Altered Oxygen Tension Modulates Cytokine-Induced Signal Transduction in Polymorphonuclear Leukocytes: Regulation of the GPLCPathway

Abstract

The purpose of these studies was to examine the sensitivity of the PIP2–PLC-transducing pathway (GPLC) and its relationship to the respiratory burst in human polymorphonuclear leukocytes (PMN) stimulated by IL-8, TNF-α, or IL-1β during sequential changes in buffer oxygen tension from normoxia (pO2= 180–200 mm Hg), to hypoxia (pO2< 30 mm Hg) and then reoxygenation (pO2> 140 mm Hg). Our specific hypothesis was that altered oxygen tensions would regulate the GPLCpathway in human PMN. GPLCactivity was assayed by investigating phospholipase C activity by measuring inositol phosphates and diacylglycerol (DAG) formation. Respiratory burst activity was assayed as O−2production and NADPH oxidase activation in intact PMN and in a cell-free system, respectively, and correlated separately to both early and late DAG production. At 1 min, DAG formation during normoxia was decreased by IL-8 plus fibronectin while hypoxia had no regulatory effect on control of DAG formation by any of the cytokines. In contrast to early DAG formation, hypoxia significantly downregulated late DAG formation induced by buffer without fibronectin, IL-8 plus fibronectin, and IL-1β with or without fibronectin. Hypoxia/reoxygenation in and of itself significantly increased DAG formation vs levels seen in the presence or absence of IL-8, TNF-α, or IL-1β with or without fibronectin. Changes in early DAG production during the alterations in oxygen tension correlated best with corresponding changes in O−2production in intact cells, whereas late DAG production correlated best with NADPH oxidase activation assayed in the cell-free system. Thus, changes in oxygen tension can directly modulate the extent of the PMN response to stimulation by IL-8, TNF-α, or IL-1β and the GPLC-receptor pathway is particularly regulated by physiologically relevant periods of hypoxia/reoxygenation.