title: The stimulatory adenosine receptor ADORA2B regulates serotonin (5-HT) synthesis and release in oxygen-depleted EC cells in inflammatory bowel disease.
abstract: OBJECTIVE
  We recently demonstrated that hypoxia, a key feature of IBD, increases enterochromaffin (EC) cell 5-HT secretion, which is also physiologically regulated by the ADORA2B mechanoreceptor. Since hypoxia is associated with increased extracellular adenosine, we wanted to examine whether this nucleotide amplifies HIF-1α-mediated 5-HT secretion.
  
  
  METHODS
  The effects of hypoxia were studied on IBD mucosa, isolated IBD-EC cells, isolated normal EC cells and the EC cell tumor derived cell line KRJ-1. Hypoxia (0.5% O2) was compared to NECA (adenosine agonist), MRS1754 (ADORA2B receptor antagonist) and SCH442146 (ADORA2A antagonist) on HIF signaling and 5-HT secretion. Antisense approaches were used to mechanistically evaluate EC cells in vitro. PCR and western blot were used to analyze transcript and protein levels of HIF-1α signaling and neuroendocrine cell function. An animal model of colitis was evaluated to confirm hypoxia:adenosine signaling in vivo.
  
  
  RESULTS
  HIF-1α is upregulated in IBD mucosa and IBD-EC cells, the majority (~90%) of which express an activated phenotype in situ. Hypoxia stimulated 5-HT release maximally at 30 mins, an effect amplified by NECA and selectively inhibited by MRS1754, through phosphorylation of TPH-1 and activation of VMAT-1. Transient transfection with Renilla luciferase under hypoxia transcriptional response element (HRE) control identified that ADORA2B activated HIF-1α signaling under hypoxic conditions. Additional signaling pathways associated with hypoxia:adenosine included MAP kinase and CREB. Antisense approaches mechanistically confirmed that ADORA2B signaling was linked to these pathways and 5-HT release under hypoxic conditions. Hypoxia:adenosine activation which could be reversed by 5'-ASA treatment was confirmed in a TNBS-model.
  
  
  CONCLUSIONS
  Hypoxia induced 5-HT synthesis and secretion is amplified by ADORA2B signaling via MAPK/CREB and TPH-1 activation. Targeting ADORA2s may decrease EC cell 5-HT production and secretion in IBD.
authors: Rikard
authors: R
authors: Dammen
authors: Gastrointestinal Pathobiology Research Group, Yale University School of Medicine, New Haven, Connecticut, United States of America.
authors: Rikard
authors: R
authors: Damen
authors: Martin
authors: M
authors: Haugen
authors: Bernhard
authors: B
authors: Svejda
authors: Daniele
authors: D
authors: Alaimo
authors: Oystein
authors: O
authors: Brenna
authors: Roswitha
authors: R
authors: Pfragner
authors: Bjorn I
authors: BI
authors: Gustafsson
authors: Mark
authors: M
authors: Kidd
mesh_terms: D000241:Adenosine / Q000494:pharmacology; D000328:Adult; D000368:Aged; D000818:Animals; D015687:Cell Hypoxia / Q000187:drug effects / Q000235:genetics; D003092:Colitis / Q000378:metabolism / Q000473:pathology; D004195:Disease Models, Animal; D004759:Enterochromaffin Cells / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D005260:Female; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism; D015212:Inflammatory Bowel Diseases / Q000378:metabolism* / Q000473:pathology; D007413:Intestinal Mucosa / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D008297:Male; D008875:Middle Aged; D010100:Oxygen / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D043704:Receptor, Adenosine A2B / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D012701:Serotonin / Q000096:biosynthesis* / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D014302:Trinitrobenzenesulfonic Acid
chemical_list: D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D012333:RNA, Messenger; D043704:Receptor, Adenosine A2B; D012701:Serotonin; D014302:Trinitrobenzenesulfonic Acid; D000241:Adenosine; D010100:Oxygen
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