers for Af diseases is therefore still an important task today.ObjectivesAf-specific IgG m3 is broadly available. Sensitivity and specificity data are contradictory and differ depending on the study population. In our prospective study on pulmonary Af diseases in CF, we determined specific IgG m3 in order to test its suitability as a biomarker for acute Af diseases and as a follow-up parameter.MethodsIn this prospective single center study, 109 patients with CF were screened from 2016 to 2019 for Af-associated diseases. According to diagnostic criteria, they were divided into four groups (control, bronchitis, ABPA, pneumonia). The groups were compared with respect to the level of Af-specific IgG (ImmunoCAP Gm3). We performed a receiver operating characteristic (ROC) curve analysis to determine cut-off, sensitivity and specificity. Twenty-one patients could be enrolled for a follow-up examination.ResultsOf the 109 patients, 36 were classified as acute Af-disease (Af bronchitis, ABPA, Af pneumonia). Of these, 21 patients completed follow up-screening. The median Af-specific Gm3 was higher in the acute Af-disease groups. There was a significant difference in Af-specific IgG m3 compared to the control group without acute Af-disease. Overall, there was a large interindividual distribution of Gm3. A cut-off value of 78.05 mg/L for Gm3 was calculated to discriminate controls and patients with ABPA/pneumonia with a specificity of 75% and a sensitivity of 74.6%. The follow up examination of 21 patients showed a decrease of Gm3 in most patients without statistical significance due to the small number of follow up patients.ConclusionAf specific IgG may be a useful biomarker for acute ABPA and Af pneumonia, but not for Af bronchitis in CF. However, due to the large interindividual variability of Gm3, it should only be interpreted alongside other biomarkers. Therefore, due to its broad availability, it could be suitable as a biomarker for ABPA and Af pneumonia in CF, if the results can be supported by a larger multicenter cohort.

Redistribution of gangliosides accompanies thermally induced Na+, K+-ATPase activity alternation and submembrane localisation in mouse brain

• Na + ,K + -ATPase activity alternates with freeze-thaw cycles. • Na + ,K + -ATPase changes membrane microlocation between lipid rafts and bulk membrane. • Na + ,K + -ATPase redistribution is accompanied by reshuffling of specific gangliosides. • Ganglioside GM1 distribution reflects Na + ,K + -ATPase protein distribution. • GD1a, GD1b and GT1b distribution mirrors the changes in Na + ,K + -ATPase activity.

Binding of SV40's Viral Capsid Protein VP1 to Its Glycosphingolipid Receptor GM1 Induces Negative Membrane Curvature: A Molecular Dynamics Study.

The binding of the pentameric capsid protein VP1 of simian virus 40 to its glycosphingolipid receptor GM1 is a key step for the entry of the virus into the host cell. Recent experimental studies have shown that the interaction of variants of soluble VP1 pentamers with giant unilamellar vesicles composed of GM1, DOPC, and cholesterol leads to the formation of tubular membrane invaginations to the inside of the vesicles, mimicking the initial steps of endocytosis. We have used coarse-grained and atomistic molecular dynamics (MD) simulations to study the interaction of VP1 with GM1/DOPC/cholesterol bilayers. In the presence of one VP1 protein, we monitor the formation of small local negative curvature and membrane thinning at the protein binding site as well as reduction of area per lipid. These membrane deformations are also observed under cholesterol-free conditions. However, here, the number of GM1 molecules attached to the VP1 binding pockets increases. The membrane curvature is slightly increased for asymmetric GM1 distribution that mimics conditions in vivo, compared to symmetric GM1 distributions which are often applied in experiments. Slightly smaller inward curvature was observed in atomistic control simulations. Binding of four VP1 proteins leads to an increase of the average intrinsic area per lipid in the protein binding leaflet. Membrane fluctuations appear to be the driving force of VP1 aggregation, as was previously shown for membrane-adhering particles because no VP1 aggregation is observed in the absence of a lipid membrane.

The glycolipid GM1 reshapes asymmetric biomembranes and giant vesicles by curvature generation

The ganglioside GM1 is present in neuronal membranes at elevated concentrations with an asymmetric spatial distribution. It is known to generate curvature and can be expected to strongly influence the neuron morphology. To elucidate these effects, we prepared giant vesicles with GM1 predominantly present in one leaflet of the membrane, mimicking the asymmetric GM1 distribution in neuronal membranes. Based on pulling inward and outward tubes, we developed a technique that allowed the direct measurement of the membrane spontaneous curvature. Using vesicle electroporation and fluorescence intensity analysis, we were able to quantify the GM1 asymmetry across the membrane and to subsequently estimate the local curvature generated by the molecule in the bilayer. Molecular-dynamics simulations confirm the experimentally determined dependence of the membrane spontaneous curvature as a function of GM1 asymmetry. GM1 plays a crucial role in connection with receptor proteins. Our results on curvature generation of GM1 point to an additional important role of this ganglioside, namely in shaping neuronal membranes.

Measuring the Intrinsic Curvature of Ganglioside GM1

The ganglioside GM1, which is present at elevated concentrations in neuron cells and is distributed asymmetrically across their membrane, is known to generate curvature and can be expected to strongly influence the neuron morphology. To elucidate these curvature effects of GM1, one would like to study appropriate model systems but the development of such systems faces several difficulties. First, mimicking the asymmetric distribution of GM1 across the cell membrane in model membranes poses a major challenge. Second, commonly used methods based on fluorescence microscopy for investigating membrane curvature modulation by proteins/lipids cannot be directly applied to GM1 because that requires GM1 molecules are labeled or complexed with suitable fluorophores. But the spontaneous curvatures of these fluorescently labeled analogs are significantly different from that of the label free GM1. Here we report the results of our investigations on measuring the intrinsic curvature of GM1 molecules. We could successfully prepare giant unilamellar vesicles from lipid mixture having 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine and varying fractions of GM1. GM1 was predominantly present at the inner leaflet. A technique was developed to pull inward nanotubes in the vesicle (Dasgupta & Dimova, J. Phys. D Appl. Phys. 47:282001, 2014), where an optically trapped latex bead was first manipulated across the lipid membrane into the vesicle interior and then used as a handle for pulling an in-tube. The force transducer property of the optical trap allowed measuring the tube pulling force with high precision. By studying the tube pulling force as a function of membrane tension imposed by an aspiration micropipette, we could estimate that the mean curvature locally induced by single GM1 molecules is ∼1/(4.78±0.29nm).

Expression of sulfatide and sulfated lactosylceramide among histological types of human ovarian carcinomas.

Among negatively charged lipids, sulfoglycolipids are known to be expressed by specific cell populations and to be involved in their functions, including in adhesion with functional proteins, modification of ion channels and induction of cellular differentiation. Accordingly, we determined their amounts in several histologically defined types of ovarian carcinoma tissues. Sulfoglycolipids were determined by TLC-immunostaining with monoclonal anti-sulfatide antibodies and the gene expression of their synthetic enzymes was by RT-PCR. All types of ovarian carcinomas were revealed to exhibit potential to synthesize sulfoglycolipids, either sulfatide (I(3)SO3-GalCer) or sulfated lactosylceramides (II(3)SO3-LacCer), which were expressed at the following frequencies, 6 out of 6 mucinous cystadenocarcinomas, 4 out of 7 serous cystadenocarcinomas, 2 out of 3 endometrioid carcinomas, and 2 out of 3 clear cell adenocarcinomas. All mucinous cystadenocarcinoma tissues preferentially contained sulfatide in amounts of 0.61-1.13μg per mg dry weight, the molecular species being similar with those of GalCer. Whereas the other carcinomas contained either sulfatide or sulfated LacCer, the latter being detected in 4 out of 6 specimens with sulfoglycolipids. The expression of sulfatide and sulfated LacCer was found to be positively correlated with the amounts of GalCer and LacCer as substrates for sulfotransferase and expression of the genes for GalCer sulfotransferase and ceramide galactosyltransferase. Sulfoglycolipids in ovarian carcinoma tissues were revealed to be expressed in morphologically defined type-characteristic manners, in contrast to the ubiquitous distribution of GM3.

Possible paraneoplastic etiology in a case of Guillain-Barré syndrome with tumoral expression of ganglioside GM3

Paraneoplastic neurological syndromes occur most frequently with small cell lung cancer. We report a 73-year-old man who had suffered from small cell lung cancer, with a fraction of the patient's malignant cells expressing membrane ganglioside GM3, and had developed Guillain–Barre syndrome with serum positivity for anti-GM3 immunoglobulin G antibody when assayed with phosphatidic acid. The distribution of GM3 in myelin labeled with anti-human GM3 antibodies corresponded well to that labeled with the patient's serum, but not with control sera. Intravenous immunoglobulins and chemotherapy for cancer resolved the symptoms of Guillain–Barre syndrome. These observations suggest that autoantibodies against conformational epitopes formed by GM3 and phosphatidic acid in cancers cross-react with neural tissue, probably as a result of molecular mimicry, and might thus underlie paraneoplastic Guillain–Barre syndrome.

Histochemical and immunoelectron microscopic analysis of ganglioside GM3 in human kidney.

Gangliosides are amphipathic lipids ubiquitously expressed in all vertebrate cells. They have been reported to play pivotal roles in cell morphology, cell adhesion, signal transduction, and modulation of immune reaction. Although human kidney contains various kinds of ganglioside, their physiological and pathophysiological roles have not been elucidated yet. As ganglioside GM3 is the most abundant ganglioside in human kidney, we tried to reveal the distribution of GM3 using histological analysis. Macroscopically normal parts of operatively resected kidney from renal cell carcinoma patients were used for analyses. Immunohistochemical and immunoelectron microscopic analyses were performed with anti-GM3 antibody. Immunohistochemical analyses showed that GM3 was observed in glomeruli and renal proximal tubules. Immunoelectron microscopy demonstrated that GM3 was localized on the foot process of podocyte and also in Golgi region of renal proximal tubule cells. Ganglioside GM3 might take a part of the negative electric charge on the surface of podocyte and its multiple physiological actions may play pivotal roles for maintaining glomerular function.

Membrane Microdomains and Cytoskeleton Organization Shape and Regulate the IL-7 Receptor Signalosome in Human CD4 T-cells

Interleukin (IL)-7 is the main homeostatic regulator of CD4 T-lymphocytes (helper) at both central and peripheral levels. Upon activation by IL-7, several signaling pathways, mainly JAK/STAT, PI3K/Akt and MAPK, induce the expression of genes involved in T-cell differentiation, activation, and proliferation. We have analyzed the early events of CD4 T-cell activation by IL-7. We have shown that IL-7 in the first few min induces the formation of cholesterol-enriched membrane microdomains that compartmentalize its activated receptor and initiate its anchoring to the cytoskeleton, supporting the formation of the signaling complex, the signalosome, on the IL-7 receptor cytoplasmic domains. Here we describe by stimulated emission depletion microscopy the key roles played by membrane microdomains and cytoskeleton transient organization in the IL-7-regulated JAK/STAT signaling pathway. We image phospho-STAT5 and cytoskeleton components along IL-7 activation kinetics using appropriate inhibitors. We show that lipid raft inhibitors delay and reduce IL-7-induced JAK1 and JAK3 phosphorylation. Drug-induced disassembly of the cytoskeleton inhibits phospho-STAT5 formation, transport, and translocation into the nucleus that controls the transcription of genes involved in T-cell activation and proliferation. We fit together the results of these quantitative analyses and propose the following mechanism. Activated IL-7 receptors embedded in membrane microdomains induce actin-microfilament meshwork formation, anchoring microtubules that grow radially from rafted receptors to the nuclear membrane. STAT5 phosphorylated by signalosomes are loaded on kinesins and glide along the microtubules across the cytoplasm to reach the nucleus 2 min after IL-7 stimulation. Radial microtubules disappear 15 min later, while transversal microtubules, independent of phospho-STAT5 transport, begin to bud from the microtubule organization center.

Ganglioside GM1 forces the redistribution of cholesterol in a biomimetic membrane

Neutron reflectivity has been applied to investigate different mixed asymmetric lipid systems, in the form of single “supported+floating” bilayers, made of phospholipids, cholesterol and GM1 ganglioside (Neu5Acα2-3(Galβ1-3GalNAcβ1-4)Galβ1-4Glcβ1Cer)) in bio-similar mole ratios. Bilayer preparation was carried out layer-by-layer with the Langmuir–Blodgett Langmuir–Schaefer techniques, allowing for compositional asymmetry in the system buildup. It is the first time that such a complex model membrane system is reported. Two important conclusions are drawn. First, it is experimentally shown that the presence of GM1 enforces an asymmetry in cholesterol distribution, opposite to what happens for a GM1-free membrane that, submitted to a similar procedure, results in a full symmetrization of cholesterol distribution. We underline that natural cholesterol has been used. Second, and most interesting, our results suggest that a preferential asymmetric distribution of GM1 and cholesterol is attained in a model membrane with biomimetic composition, revealing that a true coupling between the two molecular species occurs.

Altered Distribution of the Gangliosides GM1 and GM2 in Alzheimer’s Disease

Background: Alzheimer’s disease (AD) is a neurodegenerative disorder where β-amyloid tends to aggregate and form plaques. Lipid raft-associated ganglioside GM1 has been suggested to facilitate β-amyloid aggregation; furthermore, GM1 and GM2 are increased in lipid rafts isolated from cerebral cortex of AD cases. Aim/Method: The distribution of GM1 and GM2 was studied by immunohistochemistry in the frontal and temporal cortex of AD cases. Frontotemporal dementia (FTD) was included as a contrast group. Results: The distribution of GM1 and GM2 changes during the process of AD (n = 5) and FTD (n = 3) compared to controls (n = 5). Altered location of the GM1-positive small circular structures seems to be associated with myelin degradation. In the grey matter, the staining of GM1-positive plasma membranes might reflect neuronal loss in the AD/FTD tissue. The GM1-positive compact bundles were only visible in cells located in the AD frontal grey matter, possibly reflecting raft formation of GM1 and thus a pathological connection. Furthermore, our results suggest GM2 to be enriched within vesicles of pyramidal neurons of the AD/FTD brain. Conclusion: Our study supports the biochemical finding of ganglioside accumulation in cellular membranes of AD patients and shows a redistribution of these molecules.

Retroviral Vectors Induce Epigenetic Chromatin Modifications and IL-10 Production in Transduced B Cells via Activation of Toll-like Receptor 2

The immune response toward viral vectors used for gene therapy and genetic vaccination appears to be critically important in determining the therapeutic outcome. However, the mechanisms that control the immune response following gene transfer are poorly understood. Unexpectedly, we found that integrating retroviral vector particles induce stable interleukin-10 (IL-10) production in murine (BALB/c H-2(d)) transduced B cells. This requires a novel mechanism whereby the interaction of retroviral vector particle with its cognate cellular receptor activates intracellular signaling pathways resulting in stable epigenetic modifications. Murine B cells exposed to retroviral vector particles triggered the colocalization of the retroviral cellular receptor [mouse cationic amino acid transporter 1 (mCAT1)] and Toll-like receptor 2 (TLR2) into lipid microrafts, which in turn activated TLR2 signaling pathways. TLR2 activation induced STAT3 phosphorylation and increased phosphorylated histone 3 (H3) at the STAT3-binding site of the IL-10 promoter. In addition, TLR2 activation during transduction activates nuclear factor of κ light polypeptide gene enhancer in B-cells inhibitor, α (NFKBIA), thereby preventing the translocation of the nuclear factor-κB (NF-κB) complex to the nucleus and the transcription of proinflammatory cytokines. These findings open new perspectives for controlling immune responses following gene therapy and genetic vaccination.

Assessment of bovine rod outer segment disk membrane heterogeneity utilizing flow cytometry

The outer segment of rod photoreceptor cells is responsible for initiating visual signal transduction when light levels are low. It consists of stacked disk membranes surrounded by the plasma membrane and is under continuous renewal. Disk membranes are synthesized at the base of the outer segment and are progressively displaced toward the apical tip where they are phagocytosed by the pigment epithelium. This process takes approximately ten days during which time the lipid composition of the disk membrane is modified. Disk membranes become progressively lower in cholesterol and higher in unsaturated phospholipids during spatial displacement. The modification of disk composition is likely important for normal function of the rod cell. Flow cytometry is shown in this study to be a powerful tool to assess differences among rod outer segment disk membranes. First, the feasibility of flow cytometry to detect individual osmotically intact disks was established. Fluorescent beads of 1.0 μm, 0.5 μm, 0.2 μm, and 0.1 μm diameter were used to demonstrate that side-scatter intensity measured on a log scale corresponds to bead diameter. The intensity of disk side scatter predominantly corresponded to 0.1–0.2 μm diameter beads. We next applied this technique to detect caveolin-1, peripherin/rds, and GM1 in disk membranes. Caveolin-1 was detected with FITC-labeled anti-caveolin-1, peripherin/rds with Cy3 labeled anti-peripherin/rds, and GM1 with FITC-labeled cholera toxin subunit B. Fluorescence due to caveolin-1 and peripherin/rds binding was detected in 80% and 70% respectively of the events that corresponded to scattering attributed to disks. Fluorescence attributed to GM1 was detected in 60% of the events that corresponded to disk scattering. The intensity of fluorescence due to caveolin-1 and peripherin/rds labeling was directly proportional to the intensity of side scatter, indicating that both caveolin-1 and peripherin/rds concentrations are uniform among all the disks. Fluorescence intensity of GM1 labeled disks was independent of side-scatter intensity indicating GM1 is heterogeneously distributed among the disks. These results are consistent with integral membrane proteins such as caveolin-1, and peripherin/rds remaining in the disk bilayer as the disks are apically displaced. The GM1 distribution may be similar to the spatial distribution of cholesterol in the outer segment.

Observations of the distribution of GM3 in membrane microdomains by atomic force microscopy

Glycosphingolipids (GSLs) in membrane microdomains participate in important biological functions. In the present paper, we propose a novel model of the distribution of GSLs in membrane microdomains composed of sphingomyelin (SM) and cholesterol. We investigated the distribution of the ganglioside GM3 in a lipid membrane reconstituted with lipid extract from mouse B16 melanoma cells using an atomic force microscope (AFM). The surface topography of the reconstituted lipid bilayer showed three areas of different heights. The highest area was confirmed to be a GM3 domain by labeling with wheat germ agglutinin. To identify the lipids which are contributed to make the topography, the topographies of the artificial lipid bilayers composed of GM3, SM, 1-palmitoyl-2-oleoyl-phosphatidylcholine, and cholesterol were investigated. AFM images of the artificial lipid bilayers showed that the GM3 domain surrounded by a SM-containing phase only occurred, and its formation was found to depend on the cholesterol content.

Fluorescence-topographic NSOM directly visualizes peak-valley polarities of GM1/GM3 rafts in cell membrane fluctuations

Simultaneous fluorescence-topographic nanoscale imaging of cell-surface molecules in the context of membrane ultra-structures has not been reported. Here, near-field scanning optical microscopy (NSOM)-based direct fluorescence-topographic imaging indicated that GM3 rafts/nanodomains (190.0 +/- 49.8 nm ranging 84.5-365.0 nm) were localized predominantly on the peaks of microvillus-like protrusions in the apical membrane of GM3 + Madin-Darby canine kidney cells, whereas GM1 rafts/nanodomains (159.5 +/- 63.8 nm ranging 42-360 nm) were distributed mainly on the slops of protrusions or the valleys between protrusions in the plasma membranes of GM1 + MDCK cells. The data demonstrated that gangliosides polarized not only in a well-known apical-basolateral manner but also in the more microscopic peak-valley manner, implicating unique distribution of GM1 or GM3 in cell-surface fluctuations on the apical membrane of polarized cells. The peak-valley polarities of gangliosides also implicated their different functions relevant to lipid rafts, microvilli, or cellular processes. Importantly, our study demonstrated for the first time that the NSOM-based direct fluorescence-topographic imaging is unique and powerful for elucidating nanoscale distribution of specific cell-surface molecules in membrane fluctuations.

GM1 DYNAMICS INDICATE MEMBRANE CHANGES ASSOCIATED WITH CAPACITATION IN MURINE SPERMATOZOA

We previously showed that in live murine and bovine sperm heads, the ganglioside GM1 localizes to the sterol-rich plasma membrane overlying the acrosome (APM). Labeling GM1 using the pentameric cholera toxin subunit B (CTB) induced a dramatic redistribution of signal from the APM to the sterol-poor post-acrosomal plasma membrane (PAPM) upon sperm death. We now show a similar phenomenon in the flagellum where CTB induced GM1 redistribution to sterol-poor membrane sub-domains of the annulus and flagellar zipper. Because sterol efflux from the plasma membrane is required for capacitation, we next examined whether GM1 localization might be useful to detect membrane changes associated with capacitation and/or acrosomal exocytosis. First, incubation of murine sperm with their respective stimuli for capacitation did not change GM1 distribution in live cells. However, incubation with specific stimuli for capacitation, followed by the use of specific fixation conditions, induced reproducible, stimulus-specific patterns of GM1 distribution. By assessing changes in GM1 distribution in response to progesterone-induced acrosomal exocytosis, we show that these patterns reflect the response of sperm populations to capacitating stimuli. These data suggest that GM1 localization can be used as a diagnostic tool for evaluating sperm response to stimuli for capacitation and/or acrosomal exocytosis. Such information could be useful when deciding between technologies of assisted reproduction, or when screening for male fertility. Furthermore, stimulus-specific changes in GM1 distribution showed that sperm could respond to NaHCO3 or mediators of sterol efflux independently, thereby refining existing models of capacitation. (poster)

Agrin elicits membrane lipid condensation at sites of acetylcholine receptor clusters in C2C12 myotubes

The formation of the neuromuscular junction is characterized by the progressive accumulation of nicotinic acetylcholine receptors (AChRs) in the postsynaptic membrane facing the nerve terminal, induced predominantly through the agrin/muscle-specific kinase (MuSK) signaling cascade. However, the cellular mechanisms linking MuSK activation to AChR clustering are still poorly understood. Here, we investigate whether lipid rafts are involved in agrin-elicited AChR clustering in a mouse C2C12 cell line. We observed that in C2C12 myotubes, both AChR clustering and cluster stability were dependent on cholesterol, because depletion by methyl-beta-cyclodextrin inhibited cluster formation or dispersed established clusters. Importantly, AChR clusters resided in ordered membrane domains, a biophysical property of rafts, as probed by Laurdan two-photon fluorescence microscopy. We isolated detergent-resistant membranes (DRMs) by three different biochemical procedures, all of which generate membranes with similar cholesterol/GM1 ganglioside contents, and these were enriched in several postsynaptic components, notably AChR, syntrophin, and raft markers flotillin-2 and caveolin-3. Agrin did not recruit AChRs into DRMs, suggesting that they are present in rafts independently of agrin activation. Consequently, in C2C12 myotubes, agrin likely triggers AChR clustering or maintains clusters through the coalescence of lipid rafts. These data led us to propose a model in which lipid rafts play a pivotal role in the assembly of the postsynaptic membrane at the neuromuscular junction upon agrin signaling.

Monosialoganglioside (GM1) immunofluorescence in rat spinal roots studied with a monoclonal antibody.

Gangliosides are characteristic glycolipid components of plasma cell membranes, especially enriched in the CNS and PNS. In some diseases involving the PNS, in particular motor neuropathies associated with conduction block, IgM autoantibodies against ganglioside GM1 have been implicated as a pathogenic factor. In order to study the GM1 distribution in peripheral nerves we have investigated its in situ localization using a new anti-GM1 monoclonal antibody, GM1:1. Immunization and production of the monoclonal antibody was made by common protocols and binding specificity was investigated by using structurally related glycolipids and modified GM1-molecules. The result showed that an alpha 2-3 bound sialic acid together with a terminal galactose moiety were essential for GM1:1 binding. In situ localization of GM1 in rat dorsal and ventral spinal roots was investigated by conventional immunomicroscopy. GM1 immunoreactivity was the same in both roots and appeared like a finely granular, in places confluent, material confined to Schmidt-Lanterman's incisures, to myelin sheath paranodal end segments and to some extent to the abaxonal Schwann cell cytoplasm; all of these structures are likely to be the target for GM1 antibodies in peripheral neuropathies. Nodal gaps and fibre contours showed a weak non-specific fluorescence. The localization of GM1 to the incisures of Schmidt-Lanterman and the paranodal end segments of the myelin sheaths might indicate a role of gangliosides as adhesion molecules.

Ganglioside GM1 and GM3 in early human brain development: An immunocytochemical study

The distribution of GM1 and GM3 gangliosides in human brain development between gestational week (g.w.) 6 and 15 was demonstrated by an immunocytochemical approach using polyclonal anti-GM1 and anti-GM3 antibodies. The first appearance of GM1- and GM3-positive cells was recorded as early as in g.w. 6. Both antibodies labeled the cells in the ventricular zone of the telencephalic wall, with radially oriented fibers toward the pial surface, which represent radial glia cells with glia fibers. The intensive GM3 immunoreactivity was also exhibited in proliferating cells in the ventricular zone between g.w. 6 and 12. During the period from g.w. 12 to 15, characterized by a rapid multiplication of neurons and glia cells, an increased number of GM1- and GM3-positive cells was observed. Prominent GM1 ganglioside staining was observed at the surface of the cell bodies in the ventricular zone. Besides surface labeling in migrating cells, GM1 immunoreactivity was identified inside the soma in the regions of cortical plate and subplate. GM1 immunoreactivity was more pronounced on the membrane of neuronal cells migrating along radial glia fibers, especially at the contact site between neuronal and glial cells. The GM3 ganglioside was localized mostly inside the soma, showing a granular immunoreactivity pattern.Our observations confirm the presence of GM1 and GM3 gangliosides in neuronal and glial cells in early human brain development. The involvement, especially of GM1 ganglioside in glia-neuronal contacts during migration of neuroblasts to their final destination, as well as the presence of GM3 ganglioside in proliferative cells in the ventricular zone of the telencephalic wall was also recorded.