TY - JOUR A1 - Rauschenberger, Vera A1 - Piro, Inken A1 - Kasaragod, Vikram Babu A1 - Hörlin, Verena A1 - Eckes, Anna-Lena A1 - Kluck, Christoph J. A1 - Schindelin, Hermann A1 - Meinck, Hans-Michael A1 - Wickel, Jonathan A1 - Geis, Christian A1 - Tüzün, Erdem A1 - Doppler, Kathrin A1 - Sommer, Claudia A1 - Villmann, Carmen T1 - Glycine receptor autoantibody binding to the extracellular domain is independent from receptor glycosylation JF - Frontiers in Molecular Neuroscience N2 - Glycine receptor (GlyR) autoantibodies are associated with stiff-person syndrome and the life-threatening progressive encephalomyelitis with rigidity and myoclonus in children and adults. Patient histories show variability in symptoms and responses to therapeutic treatments. A better understanding of the autoantibody pathology is required to develop improved therapeutic strategies. So far, the underlying molecular pathomechanisms include enhanced receptor internalization and direct receptor blocking altering GlyR function. A common epitope of autoantibodies against the GlyRα1 has been previously defined to residues 1A-33G at the N-terminus of the mature GlyR extracellular domain. However, if other autoantibody binding sites exist or additional GlyR residues are involved in autoantibody binding is yet unknown. The present study investigates the importance of receptor glycosylation for binding of anti-GlyR autoantibodies. The glycine receptor α1 harbors only one glycosylation site at the amino acid residue asparagine 38 localized in close vicinity to the identified common autoantibody epitope. First, non-glycosylated GlyRs were characterized using protein biochemical approaches as well as electrophysiological recordings and molecular modeling. Molecular modeling of non-glycosylated GlyRα1 did not show major structural alterations. Moreover, non-glycosylation of the GlyRα1N38Q did not prevent the receptor from surface expression. At the functional level, the non-glycosylated GlyR demonstrated reduced glycine potency, but patient GlyR autoantibodies still bound to the surface-expressed non-glycosylated receptor protein in living cells. Efficient adsorption of GlyR autoantibodies from patient samples was possible by binding to native glycosylated and non-glycosylated GlyRα1 expressed in living not fixed transfected HEK293 cells. Binding of patient-derived GlyR autoantibodies to the non-glycosylated GlyRα1 offered the possibility to use purified non-glycosylated GlyR extracellular domain constructs coated on ELISA plates and use them as a fast screening readout for the presence of GlyR autoantibodies in patient serum samples. Following successful adsorption of patient autoantibodies by GlyR ECDs, binding to primary motoneurons and transfected cells was absent. Our results indicate that the glycine receptor autoantibody binding is independent of the receptor’s glycosylation state. Purified non-glycosylated receptor domains harbouring the autoantibody epitope thus provide, an additional reliable experimental tool besides binding to native receptors in cell-based assays for detection of autoantibody presence in patient sera. KW - glycine receptor KW - autoantibodies KW - glycosylation KW - extracellular domain KW - adsorption Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-304206 VL - 16 ER - TY - JOUR A1 - Hecker, Katharina A1 - Grüner, Julia A1 - Hartmannsberger, Beate A1 - Appeltshauser, Luise A1 - Villmann, Carmen A1 - Sommer, Claudia A1 - Doppler, Kathrin T1 - Different binding and pathogenic effect of neurofascin and contactin–1 autoantibodies in autoimmune nodopathies JF - Frontiers in Immunology N2 - Introduction IgG4 autoantibodies against paranodal proteins are known to induce acute-onset and often severe sensorimotor autoimmune neuropathies. How autoantibodies reach their antigens at the paranode in spite of the myelin barrier is still unclear. Methods We performed in vitro incubation experiments with patient sera on unfixed and unpermeabilized nerve fibers and in vivo intraneural and intrathecal passive transfer of patient IgG to rats, to explore the access of IgG autoantibodies directed against neurofascin-155 and contactin-1 to the paranodes and their pathogenic effect. Results We found that in vitro incubation resulted in weak paranodal binding of anti-contactin-1 autoantibodies whereas anti-neurofascin-155 autoantibodies bound to the nodes more than to the paranodes. After short-term intraneural injection, no nodal or paranodal binding was detectable when using anti-neurofascin-155 antibodies. After repeated intrathecal injections, nodal more than paranodal binding could be detected in animals treated with anti-neurofascin-155, accompanied by sensorimotor neuropathy. In contrast, no paranodal binding was visible in rats intrathecally injected with anti-contactin-1 antibodies, and animals remained unaffected. Conclusion These data support the notion of different pathogenic mechanisms of anti-neurofascin-155 and anti-contactin-1 autoantibodies and different accessibility of paranodal and nodal structures. KW - autoimmune nodopathy KW - IgG4 KW - neurofascin KW - contactin KW - node of ranvier KW - inflammatory neuropathy KW - passive transfer Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-320395 VL - 14 ER - TY - JOUR A1 - García-Fernández, Patricia A1 - Höfflin, Klemens A1 - Rausch, Antonia A1 - Strommer, Katharina A1 - Neumann, Astrid A1 - Cebulla, Nadine A1 - Reinhold, Ann-Kristin A1 - Rittner, Heike A1 - Üçeyler, Nurcan A1 - Sommer, Claudia T1 - Systemic inflammatory markers in patients with polyneuropathies JF - Frontiers in Immunology N2 - Introduction In patients with peripheral neuropathies (PNP), neuropathic pain is present in 50% of the cases, independent of the etiology. The pathophysiology of pain is poorly understood, and inflammatory processes have been found to be involved in neuro-degeneration, -regeneration and pain. While previous studies have found a local upregulation of inflammatory mediators in patients with PNP, there is a high variability described in the cytokines present systemically in sera and cerebrospinal fluid (CSF). We hypothesized that the development of PNP and neuropathic pain is associated with enhanced systemic inflammation. Methods To test our hypothesis, we performed a comprehensive analysis of the protein, lipid and gene expression of different pro- and anti-inflammatory markers in blood and CSF from patients with PNP and controls. Results While we found differences between PNP and controls in specific cytokines or lipids, such as CCL2 or oleoylcarnitine, PNP patients and controls did not present major differences in systemic inflammatory markers in general. IL-10 and CCL2 levels were related to measures of axonal damage and neuropathic pain. Lastly, we describe a strong interaction between inflammation and neurodegeneration at the nerve roots in a specific subgroup of PNP patients with blood-CSF barrier dysfunction. Conclusion In patients with PNP systemic inflammatory, markers in blood or CSF do not differ from controls in general, but specific cytokines or lipids do. Our findings further highlight the importance of CSF analysis in patients with peripheral neuropathies. KW - cytokines KW - polyneuropathy KW - cerebrospinal fluid KW - neurofilament light chain KW - blood CSF barrier Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-304217 VL - 14 ER - TY - JOUR A1 - García-Fernández, Patricia A1 - Reinhold, Colette A1 - Üçeyler, Nurcan A1 - Sommer, Claudia T1 - Local inflammatory mediators involved in neuropathic pain JF - International Journal of Molecular Sciences N2 - Polyneuropathy (PNP) is a term to describe diseases of the peripheral nervous system, 50% of which present with neuropathic pain. In some types of PNP, pain is restricted to the skin distally in the leg, suggesting a local regulatory process leading to pain. In this study, we proposed a pro-inflammatory pathway mediated by NF-κB that might be involved in the development of pain in patients with painful PNP. To test this hypothesis, we have collected nerve and skin samples from patients with different etiologies and levels of pain. We performed RT-qPCR to analyze the gene expression of the proposed inflammatory pathway components in sural nerve and in distal and proximal skin samples. In sural nerve, we showed a correlation of TLR4 and TNFα to neuropathic pain, and an upregulation of TNFα in patients with severe pain. Patients with an inflammatory PNP also presented a lower expression of TRPV1 and SIRT1. In distal skin, we found a reduced expression of TLR4 and miR-146-5p, in comparison to proximal skin. Our findings thus support our hypothesis of local inflammatory processes involved in pain in PNP, and further show disturbed anti-inflammatory pathways involving TRPV1 and SIRT1 in inflammatory PNP. KW - polyneuropathy KW - pain KW - inflammation KW - NF-κB KW - TNFα Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-313613 SN - 1422-0067 VL - 24 IS - 9 ER - TY - JOUR A1 - Kreß, Luisa A1 - Egenolf, Nadine A1 - Sommer, Claudia A1 - Üçeyler, Nurcan T1 - Cytokine expression profiles in white blood cells of patients with small fiber neuropathy JF - BMC Neuroscience N2 - Background The role of cytokines in the pathophysiology, diagnosis, and prognosis of small fiber neuropathy (SFN) is incompletely understood. We studied expression profiles of selected pro- and anti-inflammatory cytokines in RNA from white blood cells (WBC) of patients with a medical history and a clinical phenotype suggestive for SFN and compared data with healthy controls. Methods We prospectively recruited 52 patients and 21 age- and sex-matched healthy controls. Study participants were characterized in detail and underwent complete neurological examination. Venous blood was drawn for routine and extended laboratory tests, and for WBC isolation. Systemic RNA expression profiles of the pro-inflammatory cytokines interleukin (IL)-1ß, IL-2, IL-8, tumor necrosis factor-alpha (TNF) and the anti-inflammatory cytokines IL-4, IL-10, transforming growth factor beta-1 (TGF) were analyzed. Protein levels of IL-2, IL-8, and TNF were measured in serum of patients and controls. Receiver operating characteristic (ROC)-curve analysis was used to determine the accuracy of IL-2, IL-8, and TNF in differentiating patients and controls. To compare the potential discriminatory efficacy of single versus combined cytokines, equality of different AUCs was tested. Results WBC gene expression of IL-2, IL-8, and TNF was higher in patients compared to healthy controls (IL-2: p = 0.02; IL-8: p = 0.009; TNF: p = 0.03) and discriminated between the groups (area under the curve (AUC) ≥ 0.68 for each cytokine) with highest diagnostic accuracy reached by combining the three cytokines (AUC = 0.81, sensitivity = 70%, specificity = 86%). Subgroup analysis revealed the following differences: IL-8 and TNF gene expression levels were higher in female patients compared to female controls (IL-8: p = 0.01; TNF: p = 0.03). The combination of TNF with IL-2 and TNF with IL-2 and IL-8 discriminated best between the study groups. IL-2 was higher expressed in patients with moderate pain compared to those with severe pain (p = 0.02). Patients with acral pain showed higher IL-10 gene expression compared to patients with generalized pain (p = 0.004). We further found a negative correlation between the relative gene expression of IL-2 and current pain intensity (p = 0.02). Serum protein levels of IL-2, IL-8, and TNF did not differ between patients and controls. Conclusions We identified higher systemic gene expression of IL-2, IL-8, and TNF in SFN patients than in controls, which may be of potential relevance for diagnostics and patient stratification. KW - gene expression KW - small fiber neuropathy KW - cytokines KW - white blood cells Y1 - 2023 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-300619 VL - 24 IS - 1 ER -