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Erschienen in: Heart and Vessels 6/2014

01.11.2014 | Original Article

Forced aggregation and defined factors allow highly uniform-sized embryoid bodies and functional cardiomyocytes from human embryonic and induced pluripotent stem cells

verfasst von: Martin Pesl, Ivana Acimovic, Jan Pribyl, Renata Hezova, Aleksandra Vilotic, Jeremy Fauconnier, Jan Vrbsky, Peter Kruzliak, Petr Skladal, Tomas Kara, Vladimir Rotrekl, Alain Lacampagne, Petr Dvorak, Albano C. Meli

Erschienen in: Heart and Vessels | Ausgabe 6/2014

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Abstract

In vitro human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) can differentiate into functional cardiomyocytes (CMs). Protocols for cardiac differentiation of hESCs and hiPSCs include formation of the three-dimensional cell aggregates called embryoid bodies (EBs). The traditional suspension method for EB formation from clumps of cells results in an EB population heterogeneous in size and shape. In this study we show that forced aggregation of a defined number of single cells on AggreWell plates gives a high number of homogeneous EBs that can be efficiently differentiated into functional CMs by application of defined growth factors in the media. For cardiac differentiation, we used three hESC lines and one hiPSC line. Our contracting EBs and the resulting CMs express cardiac markers, namely myosin heavy chain α and β, cardiac ryanodine receptor/calcium release channel, and cardiac troponin T, shown by real-time polymerase chain reaction and immunocytochemistry. Using Ca2+ imaging and atomic force microscopy, we demonstrate the functionality of RyR2 to release Ca2+ from the sarcoplasmic reticulum as well as reliability in contractile and beating properties of hESC-EBs and hiPSC-EBs upon the stimulation or inhibition of the β-adrenergic pathway.
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Literatur
1.
Zurück zum Zitat Doetschman TC, Eistetter H, Katz M, Schmidt W, Kemler R (1985) The in vitro development of blastocyst-derived embryonic stem cell lines: formation of visceral yolk sac, blood islands and myocardium. J Embryol Exp Morphol 87:27–45PubMed Doetschman TC, Eistetter H, Katz M, Schmidt W, Kemler R (1985) The in vitro development of blastocyst-derived embryonic stem cell lines: formation of visceral yolk sac, blood islands and myocardium. J Embryol Exp Morphol 87:27–45PubMed
2.
Zurück zum Zitat Doi K, Itoh H, Nakagawa O, Igaki T, Yamashita J, Chun TH, Inoue M, Masatsugu K, Nakao K (1997) Expression of natriuretic peptide system during embryonic stem cell vasculogenesis. Heart Vessels Suppl 12:18–22 Doi K, Itoh H, Nakagawa O, Igaki T, Yamashita J, Chun TH, Inoue M, Masatsugu K, Nakao K (1997) Expression of natriuretic peptide system during embryonic stem cell vasculogenesis. Heart Vessels Suppl 12:18–22
3.
Zurück zum Zitat Burridge PW, Anderson D, Priddle H, Barbadillo Munoz MD, Chamberlain S, Allegrucci C, Young LE, Denning C (2007) Improved human embryonic stem cell embryoid body homogeneity and cardiomyocyte differentiation from a novel V-96 plate aggregation system highlights interline variability. Stem Cells 25:929–938PubMedCrossRef Burridge PW, Anderson D, Priddle H, Barbadillo Munoz MD, Chamberlain S, Allegrucci C, Young LE, Denning C (2007) Improved human embryonic stem cell embryoid body homogeneity and cardiomyocyte differentiation from a novel V-96 plate aggregation system highlights interline variability. Stem Cells 25:929–938PubMedCrossRef
4.
Zurück zum Zitat Burridge PW, Thompson S, Millrod MA, Weinberg S, Yuan X, Peters A, Mahairaki V, Koliatsos VE, Tung L, Zambidis ET (2011) A universal system for highly efficient cardiac differentiation of human induced pluripotent stem cells that eliminates interline variability. PLoS One 6:e18293PubMedCentralPubMedCrossRef Burridge PW, Thompson S, Millrod MA, Weinberg S, Yuan X, Peters A, Mahairaki V, Koliatsos VE, Tung L, Zambidis ET (2011) A universal system for highly efficient cardiac differentiation of human induced pluripotent stem cells that eliminates interline variability. PLoS One 6:e18293PubMedCentralPubMedCrossRef
5.
Zurück zum Zitat Kattman SJ, Witty AD, Gagliardi M, Dubois NC, Niapour M, Hotta A, Ellis J, Keller G (2011) Stage-specific optimization of activin/nodal and BMP signaling promotes cardiac differentiation of mouse and human pluripotent stem cell lines. Cell Stem Cell 8:228–240PubMedCrossRef Kattman SJ, Witty AD, Gagliardi M, Dubois NC, Niapour M, Hotta A, Ellis J, Keller G (2011) Stage-specific optimization of activin/nodal and BMP signaling promotes cardiac differentiation of mouse and human pluripotent stem cell lines. Cell Stem Cell 8:228–240PubMedCrossRef
6.
Zurück zum Zitat Xu XQ, Graichen R, Soo SY, Balakrishnan T, Rahmat SN, Sieh S, Tham SC, Freund C, Moore J, Mummery C, Colman A, Zweigerdt R, Davidson BP (2008) Chemically defined medium supporting cardiomyocyte differentiation of human embryonic stem cells. Differentiation 76:958–970PubMed Xu XQ, Graichen R, Soo SY, Balakrishnan T, Rahmat SN, Sieh S, Tham SC, Freund C, Moore J, Mummery C, Colman A, Zweigerdt R, Davidson BP (2008) Chemically defined medium supporting cardiomyocyte differentiation of human embryonic stem cells. Differentiation 76:958–970PubMed
7.
Zurück zum Zitat Yang L, Soonpaa MH, Adler ED, Roepke TK, Kattman SJ, Kennedy M, Henckaerts E, Bonham K, Abbott GW, Linden RM, Field LJ, Keller GM (2008) Human cardiovascular progenitor cells develop from a KDR + embryonic-stem-cell-derived population. Nature 453:524–528PubMedCrossRef Yang L, Soonpaa MH, Adler ED, Roepke TK, Kattman SJ, Kennedy M, Henckaerts E, Bonham K, Abbott GW, Linden RM, Field LJ, Keller GM (2008) Human cardiovascular progenitor cells develop from a KDR + embryonic-stem-cell-derived population. Nature 453:524–528PubMedCrossRef
8.
Zurück zum Zitat Dvorak P, Dvorakova D, Koskova S, Vodinska M, Najvirtova M, Krekac D, Hampl A (2005) Expression and potential role of fibroblast growth factor 2 and its receptors in human embryonic stem cells. Stem Cells 23:1200–1211PubMedCrossRef Dvorak P, Dvorakova D, Koskova S, Vodinska M, Najvirtova M, Krekac D, Hampl A (2005) Expression and potential role of fibroblast growth factor 2 and its receptors in human embryonic stem cells. Stem Cells 23:1200–1211PubMedCrossRef
9.
Zurück zum Zitat Eiselleova L, Peterkova I, Neradil J, Slaninova I, Hampl A, Dvorak P (2008) Comparative study of mouse and human feeder cells for human embryonic stem cells. Int J Dev Biol 52:353–363PubMedCrossRef Eiselleova L, Peterkova I, Neradil J, Slaninova I, Hampl A, Dvorak P (2008) Comparative study of mouse and human feeder cells for human embryonic stem cells. Int J Dev Biol 52:353–363PubMedCrossRef
10.
Zurück zum Zitat Armstrong L, Tilgner K, Saretzki G, Atkinson SP, Stojkovic M, Moreno R, Przyborski S, Lako M (2010) Human induced pluripotent stem cell lines show stress defense mechanisms and mitochondrial regulation similar to those of human embryonic stem cells. Stem Cells 28:661–673PubMedCrossRef Armstrong L, Tilgner K, Saretzki G, Atkinson SP, Stojkovic M, Moreno R, Przyborski S, Lako M (2010) Human induced pluripotent stem cell lines show stress defense mechanisms and mitochondrial regulation similar to those of human embryonic stem cells. Stem Cells 28:661–673PubMedCrossRef
11.
Zurück zum Zitat Dubois NC, Craft AM, Sharma P, Elliott DA, Stanley EG, Elefanty AG, Gramolini A, Keller G (2011) SIRPA is a specific cell-surface marker for isolating cardiomyocytes derived from human pluripotent stem cells. Nat Biotechnol 29:1011–1018PubMedCrossRef Dubois NC, Craft AM, Sharma P, Elliott DA, Stanley EG, Elefanty AG, Gramolini A, Keller G (2011) SIRPA is a specific cell-surface marker for isolating cardiomyocytes derived from human pluripotent stem cells. Nat Biotechnol 29:1011–1018PubMedCrossRef
12.
Zurück zum Zitat Lee MY, Cagavi Bozkulak E, Schliffke S, Amos PJ, Ren Y, Ge X, Ehrlich BE, Qyang Y (2011) High density cultures of embryoid bodies enhanced cardiac differentiation of murine embryonic stem cells. Biochem Biophys Res Commun 416:51–57PubMedCrossRef Lee MY, Cagavi Bozkulak E, Schliffke S, Amos PJ, Ren Y, Ge X, Ehrlich BE, Qyang Y (2011) High density cultures of embryoid bodies enhanced cardiac differentiation of murine embryonic stem cells. Biochem Biophys Res Commun 416:51–57PubMedCrossRef
13.
Zurück zum Zitat Fujita E, Nakanishi T, Nishizawa T, Hagiwara N, Matsuoka R (2013) Mutations in the cardiac troponin T gene show various prognoses in Japanese patients with hypertrophic cardiomyopathy. Heart Vessels. doi:10.1007/s00380-013-0332-3 Fujita E, Nakanishi T, Nishizawa T, Hagiwara N, Matsuoka R (2013) Mutations in the cardiac troponin T gene show various prognoses in Japanese patients with hypertrophic cardiomyopathy. Heart Vessels. doi:10.​1007/​s00380-013-0332-3
14.
Zurück zum Zitat Zwi-Dantsis L, Huber I, Habib M, Winterstern A, Gepstein A, Arbel G, Gepstein L (2013) Derivation and cardiomyocyte differentiation of induced pluripotent stem cells from heart failure patients. Eur Heart J 34:1575–1586PubMedCrossRef Zwi-Dantsis L, Huber I, Habib M, Winterstern A, Gepstein A, Arbel G, Gepstein L (2013) Derivation and cardiomyocyte differentiation of induced pluripotent stem cells from heart failure patients. Eur Heart J 34:1575–1586PubMedCrossRef
15.
16.
Zurück zum Zitat Itzhaki I, Rapoport S, Huber I, Mizrahi I, Zwi-Dantsis L, Arbel G, Schiller J, Gepstein L (2011) Calcium handling in human induced pluripotent stem cell derived cardiomyocytes. PLoS One 6:e18037PubMedCentralPubMedCrossRef Itzhaki I, Rapoport S, Huber I, Mizrahi I, Zwi-Dantsis L, Arbel G, Schiller J, Gepstein L (2011) Calcium handling in human induced pluripotent stem cell derived cardiomyocytes. PLoS One 6:e18037PubMedCentralPubMedCrossRef
17.
Zurück zum Zitat Lee YK, Ng KM, Lai WH, Chan YC, Lau YM, Lian Q, Tse HF, Siu CW (2011) Calcium homeostasis in human induced pluripotent stem cell-derived cardiomyocytes. Stem Cell Rev 7:976–986PubMedCentralPubMedCrossRef Lee YK, Ng KM, Lai WH, Chan YC, Lau YM, Lian Q, Tse HF, Siu CW (2011) Calcium homeostasis in human induced pluripotent stem cell-derived cardiomyocytes. Stem Cell Rev 7:976–986PubMedCentralPubMedCrossRef
18.
Zurück zum Zitat Liu B, Wohlfart B, Johansson BW (1990) Effects of low temperature on contraction in papillary muscles from rabbit, rat, and hedgehog. Cryobiology 27:539–546PubMedCrossRef Liu B, Wohlfart B, Johansson BW (1990) Effects of low temperature on contraction in papillary muscles from rabbit, rat, and hedgehog. Cryobiology 27:539–546PubMedCrossRef
19.
Zurück zum Zitat Fu Y, Zhang GQ, Hao XM, Wu CH, Chai Z, Wang SQ (2005) Temperature dependence and thermodynamic properties of Ca2+ sparks in rat cardiomyocytes. Biophys J 89:2533–2541PubMedCentralPubMedCrossRef Fu Y, Zhang GQ, Hao XM, Wu CH, Chai Z, Wang SQ (2005) Temperature dependence and thermodynamic properties of Ca2+ sparks in rat cardiomyocytes. Biophys J 89:2533–2541PubMedCentralPubMedCrossRef
20.
Zurück zum Zitat Wang SQ, Huang YH, Liu KS, Zhou ZQ (1997) Dependence of myocardial hypothermia tolerance on sources of activator calcium. Cryobiology 35:193–200PubMedCrossRef Wang SQ, Huang YH, Liu KS, Zhou ZQ (1997) Dependence of myocardial hypothermia tolerance on sources of activator calcium. Cryobiology 35:193–200PubMedCrossRef
21.
Zurück zum Zitat Degubareff T, Sleator W Jr (1965) Effects of caffeine on mammalian atrial muscle, and its interaction with adenosine and calcium. J Pharmacol Exp Ther 148:202–214PubMed Degubareff T, Sleator W Jr (1965) Effects of caffeine on mammalian atrial muscle, and its interaction with adenosine and calcium. J Pharmacol Exp Ther 148:202–214PubMed
22.
Zurück zum Zitat Axelsson J, Thesleff S (1958) Activation of the contractile mechanism in striated muscle. Acta Physiol Scand 44:55–66PubMedCrossRef Axelsson J, Thesleff S (1958) Activation of the contractile mechanism in striated muscle. Acta Physiol Scand 44:55–66PubMedCrossRef
23.
Zurück zum Zitat Weber A, Herz R (1968) The relationship between caffeine contracture of intact muscle and the effect of caffeine on reticulum. J Gen Physiol 52:750–759PubMedCentralPubMedCrossRef Weber A, Herz R (1968) The relationship between caffeine contracture of intact muscle and the effect of caffeine on reticulum. J Gen Physiol 52:750–759PubMedCentralPubMedCrossRef
24.
Zurück zum Zitat Bellin M, Marchetto MC, Gage FH, Mummery CL (2012) Induced pluripotent stem cells: the new patient? Nat Rev Mol Cell Biol 13:713–726PubMedCrossRef Bellin M, Marchetto MC, Gage FH, Mummery CL (2012) Induced pluripotent stem cells: the new patient? Nat Rev Mol Cell Biol 13:713–726PubMedCrossRef
25.
Zurück zum Zitat Bratt-Leal AM, Carpenedo RL, McDevitt TC (2009) Engineering the embryoid body microenvironment to direct embryonic stem cell differentiation. Biotechnol Prog 25:43–51PubMedCentralPubMedCrossRef Bratt-Leal AM, Carpenedo RL, McDevitt TC (2009) Engineering the embryoid body microenvironment to direct embryonic stem cell differentiation. Biotechnol Prog 25:43–51PubMedCentralPubMedCrossRef
26.
Zurück zum Zitat Mohr JC, Zhang J, Azarin SM, Soerens AG, de Pablo JJ, Thomson JA, Lyons GE, Palecek SP, Kamp TJ (2010) The microwell control of embryoid body size in order to regulate cardiac differentiation of human embryonic stem cells. Biomaterials 31:1885–1893PubMedCentralPubMedCrossRef Mohr JC, Zhang J, Azarin SM, Soerens AG, de Pablo JJ, Thomson JA, Lyons GE, Palecek SP, Kamp TJ (2010) The microwell control of embryoid body size in order to regulate cardiac differentiation of human embryonic stem cells. Biomaterials 31:1885–1893PubMedCentralPubMedCrossRef
27.
Zurück zum Zitat Bauwens CL, Peerani R, Niebruegge S, Woodhouse KA, Kumacheva E, Husain M, Zandstra PW (2008) Control of human embryonic stem cell colony and aggregate size heterogeneity influences differentiation trajectories. Stem Cells 26:2300–2310PubMedCrossRef Bauwens CL, Peerani R, Niebruegge S, Woodhouse KA, Kumacheva E, Husain M, Zandstra PW (2008) Control of human embryonic stem cell colony and aggregate size heterogeneity influences differentiation trajectories. Stem Cells 26:2300–2310PubMedCrossRef
28.
Zurück zum Zitat Antonchuk J (2013) Formation of embryoid bodies from human pluripotent stem cells using AggreWell plates. Methods Mol Biol 946:523–533PubMedCrossRef Antonchuk J (2013) Formation of embryoid bodies from human pluripotent stem cells using AggreWell plates. Methods Mol Biol 946:523–533PubMedCrossRef
29.
Zurück zum Zitat Preda MB, Burlacu A, Simionescu M (2013) Defined-size embryoid bodies formed in the presence of serum replacement increases the efficiency of the cardiac differentiation of mouse embryonic stem cells. Tissue Cell 45:54–60PubMedCrossRef Preda MB, Burlacu A, Simionescu M (2013) Defined-size embryoid bodies formed in the presence of serum replacement increases the efficiency of the cardiac differentiation of mouse embryonic stem cells. Tissue Cell 45:54–60PubMedCrossRef
30.
Zurück zum Zitat Ren Y, Lee MY, Schliffke S, Paavola J, Amos PJ, Ge X, Ye M, Zhu S, Senyei G, Lum L, Ehrlich BE, Qyang Y (2011) Small molecule Wnt inhibitors enhance the efficiency of BMP-4-directed cardiac differentiation of human pluripotent stem cells. J Mol Cell Cardiol 51:280–287PubMedCentralPubMedCrossRef Ren Y, Lee MY, Schliffke S, Paavola J, Amos PJ, Ge X, Ye M, Zhu S, Senyei G, Lum L, Ehrlich BE, Qyang Y (2011) Small molecule Wnt inhibitors enhance the efficiency of BMP-4-directed cardiac differentiation of human pluripotent stem cells. J Mol Cell Cardiol 51:280–287PubMedCentralPubMedCrossRef
31.
Zurück zum Zitat Gutstein DE, Marks AR (1997) Role of inositol 1,4,5-trisphosphate receptors in regulating apoptotic signaling and heart failure. Heart Vessels Suppl 12:53–57CrossRef Gutstein DE, Marks AR (1997) Role of inositol 1,4,5-trisphosphate receptors in regulating apoptotic signaling and heart failure. Heart Vessels Suppl 12:53–57CrossRef
32.
Zurück zum Zitat Satin J, Itzhaki I, Rapoport S, Schroder EA, Izu L, Arbel G, Beyar R, Balke CW, Schiller J, Gepstein L (2008) Calcium handling in human embryonic stem cell-derived cardiomyocytes. Stem Cells 26:1961–1972PubMedCrossRef Satin J, Itzhaki I, Rapoport S, Schroder EA, Izu L, Arbel G, Beyar R, Balke CW, Schiller J, Gepstein L (2008) Calcium handling in human embryonic stem cell-derived cardiomyocytes. Stem Cells 26:1961–1972PubMedCrossRef
33.
Zurück zum Zitat Shibata S, Hollander PB (1967) Effects of caffeine on the contractility and membrane potentials of rat atrium. Experientia 23:559PubMedCrossRef Shibata S, Hollander PB (1967) Effects of caffeine on the contractility and membrane potentials of rat atrium. Experientia 23:559PubMedCrossRef
34.
Zurück zum Zitat Shinohara T, Kim D, Joung B, Maruyama M, Vembaiyan K, Back TG, Wayne Chen SR, Chen PS, Lin SF (2013) Carvedilol analog modulates both basal and stimulated sinoatrial node automaticity. Heart Vessels doi. doi:10.1007/s00380-013-0378-2 Shinohara T, Kim D, Joung B, Maruyama M, Vembaiyan K, Back TG, Wayne Chen SR, Chen PS, Lin SF (2013) Carvedilol analog modulates both basal and stimulated sinoatrial node automaticity. Heart Vessels doi. doi:10.​1007/​s00380-013-0378-2
Metadaten
Titel
Forced aggregation and defined factors allow highly uniform-sized embryoid bodies and functional cardiomyocytes from human embryonic and induced pluripotent stem cells
verfasst von
Martin Pesl
Ivana Acimovic
Jan Pribyl
Renata Hezova
Aleksandra Vilotic
Jeremy Fauconnier
Jan Vrbsky
Peter Kruzliak
Petr Skladal
Tomas Kara
Vladimir Rotrekl
Alain Lacampagne
Petr Dvorak
Albano C. Meli
Publikationsdatum
01.11.2014
Verlag
Springer Japan
Erschienen in
Heart and Vessels / Ausgabe 6/2014
Print ISSN: 0910-8327
Elektronische ISSN: 1615-2573
DOI
https://doi.org/10.1007/s00380-013-0436-9

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