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Abstract: SA-OR040

BB-Cl-Amidine Limits Neutrophil Extracellular Trap Formation and Inflammation in Murine Experimental Myeloperoxidase Anti-Neutrophil Cytoplasmic Antibody Associated Glomerulonephritis

Session Information

Category: Glomerular Diseases

  • 1202 Glomerular Diseases: Immunology and Inflammation

Authors

  • O'Sullivan, Kim M., Monash University, Monash Medical Centre, Melbourne, Victoria, Australia
  • Gan, Poh-Yi, Monash University, Monash Medical Centre, Melbourne, Victoria, Australia
  • Kitching, A. Richard, Monash University, Monash Health, Clayton, Victoria, Australia
  • Holdsworth, Stephen R., Monash University, Monash Health, Clayton, Victoria, Australia

Group or Team Name

  • Centre for Inflammatory Diseases
Background

Accumulating evidence suggests that dysregulation of neutrophil extracellular traps (NETs) could be associated with the pathogenesis of anti-neutrophil cytoplasmic antibody (ANCA) vasculitis. Peptidyl arginine deiminases (PADs) are enzymes that generate citrullinated proteins, which are pro-inflammatory in both innate and adaptive immune responses. Citrullination of histone 3 (H3Cit) by PAD2 and PAD4 facilitates generation of both macrophage extracellular traps (METs) and NETs. BB-Cl-amidine is a second generation pan (PAD) inhibitor which reduces citrullination of H3Cit and subsequent NET formation. This study investigates the contribution of NETs in the pathogenesis of experimental anti-myeloperoxidase ANCA associated glomerulonephritis (MPO-ANCA GN) and the therapeutic possibility of BB-Cl amidine disrupting NET formation in vivo, and attenuating anti-MPO GN.

Methods

Experimental anti-MPO GN was induced in C57BL/6 mice by myeloperoxidase (MPO) immunisation and GN triggered using a subnephritogenic dose of anti-glomerular basement membrane globulin. BB-Cl-amidine treatment (1mg/kg/daily, n=6 or vehicle, n=7) was administered to disrupt NET formation after the establishment of MPO autoimmunity until termination of the experiment.

Results

NET accumulation identified by co-localisation of extracellular DNA, H3Cit, PAD4, and MPO, was prominent in glomeruli of untreated (vehicle) mice compared to BB-Cl-amidine treated mice (2.1 ± 0.4 vs 0.4 ± 0.1, P=0.004). Histological assessment of kidneys demonstrated prominent glomerular segmental necrosis in the untreated group versus a reduction in mice receiving BB-Cl-amidine (88 ± 3% vs 41 ± 3%, P=0.004). BB-Cl-amidine significantly attenuated glomerular leukocyte infiltration; CD4 T cells (1.2 ± 0.3 vs 0.3 ± 0.1, P=0.03), macrophages (1 ± 0.1 vs 0.4 ± 0.06, P=0.008) and neutrophils (3 ± 0.5 vs 1.6 ±0.3, P=0.03), compared to untreated mice. BB-Cl-amidine treated mice had increased numbers of CD4+FoxP3+ T regulatory cells in lymph nodes compared to the untreated mice (7.1 ± 0.9% vs 3.7 ± 0.3% P= 0.001).

Conclusion

PAD inhibtion with BB-Cl-amidine successfully attenuates in vivo formation of NET formation in experimental anti-MPO GN, reduces kidney injury, glomerular pathology, and supresses inflammation.

Funding

  • Government Support - Non-U.S.