In Vitro Modeling of Paraxial and Lateral Mesoderm Differentiation Reveals Early Reversibility

Sakurai, Hiroyuki; Era, Takumi; Jakt, Lars Martin; Okada, Mitsuhiro; Nakai, Shigeru; Nishikawa, Satomi; Nishikawa, Shin Ichi

doi:10.1634/stemcells.2005-0256

Cited by 124 publications

(161 citation statements)

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“…Three days later, cells were gently harvested and analyzed for the hematopoietic colony-forming activity as described previously [15,18,23]. Endothelial colony formation was performed as described previously [18,23].…”

Section: Hematopoietic and Endothelial Colony Formationmentioning

confidence: 99%

“…Under the serum-containing condition, induction of ES cell differentiation was performed in either ␣-minimal essential medium (␣MEM) or SF-O3 supplemented with 10% fetal calf serum (FCS) and 50 M 2-mercaptoethanol (2ME) as described previously [18,19].…”

Section: In Vitro Es Cell Differentiationmentioning

confidence: 99%

“…For induction of VE-cadherin ϩ ECs and CD45 ϩ hematopoietic cells, 2 ϫ 10 5 FACS-purified cells were recultured in one well of six-well type IV collagen-coated plates with ␣MEM containing 10% FCS, 50 M 2ME, 3 ng/ml vascular endothelial growth factor, and 100 ng/ml stem cell factor (Kit ligands) [14,18]. Three days later, the cells were harvested and collected for the examination of expression of the surface markers VE-cadherin and CD45.…”

Section: In Vitro Es Cell Differentiationmentioning

confidence: 99%

“…Rat monoclonal antibodies (MoAbs) APA5 (anti-PDGFR␣), ECCD2 (anti-E-cadherin), AVAS12 (anti-VEGFR2), APB5 (anti-PDGFR␤), and VECD1 (anti-VE-cadherin) were prepared as reported previously [14,18,19]. Anti-Cadherin 11 MoAb was kindly provided by Dr. A. Kudo (Tokyo Institute of Technology, Yokohama, Japan).…”

Section: Antibodies and Cell Staining For Facsmentioning

confidence: 99%

“…To overcome the problem of ES cell differentiation cultures in which multiple events are occurring simultaneously, we have attempted to establish molecular markers for defining the cell lineages generated in cultures [14 -16]. Using such markers for identifying the point of lineage divergence, we were able to show that vascular endothelial growth factor receptor 2 (VEGFR2; FLK1) and platelet-derived growth factor receptor-␣ (PDGFR␣) are useful surface markers that prospectively distinguish lateral and paraxial mesoderm, respectively [17,18]. These markers enabled us to further dissect the course of mesoderm differentiation to HPCs, revealing the presence of two distinct pathways for HPC differentiation, one leading directly from VEGFR2 ϩ mesoderm and the other from VEGFR2 ϩ vascular endothelial cell cadherin (VE-cadherin ϩ ) ECs derived from VEGFR2 ϩ mesoderm [14].…”

Section: Introductionmentioning

confidence: 99%

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Multiple Mesoderm Subsets Give Rise to Endothelial Cells, Whereas Hematopoietic Cells Are Differentiated Only from a Restricted Subset in Embryonic Stem Cell Differentiation Culture

Era¹,

Izumi²,

Hayashi³

et al. 2007

Stem Cells

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In the developing mouse, vascular endothelial cell (EC) and hematopoietic cell (HPC) lineages are two initial cell lineages that diverge from mesodermal cells, which have been roughly subdivided into three subtypes according to their geographical location: the organizer, embryonic mesoderm in the primitive streak, and extraembryonic mesoderm during gastrulation. Although the initial progenitors that become the two lineages appear in both vascular endothelial growth factor receptor 2 ؉ (VEGFR2 ؉ ) lateral and extraembryonic mesoderm, little is known about the underlying molecular events that regulate the derivation of ECs and HPCs. Here, we describe an experimental system consisting of two types of embryonic stem cell lines capable of distinguishing between organizer and the middle section of the primitive streak region. Using this system, we were able to establish a defined culture condition that can separately induce distinct types of mesoderm. Although we were able to differentiate ECs from all mesoderm subsets, however, the potential of HPCs was restricted to the VEGFR2 ؉ cells derived from primitive streak-type mesodermal cells. We also show that the culture condition for the progenitors of primitive erythrocytes is separated from that for the progenitors of definitive erythrocytes. These results suggest the dominant role of extrinsic regulation during diversification of mesoderm. STEM CELLS 2008;26:401-411 Disclosure of potential conflicts of interest is found at the end of this article.

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Section: Hematopoietic and Endothelial Colony Formationmentioning

confidence: 99%

Section: In Vitro Es Cell Differentiationmentioning

confidence: 99%

Section: In Vitro Es Cell Differentiationmentioning

confidence: 99%

Section: Antibodies and Cell Staining For Facsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Multiple Mesoderm Subsets Give Rise to Endothelial Cells, Whereas Hematopoietic Cells Are Differentiated Only from a Restricted Subset in Embryonic Stem Cell Differentiation Culture

Era¹,

Izumi²,

Hayashi³

et al. 2007

Stem Cells

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High‐Throughput Screening Assay for Embryoid Body Differentiation of Human Embryonic Stem Cells

Outten

Gadue

French

et al. 2012

CP Stem Cell Biology

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Serum‐free human pluripotent stem cell media offer the potential to develop reproducible clinically applicable differentiation strategies and protocols. The vast array of possible growth factor and cytokine combinations for media formulations makes differentiation protocol optimization both labor and cost‐intensive. This unit describes a 96‐well plate, 4‐color flow cytometry–based screening assay to optimize pluripotent stem cell differentiation protocols. We provide conditions both to differentiate human embryonic stem cells (hESCs) into the three primary germ layers, ectoderm, endoderm, and mesoderm, and to utilize flow cytometry to distinguish between them. This assay exhibits low inter‐well variability and can be utilized to efficiently screen a variety of media formulations, reducing cost, incubator space, and labor. Protocols can be adapted to a variety of differentiation stages and lineages. Curr. Protoc. Stem Cell Biol. 20:1D.6.1‐1D.6.13. © 2012 by John Wiley & Sons, Inc.

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PDGF Receptor Alpha+ Mesoderm Contributes to Endothelial and Hematopoietic Cells in Mice

Ding¹,

Tanaka²,

Hayashi³

et al. 2013

Developmental Dynamics

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Background: Early mesoderm can be classified into Flk-1+ or PDGF receptor alpha (PDGFRα)+ population, grossly representing lateral and paraxial mesoderm, respectively. It has been demonstrated that all endothelial (EC) and hematopoietic (HPC) cells are derived from Flk-1+ cells. Although PDGFRα+ cells give rise to ECs/HPCs in in vitro ES differentiation, whether PDGFRα+ population can become hemato-endothelial lineages has not been proved in mouse embryos. Results: Using PDGFRαMerCreMer mice, PDGFRα+ early mesoderm was shown to contribute to endothelial cells including hemogenic ECs, fetal liver B lymphocytes, and Lin-Kit+Sca-1+ (KSL) cells. Contribution of PDGFRα+ mesoderm into ECs and HPCs was limited until E8.5, indicating that PDGFRα+/Flk-1+ population that exists until E8.5 may be the source for hemato-endothelial lineages from PDGFRα+ population. The functional significance of PDGFRα+ mesoderm in vascular development and hematopoiesis was confirmed by genetic deletion of Etv2 or restoration of Runx1 in PDGFRα+ cells. Etv2 deletion and Runx1 restoration in PDGFRα+ cells resulted in abnormal vascular remodeling and rescue of fetal liver CD45+ and Lin-Kit+Sca-1+ (KSL) cells, respectively. Conclusions: Endothelial and hematopoietic cells can be derived from PDGFRα+ early mesoderm in mice. PDGFRα+ mesoderm is functionally significant in vascular development and hematopoiesis from phenotype analysis of genetically modified embryos. Developmental Dynamics 242:254–268, 2013. © 2013 Wiley Periodicals, Inc.Key FindingsPDGF receptor alpha–positive mesoderm contributes to endothelial and hematopoietic cells in physiological mouse embryogenesis.PDGF receptor alpha–positive mesoderm in early embryo is distinct from yolk sac blood island mesoderm, representing a source of hematopoietic cells on the embryo proper side.Genetic manipulation of Etv2 or Runx1 in PDGF receptor alpha–positive mesoderm demonstrates the functional significance of this mesoderm subset in vascular development and hematopoiesis.

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In Vitro Modeling of Paraxial and Lateral Mesoderm Differentiation Reveals Early Reversibility

Cited by 124 publications

References 50 publications

Multiple Mesoderm Subsets Give Rise to Endothelial Cells, Whereas Hematopoietic Cells Are Differentiated Only from a Restricted Subset in Embryonic Stem Cell Differentiation Culture

Multiple Mesoderm Subsets Give Rise to Endothelial Cells, Whereas Hematopoietic Cells Are Differentiated Only from a Restricted Subset in Embryonic Stem Cell Differentiation Culture

High‐Throughput Screening Assay for Embryoid Body Differentiation of Human Embryonic Stem Cells

PDGF Receptor Alpha+ Mesoderm Contributes to Endothelial and Hematopoietic Cells in Mice

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