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Activation and Measurement of NLRP3 Inflammasome Activity Using IL-1β in Human Monocyte-derived Dendritic Cells
09:04

Activation and Measurement of NLRP3 Inflammasome Activity Using IL-1β in Human Monocyte-derived Dendritic Cells

Published on: May 22, 2014

A key role for redox signaling in rapid P2X7 receptor-induced IL-1 beta processing in human monocytes

James Hewinson1, Samantha F Moore, Christian Glover

  • 1Department of Pharmacy and Pharmacology, University of Bath, Bath, UK.

Journal of Immunology (Baltimore, Md. : 1950)
|June 5, 2008
PubMed

Insights

Cellular oxidation via NADPH oxidase is essential for P2X(7) receptor-mediated caspase-1 activation and IL-1beta processing. Blocking superoxide formation or using antioxidants prevents this inflammatory response.

Area of Science:

  • Immunology
  • Cellular Signaling
  • Inflammation Research

Background:

  • P2X(7) receptors (P2X(7)Rs) are ATP-gated ion channels involved in inflammatory responses.
  • Caspase-1 activation, triggered by P2X(7)Rs and TLR ligands, is crucial for processing IL-1beta.
  • The precise signaling pathway linking P2X(7)Rs to caspase-1 activation is not fully understood.

Purpose of the Study:

  • To elucidate the signaling events connecting P2X(7) receptor stimulation to caspase-1 activation.
  • To investigate the role of cellular oxidation in P2X(7)R-mediated IL-1beta processing.

Main Methods:

  • Stimulation of human monocytes with ATP and purinergic receptor agonists.
  • Assessment of NADPH oxidase activity and reactive oxygen species (ROS) formation.
  • Measurement of protein nitrotyrosylation and peroxynitrite levels.
  • Inhibition of ROS, peroxynitrite, or NADPH oxidase activity.

Main Results:

  • ATP-induced P2X(7)R stimulation augments NADPH oxidase activity in monocytes.
  • This leads to protein denitrosylation, peroxynitrite formation, and subsequent caspase-1 activation.
  • IL-1beta cleavage is inhibited by blocking superoxide anion, using antioxidants, or employing a peroxynitrite scavenger.
  • Nigericin also increased NADPH oxidase activity, leading to IL-1beta and caspase-1 processing, which was blocked by peroxynitrite scavenger or NADPH oxidase inhibition.

Conclusions:

  • ATP-induced cellular oxidation, mediated by NADPH oxidase, is critical for P2X(7)R-induced caspase-1 activation.
  • NADPH oxidase signaling is fundamental for processing mature IL-1beta following P2X(7)R stimulation.
  • This pathway highlights a key mechanism in inflammatory responses involving P2X(7)Rs.

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