title: Cardiomyogenesis of embryonic stem cells upon purinergic receptor activation by ADP and ATP.
abstract: Purinergic signaling may be involved in embryonic development of the heart. In the present study, the effects of purinergic receptor stimulation on cardiomyogenesis of mouse embryonic stem (ES) cells were investigated. ADP or ATP increased the number of cardiac clusters and cardiac cells, as well as beating frequency. Cardiac-specific genes showed enhanced expression of α-MHC, MLC2v, α-actinin, connexin 45 (Cx45), and HCN4, on both gene and protein levels upon ADP/ATP treatment, indicating increased cardiomyogenesis and pacemaker cell differentiation. Real-time RT-PCR analysis of purinergic receptor expression demonstrated presence of P2X1, P2X4, P2X6, P2X7, P2Y1, P2Y2, P2Y4, and P2Y6 on differentiating ES cells. ATP and ADP as well as the P2X agonists β,γ-methylenadenosine 5'-triphosphate (β,γ-MetATP) and 8-bromoadenosine 5'-triphosphate (8-Br-ATP) but not UTP or UDP transiently increased the intracellular calcium concentration ([Ca(2+)](i)) as evaluated by the calcium indicator Fluo-4, whereas no changes in membrane potential were observed. [Ca(2+)](i) transients induced by ADP/ATP were abolished by the phospholipase C-β (PLC-β) inhibitor U-73122, suggesting involvement of metabotropic P2Y receptors. Furthermore, partial inhibition of [Ca(2+)](i) transients was achieved in presence of MRS2179, a selective P2Y1 receptor antagonist, whereas PPADS, a non-selective P2 receptor inhibitor, completely abolished the [Ca(2+)](i) response. Consequently, cardiomyocyte differentiation was decreased upon long term co-incubation of cells with ADP and P2 receptor antagonists. In summary, activation of purinoceptors and the subsequent [Ca(2+)](i) transients enhance the differentiation of ES cells toward cardiomyocytes. Purinergic receptor stimulation may be a promising strategy to drive the fate of pluripotent ES cells into a particular population of cardiomyocytes.
authors.affiliation: University Heart Center, Clinic of Internal Medicine I, Friedrich Schiller University Jena, Jena, Germany.
authors.forename: Safoura
authors.initials: S
authors.lastname: Mazrouei
authors.affiliation: Department of Physiology, Faculty of Medicine, Justus Liebig University, Giessen, Germany.
authors.forename: Fatemeh
authors.initials: F
authors.lastname: Sharifpanah
authors.affiliation: University Heart Center, Clinic of Internal Medicine I, Friedrich Schiller University Jena, Jena, Germany.|Department of Zoology, Faculty of Science, Tanta University, Tanta, Egypt.
authors.forename: Mohamed M
authors.initials: MM
authors.lastname: Bekhite
authors.affiliation: University Heart Center, Clinic of Internal Medicine I, Friedrich Schiller University Jena, Jena, Germany.
authors.forename: Hans-Reiner
authors.initials: HR
authors.lastname: Figulla
authors.affiliation: Department of Physiology, Faculty of Medicine, Justus Liebig University, Giessen, Germany.
authors.forename: Heinrich
authors.initials: H
authors.lastname: Sauer
authors.affiliation: University Heart Center, Clinic of Internal Medicine I, Friedrich Schiller University Jena, Jena, Germany. Maria.Wartenberg@med.uni-jena.de.
authors.forename: Maria
authors.initials: M
authors.lastname: Wartenberg
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology*; D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D053595:Embryonic Stem Cells / Q000187:drug effects*; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D024510:Muscle Development / Q000187:drug effects*; D009200:Myocardial Contraction / Q000187:drug effects; D032383:Myocytes, Cardiac / Q000187:drug effects*; D011247:Pregnancy; D058911:Purinergic P2X Receptor Agonists / Q000494:pharmacology; D058921:Purinergic P2Y Receptor Antagonists / Q000494:pharmacology; D058469:Receptors, Purinergic P2X / Q000096:biosynthesis / Q000187:drug effects / Q000235:genetics; D058472:Receptors, Purinergic P2Y1 / Q000187:drug effects; D014530:Uridine Diphosphate / Q000494:pharmacology; D014544:Uridine Triphosphate / Q000494:pharmacology
chemical_list: D058911:Purinergic P2X Receptor Agonists; D058921:Purinergic P2Y Receptor Antagonists; D058469:Receptors, Purinergic P2X; D058472:Receptors, Purinergic P2Y1; D014530:Uridine Diphosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D002118:Calcium; D014544:Uridine Triphosphate
keywords: ADP; ATP; Cardiomyocyte differentiation; Embryonic stem cell; Intracellular [Ca2+]i signaling; Purinergic receptors