title: Influence of variable domain glycosylation on anti-neutrophil cytoplasmic autoantibodies and anti-glomerular basement membrane autoantibodies.
abstract: BACKGROUND
  The pathophysiological significance of variable region glycosylation of autoantibodies is still unclear. In the current study, the influence of the variable region N-linked oligosaccharides on the reactivity of three autoantibody specificities was investigated with Sambucus nigra agglutinin (SNA), which mainly binds to oligosaccharides with terminal α2, 6-linked sialic acid on the variable region of IgG.
  
  
  METHODS
  Twenty-seven patients with serum positive anti-neutrophil cytoplasmic autoantibodies (ANCA) against myeploperoxidase (MPO) or proteinase 3 (PR3), or autoantibodies against glomerular basement membrane (GBM) were included. Total IgG was isolated and separated into non-SNA-binding and SNA-binding fractions with SNA affinity chromatography. Antigen-specific IgG was purified by immunoaffinity chromatography.
  
  
  RESULTS
  At the same concentration of IgG, the antigen binding level of non-SNA-binding IgG was significantly lower than that of SNA-binding IgG for MPO-ANCA (absorbance value at 405 nm, 0.572 ± 0.590 vs. 0.962 ± 0.670, P < 0.001) and for PR3-ANCA (0.362 ± 0.530 vs. 0.560 ± 0.531, P = 0.003). The antigen binding level of non-SNA-binding IgG was significantly higher than that of SNA-binding IgG for anti-GBM antibodies (1.301 ± 0.594 vs. 1.172 ± 0.583, P = 0.044). The level of variable region glycosylation of total IgG was significantly lower than that of affinity-purified MPO-ANCA (1.021 ± 0.201 vs. 1.434 ± 0.134, P = 0.004). The level of variable region glycosylation of total IgG was significantly higher than that of affinity-purified anti-GBM antibodies (1.034 ± 0.340 vs. 0.734 ± 0.333, P = 0.007). The SNA-binding fraction of MPO-ANCA-containing IgG and PR3-ANCA-containing IgG induced higher levels of neutrophil oxygen radical production than the corresponding non-SNA-binding fractions (P < 0.001 and P = 0.043, respectively). The level of variable region glycosylation of affinity-purified MPO-ANCA was higher in active AAV than the same patients in remission (P = 0.001).
  
  
  CONCLUSIONS
  Characteristics of variable region glycosylation of ANCA and anti-GBM antibodies were different from that of total IgG, which might influence the antigen-binding ability of these antibodies. Variable region glycosylation of ANCA might influence the effect of ANCA-induced neutrophils respiratory burst.
authors.affiliation: Renal Division, Department of Medicine, Peking University First Hospital, Institute of Nephrology, Peking University, Key Laboratory of Renal Disease, Ministry of Health of China, Beijing 100034, China.
authors.forename: Peng-Cheng
authors.initials: PC
authors.lastname: Xu
authors.forename: Shen-Ju
authors.initials: SJ
authors.lastname: Gou
authors.forename: Xiao-Wei
authors.initials: XW
authors.lastname: Yang
authors.forename: Zhao
authors.initials: Z
authors.lastname: Cui
authors.forename: Xiao-Yu
authors.initials: XY
authors.lastname: Jia
authors.forename: Min
authors.initials: M
authors.lastname: Chen
authors.forename: Ming-Hui
authors.initials: MH
authors.lastname: Zhao
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D056648:Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis / Q000276:immunology; D019268:Antibodies, Antineutrophil Cytoplasmic / Q000737:chemistry / Q000276:immunology*; D000915:Antibody Affinity / Q000276:immunology; D000936:Antigen-Antibody Complex / Q000276:immunology; D001323:Autoantibodies / Q000737:chemistry / Q000276:immunology*; D016022:Case-Control Studies; D005260:Female; D006031:Glycosylation; D016179:Granulocyte Colony-Stimulating Factor / Q000276:immunology; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D007377:Interleukin-3 / Q000276:immunology; D008297:Male; D008875:Middle Aged; D009504:Neutrophils / Q000276:immunology; D037121:Plant Lectins / Q000378:metabolism; D011485:Protein Binding / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology; D016897:Respiratory Burst / Q000276:immunology; D054788:Ribosome Inactivating Proteins / Q000378:metabolism; D055815:Young Adult
chemical_list: D019268:Antibodies, Antineutrophil Cytoplasmic; D000936:Antigen-Antibody Complex; D001323:Autoantibodies; D007074:Immunoglobulin G; D007135:Immunoglobulin Variable Region; D007377:Interleukin-3; D037121:Plant Lectins; D011993:Recombinant Fusion Proteins; C050600:Sambucus nigra lectins; C051951:antiglomerular basement membrane antibody; C404145:myelopoietin; D016179:Granulocyte Colony-Stimulating Factor; D054788:Ribosome Inactivating Proteins