scholarly journals Activin A Modulates CRIPTO-1/HNF4α+ Cells to Guide Cardiac Differentiation from Human Embryonic Stem Cells

2017 ◽  
Vol 2017 ◽  
pp. 1-17 ◽  
Author(s):  
Robin Duelen ◽  
Guillaume Gilbert ◽  
Abdulsamie Patel ◽  
Nathalie de Schaetzen ◽  
Liesbeth De Waele ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Fate ◽  
Heart Development ◽  
Embryoid Body ◽  
Embryonic Stem ◽  
Activin A ◽  
Embryoid Bodies ◽  
Cardiac Differentiation ◽  
Potential Applications

The use of human pluripotent stem cells in basic and translational cardiac research requires efficient differentiation protocols towards cardiomyocytes. In vitro differentiation yields heterogeneous populations of ventricular-, atrial-, and nodal-like cells hindering their potential applications in regenerative therapies. We described the effect of the growth factor Activin A during early human embryonic stem cell fate determination in cardiac differentiation. Addition of high levels of Activin A during embryoid body cardiac differentiation augmented the generation of endoderm derivatives, which in turn promoted cardiomyocyte differentiation. Moreover, a dose-dependent increase in the coreceptor expression of the TGF-β superfamily member CRIPTO-1 was observed in response to Activin A. We hypothesized that interactions between cells derived from meso- and endodermal lineages in embryoid bodies contributed to improved cell maturation in early stages of cardiac differentiation, improving the beating frequency and the percentage of contracting embryoid bodies. Activin A did not seem to affect the properties of cardiomyocytes at later stages of differentiation, measuring action potentials, and intracellular Ca2+ dynamics. These findings are relevant for improving our understanding on human heart development, and the proposed protocol could be further explored to obtain cardiomyocytes with functional phenotypes, similar to those observed in adult cardiac myocytes.

scholarly journals Analysis of cardiac differentiation at single cell resolution reveals a requirement of hypertrophic signaling for HOPX transcription

2017 ◽  
Author(s):  
Clayton E Friedman ◽  
Quan Nguyen ◽  
Samuel W Lukowski ◽  
Han Sheng Chiu ◽  
Abbigail Helfer ◽  
...  
Keyword(s):  
Stem Cells ◽  
Single Cell ◽  
Cell Fate ◽  
Heart Development ◽  
Time Course ◽  
Cardiac Differentiation ◽  
Transcriptional Networks ◽  
Hypertrophic Signaling

AbstractDifferentiation into diverse cell lineages requires the orchestration of gene regulatory networks guiding diverse cell fate choices. Utilizing human pluripotent stem cells, we measured expression dynamics of 17,718 genes from 43,168 cells across five time points over a thirty day time-course of in vitro cardiac-directed differentiation. Unsupervised clustering and lineage prediction algorithms were used to map fate choices and transcriptional networks underlying cardiac differentiation. We leveraged this resource to identify strategies for controlling in vitro differentiation as it occurs in vivo. HOPX, a non-DNA binding homeodomain protein essential for heart development in vivo was identified as dys-regulated in in vitro derived cardiomyocytes. Utilizing genetic gain and loss of function approaches, we dissect the transcriptional complexity of the HOPX locus and identify the requirement of hypertrophic signaling for HOPX transcription in hPSC-derived cardiomyocytes. This work provides a single cell dissection of the transcriptional landscape of cardiac differentiation for broad applications of stem cells in cardiovascular biology.

scholarly journals Activation of the amino acid response modulates lineage specification during differentiation of murine embryonic stem cells

2013 ◽  
Vol 305 (3) ◽  
pp. E325-E335 ◽  
Author(s):  
Jixiu Shan ◽  
Takashi Hamazaki ◽  
Tiffany A. Tang ◽  
Naohiro Terada ◽  
Michael S. Kilberg
Keyword(s):  
Stem Cells ◽  
Amino Acid ◽  
Embryonic Stem Cells ◽  
Embryoid Body ◽  
Embryonic Stem ◽  
Embryoid Bodies ◽  
Specific Cell ◽  
Lineage Specification ◽  
Amino Acid Response

In somatic cells, a collection of signaling pathways activated by amino acid limitation have been identified and referred to as the amino acid response (AAR). Despite the importance of possible detrimental effects of nutrient limitation during in vitro culture, the AAR has not been investigated in embryonic stem cells (ESC). AAR activation caused the expected increase in transcription factors that mediate specific AAR pathways, as well as the induction of asparagine synthetase, a terminal AAR target gene. Neither AAR activation nor stable knockdown of activating transcription factor (Atf) 4, a transcriptional mediator of the AAR, adversely affected ESC self-renewal or pluripotency. Low-level induction of the AAR over a 12-day period of embryoid body differentiation did alter lineage specification such that the primitive endodermal, visceral endodermal, and endodermal lineages were favored, whereas mesodermal and certain ectodermal lineages were suppressed. Knockdown of Atf4 further enhanced the AAR-induced increase in endodermal formation, suggesting that this phenomenon is mediated by an Atf4-independent mechanism. Collectively, the results indicate that, during differentiation of mouse embryoid bodies in culture, the availability of nutrients, such as amino acids, can influence the formation of specific cell lineages.

scholarly journals Reorganization of Metabolism during Cardiomyogenesis Implies Time-Specific Signaling Pathway Regulation

2021 ◽  
Vol 22 (3) ◽  
pp. 1330
Author(s):  
María Julia Barisón ◽  
Isabela Tiemy Pereira ◽  
Anny Waloski Robert ◽  
Bruno Dallagiovanna
Keyword(s):  
Stem Cells ◽  
Embryoid Body ◽  
Embryonic Stem ◽  
Cell Lineage ◽  
Cardiac Differentiation ◽  
Lipid Homeostasis ◽  
Specific Cell ◽  
Regulatory Pathways ◽  
Time Specific

Understanding the cell differentiation process involves the characterization of signaling and regulatory pathways. The coordinated action involved in multilevel regulation determines the commitment of stem cells and their differentiation into a specific cell lineage. Cellular metabolism plays a relevant role in modulating the expression of genes, which act as sensors of the extra-and intracellular environment. In this work, we analyzed mRNAs associated with polysomes by focusing on the expression profile of metabolism-related genes during the cardiac differentiation of human embryonic stem cells (hESCs). We compared different time points during cardiac differentiation (pluripotency, embryoid body aggregation, cardiac mesoderm, cardiac progenitor and cardiomyocyte) and showed the immature cell profile of energy metabolism. Highly regulated canonical pathways are thoroughly discussed, such as those involved in metabolic signaling and lipid homeostasis. We reveal the critical relevance of retinoic X receptor (RXR) heterodimers in upstream retinoic acid metabolism and their relationship with thyroid hormone signaling. Additionally, we highlight the importance of lipid homeostasis and extracellular matrix component biosynthesis during cardiomyogenesis, providing new insights into how hESCs reorganize their metabolism during in vitro cardiac differentiation.

VEGF-A165 augments erythropoietic development from human embryonic stem cells

Blood ◽  
2004 ◽  
Vol 103 (7) ◽  
pp. 2504-2512 ◽  
Author(s):  
Chantal Cerdan ◽  
Anne Rouleau ◽  
Mickie Bhatia
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Cell Fate ◽  
Human Embryonic Stem Cells ◽  
Embryoid Body ◽  
Embryonic Stem ◽  
Vascular Endothelial ◽  
Erythroid Progenitor ◽  
Endothelial Growth Factor

Abstract Combinations of hematopoietic cytokines and the ventral mesoderm inducer BMP-4 have recently been shown to augment hematopoietic cell fate of human embryonic stem cells (hESCs) during embryoid body (EB) development. However, factors capable of regulating lineage commitment of hESC-derived hematopoiesis have yet to be reported. Here we show that vascular endothelial growth factor (VEGF-A165) selectively promotes erythropoietic development from hESCs. Effects of VEGF-A165 were dependent on the presence of hematopoietic cytokines and BMP-4, and could be augmented by addition of erythropoietin (EPO). Treatment of human EBs with VEGF-A165 increased the frequency of cells coexpressing CD34 and the VEGF-A165 receptor KDR, as well as cells expressing erythroid markers. Although fetal/adult globins were unaffected, VEGF-A165 induced the expression of embryonic zeta (ζ) and epsilon (ϵ) globins, and was accompanied by expression of the hematopoietic transcription factor SCL/Tal-1. In addition to promoting erythropoietic differentiation from hESCs, the presence of VEGF-A165 enhanced the in vitro self-renewal potential of primitive hematopoietic cells capable of erythroid progenitor capacity. Our study demonstrates a role for VEGF-A165 during erythropoiesis of differentiating hESCs, thereby providing the first evidence for a factor capable of regulating hematopoietic lineage development of hESCs.

scholarly journals Thyrotropin-Independent Induction of Thyroid Endoderm from Embryonic Stem Cells by Activin A

Endocrinology ◽  
2008 ◽  
Vol 150 (4) ◽  
pp. 1970-1975 ◽  
Author(s):  
Risheng Ma ◽  
Rauf Latif ◽  
Terry F. Davies
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Embryonic Stem ◽  
Activin A ◽  
Embryoid Bodies ◽  
Sodium Iodide Symporter ◽  
Thyroid Cells ◽  
Gene And Protein Expression ◽  
Igf I

To model the differentiation of thyroid epithelial cells, we examined embryoid bodies derived from undifferentiated murine embryonic stem cells treated with activin A to induce endoderm differentiation, the germ layer from which thyroid cells occur. The resulting endodermal cells were then further exposed to TSH and/or IGF-I for up to 21 d. Oct-4 and REX1 expression, required to sustain stem cell self-renewal and pluripotency, were appropriately down-regulated, whereas GATA-4, and α-fetoprotein, both endodermal-specific markers, increased as the embryonic stem cells were exposed to activin A. By d 5 culture, TSH receptor (TSHR) and sodium iodide symporter (NIS) gene and protein expression were markedly induced. Cells isolated by the fluorescence-activated cell sorter simultaneously expressed not only TSHR and NIS proteins but also PAX8 mRNA, an expression pattern unique to thyroid cells and expected in committed thyroid progenitor cells. Such expression continued until d 21 with no influence seen by the addition of TSH or IGF-I. The sequence of gene expression changes observed in these experiments demonstrated the emergence of definitive thyroid endoderm. The activin A induction of thyroid-specific markers, NIS and TSHR, occurred in the absence of TSH stimulation, and, therefore, the emergence of thyroid endoderm in vitro paralleled the emergence of thyroid cells in TSHR-knockout mice. Activin A is clearly a major regulator of thyroid endoderm.

AW551984: a novel regulator of cardiomyogenesis in pluripotent embryonic cells

2011 ◽  
Vol 437 (2) ◽  
pp. 345-355 ◽  
Author(s):  
Satoshi Yasuda ◽  
Tetsuya Hasegawa ◽  
Tetsuji Hosono ◽  
Mitsutoshi Satoh ◽  
Kei Watanabe ◽  
...  
Keyword(s):  
Stem Cells ◽  
Transcription Factor ◽  
Embryoid Body ◽  
Es Cells ◽  
Embryonic Stem ◽  
Cardiac Differentiation ◽  
Marker Genes ◽  
Embryonic Cells ◽  
Transcript Levels

An understanding of the mechanism that regulates the cardiac differentiation of pluripotent stem cells is necessary for the effective generation and expansion of cardiomyocytes as cell therapy products. In the present study, we have identified genes that modulate the cardiac differentiation of pluripotent embryonic cells. We isolated P19CL6 cell sublines that possess distinct properties in cardiomyogenesis and extracted 24 CMR (cardiomyogenesis-related candidate) genes correlated with cardiomyogenesis using a transcriptome analysis. Knockdown of the CMR genes by RNAi (RNA interference) revealed that 18 genes influence spontaneous contraction or transcript levels of cardiac marker genes in EC (embryonal carcinoma) cells. We also performed knockdown of the CMR genes in mouse ES (embryonic stem) cells and induced in vitro cardiac differentiation. Three CMR genes, AW551984, 2810405K02Rik (RIKEN cDNA 2810405K02 gene) and Cd302 (CD302 antigen), modulated the cardiac differentiation of both EC cells and ES cells. Depletion of AW551984 attenuated the expression of the early cardiac transcription factor Nkx2.5 (NK2 transcription factor related locus 5) without affecting transcript levels of pluripotency and early mesoderm marker genes during ES cell differentiation. Activation of Wnt/β-catenin signalling enhanced the expression of both AW551984 and Nkx2.5 in ES cells during embryoid body formation. Our findings indicate that AW551984 is a novel regulator of cardiomyogenesis from pluripotent embryonic cells, which links Wnt/β-catenin signalling to Nkx2.5 expression.

scholarly journals Cardiopoietic programming of embryonic stem cells for tumor-free heart repair

2007 ◽  
Vol 204 (2) ◽  
pp. 405-420 ◽  
Author(s):  
Atta Behfar ◽  
Carmen Perez-Terzic ◽  
Randolph S. Faustino ◽  
D. Kent Arrell ◽  
Denice M. Hodgson ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Embryonic Stem Cells ◽  
Nuclear Translocation ◽  
Embryonic Stem ◽  
Embryoid Bodies ◽  
Cardiac Differentiation ◽  
Cardiac Progenitors ◽  
Tnf Α

Embryonic stem cells have the distinct potential for tissue regeneration, including cardiac repair. Their propensity for multilineage differentiation carries, however, the liability of neoplastic growth, impeding therapeutic application. Here, the tumorigenic threat associated with embryonic stem cell transplantation was suppressed by cardiac-restricted transgenic expression of the reprogramming cytokine TNF-α, enhancing the cardiogenic competence of recipient heart. The in vivo aptitude of TNF-α to promote cardiac differentiation was recapitulated in embryoid bodies in vitro. The procardiogenic action required an intact endoderm and was mediated by secreted cardio-inductive signals. Resolved TNF-α–induced endoderm-derived factors, combined in a cocktail, secured guided differentiation of embryonic stem cells in monolayers produce cardiac progenitors termed cardiopoietic cells. Characterized by a down-regulation of oncogenic markers, up-regulation, and nuclear translocation of cardiac transcription factors, this predetermined population yielded functional cardiomyocyte progeny. Recruited cardiopoietic cells delivered in infarcted hearts generated cardiomyocytes that proliferated into scar tissue, integrating with host myocardium for tumor-free repair. Thus, cardiopoietic programming establishes a strategy to hone stem cell pluripotency, offering a tumor-resistant approach for regeneration.

scholarly journals Mitochondrial Fusion by M1 Promotes Embryoid Body Cardiac Differentiation of Human Pluripotent Stem Cells

2019 ◽  
Vol 2019 ◽  
pp. 1-12 ◽  
Author(s):  
Jarmon G. Lees ◽  
Anne M. Kong ◽  
Yi C. Chen ◽  
Priyadharshini Sivakumaran ◽  
Damián Hernández ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Embryoid Body ◽  
Mitochondrial Dynamics ◽  
Embryoid Bodies ◽  
Mitochondrial Fusion ◽  
Cardiac Differentiation ◽  
Patient Specific ◽  
Mitochondrial Morphology ◽  
Human Ipscs

Human induced pluripotent stem cells (iPSCs) can be differentiated in vitro into bona fide cardiomyocytes for disease modelling and personalized medicine. Mitochondrial morphology and metabolism change dramatically as iPSCs differentiate into mesodermal cardiac lineages. Inhibiting mitochondrial fission has been shown to promote cardiac differentiation of iPSCs. However, the effect of hydrazone M1, a small molecule that promotes mitochondrial fusion, on cardiac mesodermal commitment of human iPSCs is unknown. Here, we demonstrate that treatment with M1 promoted mitochondrial fusion in human iPSCs. Treatment of iPSCs with M1 during embryoid body formation significantly increased the percentage of beating embryoid bodies and expression of cardiac-specific genes. The pro-fusion and pro-cardiogenic effects of M1 were not associated with changes in expression of the α and β subunits of adenosine triphosphate (ATP) synthase. Our findings demonstrate for the first time that hydrazone M1 is capable of promoting cardiac differentiation of human iPSCs, highlighting the important role of mitochondrial dynamics in cardiac mesoderm lineage specification and cardiac development. M1 and other mitochondrial fusion promoters emerge as promising molecular targets to generate lineages of the heart from human iPSCs for patient-specific regenerative medicine.

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