Neutrophils exposed to bacterial lipopolysaccharide upregulate NADPH oxidase assembly

J Clin Invest. 1998 Jan 15;101(2):455-63. doi: 10.1172/JCI949.

Abstract

Bacterial LPS is a pluripotent agonist for PMNs. Although it does not activate the NADPH-dependent oxidase directly, LPS renders PMNs more responsive to other stimuli, a phenomenon known as "priming." Since the mechanism of LPS-dependent priming is incompletely understood, we investigated its effects on assembly and activation of the NADPH oxidase. LPS pretreatment increased superoxide (O2-) generation nearly 10-fold in response to N-formyl methionyl leucyl phenylalanine (fMLP). In a broken-cell O2--generating system, activity was increased in plasma membrane-rich fractions and concomitantly decreased in specific granule-rich fractions from LPS-treated cells. Oxidation-reduction spectroscopy and flow cytometry indicated LPS increased plasma membrane association of flavocytochrome b558. Immunoblots of plasma membrane vesicles from LPS-treated PMNs demonstrated translocation of p47-phox but not of p67-phox or Rac2. However, PMNs treated sequentially with LPS and fMLP showed a three- to sixfold increase (compared with either agent alone) in plasma membrane-associated p47-phox, p67-phox, and Rac2, and translocation paralleled augmented O2- generation by intact PMNs. LPS treatment caused limited phosphorylation of p47-phox, and plasma membrane-enriched fractions from LPS- and/or fMLP-treated cells contained fewer acidic species of p47-phox than did those from cells treated with PMA. Taken together, these studies suggest that redistribution of NADPH oxidase components may underlie LPS priming of the respiratory burst.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Cell Membrane / enzymology
  • Cytochrome b Group / metabolism
  • Cytosol / enzymology
  • Humans
  • Lipopolysaccharides / pharmacology*
  • N-Formylmethionine Leucyl-Phenylalanine / pharmacology
  • NADPH Oxidases / chemistry
  • NADPH Oxidases / drug effects*
  • Neutrophils / drug effects*
  • Neutrophils / metabolism
  • Phosphoproteins / metabolism
  • Phosphorylation
  • Respiratory Burst / drug effects
  • Superoxides / metabolism

Substances

  • Cytochrome b Group
  • Lipopolysaccharides
  • Phosphoproteins
  • neutrophil cytosol factor 67K
  • Superoxides
  • N-Formylmethionine Leucyl-Phenylalanine
  • cytochrome b558
  • NADPH Oxidases
  • neutrophil cytosolic factor 1