Phosphatidylinositol 3-phosphate-dependent and -independent functions of p40phox in activation of the neutrophil NADPH oxidase.

In response to bacterial infection, the neutrophil NADPH oxidase assembles on phagolysosomes to catalyze the transfer of electrons from NADPH to oxygen, forming superoxide and downstream reactive oxygen species (ROS). The active oxidase is composed of a membrane-bound cytochrome together with three cytosolic phox proteins, p40(phox), p47(phox), and p67(phox), and the small GTPase Rac2, and is regulated through a process involving protein kinase C, MAPK, and phosphatidylinositol 3-kinase. The role of p40(phox) remains less well defined than those of p47(phox) and p67(phox).

Hem-1 complexes are essential for Rac activation, actin polymerization, and myosin regulation during neutrophil chemotaxis.

Migrating cells need to make different actin assemblies at the cell's leading and trailing edges and to maintain physical separation of signals for these assemblies. This asymmetric control of activities represents one important form of cell polarity. There are significant gaps in our understanding of the components involved in generating and maintaining polarity during chemotaxis.

Cytosolic phospholipase A2-alpha is necessary for platelet-activating factor biosynthesis, efficient neutrophil-mediated bacterial killing, and the innate immune response to pulmonary infection: cPLA2-alpha does not regulate neutrophil NADPH oxidase activity.

The role of a cytosolic phospholipase A(2)-alpha (cPLA(2)-alpha) in neutrophil arachidonic acid release, platelet-activating factor (PAF) biosynthesis, NADPH oxidase activation, and bacterial killing in vitro, and the innate immune response to bacterial infection in vivo was examined. cPLA(2)-alpha activity was blocked with the specific cPLA(2)-alpha inhibitor, Pyrrolidine-1 (human cells), or by cPLA(2) -alpha gene disruption (mice). cPLA(2)-alpha inhibition or gene disruption led to complete suppression of neutrophil arachidonate release and PAF biosynthesis but had no effect on neutrophil NADPH oxidase activation, FcgammaII/III or CD11b surface expression, primary or secondary granule secretion, or phagocytosis of Escherichia coli in vitro.

Distinct ligand-dependent roles for p38 MAPK in priming and activation of the neutrophil NADPH oxidase.

In response to certain cytokines and inflammatory mediators, the activity of the neutrophil NADPH oxidase enzyme is primed for enhanced superoxide production when the cells receive a subsequent oxidase-activating stimulus. The relative role of p38 MAPK in the priming and activation processes is incompletely understood. We have developed a 2-step assay that allows the relative contributions of p38 MAPK activity in priming to be distinguished from those involved in oxidase activation.

Phagocyte priming as a prognostic indicator in the intensive care unit.

Background: The purpose of this study was to study the temporal changes in circulating phagocyte respiratory burst activity and its relationship to mortality in intensive care unit (ICU) patients.

Methods: Thirty-seven consecutive patients over a 3-week period were studied on their first, third, and seventh day of admission to the regional ICU in Northern Ireland. Blood samples were assayed for respiratory burst activity using luminol-enhanced whole blood chemiluminescence.

A novel assay system implicates PtdIns(3,4)P(2), PtdIns(3)P, and PKC delta in intracellular production of reactive oxygen species by the NADPH oxidase.

Activated neutrophils assemble an NADPH oxidase enzyme complex to produce superoxide for microbial killing. Much of the initial oxidase assembly occurs on intracellular granules, followed by movement of the oxidase to phagolysosomes and the plasma membrane. We have developed a novel assay system using Streptolysin-O permeabilized neutrophils that recapitulates the initial intracellular activation process while maintaining the ultrastructural features of this granulocytic cell type.