Molecular and Functional Characterizations of Gastrula Organizer Cells Derived from Human Embryonic Stem Cells

Sharon, Nadav; Mor, Ishay; Golan‐Lev, Tamar; Fainsod, Abraham; Benvenisty, Nissim

doi:10.1002/stem.621

Cited by 34 publications

(25 citation statements)

References 43 publications

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“…For example, cells that resemble the gastrula organizer were identified in human EBs, and these cells induced a secondary axis when transplanted into frog embryos (Sharon et al, 2011). These findings suggest that human EBs can offer insights into the establishment of body axis in human embryos.…”

Section: Embryoid Bodies: a Potential Model Of Early Embryogenesismentioning

confidence: 83%

Human pluripotent stem cells: an emerging model in developmental biology

2013

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SummaryDevelopmental biology has long benefited from studies of classic model organisms. Recently, human pluripotent stem cells (hPSCs), including human embryonic stem cells and human induced pluripotent stem cells, have emerged as a new model system that offers unique advantages for developmental studies. Here, we discuss how studies of hPSCs can complement classic approaches using model organisms, and how hPSCs can be used to recapitulate aspects of human embryonic development 'in a dish'. We also summarize some of the recently developed genetic tools that greatly facilitate the interrogation of gene function during hPSC differentiation. With the development of high-throughput screening technologies, hPSCs have the potential to revolutionize gene discovery in mammalian development. Key words: Human pluripotent stem cells, Directed differentiation, Gene targeting, High-throughput screening IntroductionHuman pluripotent stem cells (hPSCs), which include human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), can self-renew indefinitely in culture while maintaining the ability to become almost any cell type in the human body (Takahashi et al., 2007;Thomson et al., 1998;Yu et al., 2007). The potential of using hPSCs for cell replacement therapy and disease modeling has been discussed extensively (Wu and Hochedlinger, 2011). This Review focuses on an equally important, yet often overlooked, aspect: using hPSCs to gain insights into human embryonic development. Although this potential has been long recognized (Keller, 2005;Pera and Trounson, 2004), it is only beginning to be realized, owing to advances in both in vitro differentiation approaches and genetic manipulation tools in hPSCs. In this Review, we summarize the latest progress in this nascent, yet rapidly advancing, field and discuss future prospects and potential challenges of using hPSCs for studies of developmental biology. First, the strengths and limitations of classic model organisms are discussed to highlight the need for a new model. Second, we illustrate how hPSCs can be used to recapitulate defined steps of embryogenesis, and we discuss how hPSC-based studies can lead to novel insights into human development. Next, we review new genetic tools that can be applied to interrogate gene function during the in vitro differentiation of hPSCs. Finally, the potential of using hPSCs for discovery-driven research is discussed.This Review focuses primarily on work performed using hPSCs, with occasional references to studies using mouse pluripotent stem cells when comparable studies are not yet available in human cells; work from both hiPSCs and hESCs is discussed, although more examples stem from hESCs, owing to their more-frequent use in differentiation experiments. Nonetheless, because of the high degree of similarity between hESCs and hiPSCs (Yamanaka, 2012), it is likely that the general strategies and most conclusions will apply to both hESCs and hiPSCs. The need for a new model systemA main challenge in biology is to understa...

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Section: Embryoid Bodies: a Potential Model Of Early Embryogenesismentioning

confidence: 83%

Human pluripotent stem cells: an emerging model in developmental biology

2013

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“…To further clarify the relationship between the Wnt pathway and SP during BMSC differentiation, four groups were created: Group A was the control group, and the same amount of PBS was added; in Group B, the culture was incubated with SP (the concentration to be determined by the results of step one); in Group C, the culture was incubated with a mixture of SP and NK1 antagonist (1 μ M Sigma CP-96345 USA) [23, 34]; and in Group D, the culture was incubated with a mixture of SP and 0.2 μ g/mL DKK1 (0.2 μ g/mL Peprotech recombinant Human DKK-1 (Dickkopf-related protein-1), USA) [35]. To identify the first changes in the expression of BMP-2 and VEGF, observations were made on 1 day, 3 days, 5 days, and 7 days; these intervals were also employed to determine the effects of SP on the migration of the BMSCs.…”

Section: Methodsmentioning

confidence: 99%

Neuropeptide Substance P Improves Osteoblastic and Angiogenic Differentiation Capacity of Bone Marrow Stem CellsIn Vitro

Mei

Wang

et al. 2014

BioMed Research International

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Our previous work showed that implanting a sensory nerve or vascular bundle when constructing vascularized and neurotized bone could promote bone osteogenesis in tissue engineering. This phenomenon could be explained by the regulatory function of neuropeptides. Neuropeptide substance P (SP) has been demonstrated to contribute to bone growth by stimulating the proliferation and differentiation of bone marrow stem cells (BMSCs). However, there have been no prior studies on the association between Wnt signaling and the mechanism of SP in the context of BMSC differentiation. Our results have shown that SP could enhance the differentiation of BMSCs by activating gene and protein expression via the Wnt pathway and by translocating β-catenin, which can be inhibited by Wnt signaling blocker treatment or by the NK-1 antagonist. SP could also increase the growth factor level of bone morphogenetic protein-2 (BMP-2). Additionally, SP could enhance the migration ability of BMSCs, and the promotion of vascular endothelial growth factor (VEGF) expression by SP has been studied. In conclusion, SP could induce osteoblastic differentiation via the Wnt pathway and promote the angiogenic ability of BMSCs. These results indicate that a vascularized and neurotized tissue-engineered construct could be feasible for use in bone tissue engineering strategies.

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“…To find optimal concentration for promotion of cellular differentiation, the cells were divided into 4 groups according to different concentrations of SP (Sigma, S6883, St. Louis, MO, USA): Group 1, incubated with SP (10 −8 M); Group 2, incubated with SP (10 −9 M); Group 3 incubated with SP (10 −10 M); and group 4, the control group, treated with the same volume of PBS. To determine the effects of SP, NK1 antagonist, and DKK1 on the differentiation of MC3T3-E1 cells, cells were divided into 4 groups: Group A, incubated with SP (10 −8 M); Group B, incubated with a mixture of SP and NK1 antagonist (1 μM, Sigma, CP-96345) [37,38]; Group C, incubated with a mixture of SP and 0.2 μg/mL DKK1 (recombinant Human DKK-1, Peprotech, Rocky Hill, CT, USA) [39]; and Group D, the control group, added with the same amount of PBS.…”

Section: Methodsmentioning

confidence: 99%

Substance P Activates the Wnt Signal Transduction Pathway and Enhances the Differentiation of Mouse Preosteoblastic MC3T3-E1 Cells

Mei

Zou

et al. 2014

IJMS

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Recent experiments have explored the impact of Wnt/β-catenin signaling and Substance P (SP) on the regulation of osteogenesis. However, the molecular regulatory mechanisms of SP on the formation of osteoblasts is still unknown. In this study, we investigated the impact of SP on the differentiation of MC3T3-E1 cells. The osteogenic effect of SP was observed at different SP concentrations (ranging from 10−10 to 10−8 M). To unravel the underlying mechanism, the MC3T3-E1 cells were treated with SP after the pretreatment by neurokinin-1 (NK1) antagonists and Dickkopf-1 (DKK1) and gene expression levels of Wnt/β-catenin signaling pathway components, as well as osteoblast differentiation markers (collagen type I, alkaline phosphatase, osteocalcin, and Runx2), were measured using quantitative polymerase chain reaction (PCR). Furthermore, protein levels of Wnt/β-catenin signaling pathway were detected using Western blotting and the effects of SP, NK1 antagonist, and DKK1 on β-catenin activation were investigated by immunofluorescence staining. Our data indicated that SP (10−9 to 10−8 M) significantly up-regulated the expressions of osteoblastic genes. SP (10−8 M) also elevated the mRNA level of c-myc, cyclin D1, and lymphocyte enhancer factor-1 (Lef1), as well as c-myc and β-catenin protein levels, but decreased the expression of Tcf7 mRNA. Moreover, SP (10−8 M) promoted the transfer of β-catenin into nucleus. The effects of SP treatment were inhibited by the NK1 antagonist and DKK1. These findings suggest that SP may enhance differentiation of MC3T3-E1 cells via regulation of the Wnt/β-catenin signaling pathway.

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Molecular and Functional Characterizations of Gastrula Organizer Cells Derived from Human Embryonic Stem Cells

Cited by 34 publications

References 43 publications

Human pluripotent stem cells: an emerging model in developmental biology

Human pluripotent stem cells: an emerging model in developmental biology

Neuropeptide Substance P Improves Osteoblastic and Angiogenic Differentiation Capacity of Bone Marrow Stem CellsIn Vitro

Substance P Activates the Wnt Signal Transduction Pathway and Enhances the Differentiation of Mouse Preosteoblastic MC3T3-E1 Cells

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