Transepithelial Migration of Neutrophils in Response to Leukotriene B4 Is Mediated by a Reactive Oxygen Species-Extracellular Signal-Regulated Kinase-Linked Cascade

Abstract

The epithelial cells that form a barrier lining the lung airway are key regulators of neutrophil trafficking into the airway lumen in a variety of lung inflammatory diseases. Although the lipid mediator leukotriene B4 (LTB4) is known to be a principal chemoattractant for recruiting neutrophils to inflamed sites across the airway epithelium, the precise signaling mechanism involved remains largely unknown. In the present study, therefore, we investigated the signaling pathway through which LTB4 induces transepithelial migration of neutrophils. We found that LTB4 induces concentration-dependent transmigration of DMSO-differentiated HL-60 neutrophils and human polymorphonuclear neutrophils across A549 human lung epithelium. This effect was mediated via specific LTB4 receptors and was inhibited by pretreating the cells with N-acetylcysteine (NAC), an oxygen free radical scavenger, with diphenylene iodonium (DPI), an inhibitor of NADPH oxidase-like flavoproteins, or with PD98059, an extracellular signal-regulated kinase (ERK) inhibitor. Consistent with those findings, LTB4-induced ERK phosphorylation was completely blocked by pretreating cells with NAC or DPI. Taken together, our observations suggest LTB4 signaling to transepithelial migration is mediated via generation of reactive oxygen species, which leads to downstream activation of ERK. The physiological relevance of this signaling pathway was demonstrated in BALB/c mice, in which intratracheal instillation of LTB4 led to acute recruitment of neutrophils into the airway across the lung epithelium. Notably, the response to LTB4 was blocked by NAC, DPI, PD98059, or CP105696, a specific LTB4 receptor antagonist.

Neutrophils and Macrophages Cooperate in Host Resistance againstLeishmania braziliensisInfection

“…Parasite killing was dependent on phagocytosis of neutrophils since coculture of infected macrophages with Jurkat cells did not decrease parasite load. Furthermore, the addition of NAC, a superoxide scavenger (26), also blocked the effect exerted by coculture with live neutrophils. Macrophages are also "primed" for enhanced superoxide release following pretreatment with neutrophil serine proteases such as cathepsin G and neutrophil elastase (49).…”

“…7D). To confirm that superoxide anion is important in eliminating parasites from Lb-infected BALB/c macrophages upon coculture with neutrophils, we performed experiments in the presence of NAC, a superoxide scavenger (26). Addition of NAC to cocultures of Lb-infected macrophages and live PMN reversed the killing effect exerted by the addition of neutrophils as seen in the percentage of infected cells (Fig.…”

Cytosolic phospholipase A2, lipoxygenase metabolites, and reactive oxygen species

“…Over the past few years, it has been reported that LTB4 treatment of fibroblasts and neutrophils results in ROS generation, and that LTB4-induced chemotaxis is mediated by a NADPH oxidase-dependent cascade. These observations suggest that LTB4-induced ROS generation occurring via NOX is crucial to cell chemotaxis (34,35). In addition, Giembycz has reported that LTB4 promotes the robust, receptor-mediated activation of NADPH oxidase in guinea pig eosinophils (21,36).…”

TNF-α-induced up-regulation of intercellular adhesion molecule-1 is regulated by a Rac-ROS-dependent cascade in human airway epithelial cells

“…Trans-airway epithelial migration of leukocyte is a critical step of host immune responses as well as inflammatory airway diseases, causing leukocytes to migration into the airway lumen (Cramer et al, 1980;Rosseau et al, 2000;Woo et al, 2003). This migration is partly due to enhanced adhesion of leukocytes onto the epithelial cells through the expression of adhesion molecules (Rosseau et al, 2000).…”