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Kidney Week

Abstract: FR-PO167

NLRP3 Inflammasome Inhibition Decreases NETosis in AKI

Session Information

  • AKI: Mechanisms - II
    November 04, 2022 | Location: Exhibit Hall, Orange County Convention Center‚ West Building
    Abstract Time: 10:00 AM - 12:00 PM

Category: Acute Kidney Injury

  • 103 AKI: Mechanisms

Authors

  • Tomaz, David Caetano Duarte Filipe, AstraZeneca PLC, Cambridge, Cambridgeshire, United Kingdom
  • Seth, Asha, AstraZeneca PLC, Cambridge, Cambridgeshire, United Kingdom
  • Parsons, Ricardo, AstraZeneca PLC, Cambridge, Cambridgeshire, United Kingdom
  • Irving, Lorraine M., AstraZeneca PLC, Cambridge, Cambridgeshire, United Kingdom
  • Miller, Lorraine, AstraZeneca PLC, Cambridge, Cambridgeshire, United Kingdom
  • Laerkegaard Hansen, Pernille B., AstraZeneca PLC, Cambridge, Cambridgeshire, United Kingdom
  • Woollard, Kevin, AstraZeneca PLC, Cambridge, Cambridgeshire, United Kingdom

Group or Team Name

  • Biosciences Renal
Background

Kidney macrophages and neutrophils are inflammasome effector cells which can drive renal injury through expression of pro-inflammatory cytokines and formation of neutrophils extracellular traps (NETs). Acute kidney injury (AKI) is associated with NETs formation (NETosis), and a significant number of neutrophils infiltrate during the early injury stage, amplifying the inflammatory response. Different studies have shown an association between inflammasome activation and NETosis. Whether and how inflammasome inhibition affects NETosis-mediated injury and mechanisms of renal dysfunction remains to be fully explored.

Methods

We studied the role of NLRP3 inhibition and its effects on the kidney neutrophil infiltrate and kidney function in-vivo using an acute lipopolysaccharide (LPS) injury mouse model. In this model, mice were dosed with LPS (6mg/kg) or a saline solution intraperitoneally for 18 hours. Blood, urine and kidneys were collected and analysed. In vitro, we studied NETs using live cell imaging with SYTOX Green and histone staining. We further investigated mechanisms of NETs injury on renal proximal tubular endothelial cells (RPTECs) in-vitro using conditioning medium indirect co-cultures.

Results

In-vitro treatment with NLRP3 inhibitor (MCC950) dose-dependently reduced NET formation over 14 hours using LPS/ Nigericin and PMA inflammasome triggers. In vivo, MCC950 decreased plasma IL-1β and IL-18 and urinary IL-18 and protected against renal dysfunction induced by LPS, as evidenced by reduced plasma creatinine and blood urea nitrogen (BUN). Flow cytometry analysis showed a decreasing trend in myeloid infiltration and activation into the kidney. Interestingly, BUN and plasma creatinine correlated with the neutrophil infiltrate. More work is underway to characterise kidney NETs ex-vivo and PTEC injury in-vitro.

Conclusion

Our preliminary data suggests neutrophil infiltrate and NETs formation may accelerate tubular cell injury and death, promoting kidney dysfunction. In conclusion, we have demonstrated that NLRP3 inflammasome inhibition reduces kidney inflammation and inhibits NETosis-mediated kidney injury. The role of neutrophils as key effector cells and NLRP3 inflammasome pathway in kidney diseases warrants further investigation.

Funding

  • Commercial Support