The n e w e ng l a n d j o u r na l of m e dic i n e

Cl inic a l I m pl ic a t ions of B a sic R e se a rch

Elizabeth G. Phimister, Ph.D., Editor

Modeling the Placenta with Stem Cells

Toshihiko Ezashi, Ph.D., Danny J. Schust, M.D., and Laura C. Schulz, Ph.D.

Pluripotent stem cells — which have the ability
to differentiate into any cell type in the body —
have enormous potential for regenerative medi-
cine and for uncovering the developmental biol-
ogy underlying health and disease. New advances
in stem-cell technologies are crucial for under-
standing the earliest stages of placental develop-
ment, a period critical to pregnancy success. If
the trophectoderm, the outer shell of the blasto-
cyst, fails to expand, differentiate, or interact
with the uterine lining, there will be no preg-
nancy; infertility is often due to loss of the con-
ceptus at this stage. Less severe disruptions to
early placentation may allow implantation but
lead to miscarriage. Finally, placentation that
can support a full pregnancy, but not optimally,
may result in common and often devastating
pregnancy disorders, such as intrauterine growth
restriction, preeclampsia, and preterm birth.
Unfortunately, the availability of placental
material from the first trimester of pregnancy is
limited. The placenta is essentially inaccessible
to researchers from approximately 1 week after
conception through the earliest possible detec-
tion of pregnancy, at 2 to 3 weeks after concep-
tion. Studies in animals have shown that there
are conserved pathways of placental development,
but the enormous diversity of mammalian pla-
centas renders them insufficient for providing a
full understanding of the human placenta. Most
of what we know of the initial development of
the human placenta comes from a handful
of archived specimens from the 1930s through
the 1960s, housed in the Carnegie collection
in Washington, D.C., and the Boyd collection in
Cambridge, United Kingdom.
Previously described stem-cell types, such as
embryonic stem cells (ESCs), can be used to ad-
dress the lack of early placental samples by reca-
pitulating many aspects of placental trophoblast
cell differentiation (Fig. 1). Primed human ESCs
are derived from cells that make up a part of the
inner cell mass of the blastocyst, called the epi-
blast. Thus, they are derived from cells that have
already separated from the trophectoderm lineage,
and they do not spontaneously differentiate into
trophoblasts in teratomas. However, on treatment
with BAP (growth factor bone morphogenetic
protein 4 [BMP4] plus the signaling inhibitors
A083-01 and PD173074), these ESCs efficiently dif-
ferentiate into placental trophoblast-type cells.2
There are drawbacks to this system. Once
ESCs have terminally differentiated into tropho-
blasts, they can no longer be propagated; thus,
ESCs must be differentiated into trophoblast-type
cells for each short-term experiment. Moreover,
although the resulting cells are uniformly tropho-
blast, they contain a mixture of trophoblast
subtypes, which makes it difficult to study spe-
cific trophoblast lineages. Short-term (24-hour)
treatment of ESCs with BAP leads to a stable
population of cells (“BAP-primed cells”) that can
spontaneously differentiate into trophoblasts as
well as into the three embryonic lineages (in the
context of induced teratomas) and can be directed
toward differentiation into specific trophoblast
sublineages; however, they cannot be maintained,
in vitro, as trophoblasts.3 Human trophoblast
stem cells were recently created from tropho-
blast cells of blastocysts, as well as from first-
trimester placental trophoblasts.4 In contrast to
ESC-derived trophoblast cells, trophoblast stem
cells can be propagated, and differentiation into
particular trophoblast subtypes can be effected
by culturing the cells in the presence of specific
factors. Trophoblast stem cells and ESCs share
the drawback of requiring derivation from con-
ceptus tissues, which raises ethical concerns for

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 The n e w e ng l a n d j o u r na l of m e dic i n e

Mixed trophoblast
cells
Induced pluripotent
Adult cells stem cells

Any cell in the body

Epiblast

Primed ESCs Expanded-potential

Blastocyst
stem cells

Extravillous
trophoblast cells

First-trimester Trophoblast stem cells

human placental
trophoblast
Syncytiotrophoblast

Figure 1. Stem Cells — Sources and Potential.

Sources and differentiation capacities of expanded potential stem cells, recently reported by Gao et al.,1 are shown
in comparison with those of previously reported embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs),
and trophoblast stem cells. Human ESCs are derived from the blastocyst inner cell mass, and iPSCs are derived from
a broad range of somatic cells (purple arrows). They share extensive molecular and cellular characteristics, as well
as similar morphology in cell culture. ESCs and iPSCs differentiate into a mixture of trophoblast subtypes when ex-
posed to BAP (bone morphogenetic protein 4 [BMP4] plus the signaling inhibitors A083-01 and PD173074) and have
the capacity to differentiate into any embryonic cell type (pluripotency). Human trophoblast stem cells are derived
from trophoblast cells of either blastocysts or first-trimester placental villi (red arrows). Trophoblast stem cells can
self-renew and can be directed toward either of the major trophoblast subtypes of the placenta (extravillous tropho-
blast and syncytiotrophoblast) on exposure to defined culture conditions. By shifting cell signaling (blue arrows),
ESCs and iPSCs can be converted into expanded-potential stem cells, which retain the ability to differentiate to any
embryonic lineage, as well as into trophoblast cells (on exposure to BAP). They are, however, unique in that they can
be converted to trophoblast stem cells.

some. The use of conceptus tissues also raises
political and regulatory uncertainties, as is evi-
dent in the recently announced human fetal tis-
sue policy of the Department of Health and
Human Services, which limits federally funded
research using human fetal tissue. Induced pluri-
potent stem cells (iPSCs) are created by repro-
gramming adult cells by transducing them with
four to six “pluripotency” genes,5 but iPSCs have
the same trophoblast-differentiation potential as
ESCs and similarly, trophoblast-like cells derived
from them cannot be propagated.

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 Clinical Implications of Basic Research

Gao and colleagues1 have recently reported a
new type of stem cell: the expanded-potential
stem cell, which combines the advantages of the
other stem cell types (Fig. 1). After screening
400 combinations of 20 factors, they arrived at a
novel cocktail that, when added to ESCs or iPSCs,
produces stable expanded-potential stem-cell cul-
tures that can be efficiently differentiated into
trophoblasts — and some lines may be con-
verted into trophoblast stem cells, something
that had not been accomplished with other types
of stem cell. Consequently, iPSCs (which can be
obtained without the use of conceptus tissues)
can be converted into expanded-potential stem
cells and then into trophoblast stem cells, which
can be maintained over many passages. Previ-
ously, trophoblast stem cells could be made only
from first-trimester placental tissue, when key
conditions of trophoblast dysfunction such as
preeclampsia, growth restriction, and signs asso-
ciated with preterm birth are not apparent. The
method described by Gao et al. makes it possible
to collect cells after delivery from a complicated
pregnancy, convert them into iPSCs, and then
convert the iPSCs into expanded-potential stem
cells that can then be differentiated into stable
trophoblast stem-cell lines to model placental
diseases.
Gao et al. also report the derivation of expanded-
potential stem cells from porcine stem cells and
blastocysts. Pigs offer a number of advantages as
a model for human health, including their simi-
larities to humans with regard to body size and
immune systems, as well as their life span,
which is long relative to that of rodents. These
well suited for the testing of personalized cell or
organ transplants derived from autologous iPSC
lines, a major goal of regenerative medicine. The
engineering of porcine expanded pluripotent stem
cells reported by Gao et al. represents a major
step in overcoming long-standing difficulties in
establishing authentic porcine pluripotent stem
cells; these difficulties have included continued
expression of exogenous pluripotency genes, poor
expression of endogenous pluripotency genes,
and limited capacity to differentiate.
The development by Gao et al. of novel stem-
cell methods that avoid the use of embryo-derived
cells while permitting research into developmen-
tal biology and regenerative and personalized
medicine is particularly welcome in the face of
continued ethical, political, and regulatory chal-
lenges to the stem-cell field.
Disclosure forms provided by the authors are available with
the full text of this article at NEJM.org.
From the Division of Animal Sciences (T.E.) and the Depart-
ment of Obstetrics, Gynecology and Women’s Health (D.J.S.,
L.C.S.), University of Missouri, Columbia.
1. Gao X, Nowak-Imialek M, Chen X, et al. Establishment of
porcine and human expanded potential stem cells. Nat Cell Biol
2019;​21:​687-99.
2. Amita M, Adachi K, Alexenko AP, et al. Complete and unidi-
rectional conversion of human embryonic stem cells to tropho-
blast by BMP4. Proc Natl Acad Sci U S A 2013;​110(13):​E1212-
E1221.
3. Yang Y, Adachi K, Sheridan MA, et al. Heightened potency of
human pluripotent stem cell lines created by transient BMP4
exposure. Proc Natl Acad Sci U S A 2015;​112(18):​E2337-E2346.
4. Okae H, Toh H, Sato T, et al. Derivation of human tropho-
blast stem cells. Cell Stem Cell 2018;​22(1):​50-63.e6.
5. Takahashi K, Tanabe K, Ohnuki M, et al. Induction of plu-
ripotent stem cells from adult human fibroblasts by defined fac-
tors. Cell 2007;​131:​861-72.

features enable testing of human surgical tech- DOI: 10.1056/NEJMcibr1907773

niques and imaging methods. They make pigs Copyright © 2019 Massachusetts Medical Society.

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Copyright © 2019 Massachusetts Medical Society. All rights reserved.