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LPS induced inflammatory responses in human peripheral blood mononuclear cells is mediated through NOX4 and Giα

Anta Ngkelo1, Koremu Meja, Mike Yeadon

  • 1Airways Disease Section, National Heart and Lung Institute, Imperial College London, London, UK. p.kirkham@imperial.ac.uk.

Journal of Inflammation (London, England)
|January 14, 2012
PubMed

Insights

Lipopolysaccharide (LPS) triggers inflammation in Chronic Obstructive Pulmonary Disease (COPD) via toll-like receptor 4 (TLR4). This study reveals a pathway involving NADPH oxidase 4 (NOX4), reactive oxygen species (ROS), and phosphatidylinositol 3-kinase (PI3K) activation, mediated by small G proteins.

Area of Science:

  • Immunology
  • Cell Biology
  • Biochemistry

Background:

  • Chronic Obstructive Pulmonary Disease (COPD) involves innate immunity, with bacterial infections exacerbating the condition and increasing mortality.
  • Pathogen-associated molecular pattern (PAMP) lipopolysaccharide (LPS) activates toll-like receptor 4 (TLR4) on immune cells, driving inflammation through NADPH oxidase (NOX) and NF-κB.
  • Understanding the molecular mechanisms of LPS-induced inflammation is crucial for developing targeted COPD therapies.

Purpose of the Study:

  • To elucidate the signaling pathway downstream of LPS-induced TLR4 activation in human peripheral blood mononuclear cells (PBMCs).
  • To investigate the roles of NADPH oxidase (NOX), reactive oxygen species (ROS), phosphatidylinositol 3-kinase (PI3K), and small G proteins in this inflammatory response.

Main Methods:

  • Human PBMCs were stimulated with LPS.
  • The effects of inhibitors (wortmannin, apocynin, N-acetylcysteine, pertussis toxin, mastoparan) on cytokine release (CXCL8, IL-6) and signaling pathways (AKT phosphorylation) were assessed.
  • Reactive oxygen species (ROS) generation was inferred through NOX4 and antioxidant inhibition.

Main Results:

  • LPS-induced release of pro-inflammatory cytokines CXCL8 and IL-6 was significantly inhibited by the PI3K inhibitor wortmannin.
  • LPS-induced PI3K activation was dependent on NOX4 and endogenous hydrogen peroxide (H2O2) production, as shown by inhibition with apocynin.
  • The antioxidant N-acetylcysteine inhibited LPS-induced PI3K activation in a concentration-dependent manner.
  • Inhibition of small G proteins by pertussis toxin blocked LPS-induced AKT phosphorylation and cytokine release.
  • Pertussis toxin and mastoparan inhibited LPS-induced CXCL8 and IL-6 release by approximately 50%.

Conclusions:

  • LPS-induced TLR4 activation in human PBMCs initiates a signaling cascade involving NOX4-dependent ROS generation and PI3K pathway activation.
  • Small G proteins mediate this pathway, facilitating the release of pro-inflammatory cytokines crucial in COPD exacerbations.
  • These findings highlight a novel mechanism in COPD pathogenesis and suggest potential therapeutic targets.

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