title: Nucleotides induce IL-6 release from human airway epithelia via P2Y2 and p38 MAPK-dependent pathways.
abstract: Extracellular nucleotides can mediate a variety of cellular functions via interactions with purinergic receptors. We previously showed that mechanical ventilation (MV) induces airway IL-6 and ATP release, modifies luminal nucleotide composition, and alters lung purinoceptor expression. Here we hypothesize that extracellular nucleotides induce secretion of IL-6 by small airway epithelial cells (SAEC). Human SAEC were stimulated with nucleotides in the presence or absence of inhibitors. Supernatants were analyzed for IL-6 and lysates for p38 MAPK activity by ELISA. RNA was analyzed by real-time RT-PCR. Rats (n=51) were randomized to groups as follows: control, small-volume MV, large-volume MV, large-volume MV-intratracheal apyrase, or small-volume MV-intratracheal adenosine 5'-O-(3-thiotriphosphate) (ATPgammaS). After 1 h of MV, bronchoalveolar lavage fluid was analyzed for ATP and IL-6 by luminometry and ELISA. ATP and ATPgammaS increased SAEC IL-6 secretion in a time- and dose-dependent manner, an effect inhibited by apyrase. Agonists were ranked in the following order: ATPgammaS>ATP=UTP>ADP=adenosine>2-methylthio-ADP=control. SB-203580, but not U-0126 or JNK1 inhibitor, decreased nucleotide effects. Additionally, nucleotides induced p38 MAPK phosphorylation. Inhibitors of Ca2+ signaling, phospholipase C, transcription, and translation decreased IL-6 release. Furthermore, nucleotides increased IL-6 expression. In vivo, large-volume MV increased airway ATP and IL-6 concentrations. IL-6 release was decreased by apyrase and increased by ATPgammaS. Extracellular nucleotides induce P2Y2-mediated secretion of IL-6 by SAEC via Ca2+, phospholipase C, and p38 MAPK-dependent pathways. This effect is dependent on transcription and translation. Our findings were confirmed in an in vivo model, thus demonstrating a novel mechanism of nucleotide-induced IL-6 secretion by airway epithelia.
authors.affiliation: Division of Trauma and Critical Care, Department of Surgery, University of North Carolina at Chapel Hill, 4008 Burnett-Womack, Chapel Hill, NC 27599-7228, USA.
authors.forename: Christelle D
authors.initials: CD
authors.lastname: Douillet
authors.forename: William P
authors.initials: WP
authors.lastname: Robinson
authors.forename: Peter M
authors.initials: PM
authors.lastname: Milano
authors.forename: Richard C
authors.initials: RC
authors.lastname: Boucher
authors.forename: Preston B
authors.initials: PB
authors.lastname: Rich
mesh_terms: D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001081:Apyrase / Q000494:pharmacology; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D015850:Interleukin-6 / Q000235:genetics / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D008297:Male; D009711:Nucleotides / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D014176:Protein Biosynthesis; D058910:Purinergic P2 Receptor Agonists; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D058473:Receptors, Purinergic P2Y2; D012121:Respiration, Artificial; D012137:Respiratory System / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*; D000093242:Interleukin-6 Inhibitors
chemical_list: D000255:Adenosine Triphosphate; D001081:Apyrase; D015850:Interleukin-6; D009711:Nucleotides; D058910:Purinergic P2 Receptor Agonists; D012333:RNA, Messenger; D018048:Receptors, Purinergic P2; D058473:Receptors, Purinergic P2Y2; D048051:p38 Mitogen-Activated Protein Kinases; D000093242:Interleukin-6 Inhibitors; C546385:P2RY2 protein, human; C022571:adenosine 5'-O-(3-thiotriphosphate)