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Human myelin oligodendrocyte glycoprotein is decorated with fucosylated N-glycans that are recognized by DC-SIGN+ DCs and microglia that control immune homeostasis., Myelin oligodendrocyte glycoprotein (MOG), a constituent of central nervous system myelin, is an important autoantigen in the neuroinflammatory disease multiple sclerosis (MS). However, its function remains unknown. Here, we show that, in healthy human myelin, MOG is decorated with fucosylated N-glycans that support recognition by the C-type lectin receptor (CLR) DC-specific intercellular adhesion molecule-3–grabbing nonintegrin (DC-SIGN) on microglia and DCs. The interaction of MOG with DC-SIGN in the context of simultaneous TLR4 activation resulted in enhanced IL-10 secretion and decreased T cell proliferation in a DC-SIGN-, glycosylation-, and Raf1-dependent manner. Exposure of oligodendrocytes to proinflammatory factors resulted in the down-regulation of fucosyltransferase expression, reflected by altered glycosylation at the MS lesion site. Indeed, removal of fucose on myelin reduced DC-SIGN–dependent homeostatic control, and resulted in inflammasome activation, increased T cell proliferation, and differentiation toward a Th17-prone phenotype. These data demonstrate a new role for myelin glycosylation in the control of immune homeostasis in the healthy human brain through the MOG–DC-SIGN homeostatic regulatory axis, which is comprised by inflammatory insults that affect glycosylation. This phenomenon should be considered as a basis to restore immune tolerance in MS.

Pathogenic mycobacteria have the ability to persist in phagocytic cells and to suppress the immune system. The glycolipid lipoarabinomannan (LAM), in particular its mannose cap, has been shown to inhibit phagolysosome fusion and to induce immunosuppressive IL-10 production via interaction with the mannose receptor or DC-SIGN. Hence, the current paradigm is that the mannose cap of LAM is a crucial factor in mycobacterial virulence. However, the above studies were performed with purified LAM, never with live bacteria. Here we evaluate the biological properties of capless mutants of Mycobacterium marinum and M.bovis BCG, made by inactivating homologues of Rv1635c. We show that its gene product is an undecaprenyl phosphomannose-dependent mannosyltransferase. Compared with parent strain, capless M.marinum induced slightly less uptake by and slightly more phagolysosome fusion in infected macrophages but this did not lead to decreased survival of the bacteria in vitro, nor in vivo in zebra fish. Loss of caps in M.bovis BCG resulted in a sometimes decreased binding to human dendritic cells or DC-SIGN-transfected Raji cells, but no differences in IL-10 induction were observed. In mice, capless M.bovis BCG did not survive less well in lung, spleen or liver and induced a similar cytokine profile. Our data contradict the current paradigm and demonstrate that mannose-capped LAM does not dominate the Mycobacterium-host interaction. © 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd.

Background and Objectives Autoantibodies specific for citrullinated antigens are highly relevant diagnostic and prognostic biomarkers in rheumatoid arthritis and have been considered to be involved in disease pathogenesis. Previous studies have indicated that the ACPA-specific immune response differs from conventional B cell responses by generating polyclonal, cross-reactive antibodies of mostly low-avidity. In addition, ACPA were found to carry aberrant glycosylation patterns at the IgG-Fc tail. The present study was undertaken to further characterise the molecular make-up of ACPA and its potential functional consequences in the context of RA. Materials and Methods Serum components of RA patients were fractionated by size exclusion chromatography and analysed for the presence of ACPA-IgG by ELISA. In addition, ACPA-IgG and non-citrulline-specific IgG were affinity purified from RA patient serum and synovial fluid and analysed by gel electrophoresis. Electrophoresis bands were excised and subsequently analysed by HPLC and mass-spectrometry. Recombinant monoclonal ACPA with variations in ACPA molecular structure were used to study binding affinity by surface plasmon resonance. Results We discovered that ACPA-IgG from RA patients have a higher apparent molecular weight as compared to other IgG molecules including antibodies against recall antigens and other autoantibodies. This higher molecular weight was explained by the overrepresentation of N-linked glycans in the variable domain (Fab region) of ACPA-IgG. Detailed structural analysis of these glycans demonstrated that ACPA-IgG linked Fab glycans are complex-type biantennary N-glycans that differ from the conventional Fc-linked N-glycans by a high degree of sialylation, galactosylation, and fucosylation together with the presence of bisecting N-acetylglucosamine. Using recombinant ACPA-IgG monoclonal antibodies with and without Fab-glycans, we found that Fab-glycans modulate binding affinity of ACPA-IgG for citrullinated antigens. Finally, lectin-immunoblotting showed that ACPA Fab-glycans can bind to sialic acid-binding immunoglobulin-type lectins. Conclusions This study describes an unusual and novel molecular feature of the citrulline-specific immune response in RA. ACPA-IgG, in contrast to non-citrulline-specific IgG, are highly glycosylated in the variable region, which modulates recognition of citrullinated antigens. Moreover, ACPA-IgG linked Fab glycans can be the target of specific lectins, suggesting additional functional features potentially involved in ACPA-mediated pathogenetic effects. This finding points to aberrations in the development of ACPA-specific B cells and further elucidates our understanding of basic disease mechanisms in RA.

Background: Prebiotic galacto- and fructo-oligosaccharides (scGOS/lcFOS) resembling non-digestible oligosaccharides in human milk reduce the development of atopic disorders. However, the underlying mechanisms are still unclear. Galectins are soluble-type lectins recognizing β-galactoside containing glycans. Galectin-9 has been shown to regulate mast cell degranulation and T-cell differentiation. In this study, the involvement of galectin-9 as a mechanism by which scGOS/lcFOS in combination with Bifidobacterium breve M-16V protects against acute allergic symptoms was investigated. Methods: Mice were sensitized orally to whey, while being fed with a diet containing scGOS/lcFOS and Bifidobacterium breve M-16V (GF/Bb) or a control diet. Galectin-9 expression was determined by immunohistochemistry in the intestine and measured in the serum by ELISA. T-cell differentiation was investigated in the mesenteric lymph nodes (MLN) as well as in galectin-9-exposed peripheral blood mononuclear cells (PBMC) cultures. Sera of the mice were evaluated for the capacity to suppress mast cell degranulation using a RBL-2H3 degranulation assay. In addition, in a double-blind, placebo-controlled multicenter trial, galectin-9 levels were measured in the sera of 90 infants with atopic dermatitis who received hydrolyzed formulae with or without GF/Bb. Results: Galectin-9 expression by intestinal epithelial cells and serum galectin-9 levels were increased in mice and humans following dietary intervention with GF/Bb and correlated with reduced acute allergic skin reaction and mast cell degranulation. In addition, GF/Bb enhanced Th1- and Treg-cell differentiation in MLN and in PBMC cultures exposed to galectin-9. Conclusions: Dietary supplementation with GF/Bb enhances serum galectin-9 levels, which associates with the prevention of allergic symptoms.