Structural differentiation, proliferation, and association of human embryonic stem cell-derived cardiomyocytes in vitro and in their extracardiac tissues

Cui, Li; Johkura, Kohei; Takei, Shunsuke; Ogiwara, Naoko; Sasaki, Katsunori

doi:10.1016/j.jsb.2006.11.009

Cited by 21 publications

(19 citation statements)

References 32 publications

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“…At early stages of differentiation, they developed intercellular junctions to form functional colonies. A localization of the fascia adherens protein cadherin and the gap junction protein connexin 43 at the cellular junctions means that there was a mechanical and electrical connection of the cardiomyocytes, consistent with our previous report on the differentiation of ES cell-derived cardiomyocyte associations (7). Although once isolated, it was difficult for the relatively differentiated cardiomyocytes to reassociate, judging from the impaired immunoreactivity for connexin 43 in the aggregates.…”

Section: Discussionsupporting

confidence: 90%

“…The amount of cTnI and ANP proteins increased sharply between days 7 and 14. Previous studies, including ours, showed that the proliferative activity of induced cardiomyocytes remains high at this stage of in vitro differentiation (7,20). The increase in the cardiac protein content confirms both the proliferation and maturation of cardiomyocytes in this period.…”

Section: Discussionsupporting

confidence: 81%

“…We have experienced that the undifferentiated human ES cells can be maintained on feeder cell layer in knockout serum replacement (KSR) medium, whereas the formation of EBs capable of generating cardiomyocytes requires FBS (7). When FBS was replaced by KSR in cultures of mouse (34), monkey (13), and human (2) ES cells, a significant decrease in the cardiac differentiation occurred.…”

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confidence: 99%

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Bone morphogenetic protein-4 promotes induction of cardiomyocytes from human embryonic stem cells in serum-based embryoid body development

Takei¹,

Ichikawa²,

Johkura³

et al. 2009

American Journal of Physiology-Heart and Circulatory Physiology

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Cardiomyocytes derived from human embryonic stem (ES) cells are a potential source for cell-based therapy for heart diseases. We studied the effect of bone morphogenetic protein (BMP)-4 in the presence of fetal bovine serum (FBS) on cardiac induction from human H1 ES cells during embryoid body (EB) development. Suspension culture for 4 days with 20% FBS produced the best results for the differentiation of early mesoderm and cardiomyocytes. The addition of Noggin reduced the incidence of beating EBs from 23.6% to 5.3%, which indicated the involvement of BMP signaling in the spontaneous cardiac differentiation. In this condition, treatment with 12.5–25 ng/ml BMP-4 during the 4-day suspension optimally promoted the cardiomyocyte differentiation. The incidence of beating EBs at 25 ng/ml BMP-4 reached 95.8% on day 6 of expansion and then plateaued until day 20. In real-time PCR analysis, the cardiac development-related genes MESP1 and Nkx2.5 were upregulated in the EB outgrowths by 25 ng/ml BMP-4. The activation of BMP signaling in EBs was confirmed by the increase in the phosphorylation of Smad1/5/8 and by the nuclear localization of phospho-Smad1/5/8 and Smad4. The addition of 150 ng/ml Noggin considerably decreased the incidence of beating EBs and Nkx2.5 expression, and Noggin alone increased Nestin expression and neural differentiation in EB outgrowths. The cardiomyocytes induced by 25 ng/ml BMP-4 showed proper cell biological characteristics and a course of differentiation as judged from isoproterenol administration, gene expression, protein assay, immunoreactivity, and subcellular structures. No remarkable change in the extent of apoptosis and proliferation in the cardiomyocytes was observed by BMP-4 treatment. These findings showed that BMP-4 in combination with FBS at the appropriate time and concentrations significantly promotes cardiomyocyte induction from human ES cells.

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Section: Discussionsupporting

confidence: 90%

Section: Discussionsupporting

confidence: 81%

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confidence: 99%

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Bone morphogenetic protein-4 promotes induction of cardiomyocytes from human embryonic stem cells in serum-based embryoid body development

Takei¹,

Ichikawa²,

Johkura³

et al. 2009

American Journal of Physiology-Heart and Circulatory Physiology

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“…Beating clusters appeared within EBs on day 4 after plating, and continued to contract in the culture for up to 194 d, which is much longer than the 106 d previously reported by Cui et al [37]. We also noted rhythmic contraction driven by pacemaker-like cells within EBs, and found that the beating rates in 58% of cases were similar to the heartbeat rates of human adults.…”

Section: Discussionsupporting

confidence: 74%

Enhancement of cardiomyocyte differentiation from human embryonic stem cells

Wang

Chen

Song

et al. 2010

Sci. China Life Sci.

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Several approaches have been used to encourage the differentiation of cardiomyocytes from human embryonic stem cells. However, the differentiation efficiency is low, and appropriate culture protocols are needed to produce adequate numbers of cardiomyocytes for therapeutic cell transplantation. This study investigated the effects of serum on cardiomyocyte differentiation in suspension culture medium during embryoid body (EB) formation by human embryonic stem cells. The addition of ascorbic acid, dimethylsulfoxide and 5-aza-2'-deoxycytidine during days 5-7 at the EB-forming stage resulted in an increase in the numbers of rhythmically contracting clusters of derived cardiomyocytes. Treatment with 0.1 mmol L(-1) ascorbic acid alone, or more notably in combination with 10 micromol L(-1) 5-aza-2'-deoxycytidine, induced the formation of beating cells within EBs. Most of the beating clusters had spontaneous contraction rates similar to those found in human adults, and their contractile activity lasted for up to 194 days.

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“…this is consistent with structural maturation in the esc-derived cardiomyocytes. 7 the analysis of spontaneous intracellular ca 2+ fluctuations in d7cMs and d21cMs showed that the mean rate of beating in d21cMs (51.5 ± 9.0 bpm, n = 6) was higher than in d7cMs (34.2 ± 8.2 bpm, n = 10) (Fig. 4A).…”

Section: Gene Expression Patterns and Ultrastructural Characteristicsmentioning

confidence: 95%

Gene Expression Profiling of Functional Murine Embryonic Stem Cell-Derived Cardiomyocytes and Comparison with Adult Heart: Profiling of Murine ESC-Derived Cardiomyocytes

et al. 2009

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although embryonic stem cell (esc)-derived cardiomyocytes may be a powerful tool in drug discovery, their potential has not yet been fully explored. nor has a detailed comparison with adult heart tissue been performed. We have developed a method for efficient production of cardiomyocyte-rich embryoid bodies (eBs) from murine escs. analysis of global gene expression profiles showed that eBs on day 7 and/or 21 of differentiation (d7cMs and d21cMs, respectively) were similar to adult heart tissue for genes categorized as regulators of muscle contraction or voltage-gated ion channel activity, although d21cMs were more mature than d7cMs for contractile components related to morphological structures. calcium and sodium channel blockers altered ca 2+ transients, and isoproterenol, a β-adrenergic compound, increased the rate of beating in d7cMs and d21cMs. our gene analytic system therefore enabled us to identify genes that are expressed in the physiological pathways associated with ion channels and structural components in d7cMs and d21cMs. We conclude that eBs might be of use for the basic screening of drugs that might affect contractile function through ion channels. (Journal of Biomolecular

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Structural differentiation, proliferation, and association of human embryonic stem cell-derived cardiomyocytes in vitro and in their extracardiac tissues

Cited by 21 publications

References 32 publications

Bone morphogenetic protein-4 promotes induction of cardiomyocytes from human embryonic stem cells in serum-based embryoid body development

Bone morphogenetic protein-4 promotes induction of cardiomyocytes from human embryonic stem cells in serum-based embryoid body development

Enhancement of cardiomyocyte differentiation from human embryonic stem cells

Gene Expression Profiling of Functional Murine Embryonic Stem Cell-Derived Cardiomyocytes and Comparison with Adult Heart: Profiling of Murine ESC-Derived Cardiomyocytes

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