Synergy between Extracellular Group IIA Phospholipase A2 and Phagocyte NADPH Oxidase in Digestion of Phospholipids of Staphylococcus aureus Ingested by Human Neutrophils

Abstract

Acute inflammatory responses to invading bacteria such as Staphylococcus aureus include mobilization of polymorphonuclear leukocytes (PMN) and extracellular group IIA phospholipase A2 (gIIA-PLA2). Although accumulating coincidentally, the in vitro anti-staphylococcal activities of PMN and gIIA-PLA2 have thus far been studied separately. We now show that degradation of S. aureus phospholipids during and after phagocytosis by human PMN requires the presence of extracellular gIIA-PLA2. The concentration of extracellular gIIA-PLA2 required to produce bacterial digestion was reduced 10-fold by PMN. The effects of added gIIA-PLA2 were greater when present before phagocytosis but even apparent when added after S. aureus were ingested by PMN. Related group V and X PLA2, which are present within PMN granules, do not contribute to bacterial phospholipid degradation during and after phagocytosis even when added at concentrations 30-fold higher than that needed for action of the gIIA-PLA2. The action of added gIIA-PLA2 required catalytically active gIIA-PLA2 and, in PMN, a functional NADPH oxidase but not myeloperoxidase. These findings reveal a novel collaboration between cellular oxygen-dependent and extracellular oxygen-independent host defense systems that may be important in the ultimate resolution of S. aureus infections.

The bactericidal/permeability-increasing protein (BPI) in infection and inflammatory disease

“…These combined actions can take place both within neutrophils after uptake of the PLA2 and complement with bacteria during phagocytosis [97,98] and in the extracellular inflammatory fluid [40,86,[99][100][101]. Levels of BPI in cell-free inflammatory fluid of neutrophil-rich exudates are functionally significant (≥10 nM) and co-exist with other antimicrobial host factors that can act in concert with BPI [76,86,[100][101][102][103][104]. Thus, in cooperation with other cellular and humoral components of the innate immune system, BPI can contribute to killing of invading GNB.…”

Effect of d-Alanylation of (Lipo)Teichoic Acids of Staphylococcus aureus on Host Secretory Phospholipase A2 Action before and after Phagocytosis by Human Neutrophils

“…In the case of gIIA PLA 2 , the effect of D-alanylation is not on initial protein binding to the bacterial surface, but, rather, on the ability of the bound enzyme to penetrate the cell wall and degrade membrane PL (24). This effect of Dalanylation is the same for the highly cationic wild-type (wt) enzyme and less cationic and less active gIIA PLA 2 mutants, raising the possibility that the resistance of ingested S. aureus to gV and gX PLA 2 in human neutrophils (13) might also depend on D-alanylation of bacterial (lipo)teichoic acids.…”

“…Purified recombinant human gV and gX PLA 2 were provided by Dr. M. Gelb (University of Washington, Seattle, WA) (13).…”

“…Despite the high concentrations of antimicrobial compounds within the phagocytic vacuoles of neutrophils, not all ingested S. aureus are killed, and most, if not all, ingested cocci remain grossly intact even after many hours within the neutrophil (13)(14)(15). Appreciable degradation of bacterial phospholipids (PL) during and after phagocytosis of S. aureus by human neutrophils in vitro requires the added presence of group IIA (gIIA) phospholipase A 2 (PLA 2 ) (13), a potent extracellular antistaphylococcal enzyme that is mobilized in addition to neutrophils during acute inflammation (4, 16 -19).…”

“…Human neutrophils lack gIIA PLA 2 (20), but contain two structurally related PLA 2 , gV and gX (20,21). To date, the gV and gX PLA 2 are not known to contribute to digestion of ingested S. aureus (13), presumably because of their relatively low intrinsic antistaphylococcal activity and low abundance within human neutrophils (20,22,23).…”