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Kidney Week

ASN / Education & Meetings / Kidney Week /
Abstract: TH-PO0273

IgA-IgG-C3-Containing Immune Complexes from Patients with IgAN, but Not the Uncomplexed Proteins, Induce Cytokine/Chemokine Production in Human Mesangial Cells

Session Information

Category: Glomerular Diseases

  • 1401 Glomerular Diseases: Mechanisms, including Podocyte Biology

Authors

  • Hirayama, Masaya, The University of Alabama at Birmingham Department of Microbiology, Birmingham, Alabama, United States
  • Hall, Stacy D., The University of Alabama at Birmingham Department of Microbiology, Birmingham, Alabama, United States
  • Moldoveanu, Zina, The University of Alabama at Birmingham Department of Microbiology, Birmingham, Alabama, United States
  • Botta, Davide, The University of Alabama at Birmingham Department of Microbiology, Birmingham, Alabama, United States
  • Huang, Zhi qiang, The University of Alabama at Birmingham Department of Microbiology, Birmingham, Alabama, United States
  • Qiu, Shihong, The University of Alabama at Birmingham Department of Microbiology, Birmingham, Alabama, United States
  • Shang, Qiao, The University of Alabama at Birmingham Department of Microbiology, Birmingham, Alabama, United States
  • Rizk, Dana V., The University of Alabama at Birmingham Department of Medicine, Birmingham, Alabama, United States
  • Julian, Bruce A., The University of Alabama at Birmingham Department of Medicine, Birmingham, Alabama, United States
  • Green, Todd J., The University of Alabama at Birmingham Department of Microbiology, Birmingham, Alabama, United States
  • Novak, Jan, The University of Alabama at Birmingham Department of Microbiology, Birmingham, Alabama, United States
Background

IgAN patients develop glomerular immune-complex deposits enriched for galactose-deficient IgA1 (Gd-IgA1) and IgG autoantibodies, usually with complement C3. A greater colocalization of C3 with IgA is associated with active disease (mesangial proliferation, endocapillary hypercellularity, and crescents). The glomerular immune-complex deposits likely originate from the circulation. However, these circulating Gd-IgA1-containing immune complexes remain poorly characterized. To address this gap, we determined the size, composition, and biological activity of immune complexes in sera of IgAN patients.

Methods

Immune complexes were isolated from sera of 6 IgAN patients by size-exclusion chromatography as fractions F1-F8 (Mr ~4 MDa to 0.7 MDa). F1-F8 of IgA-depleted sera served as negative controls. Biological activity was tested using cultured primary human mesangial cells (MC). Cytokines were quantified using magnetic bead-based multiplex immunoassays and MC proliferation was determined by click-iT EdU cell-proliferation assay. Composition of immune complexes was analyzed by ELISA, immunoglobulin multiplex assay, and SDS-PAGE immunoblotting with antibodies specific for IgA, IgG, and C3.

Results

Fractions F6-F8 from native IgAN sera increased production of IL-6, IL-8, and MCP-1 by 2- to 10-fold; IgA depletion reduced this activity. IL-6 and MCP-1 production correlated with IgA concentration (p=0.0008, p=3.5e-5, respectively). F6 and F8 from all sera increased cellular proliferation, whereas F7 from 2 patients inhibited proliferation. Fractions F6-F8 from native sera, but not from IgA-depleted sera, contained IgG-IgA and C3-IgA immune complexes. Immunoblotting confirmed covalent association of C3 with IgG and IgA. These fractions contained C3(H2O), C3b, and iC3b. Immune complexes from sera of different IgAN patients exhibited variability in the amounts of IgA, IgG, and molecular forms of C3. In contrast, individual purified proteins, Gd-IgA1, IgG autoantibody, C3, C3a, C3b, or iC3b did not significantly increase cellular proliferation of MC or production of IL-6, IL-8, and MCP-1.

Conclusion

Circulating IgA-IgG-C3-containing immune complexes from sera of IgAN patients, but not their individual components, activated MC and induced overproduction of cytokines/chemokines that accentuate disease activity in IgAN.

Funding

  • NIDDK Support