Changes in protein synthesis during differentiation of embryonal carcinoma cells, and a comparison with embryo cells

Development ◽  
1980 ◽  
Vol 59 (1) ◽  
pp. 187-206
Author(s):  
R. H. Lovell-Badge ◽  
M. J Evans
Keyword(s):  
Protein Synthesis ◽  
Embryonal Carcinoma ◽  
Inner Cell Mass ◽  
Embryoid Body ◽  
Cell Mass ◽  
Cell Types ◽  
Embryonic Cell ◽  
Embryoid Bodies ◽  
Ec Cells

Two-dimensional electrophoresis was used to find changes in protein synthesis occurring as pluripotent embryonal carcinoma (EC) cells differentiate to give embryoid bodies in vitro. 2-D patterns from other embryonic cell lines, and from the inner cell mass (ICM) cells of mouse embryos, were also analysed for the expression of those proteins showing some change during embryoid body formation and for overall differences between these and the EC cells. Most changes in protein synthesis occurred before 12 h but endoderm was not discerned morphologically on the outside of EC cell clumps until at least 18 h after their suspension. The number of changes occurring is small compared with the number of polypeptides resolved, but is in line with similar studies. Comparisons with nullipotent EC cells and an endodermal cell line have allowed these changes to be assigned, tentatively, to the different cell types within embryoid bodies, and may allow them to be used as markers of differentiation. Comparisons between the 2-D patterns derived from ICMs and EC cells reveal substantial differences between the two that might not have been expected from their developmental homology. The importance of these differences to their pluripotentiality is discussed.

The effect of retinoic acid pretreatment on the ability of murine embryonal carcinoma and inner cell mass cells to participate in chimaera development

Development ◽  
1986 ◽  
Vol 98 (1) ◽  
pp. 99-110
Author(s):  
B. K. Waters ◽  
J. Rossant
Keyword(s):  
Retinoic Acid ◽  
Embryonal Carcinoma ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Types ◽  
P19 Cells ◽  
Acid Pretreatment ◽  
Preimplantation Mouse ◽  
Embryo Cells

Certain embryonal carcinoma (EC) cell lines can colonize the embryo following blastocyst injection or embryo aggregation, giving rise to EC-embryo chimaeras. However, such chimaeras often develop abnormally. For example, diploid P19 cells colonize the embryo readily but resulting chimaeras are usually abnormal, with persistence of tumour cells. Retinoic acid (RA) induces differentiation of EC cells to a variety of cell types in vitro but, in this study, it was shown that pretreatment of P19 cells with RA did not result in more normal development of P19-embryo chimaeras. The only significant effect of RA was to reduce the ability of P19 cells to participate in embryonic development at all after blastocyst injection. RA did not have a direct toxic or teratogenic effect on preimplantation mouse embryos and did not affect the ability of pluripotent embryo cells to colonize chimaeras. Therefore, RA may not be the normal inducer of differentiation in early embryogenesis.

Embryonic stem (ES) cells and embryonal carcinoma (EC) cells: opposite sides of the same coin

2005 ◽  
Vol 33 (6) ◽  
pp. 1526-1530 ◽  
Author(s):  
P.W. Andrews ◽  
M.M. Matin ◽  
A.R. Bahrami ◽  
I. Damjanov ◽  
P. Gokhale ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonal Carcinoma ◽  
Inner Cell Mass ◽  
Tumour Progression ◽  
Es Cells ◽  
Cell Mass ◽  
Embryonic Stem ◽  
Blastocyst Stage ◽  
Ec Cells

Embryonal carcinoma (EC) cells are the stem cells of teratocarcinomas, and the malignant counterparts of embryonic stem (ES) cells derived from the inner cell mass of blastocyst-stage embryos, whether human or mouse. On prolonged culture in vitro, human ES cells acquire karyotypic changes that are also seen in human EC cells. They also ‘adapt’, proliferating faster and becoming easier to maintain with time in culture. Furthermore, when cells from such an ‘adapted’ culture were inoculated into a SCID (severe combined immunodeficient) mouse, we obtained a teratocarcinoma containing histologically recognizable stem cells, which grew out when the tumour was explanted into culture and exhibited properties of the starting ES cells. In these features, the ‘adapted’ ES cells resembled malignant EC cells. The results suggest that ES cells may develop in culture in ways that mimic changes occurring in EC cells during tumour progression.

Use of F9 teratocarcinoma STEM cells to study cell-matrix interactions in the early mouse embryo

1992 ◽  
Vol 50 (1) ◽  
pp. 602-603
Author(s):  
Marc Lenburg ◽  
Rulang Jiang ◽  
Lengya Cheng ◽  
Laura Grabel
Keyword(s):  
Mouse Embryo ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Types ◽  
Embryoid Bodies ◽  
F9 Cells ◽  
Cell Matrix ◽  
Matrix Interactions ◽  
Cell Matrix Interactions ◽  
F9 Teratocarcinoma

We are interested in defining the cell-cell and cell-matrix interactions that help direct the differentiation of extraembryonic endoderm in the peri-implantation mouse embryo. At the blastocyst stage the mouse embryo consists of an outer layer of trophectoderm surrounding the fluid-filled blastocoel cavity and an eccentrically located inner cell mass. On the free surface of the inner cell mass, facing the blastocoel cavity, a layer of primitive endoderm forms. Primitive endoderm then generates two distinct cell types; parietal endoderm (PE) which migrates along the inner surface of the trophectoderm and secretes large amounts of basement membrane components as well as tissue-type plasminogen activator (tPA), and visceral endoderm (VE), a columnar epithelial layer characterized by tight junctions, microvilli, and the synthesis and secretion of α-fetoprotein. As these events occur after implantation, we have turned to the F9 teratocarcinoma system as an in vitro model for examining the differentiation of these cell types. When F9 cells are treated in monolayer with retinoic acid plus cyclic-AMP, they differentiate into PE. In contrast, when F9 cells are treated in suspension with retinoic acid, they form embryoid bodies (EBs) which consist of an outer layer of VE and an inner core of undifferentiated stem cells. In addition, we have established that when VE containing embryoid bodies are plated on a fibronectin coated substrate, PE migrates onto the matrix and this interaction is inhibited by RGDS as well as antibodies directed against the β1 integrin subunit. This transition is accompanied by a significant increase in the level of tPA in the PE cells. Thus, the outgrowth system provides a spatially appropriate model for studying the differentiation and migration of PE from a VE precursor.

Metabolic co-operation between embryonic and embryonal carcinoma cells of the mouse

Development ◽  
1979 ◽  
Vol 54 (1) ◽  
pp. 263-275
Author(s):  
Stephen J. Gaunt ◽  
Virginia E. Papaioannou
Keyword(s):  
Embryonal Carcinoma ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Embryonic Cells ◽  
Cone Cells ◽  
Ec Cells ◽  
Hypoxanthine Guanine Phosphoribosyltransferase ◽  
Early Mouse ◽  
Intercellular Exchange ◽  
Embryonic Ectoderm

Mouse embryonal carcinoma (EC) cells form permeable junctions at their homotypic cell-to-cell contacts which permit intercellular exchange of metabolites (metabolic co-operation). Hooper & Slack (1977) showed how this exchange could be detected by autoradiography as the transfer of [3H]nucleotides between PCI3 (a pluripotential EC line) and PCI 3- TG8 (a variant of PC13 which is deficient in hypoxanthine guanine phosphoribosyltransferase). We now show that cells taken from several different tissues of early mouse embryos, that is, from the morula, the inner cell mass of the blastocyst, and the endoderm, mesoderm and embryonic ectoderm of the 8th day egg cylinder, are able to serve as donors of [3H] ucleotides to PC13TG8. In contrast, trophectodermal cells of cultured blastocysts, and the trophectodermal derivatives in the 8th day egg cylinder, that is, extra-embryonic ectoderm and ectoplacental cone cells, showed little or no metabolic co-operation with PC13TG8. With reference to some common properties of EC and embryonic cells, we suggest how our findings may provide insight into cell-to-cell interactions in the early mouse embryo.

Developmental complexity of early mammalian pluripotent cell populations in vivo and in vitro

1998 ◽  
Vol 10 (8) ◽  
pp. 535 ◽  
Author(s):  
T. A. Pelton ◽  
M. D. Bettess ◽  
J. Lake ◽  
J. Rathjen ◽  
P. D. Rathjen
Keyword(s):  
Cell Biology ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Cell Types ◽  
Experimental Manipulation ◽  
Pluripotent Cell ◽  
Pluripotent Cells ◽  
Cellular Origin

Early mammalian embryogenesis is characterised by the coordinated proliferation, differentiation, migration and apoptosis of a pluripotent cell pool that is able to give rise to extraembryonic lineages and all the cell types of the embryo proper. These cells retain pluripotent differentiation capability, defined in this paper as the ability to form all cell types of the embryo and adult, until differentiation into the three embryonic germ layers at gastrulation. Our understanding of pluripotent cell biology and molecular regulation has been hampered by the difficulties associated with experimental manipulation of these cells in vivo. However, a more detailed understanding of pluripotent cell behaviour is emerging from the application of molecular technologies to early mouse embryogenesis. The construction of mouse mutants by gene targeting, mapping of gene expression in vivo, and modelling of cell decisions in vitro are providing insight into the cellular origin, identity and action of key developmental regulators, and the nature of pluripotent cells themselves. In this review we discuss the properties of early embryonic pluripotent cells in vitro and in vivo, focusing on progression from inner cell mass (ICM) cells in the blastocyst to the onset of gastrulation.

scholarly journals Generation of embryonic stem cell lines from mouse blastocysts developed in vivo and in vitro: relation to Oct-4 expression

Reproduction ◽  
2006 ◽  
Vol 132 (1) ◽  
pp. 59-66 ◽  
Author(s):  
S Tielens ◽  
B Verhasselt ◽  
J Liu ◽  
M Dhont ◽  
J Van Der Elst ◽  
...  
Keyword(s):  
Cell Lines ◽  
Inner Cell Mass ◽  
Es Cells ◽  
Cell Mass ◽  
Embryonic Stem ◽  
Embryoid Bodies ◽  
Differentiation Potential ◽  
Stem Cell Lines

Embryonic stem (ES) cells are the source of all embryonic germ layer tissues. Oct-4 is essential for their pluripotency. Sincein vitroculture may influence Oct-4 expression, we investigated to what extent blastocysts culturedin vitrofrom the zygote stage are capable of expressing Oct-4 and generating ES cell lines. We comparedin vivowithin vitroderived blastocysts from B6D2 mice with regard to Oct-4 expression in inner cell mass (ICM) outgrowths and blastocysts. ES cells were characterized by immunostaining for alkaline phosphatase (ALP), stage-specific embryonic antigen-1 (SSEA-1) and Oct-4. Embryoid bodies were made to evaluate the ES cells’ differentiation potential. ICM outgrowths were immunostained for Oct-4 after 6 days in culture. A quantitative real-time PCR assay was performed on individual blastocysts. Of thein vitroderived blastocysts, 17% gave rise to ES cells vs 38% of thein vivoblastocysts. Six-day old outgrowths fromin vivodeveloped blastocysts expressed Oct-4 in 55% of the cases vs 31% of thein vitroderived blastocysts. The amount of Oct-4 mRNA was significantly higher for freshly collectedin vivoblastocysts compared toin vitrocultured blastocysts.In vitrocultured mouse blastocysts retain the capacity to express Oct-4 and to generate ES cells, be it to a lower level thanin vivoblastocysts.

BMPs regulate differentiation of a putative visceral endoderm layer within human embryonic stem-cell-derived embryoid bodies

2007 ◽  
Vol 85 (1) ◽  
pp. 121-132 ◽  
Author(s):  
Brock J Conley ◽  
Sarah Ellis ◽  
Lerna Gulluyan ◽  
Richard Mollard
Keyword(s):  
Inner Cell Mass ◽  
Cell Mass ◽  
Embryonic Stem ◽  
Embryoid Bodies ◽  
Visceral Endoderm ◽  
Transmission Electron ◽  
Spherical Structures ◽  
Bmp Signalling ◽  
Serum Free Conditions

Human embryonic stem cells (HESCs), pluripotent cells derived from the inner cell mass (ICM) of human blastocysts, represent a novel tool for the study of early human developmental events. When cultured in suspension with serum, HESCs form spherical structures resembling embryoid bodies (EBs). We show that differentiation of HESCs within EBs occurs radially, with central cells then undergoing apoptosis in association with EB cavitation. Cells within the outer layer of cavitating EBs display stage-specific immunoreactivity to pan-keratin, cytokeratin-8, GATA6, α-fetoprotein, and transthyretin specific antibodies, and hybridization to disabled-2, GATA4, and GATA6 specific riboprobes. Transmission electron microscopy of these cells reveals clathrin-coated micropinocytotic vesicles, microvilli, and many vacuoles, a phenotype consistent with mouse visceral endoderm (VE) rather than mouse definitive or parietal endoderm. When cultured in media supplemented with the BMP inhibitor noggin, or in the absence of serum, HESC derivatives do not develop the mouse VE-like phenotype. The addition of BMP-4 to noggin-treated HESCs cultured in serum or in serum-free conditions reconstituted development of the VE-like phenotype. These data demonstrate that human EBs undergo developmental events similar to those of mouse EBs and that in vitro BMP signalling induces derivatives of the human ICM to express a phenotype similar to mouse VE.

A transfected H-ras oncogene does not inhibit differentiation of cardiac and skeletal muscle from embryonal carcinoma cells

1989 ◽  
Vol 67 (9) ◽  
pp. 590-596 ◽  
Author(s):  
Michael A. Rudnicki ◽  
Kenneth R. Reuhl ◽  
Michael W. McBurney
Keyword(s):  
Skeletal Muscle ◽  
Cell Lines ◽  
Embryonal Carcinoma ◽  
Cell Types ◽  
P19 Cells ◽  
Ras Oncogene ◽  
Specific Expression ◽  
Ec Cells ◽  
Myoblast Cell

P19 embryonal carcinoma (EC) cells can be induced to differentiate in vitro into a variety of cell types, including cardiac and skeletal myocytes. We have isolated P19 cells stably transformed with either the activated human H-ras oncogene or with a chimeric gene in which the H-ras oncogene was controlled by a muscle-specific promoter. These P19 lines exhibited ubiquitous and muscle-specific expression of the activated H-ras protein, respectively. In both lines of P19 cells, normal cardiac and skeletal muscle differentiation was observed. Since the activated H-ras prevents differentiation of myoblast cell lines, our results suggest that the EC-derived muscle progenitor cell differs from continuous myoblast cell lines, perhaps by lacking a complementing oncogene responsible for myoblast immortalization.Key words: embryonal carcinoma, oncogene, ras, differentiation, myogenesis.

001. ISOLATION AND CHARACTERISATION OF PORCINE ES CELLS

2009 ◽  
Vol 21 (9) ◽  
pp. 1
Author(s):  
M. B. Nottle ◽  
I. M. Vassiliev ◽  
S. Vassilieva ◽  
L. F. S. Beebe ◽  
S. J. Harrison ◽  
...  
Keyword(s):  
Inner Cell Mass ◽  
Es Cells ◽  
Cell Mass ◽  
Embryonic Stem ◽  
Cell Types ◽  
Embryoid Bodies ◽  
The Body ◽  
Domestic Species ◽  
Serum Replacement

Embryonic stem (ES) cellshave the capacity for self renewal, can remain undifferentiated in long term culture and can contribute to all the cells in the body including the germ cells. EScells have been isolated in mice and have also been described for humans. However despite considerable effort for more than two decades ES cellswhich can contribute to the germline are yet to be isolated for the pig or any domestic species for that matter. We have developed a new method for isolating porcine ES cells which uses whole embryos cultured in alpha MEM with 10% serum replacement plus additives under 5% O2. Unlike methods employed previously this method results in homogenous outgrowths whose cells resemble ES cells and which express Oct 4 and Nanog and SSEA-1 [1]. These cells can be passaged and cryopreserved repeatedly resulting in the establishment of cell lines at similar efficiencies to that reported previously for 129Sv mice [2]. These cells can form embryoid bodies and can be differentiated to various cell types representative of all three germ layers [3]. Following their injection into blastocysts these cells localise /become incorporated in the inner cell mass and can be used to produce chimaeras when these embryos are transferred to recipient animals [2]. To date we have produced chimaeric pigs from one male ES cell line [2]. These are currently being mated to demonstrate germline transmission. Future studies will examine the applicability of our method to other species commencing with mice and cattle before extending these to humans.

scholarly journals Absence of Basement Membranes after Targeting the LAMC1 Gene Results in Embryonic Lethality Due to Failure of Endoderm Differentiation

1999 ◽  
Vol 144 (1) ◽  
pp. 151-160 ◽  
Author(s):  
Neil Smyth ◽  
H. Seda Vatansever ◽  
Patricia Murray ◽  
Michael Meyer ◽  
Christian Frie ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Basement Membrane ◽  
Inner Cell Mass ◽  
Cell Mass ◽  
Embryonic Stem ◽  
Basement Membranes ◽  
Embryoid Bodies ◽  
Endoderm Differentiation

The LAMC1 gene coding for the laminin γ1 subunit was targeted by homologous recombination in mouse embryonic stem cells. Mice heterozygous for the mutation had a normal phenotype and were fertile, whereas homozygous mutant embryos did not survive beyond day 5.5 post coitum. These embryos lacked basement membranes and although the blastocysts had expanded, primitive endoderm cells remained in the inner cell mass, and the parietal yolk sac did not develop. Cultured embryonic stem cells appeared normal after targeting both LAMC1 genes, but the embryoid bodies derived from them also lacked basement membranes, having disorganized extracellular deposits of the basement membrane proteins collagen IV and perlecan, and the cells failed to differentiate into stable myotubes. Secretion of the linking protein nidogen and a truncated laminin α1 subunit did occur, but these were not deposited in the extracellular matrix. These results show that the laminin γ1 subunit is necessary for laminin assembly and that laminin is in turn essential for the organization of other basement membrane components in vivo and in vitro. Surprisingly, basement membranes are not necessary for the formation of the first epithelium to develop during embryogenesis, but first become required for extra embryonic endoderm differentiation.

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