Cardiomyocytes derived from embryonic stem cells resemble cardiomyocytes of the embryonic heart tube.
OBJECTIVE
After formation of the linear heart tube a chamber-specific program of gene expression becomes active that underlies the formation of the chamber myocardium. To assess whether this program is recapitulated in in vitro differentiated embryonic stem cells, we performed qualitative and quantitative analyses of cardiogenesis in vivo and in vitro.


METHODS
Gene expression profiles were made by in situ hybridisation and real-time PCR and electrophysiological profiles by patch clamp analyses of cardiomyocytes derived from time series of differentiating HM1 mouse embryonic stem cells and from embryonic and adult mouse hearts.


RESULTS
In embryoid bodies the in situ patterns of expression of alpha-myosin heavy chain, myosin light chain 2a and sarcoendoplasmic reticulum calcium ATPase 2a were similar to that of the heart muscle-specific marker gene cardiac troponin I. Myosin light chain 2v was expressed in part of the cardiac troponin I-expressing area, indicating heterogeneity within the cardiac cell population. Atrial natriuretic factor expression, indicative of the chamber-type program, could only very occasionally be detected by in situ hybridisation. Quantitative reverse transcriptase PCR showed that all cardiac genes, most notably atrial natriuretic factor, were expressed at relatively low levels, similar to those in embryonic hearts at embryonic day 8.75-9. Analysis of the electrophysiological characteristics of embryonic stem cell-derived cardiomyocytes showed an increase of the upstroke velocity and a shorter duration of the action potential during prolonged differentiation in vitro. When embryonic mouse heart compartments of embryonic day 12.5 were used as a reference, the electrophysiological characteristics of a substantial part of the embryonic stem cell-derived cardiomyocytes were most reminiscent to those observed in the embryonic outflow tract.


CONCLUSIONS
Together, these data suggest that most cardiomyocytes acquired by differentiation of embryonic stem cells maintain a phenotype reminiscent of that of the cardiomyocytes of the primary heart tube, and hardly any myocytes develop a chamber myocardial phenotype.
Fijnvandraat
Arnoud C
AC
Experimental and Molecular Cardiology Group, Academic Medical Center, University of Amsterdam, Meibergdreef 15, 1105 AZ, Amsterdam, The Netherlands.
van Ginneken
Antoni C G
AC
de Boer
Piet A J
PA
Ruijter
Jan M
JM
Christoffels
Vincent M
VM
Moorman
Antoon F M
AF
Lekanne Deprez
Ronald H
RH
D000200:Action Potentials; D000818:Animals; D009320:Atrial Natriuretic Factor / Q000235:genetics; D000252:Calcium-Transporting ATPases / Q000235:genetics; D024722:Cardiac Myosins / Q000235:genetics; D002454:Cell Differentiation; D002469:Cell Separation; D004151:Dipeptides; D004627:Embryonic Induction / Q000235:genetics*; D015870:Gene Expression; D020869:Gene Expression Profiling; D005865:Gestational Age; D006321:Heart / Q000196:embryology*; D007150:Immunohistochemistry / Q000379:methods; D017403:In Situ Hybridization / Q000379:methods; D051379:Mice; D008815:Mice, Inbred Strains; D032383:Myocytes, Cardiac / Q000502:physiology; D018994:Myosin Light Chains / Q000235:genetics; D018408:Patch-Clamp Techniques; D020033:Protein Isoforms / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D019831:Sodium-Calcium Exchanger / Q000235:genetics; D013234:Stem Cells / Q000166:cytology* / Q000502:physiology; D019210:Troponin I / Q000235:genetics
D004151:Dipeptides; D018994:Myosin Light Chains; D020033:Protein Isoforms; D019831:Sodium-Calcium Exchanger; D019210:Troponin I; C075385:Trp(Nps)-psi(COCH2)-Lys; C107184:myosin light chain 2; C110374:sodium-calcium exchanger 1; D009320:Atrial Natriuretic Factor; D024722:Cardiac Myosins; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases