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Abstract: PO1968

The Glomerulus-on-a-Chip as a System to Unravel Novel Membrane Attack Complex (MAC)-Independent Role of Complement in Membranous Nephropathy

Session Information

  • Podocyte Biology
    October 22, 2020 | Location: On-Demand
    Abstract Time: 10:00 AM - 12:00 PM

Category: Glomerular Diseases

  • 1204 Podocyte Biology

Authors

  • Da Sacco, Stefano, Children's Hospital of Los Angeles, Los Angeles, California, United States
  • Petrosyan, Astgik, Children's Hospital of Los Angeles, Los Angeles, California, United States
  • Hartzell, Susan, Icahn School of Medicine at Mount Sinai, New York, New York, United States
  • Villani, Valentina, Children's Hospital of Los Angeles, Los Angeles, California, United States
  • Dedhia, Charmi, Children's Hospital of Los Angeles, Los Angeles, California, United States
  • Angeletti, Andrea, Universita di Bologna, Bologna, Emilia-Romagna, Italy
  • Manrique, Joaquin, Complejo Hospitalario de Navarra, Pamplona, Navarra, Spain
  • De Filippo, Roger E., Children's Hospital of Los Angeles, Los Angeles, California, United States
  • Perin, Laura, Children's Hospital of Los Angeles, Los Angeles, California, United States
  • Cravedi, Paolo, Icahn School of Medicine at Mount Sinai, New York, New York, United States
Background

Primary membranous nephropathy (MN) is a leading cause of nephrotic syndrome in adults worldwide due to the deposition of anti-podocyte-antibodies against in the glomerular subepithelial space. While complement deposition is thought to play a crucial pathogenic role, the exact effector mechanism of complement in MN is unclear due to the lack of in vitro and in vivo systems that recapitulate human disease. We have developed a novel glomerulus-on-a-chip system (GOAC) using human primary podocytes and human glomerular endothelial cells (GEC) in combination with OrganoPlates and assessed the functional response to human MN serum and the role of MAC deposition and C3a/C3aR1 signaling in MN pathogenesis.

Methods

Glomerular chips were cultured with serum from anti-PLA2R+ MN patients or healthy individuals. Functional response was assessed by albumin permeability assay to evaluate selective-permeability. Role of MAC and C3a/C3aR1 signaling pathway in glomerular filtration barrier damage was assessed by immunofluorescence and functional analysis while mechanisms of action were explored by PCR arrays, Western Blotting and immunostaining. Results were confirmed in vitro using podocytes on which C3aR1 was silenced and in vivo using a C3aR1 KO mice model.

Results

Following exposure to sera from MN patients, we have confirmed deposition of human IgG on podocytes and formation of MAC complex, accompanied by albumin leakage. GOAC supplemented with C3aR1 antagonists as well as GOAC using podocytes in which C3aR1 was silenced were able to prevent glomerular filtration damage on the GOAC as confirmed by rescue of permselectivity efficiency, while inhibition of MAC formation by protein S (an inhibitor of MAC formation) did not significantly reduce GOAC permeability.

Conclusion

We have successfully developed a glomerulus-on-a-chip system that closely mimics the GFB structure and provides a powerful tool for studying renal regenerative and disease mechanisms in proteinuric diseases. Using this model, w showed that C3a/C3aR signaling plays a dominant role in complement-mediated MN pathogenesis.

Funding

  • Private Foundation Support