title: ATP release from activated neutrophils occurs via connexin 43 and modulates adenosine-dependent endothelial cell function.
abstract: Extracellular ATP liberated during hypoxia and inflammation can either signal directly on purinergic receptors or can activate adenosine receptors following phosphohydrolysis to adenosine. Given the association of polymorphonuclear leukocytes (PMNs) with adenine-nucleotide/nucleoside signaling in the inflammatory milieu, we hypothesized that PMNs are a source of extracellular ATP. Initial studies using high-performance liquid chromatography and luminometric ATP detection assays revealed that PMNs release ATP through activation-dependent pathways. In vitro models of endothelial barrier function and neutrophil/endothelial adhesion indicated that PMN-derived ATP signals through endothelial adenosine receptors, thereby promoting endothelial barrier function and attenuating PMN/endothelial adhesion. Metabolism of extracellular ATP to adenosine required PMNs, and studies addressing these metabolic steps revealed that PMN express surface ecto-apyrase (CD39). In fact, studies with PMNs derived from cd39(-/-) mice showed significantly increased levels of extracellular ATP and lack of ATP dissipation from their supernatants. After excluding lytic ATP release, we used pharmacological strategies to reveal a potential mechanism involved in PMN-dependent ATP release (eg, verapamil, dipyridamole, brefeldin A, 18-alpha-glycyrrhetinic acid, connexin-mimetic peptides). These studies showed that PMN ATP release occurs through connexin 43 (Cx43) hemichannels in a protein/phosphatase-A-dependent manner. Findings in human PMNs were confirmed in PMNs derived from induced Cx43(-/-) mice, whereby activated PMNs release less than 15% of ATP relative to littermate controls, whereas Cx43 heterozygote PMNs were intermediate in their capacity for ATP release (P<0.01). Taken together, our results identify a previously unappreciated role for Cx43 in activated PMN ATP release, therein contributing to the innate metabolic control of the inflammatory milieu.
authors.affiliation: Department of Anesthesiology and Intensive Care Medicine, Tübingen University Hospital, Germany. heltzschig@partners.org
authors.forename: Holger K
authors.initials: HK
authors.lastname: Eltzschig
authors.forename: Tobias
authors.initials: T
authors.lastname: Eckle
authors.forename: Alice
authors.initials: A
authors.lastname: Mager
authors.forename: Natalie
authors.initials: N
authors.lastname: Küper
authors.forename: Christian
authors.initials: C
authors.lastname: Karcher
authors.forename: Thomas
authors.initials: T
authors.lastname: Weissmüller
authors.forename: Kerstin
authors.initials: K
authors.lastname: Boengler
authors.forename: Rainer
authors.initials: R
authors.lastname: Schulz
authors.forename: Simon C
authors.initials: SC
authors.lastname: Robson
authors.forename: Sean P
authors.initials: SP
authors.lastname: Colgan
mesh_terms: D000241:Adenosine / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism / Q000502:physiology*; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D001081:Apyrase / Q000172:deficiency / Q000378:metabolism; D018031:Connexin 43 / Q000172:deficiency / Q000378:metabolism / Q000502:physiology*; D042783:Endothelial Cells / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D051379:Mice; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D018375:Neutrophil Activation / Q000502:physiology*; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D010766:Phosphorylation
chemical_list: D015703:Antigens, CD; D018031:Connexin 43; D009240:N-Formylmethionine Leucyl-Phenylalanine; D000255:Adenosine Triphosphate; D001081:Apyrase; C067869:CD39 antigen; D000241:Adenosine