Abstract: TH-PO375

ANCA Stimulation of Monocytes Changes Their Cellular Metabolism

Session Information

Category: Cell Biology

  • 201 Cell Signaling, Oxidative Stress

Authors

  • O'Brien, Eóin, Trinity College Dublin, Dublin, Ireland
  • White, Carla A, Trinity College Dublin, Dublin, Ireland
  • Mohamed, Safa Hasab Elrasoul, Trinity College Dublin, Dublin, Ireland
  • Little, Mark Alan, Trinity College Dublin, Dublin, Ireland
  • Hickey, Fionnuala B., Trinity College Dublin, Dublin, Ireland
Background

Anti-neutrophil cytoplasmic antibody (ANCA) vasculitis causes rapidly progressive glomerulonephritis and is characterised by autoantibodies against myeloperoxidase (MPO) or proteinase-3 (PR3). We have previously shown that stimulation of monocytes with anti-MPO, but not anti-PR3, antibodies results in increased pro-inflammatory cytokine release, suggesting a pathogenic role for this cell type. Cellular metabolism (particularly a switch to aerobic glycolysis), has recently been shown to be important in the immune response, and targeting metabolic pathways postulated as a potential treatment in autoimmunity. We investigated the effect of ANCA on the metabolism of monocytes.

Methods

Monocytes were isolated from healthy donors. Following stimulation with anti-MPO/PR3 antibodies, with or without a range of metabolic pathway inhibitors, changes in metabolism were measured using Seahorse extracellular flux analysis. Changes in cytokine production were assessed by ELISA.

Results

In accordance with prior data indicating that pro-inflammatory leukocytes preferentially use glycolysis for metabolism, we found increased glycolysis in ANCA-stimulated monocytes. However, we also observed upregulated oxidative respiration. These changes occur within minutes of exposure to ANCA. Cells treated with anti-PR3 antibodies displayed different oxygen consumption kinetics compared to those treated with anti-MPO; changes in these metabolic pathways were linked to the previously observed differences in cytokine production. To further investigate the mechanism by which metabolic pathways are involved in the response to ANCA, we used pharmacological inhibitors to block elements of glucose metabolism. Blocking oxidative phosphorylation, as measured by reduced oxygen consumption, resulted in no change in IL-1β production, but blocking glycolysis resulted in complete inhibition. Pyruvate dehydrogenase, a major point-of-no-return in the metabolism of glucose, was required for IL-1β production. In addition, using a specific scavenger of mitochondrial reactive oxygen species (mROS) we showed an inhibition of the IL-1β induced by anti-MPO treatment indicating that mROS has a role in activation of this inflammatory pathway.

Conclusion

These data indicate an important role for the upregulation of cellular metabolism in the pro-inflammatory activation of monocytes in response to ANCA.

Funding

  • Government Support - Non-U.S.