1 University of Wisconsin - Medical School and The National Primate Research
Center, University of Wisconsin, Madison, WI 53715, USA
2 WiCell Research Institute, Madison, WI 53715, USA
* Author for correspondence (e-mail: thomson{at}primate.wisc.edu)
SUMMARY
Because embryonic stem (ES) cells are generally derived by the culture of inner cell mass (ICM) cells, they are often assumed to be the equivalent of ICM cells. However, various evidence indicates that ICM cells transition to a different cell type during ES-cell derivation. Historically, ES cells have been believed to most closely resemble pluripotent primitive ectoderm cells derived directly from the ICM. However, differences between ES cells and primitive ectoderm cells have caused developmental biologists to question whether ES cells really have an in vivo equivalent, or whether their properties merely reflect their tissue culture environment. Here, we review recent evidence that the closest in vivo equivalent of an ES cell is an early germ cell.
Introduction
Embryonic stem (ES) cells are pluripotent (see
Box 1) and can be expanded
without limit in vitro (Evans and Kaufman,
1981; Martin,
1981
; Thomson et al.,
1998
). It is remarkable that permanent pluripotent stem cell lines
can be derived from preimplantation embryos at all, because, in vivo,
pluripotent cells of the early mammalian embryo proliferate only briefly
before becoming cells with a more restricted developmental potential. A few
years after the initial derivation of mouse ES cells, it was suggested that
they be called `embryo-derived stem cells', a more precise term that would
distinguish between these new pluripotent cell lines and cells within the
embryo (Rossant and Papaioannou,
1984
). However, this term was never adopted, and the extent to
which these pluripotent stem cell lines represent any specific embryonic cell
type or reflect their artificial tissue culture environment is still an open
issue today - two decades later. Elucidating the origin of ES cells is of
importance because it may help us to identify genes that are essential for the
long-term maintenance of the pluripotent state. It could also assist with the
derivation of ES cells from species whose ES cells have proved difficult to
isolate. It will also help us to assess how accurately ES cell differentiation
reflects events that normally occur in vivo. Here, we review the origin of ES
cells, and explore recent evidence that ES cells are closely related to early
germ cells.
The historical origins of ES cells: embryonal carcinoma cells
Historically, work with mouse teratocarcinomas paved the way for the
derivation of ES cells. These germ cell tumors contain multiple differentiated
tissues and undifferentiated stem cells, called embryonal carcinoma (EC) cells
(Damjanov and Solter, 1974;
Dixon and Moore, 1952
;
Kleinsmith and Pierce, 1964
).
Although teratocarcinomas had been known as medical curiosities for centuries
(Wheeler, 1983
), it was the
discovery that male mice of strain 129 had a high incidence of testicular
teratocarcinomas (Stevens and Little,
1954
) that made these tumors more routinely amenable to
experimental analysis. Because their growth is sustained by the persistent EC
cell component (Stevens and Little,
1954
), teratocarcinomas can be serially transplanted between mice.
Eventually, conditions were developed that allowed the culture of EC cells in
vitro, establishing them as an in vitro model of mammalian development
(Kahan and Ephrussi,
1970
).
As pluripotent cells of the intact early embryo proliferate for only a
limited period of time, it was not initially obvious whether pluripotent cell
lines could be established without undergoing malignant transformation.
However, the transplantation of genital ridges or of egg-cylinder-stage
embryos into ectopic sites, such as under the kidney capsule of adult mice,
gave rise to teratocarcinomas at a high frequency in strains that did not
spontaneously produce these tumors (Solter
et al., 1970; Stevens,
1970a
; Stevens,
1970b
). These teratocarcinomas could be serially transplanted
between adult mice, depending on whether the EC cell component persisted or
differentiated (Solter et al.,
1981
). If the EC compartment disappears, the resulting tumor
develops as a benign teratoma. Indeed, the malignant phenotype of EC cells
often depends on the strain of the host mouse, and not on the tumor strain. EC
cells injected into mouse blastocysts can contribute to either the normal
tissues of the resulting chimera (Brinster,
1974
) or, in some cases, to tumors
(Rossant and McBurney, 1982
).
Because the ectopic transplantation of normal peri-implantation embryos can
give rise to pluripotent cell lines, the direct derivation of pluripotent cell
lines in vitro was attempted without the teratocarcinoma step. The culture
conditions that were established to support mouse EC cells, including the use
of feeder cell layers, were essentially those used to isolate mouse, and
eventually human, ES cells (Evans and
Kaufman, 1981
; Martin,
1981
; Thomson et al.,
1998
).
One indication that these early EC cell lines may be derived from germ
cells (Solter et al., 1970;
Stevens, 1967
;
Stevens, 1970a
) came from
mouse genital ridge-transplantation experiments. These experiments showed that
genital ridges
Are ES cells a tissue culture artifact?
ES cells clearly exhibit some properties that are not normally shown by
cells of the intact embryo. For example, although ES cells retain properties
of early embryonic cells in vitro, no pluripotent cell demonstrates long-term
self-renewal in vivo. Embryonic cells, once brought into tissue culture, are
exposed to numerous extrinsic signals to which they never would be exposed to
in vivo. ES cells certainly adapt to selective tissue culture conditions and
acquire novel functions that allow them to proliferate in an undifferentiated
state indefinitely, and, because of this, ES cells are in some sense tissue
culture artifacts (Buehr and Smith,
2003; Rossant,
2001
; Smith,
2001
).
As these changes are inevitable, the issue is not whether ES cells exhibit some properties that merely reflect their tissue culture environment, but rather whether they are most closely related to a specific in vivo cell type in the embryo, or if the influence of the culture environment is so dominant that it is impossible to relate ES cells to a single, in vivo cell type. We will certainly not completely resolve this issue here, but will re-explore the relationship of ES cells to specific early embryonic cell types.
Are ES cells most closely related to primitive ectoderm?
Although ES cell lines are generally derived from the culture of the ICM,
some experiments suggest that ES cells more closely resemble cells from the
primitive ectoderm. For example, isolated primitive ectoderm from the mouse
gives rise to ES cell lines at a higher frequency than does isolated ICM.
Moreover, the culture of primitive ectoderm allows the isolation of ES cell
lines from mouse strains that have been previously refractory to ES cell
isolation (Brook and Gardner,
1997). Indeed, ES cell lines can be derived from single, isolated,
mouse primitive ectoderm cells, which is not possible with ICM cells
(Gardner and Brook, 1997
).
Although these experiments suggest that ES cells are more closely related to
primitive ectoderm than to ICM, they do not reveal whether ES cells more
closely resemble primitive ectoderm or a cell derived from it in vitro.
A maximum of three individual cultured primitive ectoderm cells per embryo
have been shown to give rise to ES cell colonies
(Gardner and Brook, 1997).
This low frequency could have been due to some variability in the potential of
primitive ectoderm cells, to some variability in the environment in which they
were placed or to damage caused by the dissociation of the primitive ectoderm
into individual cells. However, by tracking the expression of the
octamer-binding transcription factor 4 (Oct4) gene, a marker of
pluripotency, in intact cultured ICM/epiblast cells, it was shown that
Oct4 expression was maintained in only a small proportion of
outgrowing cells (Buehr et al.,
2003
), which also suggests that only a minority of primitive
ectoderm cells can transit to a new stable, proliferative pluripotent state,
and, subsequently, be expanded as ES cells. These results could be due to a
requirement for a relatively rare intrinsic or extrinsic stochastic event, or
to an inherent heterogeneity of the primitive ectodermal cell population.
Recent data indicate that even the earliest ICM is heterogeneous and consists
of a mixture of cells that express either Oct4 or Gata6
(Rossant et al., 2003
), and a
similar later heterogeneity could account for the fact that only a minority of
primitive ectoderm cells generally give rise to ES cells in culture.
Established mouse ES cell lines express some specific markers of primitive
ectoderm at a very low level, if at all
(Table 1), such as fibroblast
growth factor 5 (Fgf5) (Haub and
Goldfarb, 1991; Hebert et al.,
1991
; Rathjen et al.,
1999
). Culture conditions have been established that convert mouse
ES cells into early primitive ectoderm-like cells that express both
Fgf5 and Oct4 (Rathjen
et al., 1999
), but these cells fail to form chimeras when injected
into mouse blastocysts. Taken together, these results suggest that ES cells
are most closely related to a subpopulation of primitive ectoderm cells, or to
a close derivative of primitive ectoderm cells.
|
Germ cells and the primitive ectoderm
In elegant, clonal-fate mapping studies in the mouse
(Lawson and Hage, 1994), germ
cells were shown to arise from a founder population in the E6.0-6.5 proximal
epiblast adjacent to the extra-embryonic ectoderm. These founder cells then
pass through the primitive streak and give rise to several extra-embryonic
mesodermal lineages and to germ cells. By E7.25, a distinct cluster of
45
tissue non-specific, alkaline phosphatase (Tnap)-positive germ cells is
present at the base of the allantois (Fig.
1) (Ginsburg et al.,
1990
). The E6.5 distal epiblast, which would not normally
contribute to germ cells, will contribute to germ cells if transplanted to a
proximal location (Tam and Zhou,
1996
), which demonstrates that location and inductive signals,
rather than germ plasm determinants, are responsible for the specification of
germ cells in mice (Extavour and Akam,
2003
). This flexibility suggests that cultured primitive ectoderm
cells could spontaneously give rise to early germ cells in culture.
|
Similarities between germ cells and ES cells
In mice, PGCs migrate and proliferate until 25,000 are present in the
genital ridge at E13.0 (Tam and Snow,
1981
). Pluripotent cell lines from pre- and post-migratory
(Resnick et al., 1992
;
Matsui et al., 1992
;
Shamblott et al., 1998
), as
well as from migratory (Durcova-Hills et
al., 2001
), germ cells have been isolated, and these cell lines
are termed embryonic germ (EG) cells to distinguish their origin. Mouse EG
cell lines are remarkably similar to mouse ES cell lines
(Donovan and de Miguel, 2003
).
During germ cell migration and maturation, however, the somatic status of
imprinted genes is progressively erased
(Yamazaki et al., 2003
), and
EG cells isolated at various stages of migration retain some of these
differences, such as the reduced methylation of many imprinted genes,
including H19 and Snrpn
(Hajkova et al., 2002
). The
analysis of mouse PGCs at E10.5 suggests that methylation erasure has already
begun by this time, as supported by studies of the expression of imprinted
genes (Yamazaki et al., 2003
).
This study showed that imprinted genes, such as H19 and
Snrpn, exhibit imprinted (somatic) expression patterns in E9.5 PGCs,
but by E10.5 have switched to a bi-allelic mode of expression
(Yamazaki et al., 2003
).
Because the genes expressed in ES cells exhibit somatic imprinting patterns
(Geijsen et al., 2004
), their
change in imprinting status suggests that if ES cells are derived from germ
cells, this derivation must occur before E9.5.
There is a paucity of known molecular markers that distinguish early germ
cells from other pluripotent cells of the early embryo. One marker, Tnap, is
strongly expressed by early germ cells and by ES cells, but is weakly
expressed by the epiblast and other surrounding embryonic cells
(Chiquoine, 1954;
Ginsburg et al., 1990
). Two
new markers for early germ cells, fragilis (Ifitm3 - Mouse Genome Informatics)
and Dppa3 (also know as stella or PGC7), have recently been identified that
allow the better separation of early germ cell precursors from their
differentiated neighboring cells (Saitou
et al., 2002
). Dppa3 is expressed in pre-implantation embryos and
in germ cells (Sato et al.,
2002
) and has recently been reported to have a role as a maternal
transcript in preimplantation embryonic development
(Bortvin et al., 2004
).
Dppa3-positive cells show increased expression of fragilis and remain positive
for Tnap (Akp2 - Mouse Genome Informatics) and Oct4
(Saitou et al., 2002
). Once
Dppa3-positive PGCs start to migrate, they begin to express additional
markers, such as steel factor receptor, followed by markers of more mature
germ cells, such as murine vasa homolog (MVH; Ddx4 - Mouse Genome Informatics)
(Saitou et al., 2002
).
Several recent reports describing the differentiation of mouse ES cells
into cells that express markers of mature male and female germ cells
(Geijsen et al., 2004;
Hubner et al., 2003
;
Toyooka et al., 2003
) are
important for our understanding of the origin of ES cells. In each of these
reports, germ cell markers were expressed by ES cells themselves, including
those, such as Dppa3, that help distinguish germ cells from primitive ectoderm
(Table 1). Only the expression
of more mature germ cell markers (such as MVH) enabled in vitro-derived germ
cells to be distinguished from ES cells themselves. In one study that examined
the differentiation of human ES cells into germ cells
(Clark et al., 2004
), the
expression of each of eight genes that are characteristic of early germ cells
was detected in human ES cells, but the expression of each of six genes that
are characteristic of later germ cells was not detected, strongly suggesting
that the expression of the early germ cell-genes was not merely a result of
the broadly `leaky' transcription that is often attributed to ES cells. Using
immunocytochemistry, it was also shown that most individual human ES cells in
a population express the early germ cell markers stella related (STELLAR) and
deleted in azoospermia-like (DAZL), indicating that a minor subset of randomly
differentiating cells in a mixed population is not responsible for the
expression of germ cell markers in ES cell cultures. Importantly, it was also
shown that at least one germ cell-specific gene, DAZL, was expressed
by human ES cells but not by human ICM. The existing gene expression data,
then, are consistent with the idea that the closest in vivo equivalent to ES
cells is not the ICM or primitive ectoderm, but an early germ cell.
Some of the properties of ES cells, however, suggest that they are not
merely the equivalent of early germ cells. For example, the earliest PGCs do
not self-renew for prolonged periods of time, but instead begin a series of
maturation steps, beginning with germ cell migration and ending in the highly
specialized development of sperm or egg
(Wylie, 1999). Although ES
cells can differentiate into more mature germ cells in vitro, they do so
relatively inefficiently. Indeed, the ability to colonize the germline of
chimeras is one of the most easily lost properties of ES cells. If ES cells
most closely represent early germ cells, it is unclear why they are not better
at giving rise to more mature germ cells. In addition, isolated PGCs have
never been demonstrated to contribute to chimeras when injected into
blastocysts, so an exact equivalence to ES cells is unlikely.
Because a comprehensive and comparative analysis of the transcriptomes of isolated ICM, primitive ectoderm and early germ cells has not yet been reported, it is not yet clear how much the particular repertoire of genes expressed by ES cells represents an early germ cell, another specific in vivo cell type, a response to the tissue culture environment, or a combination of all three. If the ICM and primitive ectoderm are inherently heterogeneous, transcriptome analysis may need to be carried out at the single-cell level to ultimately understand these relationships. However, at the moment, the greatest concordance of known markers appears to be between ES cells and early germ cells.
Conclusions
We hypothesize that ES, EC and EG cells represent a family of related pluripotent cell lines, whose common properties reflect a common origin from germ cells (Fig. 2). Although a more detailed transcriptional analysis could ultimately refute the proposed relationship between ES cells and early germ cells, we hope this idea will at least help to stimulate a healthy re-evaluation of what is actually being studied when ES cells differentiate in vitro.
|
Another implication of our hypothesis is that when looking for evolutionary
clues to understand the pluripotent state, the comparative germ cell
literature will be the most instructive. In a species such as the zebrafish,
which has a germ plasm that strictly separates germ cells from somatic cells,
it makes sense that pluripotent cell lines that can contribute to the germline
in chimeras (Ma et al., 2001)
would have to be derived from germ line-lineage cells.
Another prediction arising from the hypothesis that ES cells most closely represent early germ cells is that the very earliest events of ES cell differentiation into somatic and extra-embryonic lineages will not accurately reflect events that normally occur in vivo. The idea that ES cells represent an in vitro equivalent to the ICM, however, is firmly entrenched and continues to strongly influence our thinking about these cells. When examining the differentiation of ES cells in vitro, the pervasive mental image is of a forward progression that recapitulates normal embryonic events. For example, one thinks of ICM cells progressing to primitive ectoderm cells, then to neural ectoderm cells, and finally to more specialized neural cell types. If ES cells most closely represent early germ cells, this mental image needs revision, as the earliest transition would appear to be more `lateral' or even `backward' than `forward'. It will be illuminating to define each of the distinct transitions that ES cells can make in a single step and to determine how much these initial transitions resemble in vivo or artificial differentiation. If ES cells really represent early germ cells, the initial events in differentiation would be expected to be transitions that do not normally occur in intact embryos, except, perhaps, when the transition is to more mature germ cells.
ACKNOWLEDGMENTS
We thank Dr Azim Surani and Dr Ivan Damjanov for critical reading of our manuscript and the helpful discussion. We thank Kimberly Smuga-Otto and David McDougal for helping us prepare the figures. Additionally, we thank Henry Yuen and the Oscar Rennebohm Foundation for their gifts to the Wisconsin Alumni Foundation that supported the authors of this work.
REFERENCES
Beddington, R. S. and Robertson, E. J. (1989). An assessment of the developmental potential of embryonic stem cells in the midgestation mouse embryo. Development 105,733 -737.[Abstract]
Bortvin, A., Goodheart, M., Liao, M. and Page, D. C. (2004). Dppa3/Pgc7/stella is a maternal factor and is not required for germ cell specification in mice. BMC Dev Biol 4,2 .[CrossRef][Medline]
Brinster, R. L. (1974). The effect of cells
transferred into the mouse blastocyst on subsequent development. J.
Exp. Med. 140,1049
-1056.
Brook, F. A. and Gardner, R. L. (1997). The
origin and efficient derivation of embryonic stem cells in the mouse.
Proc. Natl. Acad. Sci. USA
94,5709
-5712.
Buehr, M., Nichols, J., Stenhouse, F., Mountford, P.,
Greenhalgh, C. J., Kantachuvesiri, S., Brooker, G., Mullins, J. and
Smith, A. G. (2003). Rapid loss of Oct-4 and pluripotency in
cultured rodent blastocysts and derivative cell lines. Biol.
Reprod. 68,222
-229.
Buehr, M. and Smith, A. (2003). Genesis of embryonic stem cells. Philos. Trans. R. Soc. Lond. B Biol. Sci. 358,1397 -1402.[CrossRef][Medline]
Chambers, I., Colby, D., Robertson, M., Nichols, J., Lee, S., Tweedie, S. and Smith, A. (2003). Functional expression cloning of Nanog, a pluripotency sustaining factor in embryonic stem cells. Cell 113,643 -655.[Medline]
Chapman, G., Remiszewski, J. L., Webb, G. C., Schulz, T. C., Bottema, C. D. and Rathjen, P. D. (1997). The mouse homeobox gene, Gbx2: genomic organization and expression in pluripotent cells in vitro and in vivo. Genomics 46,223 -233.[CrossRef][Medline]
Chiquoine, A. D. (1954). The identification, origin, and migration of the primordial germ cells in the mouse embryo. Anat. Rec. 118,135 -146.
Clark, A. T., Bodnar, M. S., Fox, M., Rodriquez, R. T., Abeyta,
M. J., Firpo, M. T. and Pera, R. A. (2004).
Spontaneous differentiation of germ cells from human embryonic stem cells in
vitro. Hum. Mol. Genet.
13,727
-739.
Damjanov, I. and Solter, D. (1974). Experimental teratoma. Curr. Top. Pathol. 59, 69-130.[Medline]
Damjanov, I., Solter, D. and Skreb, N. (1971). Teratocarcinogenesis as related to the age of embryos grafted under the kidney capsule. Wilhelm Roux Arch. Entwicklungsmech. Org. 173,282 -284.
Diwan, S. B. and Stevens, L. C. (1976). Development of teratomas from the ectoderm of mouse egg cylinders. J. Natl. Cancer Inst. 57,937 -942.[Medline]
Dixon, F. S. and Moore, R. A. (1952). Tumors of the male sex organs. In Atlas of Tumor Pathology, Vol.8 (fascicles 31b and 32). Washington, DC: Armed Forces Institute of Pathology.
Donovan, P. J. and de Miguel, M. P. (2003). Turning germ cells into stem cells. Curr. Opin. Genet. Dev. 13,463 -471.[CrossRef][Medline]
Durcova-Hills, G., Ainscough, J. and McLaren, A. (2001). Pluripotential stem cells derived from migrating primordial germ cells. Differentiation 68,220 -226.[CrossRef][Medline]
Evans, M. J. and Kaufman, M. H. (1981). Establishment in culture of pluripotential cells from mouse embryos. Nature 292,154 -156.[Medline]
Extavour, C. G. and Akam, M. (2003). Mechanisms
of germ cell specification across the metazoans: epigenesis and preformation.
Development 130,5869
-5884.
Gardner, R. L. and Brook, F. A. (1997). Reflections on the biology of embryonic stem (ES) cells. Int. J. Dev. Biol. 41,235 -243.[Medline]
Geijsen, N., Horoschak, M., Kim, K., Gribnau, J., Eggan, K. and Daley, G. Q. (2004). Derivation of embryonic germ cells and male gametes from embryonic stem cells. Nature 427,148 -154.[CrossRef][Medline]
Ginsburg, M., Snow, M. H. and McLaren, A. (1990). Primordial germ cells in the mouse embryo during gastrulation. Development 110,521 -528.[Abstract]
Hajkova, P., Erhardt, S., Lane, N., Haaf, T., El-Maarri, O., Reik, W., Walter, J. and Surani, M. A. (2002). Epigenetic reprogramming in mouse primordial germ cells. Mech. Dev. 117,15 -23.[CrossRef][Medline]
Haub, O. and Goldfarb, M. (1991). Expression of the fibroblast growth factor-5 gene in the mouse embryo. Development 112,397 -406.[Abstract]
Hebert, J. M., Boyle, M. and Martin, G. R. (1991). mRNA localization studies suggest that murine FGF-5 plays a role in gastrulation. Development 112,407 -415.[Abstract]
Horie, K., Takakura, K., Taii, S., Narimoto, K., Noda, Y., Nishikawa, S., Nakayama, H., Fujita, J. and Mori, T. (1991). The expression of c-kit protein during oogenesis and early embryonic development. Biol. Reprod. 45,547 -552.[Abstract]
Hubner, K., Fuhrmann, G., Christenson, L. K., Kehler, J.,
Reinbold, R., de la Fuente, R., Wood, J., Strauss, J. F., 3rd, Boiani, M. and
Scholer, H. R. (2003). Derivation of oocytes from mouse
embryonic stem cells. Science
300,1251
-1256.
Kahan, B. W. and Ephrussi, B. (1970). Developmental potentialities of clonal in vitro cultures of mouse testicular teratoma. J. Natl. Cancer Inst. 44,1015 -1036.[Medline]
Kleinsmith, L. J. and Pierce, G. B., Jr (1964). Multipotentiality of single embryonal carcinoma cells. Cancer Res. 24,1544 -1551.[Medline]
Lawson, K. A. and Hage, W. J. (1994). Clonal analysis of the origin of primordial germ cells in the mouse. Ciba Found. Symp. 182,68 -91.[Medline]
Lawson, K. A., Dunn, N. R., Roelen, B. A., Zeinstra, L. M.,
Davis, A. M., Wright, C. V., Korving, J. P. and Hogan, B. L.
(1999). Bmp4 is required for the generation of primordial germ
cells in the mouse embryo. Genes Dev.
13,424
-436.
Ma, C., Fan, L., Ganassin, R., Bols, N. and Collodi, P.
(2001). Production of zebrafish germ-line chimeras from embryo
cell cultures. Proc. Natl. Acad. Sci. USA
98,2461
-2466.
Martin, G. R. (1981). Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells. Proc. Natl. Acad. Sci. USA 78,7634 -7638.[Abstract]
Matsui, Y., Zsebo, K. and Hogan, B. L. (1992). Derivation of pluripotential embryonic stem cells from murine primordial germ cells in culture. Cell 70,841 -847.[Medline]
Pesce, M. and Scholer, H. R. (2001). Oct-4:
gatekeeper in the beginnings of mammalian development. Stem
Cells 19,271
-278.
Rathjen, J., Lake, J. A., Bettess, M. D., Washington, J. M.,
Chapman, G. and Rathjen, P. D. (1999). Formation of a
primitive ectoderm like cell population, EPL cells, from ES cells in response
to biologically derived factors. J. Cell Sci.
112,601
-612.
Resnick, J. L., Bixler, L. S., Cheng, L. and Donovan, P. J. (1992). Long-term proliferation of mouse primordial germ cells in culture. Nature 359,550 -551.[CrossRef][Medline]
Rogers, M. B., Hosler, B. A. and Gudas, L. J. (1991). Specific expression of a retinoic acid-regulated, zinc-finger gene, Rex-1, in preimplantation embryos, trophoblast and spermatocytes. Development 113,815 -824.[Abstract]
Rossant, J. (2001). Stem cells from the
Mammalian blastocyst. Stem Cells
19,477
-482.
Rossant, J., Chazaud, C. and Yamanaka, Y. (2003). Lineage allocation and asymmetries in the early mouse embryo. Philos. Trans. R. Soc. Lond. B Biol. Sci. 358,1341 -1349.[CrossRef][Medline]
Rossant, J. and McBurney, M. W. (1982). The developmental potential of a euploid male teratocarcinoma cell line after blastocyst injection. J. Embryol. Exp. Morphol. 70, 99-112.[Medline]
Rossant, J. and Papaioannou, V. E. (1984). The relationship between embryonic, embryonal carcinoma and embryo-derived stem cells. Cell Differ 15,155 -161.[CrossRef][Medline]
Saitou, M., Barton, S. C. and Surani, M. A. (2002). A molecular programme for the specification of germ cell fate in mice. Nature 418,293 -300.[CrossRef][Medline]
Sato, M., Kimura, T., Kurokawa, K., Fujita, Y., Abe, K., Masuhara, M., Yasunaga, T., Ryo, A., Yamamoto, M. and Nakano, T. (2002). Identification of PGC7, a new gene expressed specifically in preimplantation embryos and germ cells. Mech. Dev. 113, 91-94.[CrossRef][Medline]
Shamblott, M. J., Axelman, J., Wang, S., Bugg, E. M.,
Littlefield, J. W., Donovan, P. J., Blumenthal, P. D., Huggins, G. R.
and Gearhart, J. D. (1998). Derivation of pluripotent stem
cells from cultured human primordial germ cells. Proc. Natl. Acad.
Sci. USA 95,13726
-13731.
Smith, A. G. (2001). Embryo-derived stem cells: of mice and men. Annu. Rev. Cell Dev. Biol. 17,435 -462.[CrossRef][Medline]
Solter, D., Dominis, M. and Damjanov, I. (1981). Embryo-derived teratocarcinoma. III. Development of tumors from teratocarcinoma-permissive and non-permissive strain embryos transplanted to F1 hybrids. Int. J. Cancer 28,479 -483.[Medline]
Solter, D., Skreb, N. and Damjanov, I. (1970). Extrauterine growth of mouse egg-cylinders results in malignant teratoma. Nature 227,503 -504.[Medline]
Stevens, L. C. (1967). Origin of testicular teratomas from primordial germ cells in mice. J. Natl. Cancer Inst. 38,549 -552.[Medline]
Stevens, L. C. (1970a). The development of transplantable teratocarcinomas from intratesticular grafts of pre- and postimplantation mouse embryos. Dev. Biol. 21,364 -382.[Medline]
Stevens, L. C. (1970b). Experimental production of testicular teratomas in mice of strains 129, A/He, and their F1 hybrids. J. Natl. Cancer Inst. 44,923 -929.[Medline]
Stevens, L. (1983). The origin and development of testicular, ovarian, and embryo-derived teratomas. Cold Spring Harb. Conf. Cell Prolif. 10,23 -36.
Stevens, L. C. and Little, C. C. (1954). Spontaneous testicular teratomas in an inbred strain of mice. Proc. Natl. Acad. Sci. USA 40,1080 -1087.
Tam, P. P. and Snow, M. H. (1981). Proliferation and migration of primordial germ cells during compensatory growth in mouse embryos. J. Embryol. Exp. Morphol. 64,133 -147.[Medline]
Tam, P. P. and Zhou, S. X. (1996). The allocation of epiblast cells to ectodermal and germ-line lineages is influenced by the position of the cells in the gastrulating mouse embryo. Dev. Biol. 178,124 -132.[CrossRef][Medline]
Thomson, J. A., Itskovitz-Eldor, J., Shapiro, S. S., Waknitz, M.
A., Swiergiel, J. J., Marshall, V. S. and Jones, J. M.
(1998). Embryonic stem cell lines derived from human blastocysts.
Science 282,1145
-1147.
Toyooka, Y., Tsunekawa, N., Akasu, R. and Noce, T.
(2003). Embryonic stem cells can form germ cells in vitro.
Proc. Natl. Acad. Sci. USA
100,11457
-11462.
Wheeler, J. E. (1983). History of teratomas. In The Human Teratomas: Experimental and Clinical Biology (ed. I. Damjanov, B. B. Knowles and D. Solter), pp.1 -22. Clifton, NJ: Humana Press.
Wylie, C. (1999). Germ cells. Cell 96,165 -174.[Medline]
Xu, R. H., Chen, X., Li, D. S., Li, R., Addicks, G. C., Glennon, C., Zwaka, T. P. and Thomson, J. A. (2002). BMP4 initiates human embryonic stem cell differentiation to trophoblast. Nat. Biotechnol. 20,1261 -1264.[CrossRef][Medline]
Yamazaki, Y., Mann, M. R., Lee, S. S., Marh, J., McCarrey, J.
R., Yanagimachi, R. and Bartolomei, M. S. (2003).
Reprogramming of primordial germ cells begins before migration into the
genital ridge, making these cells inadequate donors for reproductive cloning.
Proc. Natl. Acad. Sci. USA
100,12207
-12212.
Ying, Q. L., Nichols, J., Chambers, I. and Smith, A. (2003). BMP induction of Id proteins suppresses differentiation and sustains embryonic stem cell self-renewal in collaboration with STAT3. Cell 115,281 -292.[CrossRef][Medline]
Ying, Y., Liu, X. M., Marble, A., Lawson, K. A. and Zhao, G.
Q. (2000). Requirement of Bmp8b for the generation of
primordial germ cells in the mouse. Mol. Endocrinol.
14,1053
-1063.
Ying, Y., Qi, X. and Zhao, G. Q. (2001).
Induction of primordial germ cells from murine epiblasts by synergistic action
of BMP4 and BMP8B signaling pathways. Proc. Natl. Acad. Sci.
USA 98,7858
-7862.