title: Prostaglandin E₂ maintains mouse ESC undifferentiated state through regulation of connexin31, connexin43 and connexin45 expression: involvement of glycogen synthase kinase 3β/β-catenin.
abstract: BACKGROUND
  Although previous reports have examined the function of prostaglandin E₂ (PGE₂) on gap junctions and undifferentiated stem cells, its effects on the reciprocal action of connexin (Cx) isoforms and undifferentiation in embryonic stem cells (ESCs) are unclear. Therefore, we investigated the role of PGE₂ on Cx isoforms and maintenance of mouse ESC undifferentiated state.
  
  
  RESULTS
  We have analysed 10 Cx genes, but found nine of them. PGE₂ (50 μM) stimulated Cx31, Cx32, Cx40, Cx43 and Cx45 mRNA expression. Amongst them, PGE₂ maximally stimulated the Cx43 mRNA expression and gap junction inter-cellular coupling. Therefore, we investigated the effect of PGE₂ on Cx43 expression. PGE₂ activated cAMP/protein kinase A (PKA) and phosphatidylinositol 3-kinase (PI3K)/Akt phosphorylation. In addition, treatments of adenylate cyclase activators increased Cx43 expression, but not PI3K/Akt phosphorylation. PGE₂ also inactivated GSK-3β and stimulated active-β-catenin. Furthermore, a ChiP assay demonstrated the association of β-catenin with the Cx26 (as control) and Cx43 promoter. Finally, down-regulation of PGE₂-induced Cx isoforms by AH 6809, Cx31-, Cx43-, Cx45 small interfering (si)RNA and 18α-glycyrrhetinic acid decreased levels of undifferentiated markers of ESCs, including Oct4, FoxD3, Sox2 and SSEA-1, but Nanog did not be down-regulated by Cx43 siRNA.
  
  
  CONCLUSIONS
  PGE₂ stimulates Cx isoforms via GSK-3β/β-catenin via EP2-receptor-dependent cAMP/PKA and PI3K/Akt in mouse ESCs, thereby partially contributing to the maintenance of their undifferentiated state.
authors.affiliation: Department of Veterinary Physiology, College of Veterinary Medicine, Chonnam National University, Gwangju 500-757, Republic of Korea.
authors.forename: Seung Pil
authors.initials: SP
authors.lastname: Yun
authors.forename: Jung Min
authors.initials: JM
authors.lastname: Ryu
authors.forename: Jae Hong
authors.initials: JH
authors.lastname: Park
authors.forename: Mi Ok
authors.initials: MO
authors.lastname: Kim
authors.forename: Jang-Hern
authors.initials: JH
authors.lastname: Lee
authors.forename: Ho Jae
authors.initials: HJ
authors.lastname: Han
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D000072259:Connexin 26; D018031:Connexin 43 / Q000235:genetics / Q000378:metabolism; D017630:Connexins / Q000378:metabolism*; D000242:Cyclic AMP / Q000096:biosynthesis; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D015232:Dinoprostone / Q000494:pharmacology*; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D017629:Gap Junctions / Q000187:drug effects / Q000378:metabolism; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D051379:Mice; D010766:Phosphorylation / Q000187:drug effects; D020033:Protein Isoforms / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D058307:Receptors, Prostaglandin E, EP2 Subtype / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D051176:beta Catenin / Q000378:metabolism*
chemical_list: D018031:Connexin 43; D017630:Connexins; C000607042:GJB2 protein, human; D020033:Protein Isoforms; D058307:Receptors, Prostaglandin E, EP2 Subtype; D051176:beta Catenin; C082920:connexin 45; D000072259:Connexin 26; C068044:GJB3 protein, human; D000242:Cyclic AMP; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D051057:Proto-Oncogene Proteins c-akt; D017868:Cyclic AMP-Dependent Protein Kinases; D038362:Glycogen Synthase Kinase 3; D015232:Dinoprostone