Identification of small signalling molecules promoting cardiac-specific differentiation of mouse embryonic stem cells.
Identification of signalling cascades involved in cardiomyogenesis is crucial for optimising the generation of cardiomyocytes from embryonic stem cells (ES cells) (in vitro). We used a transgenic ES cell lineage expressing enhanced green fluorescent protein (EGFP) under the control of the alpha-myosin heavy chain (alpha-MHC) promoter (palphaMHC-EGFP) to investigate the effects of 33 small molecules interfering with several signalling cascades on cardiomyogenesis. Interestingly, the L-Type Ca2+ channel blocker Verapamil as well as Cyclosporin, an inhibitor of the protein phosphatase 2B, exerted the most striking pro-cardiomyogenic effect. Forskolin (adenylate cyclase stimulator) exerted the most striking anti-cardiomyogenic effect. The cardiomyogenic effect of Cyclosporin and Verapamil correlated with an expression of early cardiac markers Nkx2.5 and GATA4. Compared to the effects on late developmental stage embryoid bodies (EBs) stimulation of early developmental stage EBs (1-day old) with Verapamil or Cyclosporin for 48 h resulted in a potent cardiomyogenic effect. Accordingly, enhanced expression of alpha-MHC mRNA and EGFP mRNA was observed after stimulation of the early developmental stage EBs for 48 h. No expression of alpha-smooth muscle actin or platelet endothelial cell adhesion molecule-1 (PECM-1) as well as of neuronal genes (Nestin, Neurofilament H) has been observed demonstrating a preferentially pro-cardiomyogenic effect by both molecules.
Sachinidis
Agapios
A
Centre of Physiology and Pathophysiology, Institute of Neurophysiology, Cologne, Germany. a.sachinidis@uni-koeln.de
Schwengberg
Silke
S
Hippler-Altenburg
Rita
R
Mariappan
Devi
D
Kamisetti
Naidu
N
Seelig
Bianca
B
Berkessel
Albrecht
A
Hescheler
Jurgen
J
D000262:Adenylyl Cyclases / Q000187:drug effects; D000818:Animals; D001483:Base Sequence; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D065095:Calcineurin Inhibitors; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002454:Cell Differentiation; D005576:Colforsin / Q000494:pharmacology; D016572:Cyclosporine / Q000494:pharmacology; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D050991:GATA4 Transcription Factor / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D032383:Myocytes, Cardiac / Q000737:chemistry / Q000166:cytology* / Q000378:metabolism; D018995:Myosin Heavy Chains / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis; D015398:Signal Transduction; D014157:Transcription Factors / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D014700:Verapamil / Q000494:pharmacology
D015415:Biomarkers; D065095:Calcineurin Inhibitors; D002121:Calcium Channel Blockers; D050991:GATA4 Transcription Factor; C484171:Gata4 protein, mouse; D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins; C484469:Nkx2-5 protein, mouse; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D012333:RNA, Messenger; D014157:Transcription Factors; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D005576:Colforsin; D016572:Cyclosporine; D014700:Verapamil; D018995:Myosin Heavy Chains; D000262:Adenylyl Cyclases