title: Bone marrow-derived regenerated cardiomyocytes (CMG Cells) express functional adrenergic and muscarinic receptors.
abstract: BACKGROUND
  We recently reported that cardiomyocytes could be differentiated from bone marrow mesenchymal stem cells in vitro by 5-azacytidine treatment. In native cardiomyocytes, adrenergic and muscarinic receptors play crucial roles in mediating heart rate, conduction velocity, contractility, and cardiac hypertrophy. We investigated whether these receptors are expressed in differentiated CMG cells, and if so, whether they have downstream signaling systems.
  
  
  RESULTS
  Reverse transcription-polymerase chain reaction revealed that CMG cells had already expressed alpha(1A)-, alpha(1B)-, and alpha(1D)-adrenergic receptor mRNA before 5-azacytidine treatment, whereas expression of beta(1)-, beta(2)-adrenergic and M(1)-, M(2)-muscarinic receptors was first detected at 1 day. Phenylephrine dose-dependently induced phosphorylation of ERK1/2, which was completely inhibited by prazosin, and significantly increased cell size. Isoproterenol augmented cAMP by 38-fold, which was fully inhibited by propranolol. Isoproterenol (10(-7) mol/L) increased the spontaneous beating rate by 47.6% (basal, 127+/-16 bpm), and propranolol and CGP20712A (beta(1)-selective blocker) reduced it by 79.0% and 71.0%, respectively, whereas ICI118551 (beta(2)-selective blocker) induced slight reduction. Cell motion, percent shortening, and contractile velocity were increased by 37.5%, 26.9%, and 50.6%, respectively, in response to isoproterenol. Phenylephrine and isoproterenol augmented ANP and BNP gene expressions. Carbachol increased IP(3) by 32-fold, which was markedly inhibited by atropine as well as AFDX116 (M(2)-selective blocker) measured by radioimmunoassay.
  
  
  CONCLUSIONS
  These findings indicate that CMG cells expressed alpha(1A), alpha(1B), and alpha(1D) receptors before differentiation and expressed beta(1), beta(2), M(1), and M(2) receptors after they obtained the cardiomyocyte phenotype. These receptors had functional signal transduction pathways and could modulate cell function.
authors.affiliation: Cardiopulmonary Division, Department of Internal Medicine, Institute for Advanced Cardiac Therapeutics, Keio University School of Medicine, Tokyo, Japan.
authors.forename: Daihiko
authors.initials: D
authors.lastname: Hakuno
authors.forename: Keiichi
authors.initials: K
authors.lastname: Fukuda
authors.forename: Shinji
authors.initials: S
authors.lastname: Makino
authors.forename: Fusako
authors.initials: F
authors.lastname: Konishi
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Tomita
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Manabe
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Suzuki
authors.forename: Akihiro
authors.initials: A
authors.lastname: Umezawa
authors.forename: Satoshi
authors.initials: S
authors.lastname: Ogawa
mesh_terms: D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000818:Animals; D001374:Azacitidine / Q000494:pharmacology; D001854:Bone Marrow Cells / Q000502:physiology*; D002454:Cell Differentiation; D002478:Cells, Cultured; D006321:Heart / Q000502:physiology; D006339:Heart Rate / Q000187:drug effects; D007545:Isoproterenol / Q000494:pharmacology; D009200:Myocardial Contraction / Q000187:drug effects; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D010656:Phenylephrine / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D011941:Receptors, Adrenergic / Q000096:biosynthesis* / Q000235:genetics / Q000502:physiology*; D018340:Receptors, Adrenergic, alpha-1 / Q000096:biosynthesis / Q000235:genetics; D011943:Receptors, Adrenergic, beta / Q000096:biosynthesis / Q000235:genetics; D011976:Receptors, Muscarinic / Q000096:biosynthesis* / Q000235:genetics / Q000502:physiology*; D015398:Signal Transduction; D014158:Transcription, Genetic
chemical_list: D000316:Adrenergic alpha-Agonists; D000318:Adrenergic beta-Agonists; D012333:RNA, Messenger; D011941:Receptors, Adrenergic; D018340:Receptors, Adrenergic, alpha-1; D011943:Receptors, Adrenergic, beta; D011976:Receptors, Muscarinic; D010656:Phenylephrine; D007545:Isoproterenol; D001374:Azacitidine