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Abstract: TH-PO455

The Role of Complement in Primary Membranous Nephropathy

Session Information

Category: Glomerular Diseases

  • 1302 Glomerular Diseases: Immunology and Inflammation

Authors

  • Bruno, Valentina, The Hospital for Sick Children, Toronto, Ontario, Canada
  • Ortiz-Sandoval, Carolina G., The Hospital for Sick Children, Toronto, Ontario, Canada
  • Moran, Sarah Margaret, University College Cork, Cork, Ireland
  • Bowen, Emily Elizabeth, The Hospital for Sick Children, Toronto, Canada
  • Cattran, Daniel C., Toronto General Research Institute, Toronto, Ontario, Canada
  • Licht, Christoph, The Hospital for Sick Children, Toronto, Ontario, Canada
Background

Primary membranous nephropathy (PMN) is the leading cause of nephrotic syndrome in adults and a common cause of end-stage kidney disease (ESKD). The Heymann’s nephritis rat model of PMN shows that proteinuria is complement-mediated. However, the pathogenetic role of complement in human PMN remains unclear.
Our preliminary data showed complement deposition on podocytes exposed to serum of PMN patients (from the Toronto GN Registry), leading to disruption of actin cytoskeleton (Figure 1a-b).
We aim to demonstrate that complement activation can have both structural and functional effects on podocytes.

Methods

An in-vitro model of immortalized human podocytes (from Moin Saleem, Bristol, UK) was used for all the experiments. Cells pre-sensitized with anti-CD59 were exposed to 50% normal human serum (NHS) to obtain complement deposition. Changes in intracellular calcium levels were monitored using a fluorescent dye (Fluo8-AM), acquiring images every 20 seconds (up to 10 minutes) by confocal microscopy. Calcium effects on mitochondrial membrane potential were measured by flow cytometry using tetramethylrhodamine, methyl ester (TMRM) dye. Intracellular adenosine triphosphate (ATP) changes were analyzed by bioluminescence. Wound healing assays were performed to study functional effects on cell migration.

Results

Complement activation led to a significant rise in the intracellular calcium levels. Loss of mitochondrial membrane potential was also observed, together with intracellular ATP decrease, disruption of the actin cytoskeleton and impaired cell migration (Figure 1c-e).

Conclusion

Complement is active in PMN, leading to both structural and functional effects on podocytes. Further studies are needed to better understand the consequences on the podocyte energy machinery and the possibility of its reversibility by using complement inhibitors. Our research of such alternate therapy could lead to improvement in outcome in PMN where, despite current therapies, up to one third of patients develops ESKD.