Human embryonic stem cells: challenges and opportunities

2006 ◽  
Vol 18 (8) ◽  
pp. 839 ◽  
Author(s):  
Steven L. Stice ◽  
Nolan L. Boyd ◽  
Sujoy K. Dhara ◽  
Brian A. Gerwe ◽  
David W. Machacek ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Stem Cells ◽  
Cell Line ◽  
Cell Fate ◽  
Neural Development ◽  
Es Cells ◽  
Embryonic Stem ◽  
Normal Karyotype ◽  
Es Cell ◽  
Cell Therapies

Human and non-human primate embryonic stem (ES) cells are invaluable resources for developmental studies, pharmaceutical research and a better understanding of human disease and replacement therapies. In 1998, subsequent to the establishment of the first monkey ES cell line in 1995, the first human ES cell line was developed. Later, three of the National Institute of Health (NIH) lines (BG01, BG02 and BG03) were derived from embryos that would have been discarded because of their poor quality. A major challenge to research in this area is maintaining the unique characteristics and a normal karyotype in the NIH-registered human ES cell lines. A normal karyotype can be maintained under certain culture conditions. In addition, a major goal in stem cell research is to direct ES cells towards a limited cell fate, with research progressing towards the derivation of a variety of cell types. We and others have built on findings in vertebrate (frog, chicken and mouse) neural development and from mouse ES cell research to derive neural stem cells from human ES cells. We have directed these derived human neural stem cells to differentiate into motoneurons using a combination of developmental cues (growth factors) that are spatially and temporally defined. These and other human ES cell derivatives will be used to screen new compounds and develop innovative cell therapies for degenerative diseases.

An in situ assessment of the routes and extents of colonisation of the mouse embryo by embryonic stem cells and their descendants

Development ◽  
1990 ◽  
Vol 110 (4) ◽  
pp. 1241-1248 ◽  
Author(s):  
Y. Lallemand ◽  
P. Brulet
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Cell Line ◽  
Mouse Embryo ◽  
Es Cells ◽  
Embryonic Stem ◽  
Es Cell ◽  
Low Degree ◽  
Morula Stage

An embryonic stem (ES) cell line stably expressing lacZ under the control of an endogenous promoter has been isolated and used as a marker to follow the fate of ES cells injected into blastocysts and morulae, before midgestation. The results show a multisite pattern of blastocyst colonization by ES cells deposited into the blastocoel cavity and a low degree of mingling between ES cells and ICM cells. Furthermore, analysis of dispersal of ES cell descendants in postimplantation chimaeric embryos showed that colonization can be highly variable from one region of the embryo to another. In contrast, a high and reproducible degree of chimaerism was obtained when the ES cells were injected at the morula stage prior to ICM formation.

scholarly journals An Inducible Expression System of the Calcium-Activated Potassium Channel 4 to Study the Differential Impact on Embryonic Stem Cells

2011 ◽  
Vol 2011 ◽  
pp. 1-12 ◽  
Author(s):  
Stefan Liebau ◽  
Michael Tischendorf ◽  
Daniel Ansorge ◽  
Leonhard Linta ◽  
Marianne Stockmann ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Differentiation ◽  
Cell Line ◽  
Es Cells ◽  
Embryonic Stem ◽  
Stem Cell Differentiation ◽  
Pacemaker Cell ◽  
Es Cell ◽  
Sk Channel ◽  
Mouse Es Cell

Rationale. The family of calcium-activated potassium channels consists of four members with varying biological functions and conductances. Besides membrane potential modulation, SK channels have been found to be involved in cardiac pacemaker cell development from ES cells and morphological shaping of neural stem cells.Objective. Distinct SK channel subtype expression in ES cells might elucidate their precise impact during cardiac development. We chose SK channel subtype 4 as a potential candidate influencing embryonic stem cell differentiation.Methods. We generated a doxycycline inducible mouse ES cell line via targeted homologous recombination of a cassette expressing a bicistronic construct encoding SK4 and a fluorophore from the murine HPRT locus.Conclusion. We characterized the mouse ES cell line iSK4-AcGFP. The cassette is readily expressed under the control of doxycycline, and the overexpression of SK4 led to an increase in cardiac and pacemaker cell differentiation thereby serving as a unique tool to characterize the cell biological variances due to specific SK channel overexpression.

scholarly journals Isolation and Initial Application of a Novel Peptide That Specifically Recognizes the Neural Stem Cells Derived from Rhesus Monkey Embryonic Stem Cells

2010 ◽  
Vol 15 (6) ◽  
pp. 687-694 ◽  
Author(s):  
Wenxiu Zhao ◽  
Hang Yuan ◽  
Xing Xu ◽  
Lan Ma
Keyword(s):  
Stem Cells ◽  
Cell Differentiation ◽  
Rhesus Monkey ◽  
Neural Stem Cells ◽  
Phage Display ◽  
Es Cells ◽  
Embryonic Stem ◽  
Peptide Sequence ◽  
Enzyme Linked Immunosorbent Assay ◽  
Es Cell

The search for new receptor ligands is important in the study of embryonic stem (ES) cell differentiation processes. In this study, a novel peptide (HGE VPRFHAVHL) with a specific ability to bind with neural stem cells derived from rhesus monkey ES cells was successfully screened out using a Ph.D-12 peptide phage display library. High affinity and specificity of the HGE phage were shown in an enzyme-linked immunosorbent assay. The binding ability of the phage could be matched with that of a chemically synthesized peptide with a sequence identical to that displayed by the phage, indicating that this binding capability manifests a peptide sequence. Combined with quantum dots, the HGE peptide can be used as a direct tool to show optical imaging of specific binding on a single cell membrane. Further results of Western blot showed that the HGE peptide interacted with 48/34-kDa proteins on the membrane of neural stem cells. This work is the first time that a phage display technique has been applied in ES cell differentiation studies. The findings extend the utilization of a targeting agent for neural stem cells and can also be used as a research tool in studying other cell lineages derived from ES cells.

scholarly journals Mouse embryonic stem cells switch migratory behaviour during early differentiation

2020 ◽  
Author(s):  
Irene M. Aspalter ◽  
Wolfram Pönisch ◽  
Kevin J. Chalut ◽  
Ewa K. Paluch
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Cell Fate ◽  
Es Cells ◽  
Embryonic Stem ◽  
Mouse Embryonic Stem Cells ◽  
Migratory Behaviour ◽  
Es Cell ◽  
Early Differentiation ◽  
Cell State

AbstractDevelopment relies on a series of precisely orchestrated cell fate changes. While studies of fate transitions often focus on changes in gene regulatory networks, most transitions are also associated with changes in cell shape and cell behaviour. Here, we investigate changes in migratory behaviour in mouse embryonic stem (ES) cells during their first developmental fate transition, exit from ES cell state. We show that naïve pluripotent ES cells cannot efficiently migrate on 2-dimensional substrates but are able to migrate in an amoeboid fashion when placed in confinement. Exit from ES cell state, typically characterised by enhanced cell spreading, is associated with decreased migration in confinement and acquisition of mesenchymal-like migration on 2D substrates. Interestingly, confined, amoeboid-like migration of ES cells strongly depends on Myosin IIA, but not Myosin IIB. In contrast mesenchymal-like migration of cells exiting the ES cell state does not depend on Myosin motor activity but relies on the activity of the Arp2/3 complex. Together, our data suggest that during early differentiation, cells undergo a switch in the regulation of the actin cytoskeleton, leading to a transition from amoeboid-to mesenchymal-like migration.Summary statementNaïve mouse embryonic stem cells display amoeboid-like migration in confinement, but switch to mesenchymal-like migration as they exit the ES cell state.

Germ line transmission of an inactive N-myc allele generated by homologous recombination in mouse embryonic stem cells

1990 ◽  
Vol 10 (12) ◽  
pp. 6755-6758
Author(s):  
B R Stanton ◽  
S W Reid ◽  
L F Parada
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Homologous Recombination ◽  
Embryonic Development ◽  
Cell Lines ◽  
Germ Line ◽  
Es Cells ◽  
Embryonic Stem ◽  
Es Cell ◽  
Germ Line Transmission

We have disrupted one allele of the N-myc locus in mouse embryonic stem (ES) cells by using homologous recombination techniques and have obtained germ line transmission of null N-myc ES cell lines with transmission of the null N-myc allele to the offspring. The creation of mice with a deficient N-myc allele will allow the generation of offspring bearing null N-myc alleles in both chromosomes and permit study of the role that this proto-oncogene plays in embryonic development.

scholarly journals Designer blood: creating hematopoietic lineages from embryonic stem cells

Blood ◽  
2006 ◽  
Vol 107 (4) ◽  
pp. 1265-1275 ◽  
Author(s):  
Abby L. Olsen ◽  
David L. Stachura ◽  
Mitchell J. Weiss
Keyword(s):  
Stem Cells ◽  
Es Cells ◽  
Embryonic Stem ◽  
Basic Research ◽  
Cell Types ◽  
Patient Specific ◽  
Es Cell ◽  
Blood Disorders ◽  
Hematopoietic Stem

Embryonic stem (ES) cells exhibit the remarkable capacity to become virtually any differentiated tissue upon appropriate manipulation in culture, a property that has been beneficial for studies of hematopoiesis. Until recently, the majority of this work used murine ES cells for basic research to elucidate fundamental properties of blood-cell development and establish methods to derive specific mature lineages. Now, the advent of human ES cells sets the stage for more applied pursuits to generate transplantable cells for treating blood disorders. Current efforts are directed toward adapting in vitro hematopoietic differentiation methods developed for murine ES cells to human lines, identifying the key interspecies differences in biologic properties of ES cells, and generating ES cell-derived hematopoietic stem cells that are competent to repopulate adult hosts. The ultimate medical goal is to create patient-specific and generic ES cell lines that can be expanded in vitro, genetically altered, and differentiated into cell types that can be used to treat hematopoietic diseases.

scholarly journals Neural Induction, Neural Fate Stabilization, and Neural Stem Cells

2002 ◽  
Vol 2 ◽  
pp. 1147-1166 ◽  
Author(s):  
Sally A. Moody ◽  
Hyun-Soo Je
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Neural Stem Cells ◽  
Neural Development ◽  
Current Knowledge ◽  
Embryonic Stem ◽  
Neural Induction ◽  
Neural Plate ◽  
Natural Rate ◽  
Neural Fate

The promise of stem cell therapy is expected to greatly benefit the treatment of neurodegenerative diseases. An underlying biological reason for the progressive functional losses associated with these diseases is the extremely low natural rate of self-repair in the nervous system. Although the mature CNS harbors a limited number of self-renewing stem cells, these make a significant contribution to only a few areas of brain. Therefore, it is particularly important to understand how to manipulate embryonic stem cells and adult neural stem cells so their descendants can repopulate and functionally repair damaged brain regions. A large knowledge base has been gathered about the normal processes of neural development. The time has come for this information to be applied to the problems of obtaining sufficient, neurally committed stem cells for clinical use. In this article we review the process of neural induction, by which the embryonic ectodermal cells are directed to form the neural plate, and the process of neural�fate stabilization, by which neural plate cells expand in number and consolidate their neural fate. We will present the current knowledge of the transcription factors and signaling molecules that are known to be involved in these processes. We will discuss how these factors may be relevant to manipulating embryonic stem cells to express a neural fate and to produce large numbers of neurally committed, yet undifferentiated, stem cells for transplantation therapies.

290 A CYNOMOLGUS MONKEY EMBRYONIC STEM CELL LINE DERIVED FROM A SINGLE BLASTOMERE

2008 ◽  
Vol 20 (1) ◽  
pp. 224
Author(s):  
J. Okahara-Narita ◽  
J. Yamasaki ◽  
C. Iwatani ◽  
H. Tsuchiya ◽  
K. Wakimoto ◽  
...  
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Cynomolgus Monkey ◽  
Es Cells ◽  
Embryonic Stem ◽  
Es Cell ◽  
Cell Stage ◽  
Vitro Assay ◽  
Single Blastomere ◽  
Cell Colony

The establishment of most embryonic stem (ES) cell lines requires the destruction of embryos. Some ES cell lines in mice and humans are currently derived from a single blastomere, so that remaining blastomeres can still develop into fetuses. However, the procedures currently in use for establishing these lines are very complicated, and other ES cell lines from the same species are needed (Chung et al. 2006 Nature 439, 216–219; Klimanskaya et al. 2006 Nature 444, 481–485). The objective of this study was to devise a method simpler than those previously described for establishing ES cell lines from a single blastomere in the cynomolgus monkey. Controlled ovarian stimulation and oocyte recovery have been described previously by Torii et al. (2000 Primates 41, 39–47). Cumulus-free mature oocytes were fertilized by intracytoplasmic sperm injection (ICSI), and then cultured at 38�C in 5% CO2, 5% O2 for 2 days. The zona pellucida of 4- to 5-cell-stage embryos was disrupted using acidic Tyrode's solution, and individual blastomeres were separated from the denuded embryos using trypsin. These blastomeres were cultured on mitomycin-C-treated mouse embryonic fibroblasts and ES medium containing adrenocorticotropic hormone (ACTH) (Ogawa et al. 2004 Genes to Cells 9, 471–477). After the formation of initial outgrowths, half of the medium was changed every other day until the outgrowths reached approximately 100 cells. Passage of putative monkey ES cells was performed by mechanical dispersion of the colonies and transfer to fresh feeders every 3–4 days until there were enough cells for enzymatic dispersion. One stable ES cell line was obtained from two 4- or 5-cell-stage embryos using ES medium containing ACTH. The morphology of this ES cell colony was consistent with the monkey ES cell colony previous described by Suemori et al. (2001 Dev. Dynamics 222, 273–279). The ES cell line was passaged more than 17 times, and the morphology of the ES cell colony did not differ between the first and seventeenth passages. The ES cells showed normal karyotype and retained pluripotency markers for primate ES cells including octamer-binding protein 4 (Oct-4), stage-specific embryonic antigen (SSEA)-4, tumor-rejection antigen (TRA)-1-60, and TRA-1-81. We are presently confirming whether this ES cell line possesses potencies to differentiate in all three embryonic germ layers using both an in vitro assay and teratoma formation. Here we showed that cynomolgus monkey ES cells can be derived from a single blastomere, without co-culturing another ES cell line, as has been done in previous studies on mice and humans. This method allows the establishment of ES cell lines from a single blastomere, leaving the other blastomeres available for embryo transfer. Thus, the method described here is simpler than previously described methods and alleviates some ethical concerns.

scholarly journals Stem cell culture on polyvinyl alcohol hydrogels having different elasticity and immobilized with ECM-derived oligopeptides

2017 ◽  
Vol 37 (7) ◽  
pp. 647-660 ◽  
Author(s):  
Saradaprasan Muduli ◽  
Li-Hua Chen ◽  
Meng-Pei Li ◽  
Zhao-wen Heish ◽  
Cheng-Hui Liu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Culture ◽  
Extracellular Matrix ◽  
Stem Cell ◽  
Cell Fate ◽  
Es Cells ◽  
Embryonic Stem ◽  
Poly Vinyl Alcohol ◽  
Hematopoietic Stem ◽  
Stem Cell Culture

Abstract The physical characteristics of cell culture materials, such as their elasticity, affect stem cell fate with respect to cell proliferation and differentiation. We systematically investigated the morphologies and characteristics of several stem cell types, including human amniotic-derived stem cells, human hematopoietic stem cells, human induced pluripotent stem (iPS) cells, and embryonic stem (ES) cells on poly(vinyl alcohol) (PVA) hydrogels immobilized with and without extracellular matrix-derived oligopeptide. Human ES cells did not adhere well to soft PVA hydrogels immobilized with oligovitronectin, whereas they did adhere well to PVA hydrogel dishes with elasticities greater than 15 kPa. These results indicate that biomaterials such as PVA hydrogels should be designed to possess minimum elasticity to facilitate human ES cell attachment. PVA hydrogels immobilized with and without extracellular matrix-derived oligopeptides are excellent candidates of cell culture biomaterials for investigations into how cell culture biomaterial elasticity affects stem cell culture and differentiation.

Serum Free Induction of a Lympho-Hematopoietic Precursor Population from Murine Embryonic Stem Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3605-3605
Author(s):  
Stefan Irion ◽  
Herve Luche ◽  
Hans J. Fehling ◽  
Gordon Keller
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
B Cell ◽  
Es Cells ◽  
Embryonic Stem ◽  
Yolk Sac ◽  
Es Cell ◽  
Erythroid Progenitors ◽  
Hematopoietic System ◽  
Serum Free

Abstract Hematopoiesis is initiated at several independent sites in the mouse embryo. The earliest site, the yolk sac, supports the development of a restricted hematopoietic program that consists of the production of primitive erythrocytes and macrophages, as well as progenitors of the definitive erythroid, megakaryocytic and mast cell lineages. Lymphoid cells are not generated during the early phase of yolk sac hematopoiesis. Following the onset of yolk sac hematopoiesis, a second hematopoietic program is initiated in a region known as the para-aortic splanchnopleura (P-Sp). The hematopoietic system generated in the P-Sp contains hematopoietic stem cells as well as progenitors of the lymphoid, myeloid and definitive erythroid lineages. The P-Sp does not give rise to primitive erythrocytes. The differentiation of embryonic stem (ES) cells in culture offers an outstanding system for modeling early hematopoietic development and for investigating the mechanisms regulating lineage commitment. While a number of different studies have provided convincing evidence that the ES cell model can recapitulate yolk sac hematopoiesis, it is unclear if the equivalent of the P-Sp hematopoietic system is established in these differentiation cultures. To address this question we induced different hematopoietic populations with a combination of activin A and BMP2 in serum-free media using an ES cell line carrying the GFP cDNA targeted to the mesoderm gene brachyury (GFP-Bry ES cells). When induced with these factors, the GFP-Bry cells generated two distinct populations with respect to expression of GFP-Bry and Flk-1, the receptor for vascular endothelial growth factor. The first expressed GFP-Bry, but no Flk-1 (GFP-Bry+/Flk-1−), whereas the second expressed both markers (GFP-Bry+/Flk-1+). If the GFP-Bry+/Flk-1− cells were allowed to reaggregate and cultured for an additional 24 hours, they generated a second GFP-Bry+/Flk-1+ population. Analysis of these two distinct Flk-1 populations revealed that both contained hematopoietic progenitors, but that their potential differed. The first Flk-1 population contained BL-CFC, the in vitro equivalent of the hemangioblast as well primitive erythroid and macrophage progenitors. It displayed limited potential to generate B and T lymphocytes when cultured on the OP9 and OP9-DL1 cells respectively. In contrast, the second Flk-1 population did generate B cell and T cells following coculture with the OP9 and OP9-DL1 stromal cells. B cell development was monitored by expression of B220, CD19 and surface IgM whereas T cells were identified by expression of CD4, CD8 and CD3. In addition to lymphoid progenitors, the second Flk-1 population contained multipotent, macrophage and definitive erythroid progenitors. It did not, however, contain significant numbers of BL-CFC or primitive erythroid progenitors. Taken together, these findings demonstrate that is it possible to generate two distinct hematopoietic populations in defined culture conditions. The developmental potential of these populations suggests that they could represent the equivalent of the yolk sac and P-Sp hematopoietic programs.

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