Purinergic receptor modulation of LPS-stimulated signaling events and nitric oxide release in RAW 264.7 macrophages

Abstract

Purinergic receptors of the P2 class are cell surface receptors which are sensitive to extracellular adenine nucleotides, such as ATP and ADP. This class of receptors is divided into the P2Y family of G protein-coupled receptors and the P2X family of ligand-gated ion channels. The P2X receptors, seven of which have been cloned, are thought to possess two transmembrane domains and function as multimeric complexes. Numerous studies have suggested a role for P2 receptors in activation of macrophages by Gram-negative bacterial endotoxin (lipopolysaccharide; LPS). LPS is thought to exert its toxic effects, in large part, by inducing macrophages to release inflammatory mediators such as tumor necrosis factor α (TNFα), interleukin-1 (IL-1) and nitric oxide (NO). Although multiple signal transduction pathways are activated by LPS in macrophages, the proximal mechanisms by which LPS exerts these effects remain unclear. The current study examines the role of the P2X₇/P2Z purinergic receptor in LPS signaling events and in nitric oxide (NO) production. The results indicate that the P2X₇ receptor is required for maximal LPS activation of the mitogenactivated protein (MAP) kinases extracellular signal-regulated kinase (ERK)1 and ERK2, for activation of nuclear factor (NF)-κB, as well as for upregulation of the inducible form of nitric oxide synthase (iNOS). These results are fortified by our recent observation that the C-terminus of the P2X₇ receptor is homologous to conserved LPS binding domains of proteins critical to host responses to Gram-negative bacterial infection, such as LPS-binding protein (LBP) and bactericidal permeability-increasing protein (BPI). Taken together, these observations suggest that the P2X ₇ receptor plays a fundamental role in LPS signal transduction and activation of macrophages, and thus may represent a therapeutic target for Gram-negative bacterial septicemia.