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Abstract: FR-PO575

Endothelial Cell Ferroptosis Promotes Renal Damage in ANCA-Induced Glomerulonephritis

Session Information

Category: Glomerular Diseases

  • 1302 Glomerular Diseases: Immunology and Inflammation

Authors

  • Rousselle, Anthony, Experimental and Clinical Research Center, Max Delbrück Center for Molecular Medicine (MDC) and Charité, Berlin, Germany
  • Lodka, Dörte, Experimental and Clinical Research Center, Max Delbrück Center for Molecular Medicine (MDC) and Charité, Berlin, Germany
  • Kling, Lovis, Charite Universitatsmedizin Berlin, Berlin, Germany
  • Kettritz, Ralph, Charite Universitatsmedizin Berlin, Berlin, Germany
  • Schreiber, Adrian, Charite Universitatsmedizin Berlin, Berlin, Germany
Background

Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitides (AAV) are systemic autoimmune diseases characterized by inflammation of small blood vessels and organ damage, including necrotizing and crescentic glomerulonephritis (NCGN). ANCA are circulating immunoglobulin G (IgG) autoantibodies binding to and activating neutrophil granulocytes and monocytes. The persistent inflammation leads to renal cell necrosis. Ferroptosis is a form of programmed cell death characterized by the production of ROS- and iron-dependent lipid peroxidation leading to membrane rupture. The Acyl-CoA Synthetase Long Chain Family Member 4 (ACSL4) enzyme regulates the generation of lipid peroxides. We tested the hypothesis that endothelial cell ferroptosis contributes to ANCA-associated NCGN.

Methods

To analyse the biological significance of ferroptosis in endothelial cells in vivo, we generated MPO-/- ACSL4EC mice, which lack ACSL4 specifically in endothelial cells. MPO-/- ACSL4EC mice were immunized with murine MPO, irradiated and subsequently transplanted with hematopoietic cells from C/57BL6J (WT) mice. Mice were sacrificed and analyzed 6-8 weeks following transplantation. Ferroptosis was investigated in vitro using human umbilical vein endothelial cells and ANCA-stimulated neutrophils. Cell death and lipid peroxidation were detected by flow cytometry. Ferrostatin-1 (Fer-1) and siRNA against ACSL4 were used to inhibit ferroptosis.

Results

We found increased lipid peroxidation (4-HNE staining) in kidney section of mice with AAV. MPO-/- ACSL4EC chimeric mice showed reduced renal damage as indicated by less necrotic and crescentic glomeruli supporting the notion that endothelial cell ferroptosis is important for the development of MPO-ANCA-induced NCGN. In vitro experiments revealed that ANCA-activated neutrophils induced endothelial cell death and this effect was prevented by ferroptosis inhibition with Fer-1 and siRNA against ACSL4. In contrast, inhibition of either necroptosis or apoptosis did not prevent endothelial cell death. Finally, ferroptosis inhibition alleviated the accumulation of lipid peroxides and endothelial dysfunction induced by ANCA-activated neutrophils.

Conclusion

ANCA-activated neutrophils induce ferroptosis in endothelial cells in vitro and endothelial cell ferroptosis contributes to ANCA-associated NCGN in a murine AAV model.

Funding

  • Government Support – Non-U.S.