Aberrantly glycosylated IgA1 in IgA nephropathy patients is recognized by IgG antibodies with restricted heterogeneity
Hitoshi Suzuki, … , Jiri Mestecky, Jan Novak
Hitoshi Suzuki, … , Jiri Mestecky, Jan Novak
Published May 26, 2009
Citation Information: J Clin Invest. 2009;119(6):1668-1677. https://doi.org/10.1172/JCI38468.
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Research Article

Aberrantly glycosylated IgA1 in IgA nephropathy patients is recognized by IgG antibodies with restricted heterogeneity

Abstract

IgA nephropathy (IgAN) is characterized by circulating immune complexes composed of galactose-deficient IgA1 and a glycan-specific IgG antibody. These immune complexes deposit in the glomerular mesangium and induce the mesangioproliferative glomerulonephritis characteristic of IgAN. To define the precise specificities and molecular properties of the IgG antibodies, we generated EBV-immortalized IgG-secreting lymphocytes from patients with IgAN and found that the secreted IgG formed complexes with galactose-deficient IgA1 in a glycan-dependent manner. We cloned and sequenced the heavy- and light-chain antigen-binding domains of IgG specific for galactose-deficient IgA1 and identified an A to S substitution in the complementarity-determining region 3 of the variable region of the gene encoding the IgG heavy chain in IgAN patients. Furthermore, site-directed mutagenesis that reverted the residue to alanine reduced the binding of recombinant IgG to galactose-deficient IgA1. Finally, we developed a dot-blot assay for the glycan-specific IgG antibody that differentiated patients with IgAN from healthy and disease controls with 88% specificity and 95% sensitivity and found that elevated levels of this antibody in the sera of patients with IgAN correlated with proteinuria. Collectively, these findings indicate that glycan-specific antibodies are associated with the development of IgAN and may represent a disease-specific marker and potential therapeutic target.

Authors

Hitoshi Suzuki, Run Fan, Zhixin Zhang, Rhubell Brown, Stacy Hall, Bruce A. Julian, W. Winn Chatham, Yusuke Suzuki, Robert J. Wyatt, Zina Moldoveanu, Jeannette Y. Lee, James Robinson, Milan Tomana, Yasuhiko Tomino, Jiri Mestecky, Jan Novak

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Figure 3

Characterization of immune-complex formation.

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Characterization of immune-complex formation. (A) Size-exclusion chromat...
(A) Size-exclusion chromatography and ELISA analysis of immune complexes formed in vitro with monomeric Gal-deficient IgA1 (50 μg) and glycan-specific IgG (50 μg) from 3 patients with IgAN (filled circles) or 3 healthy controls (open circles). IgG and monomeric (m) and dimeric (d) IgA1 standards were used to calibrate the column. Glycan-specific IgG from IgAN patients exhibited more binding to Gal-deficient IgA1 as compared with the binding of IgG from healthy controls. Immune complexes likely contained 1 or 2 molecules of IgA1 bound to 1 molecule of IgG. Data are shown as mean ± SD. (B) Dot-blot analysis showed that IgG secreted by cell lines from 5 of the 6 IgAN patients exhibited high binding to Gal-deficient IgA1; cell line no. 3081 from an IgAN patient and cells from 5 of the 6 healthy controls exhibited low binding. (C) Findings shown in Table 1 were confirmed by densitometrical analysis. P < 0.01; P values were generated using the 2-tailed Student’s t test. Data are shown as individual values and mean ± SD. Experiments were repeated 3 times with similar results.