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Biological characteristics of stem cells from foetal, cord blood

and extraembryonic tissues
Hassan Abdulrazzak, Dafni Moschidou, Gemma Jones and Pascale V. Guillot
J. R. Soc. Interface published online 25 August 2010
doi: 10.1098/rsif.2010.0347.focus

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 Downloaded from rsif.royalsocietypublishing.org on August 28, 2010

J. R. Soc. Interface
doi:10.1098/rsif.2010.0347.focus
Published online

REVIEW

Biological characteristics of stem cells

from foetal, cord blood and
extraembryonic tissues
Hassan Abdulrazzak, Dafni Moschidou, Gemma Jones
and Pascale V. Guillot*
Institute of Reproductive and Developmental Biology, Imperial College London,
London W12 0NN, UK
Foetal stem cells (FSCs) can be isolated during gestation from many different tissues such as
blood, liver and bone marrow as well as from a variety of extraembryonic tissues such as
amniotic fluid and placenta. Strong evidence suggests that these cells differ on many biolo-
gical aspects such as growth kinetics, morphology, immunophenotype, differentiation
potential and engraftment capacity in vivo. Despite these differences, FSCs appear to be
more primitive and have greater multi-potentiality than their adult counterparts. For
example, foetal blood haemopoietic stem cells proliferate more rapidly than those found in
cord blood or adult bone marrow. These features have led to FSCs being investigated for
pre- and post-natal cell therapy and regenerative medicine applications. The cells have
been used in pre-clinical studies to treat a wide range of diseases such as skeletal dysplasia,
diaphragmatic hernia and respiratory failure, white matter damage, renal pathologies as
well as cancers. Their intermediate state between adult and embryonic stem cells also
makes them an ideal candidate for reprogramming to the pluripotent status.

Keywords: stem cell; foetal; regenerative medicine; induced pluripotent stem

1. INTRODUCTION work has shown that ES cells can be derived from

single blastomeres isolated using procedures similar to
Stem cells are undifferentiated cells with the capacity to
those routinely used for pre-implantation genetic diag-
self-renew, differentiate and repopulate a host in vivo
nosis (Klimanskaya et al. 2007). However, safety
(Weissman et al. 2001). Their plasticity or potency is
concerns still remain because of the tumorigenicity of
hierarchical ranging from totipotent (differentiating
ES cells. Alternatively, adult stem cells can be found
into all cell types including placenta), pluripotent (dif-
in almost all tissues examined including brain, dental
ferentiating into cells of the three germ layers,
pulp, muscle, bone marrow, skin and pancreas and
ectoderm, mesoderm and endoderm, but not tropho-
have been extensively characterized for their thera-
blastic cells), multipotent (differentiating into cells of
peutic potential. The adult stem cell could be multi-
more than one type but not necessarily into all the
potent (e.g. haematopoietic stem cells (HSCs) giving
cells of a given germ layer) to unipotent (differentiating
rise to all blood cells and adherent stromal/mesenchy-
into one type of cell only, e.g. muscle or neuron).
mal stem cells (MSCs) that give rise to bone, fat,
Pluripotent stem cells can be derived from the inner
cartilage and muscle) or unipotent (e.g. progenitor
cell mass of the pre-implantation embryo (i.e. embryo-
cells). Adult MSCs have the problem of being difficult
nic stem (ES) cells) or isolated from the foetal
to extract in sufficient numbers for therapy and/or pre-
primordial germ cell pool (PGC) above the allantois
senting restricted plasticity and limited proliferative
(i.e. embryonic germ (EG) cells and embryonic carci-
capacity compared with ES cells.
noma (EC) cells; Thomson et al. 1998; Andrews et al.
In recent years, foetal stem cells (FSCs) and stem
2005). The destruction of the blastocyst or early
cells isolated from cord blood or extraembryonic tissues
foetus necessary for their derivation/isolation raises
have emerged as a potential ‘half way house’ between
ethical concerns (Lo & Parham 2009), although recent
ES cells and adult stem cells. FSCs can be found
in foetal tissues such as blood, liver, bone marrow,
*Author for correspondence ( pascale.guillot@imperial.ac.uk). pancreas, spleen and kidney, and stem cells are also
One contribution to a Theme Supplement ‘Translation and found in cord blood and extraembryonic tissues such
commercialization of regenerative medicines’. as amniotic fluid, placenta and amnion (Marcus &

Received 30 June 2010

Accepted 5 August 2010 1 This journal is # 2010 The Royal Society
 Downloaded from rsif.royalsocietypublishing.org on August 28, 2010
2 Review. Characteristics of stem cells H. Abdulrazzak et al.
Woodbury 2008). Their primitive properties, expansion
potential and lack of tumorogenicity make them an
attractive option for regenerative medicine in cell
therapy and tissue engineering settings. While extraem-
bryonic tissues could be used with few ethical
reservations, the isolation of FSCs from abortuses is
subject to significant public unease. We review here
the phenotypic characteristics of stem cells from
foetal, cord blood and extraembryonic tissues, their
application in cell therapy and their potential for repro-
gramming towards pluripotentiality.
2. PHENOTYPIC CHARACTERISTICS OF
FOETAL STEM CELLS
Stem cells are collected from abortal foetal tissue,
surplus of pre-natal diagnostic tissues or tissues at
delivery, subject to informed consent, institutional
ethics approval and compliance with national guidelines
covering foetal tissue research. Until recently they
were thought to be multi-potent (O’Donoghue & Fisk
2004), but this picture is now changing as evidence is
mounting regarding the existence of pluripotent sub-
populations in some foetal and extraembryonic tissues
(Cananzi et al. 2009). Some of the properties of the
cells are summarized in table 1.
2.1. Foetal tissues
2.1.1. Foetal MSCs. MSCs are multi-potent stem cells
that can differentiate towards mesoderm-derived
lineages (i.e. osteogenic, adipogenic, chondrogenic and
myogenic; Pittenger et al. 1999). Their potential to
repair damaged tissue and their immunomodulatory
properties make them very attractive for a wide range
of regenerative medicine applications such as cell
therapy and tissue engineering for hollow and solid
organs (Horwitz et al. 2002; Abdallah & Kassem
2008; Le Blanc et al. 2008). They were first identified
in adult bone marrow where they represent 0.001–
0.01% of total nucleated cells (Owen & Friedenstein
1988). Adherent stromal cells with similar characteristics
were subsequently isolated from other tissues such as adi-
pose, dental pulp, muscle, liver and brain (Porada et al.
2006; Huang et al. 2009). They are also found in foetal
and extraembryonic tissues, where they seem to possess
greater proliferation capacity and differentiation poten-
tial than their adult counterparts (Campagnoli et al.
2001; Guillot et al. 2007b; Pappa & Anagnou 2009).
2.1.1.1. Common characteristics. MSCs isolated from
foetal tissues such as blood, liver, bone marrow, lung
and pancreas all share common characteristics. For
example, they are spindle-shaped cells with the capacity
to differentiate into the standard mesenchymal
lineages, i.e. bone, fat and cartilage. They do not
express haematopoietic or endothelial markers (e.g.
they are CD452/342/142 and von Willebrand factor
negative; Gucciardo et al. 2009), but express stroma-
associated markers CD29 (b1-integrin), CD73 (SH3
and SH4), CD105 (SH2), CD44 (HCAM1), the early
bone marrow progenitor cell marker CD90 (thy-1) and
the extracellular matrix proteins vimentin, laminin and
J. R. Soc. Interface
fibronectin (Guillot et al. 2006, 2007a). Contrary to
adult bone marrow MSCs, first-trimester foetal blood,
liver and bone marrow MSCs express baseline levels of
the pluripotency stem cell markers Oct-4, Nanog, Rex-1,
SSEA-3, SSEA-4, Tra-1-60 and Tra-1-81 (Guillot et al.
2007b; Zhang et al. 2009). Regardless of their tissue of
origin, first-trimester foetal MSCs self-renew faster in cul-
ture than adult MSCs (30–35 h versus 80–100 h) and
senesce later while retaining a stable phenotype (Guillot
et al. 2007b). Foetal MSCs are therefore more readily
expandable to therapeutic scales for either pre- or post-
natal ex vivo gene or cell therapy and tissue engineering.
Having greater multi-potentiality and differentiating
more readily than adult MSCs into cells of mesodermal
origin such as bone and muscle, they can also differentiate
into cells from other lineages such as oligodendrocytes
and haematopoietic cells (MacKenzie et al. 2001; Chan
et al. 2006; Kennea et al. 2009). In terms of engraftment,
they have a competitive advantage compared with adult
cells (Rebel et al. 1996; Harrison et al. 1997; Taylor et al.
2002). Other advantageous characteristics relevant to cell
therapy include having active telomerase (Guillot et al.
2007b), expressing low levels of HLA I and lacking intra-
cellular HLA II, and taking longer to express this upon
interferon g stimulation than adult MSCs (Gotherstrom
et al. 2004). Foetal MSCs also express a shared a2, a4
and a5b1 integrin profile with first-trimester HSCs, impli-
cating them in homing and engraftment, and, consistent
with this, they have significantly greater binding to
their respective extracellular matrix ligands than adult
MSCs (de la Fuente et al. 2003). Finally, human foetal
MSCs are readily transducible with transduction efficien-
cies of greater than 95 per cent using lentiviral vectors
with stable gene expression at both short- and long-time
points without affecting self-renewal or multi-potentiality
(Chan et al. 2005).
2.1.1.2. Specific characteristics. Although the above
characteristics are shared among the various types of
foetal stem, they also have specific traits that are depen-
dent on tissue of origin and gestational age. For example,
foetal MSCs are present in foetal blood from the earliest
gestation tested, seven weeks, where they account for
approximately 0.4 per cent of foetal nucleated cells.
Their numbers decline to very low levels after 13
weeks’ gestation (Campagnoli et al. 2001). MSC derived
from foetal bone marrow also decline with age with one
MSC found among 10 000 in mid-trimester as compared
with one MSC per 250 000 cells in adult marrow.
Significant variations have been found among foetal
tissue cells in terms of cell surface markers. For
example, foetal lung has a higher proportion of
CD34þ/CD452 cells (44%) than bone marrow (4.8%),
spleen (12.6%) and liver (7.5%) (in’t Anker et al.
2003a,b). Also foetal metanephric MSCs do not express
CD45, CD34 or other haematopoietic markers; instead,
they express high levels of mesenchymal markers such
as vimentin, laminin and type I collagen.
There is also variation among the cells in terms of
differentiation capacity. MSCs derived from foetal
liver during the first and second trimester have a
reduced osteogenic differentiation potential in
 Table 1. Characteristics of stem cells. E, embryonic; MSC, mesenchymal; H, haematopoietic; EP, epithelial; VSEL, very small embryonic like; P, pluripotent; M, multipotent; U,
unipotent; n.d., not determined. Asterisk denotes that the characteristics of these cells are not listed in the table (Kyo et al. 1997; Campagnoli et al. 2001; Gammaitoni et al. 2004; Miki
et al. 2005, 2007; Pranke et al. 2005; Dan et al. 2006; Habich et al. 2006; Miki & Strom 2006; Portmann-Lanz et al. 2006; Zhao et al. 2006; De Coppi et al. 2007a; Fong et al. 2007; Guillot

J. R. Soc. Interface
et al. 2007b; Ilancheran et al. 2007; Karahuseyinoglu et al. 2007; Kim et al. 2007; Li et al. 2007; Musina et al. 2007; Prat-Vidal et al. 2007; Degistirici et al. 2008; Moon et al. 2008;
Mountford 2008; Poloni et al. 2008; Sessarego et al. 2008; Troyer & Weiss 2008; Brooke et al. 2009; Cananzi et al. 2009; Riekstina et al. 2009; Zhang et al. 2009; Alaminos et al. 2010;
Anzalone et al. 2010; Castrechini et al. 2010; Hsieh et al. 2010; Insausti et al. 2010; Vaziri et al. 2010).

foetal tissues extraembryonic

embryonic stem cord Wharton’s amnion-derived amnion- chorion-

cells blood liver bone marrow blood jelly amniotic fluid epithelial cells derived MSC derived MSC adult BM MSC

cell phenotype (s) E MSC/H* MSC/H* MSC/H* MSC/H* MSC MSC/AFS/VSEL* EP MSC MSC MSC
doubling time (DT)/ 48– 24 h (DT)/ 21.9 + 1.1 h 46 h (second- 32 h (second- 39– 43 h 85– 11 h 25–38 h (second- 38.4 h (DT) 30c (PD) 21.25 + 3.2 h 54–111 h (DT)/
population doublings indefinite (first-trimester trimester trimester (DT)/ (DT) trimester MSC) (DT)/30d 7.1a (PD)
(PD) (PD) MSC) (DT)/ MSC) (DT) MSC) 20b 36 h (AFS) (DT)/
28.4a (PD) (DT) (PD) 66 (MSC)/over
250 AFS (PD)
feeder/matrigel þ 2 2 2 2 2 2 2 2 2 2
requirement
potency P P/M P/M P/M P/M P/M P/M P/M P/M P/M M
Oct-4 þþþ þþ þþ þ þ þ þ þ þ þ +
Sox2 þþþ n.d. n.d. n.d. þ 2 þ n.d. n.d. +
Nanog þþþ þþ þþ þþ þ þ + þ n.d. þ +
c-Myc þþþ n.d. n.d. n.d. n.d. n.d. þ þ n.d. n.d. n.d.
Klf-4 þþþ n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d.
TRF length/telomerase þþþ þþ þþ þþ þþ þ n.d. 2 n.d. þ 2
activity
alkaline phosphatase þþþ n.d. n.d. þþ þ þ þ 2 n.d. +
c-Kit (CD117) þþþ þ þ þ + þ þ + + 2 2
Rex-1 þþþ þþ þ þþ þ n.d. þ þ 2 þ +
SSEA-4 þþþ þþ þþ þþ þ þ þ þ 2 þ 2
SSEA-3 þþþ þþ þþ þþ þ 2 2 þ(low) 2 þ 2
Downloaded from rsif.royalsocietypublishing.org on August 28, 2010

Tra-1-81 þþþ þþ þþ þþ n.d. þ 2 þ 2 þ 2

Tra-1-61 þþþ þþ þþ þþ n.d. þ + þ 2 þ 2
Review. Characteristics of stem cells

MHC class I þ(low) þ(low) þ(low) þ(low) þ þ þ þ(low) þ(low) þ þ

MHC class II (HLA-DR) 2 2 2 2 + 2 + 2 2 2 2
teratoma formation in yes n.d. n.d. n.d. n.d. n.d. no no n.d. n.d. no
immune compromised
mice
a
Over a period of 50 days.
b
Over eight passages.
c
Over 14 passages.
d
Over five passages.
H. Abdulrazzak et al. 3
 Downloaded from rsif.royalsocietypublishing.org on August 28, 2010
4 Review. Characteristics of stem cells H. Abdulrazzak et al.
comparison with first-trimester foetal blood and with
second-trimester spleen, lung and bone marrow MSCs,
possibly because of a reduction in osteogenic progenitor
numbers (in‘t Anker et al. 2003a,b). Foetal pancreatic
MSCs, shown to be capable of differentiating into chon-
drogenic, osteogenic or adipogenic lineages (Hu et al.
2003), can also engraft, differentiate and secrete
human insulin in a sheep model (Ersek et al. 2010).
Moreover, foetal pancreatic ductal stem cells can
differentiate into insulin-producing cells in vitro (Yao
et al. 2004).
Foetal metanephric MSCs can be induced in vitro to
adopt an osteogenic or myogenic phenotype and, when
transplanted in a foetal lamb model, they show long-
term persistence and site-specific differentiation
(Almeida-Porada et al. 2002).
Altogether these data indicate a high level of
heterogeneity within the foetal stem cell pool.
2.1.2. Foetal HSCs. HSCs are multi-potent stem cells
that maintain functional haematopoiesis by generation
of all haematopoietic lineages throughout foetal and
adult life (Weissman & Shizuru 2008). They are charac-
terized by the expression of CD34 and CD45 antigens,
and the absence of markers such as CD38 and human
leucocyte antigen (HLA)/DRE (Huss 2000; Taylor
et al. 2002). During ontogeny the site of haematopoiesis
is modified several times (Cumano et al. 2001). An area
along the dorsal embryonic aorta termed the aorta–
gonad–mesonephros (AGM) is a rich source of HSCs,
which then migrate to the embryonic liver and later
seed other haematopoietic tissues such as the bone
marrow (McGrath & Palis 2008). HSCs are usually
assayed in animal models based on their capacity to
repopulate the entire haematopoietic system in con-
ditioned recipients after transplantation. Similar to
MSCs, it is possible to transplant human cells into xeno-
geneic recipients such as foetal sheep that act as models
of human haematopoiesis (O’Donoghue & Fisk 2004).
First-trimester foetal blood contains more CD34þ
cells than term gestation blood, yet CD34þ cells
account for only 5 per cent of CD45þ cells in the
blood at that stage (Campagnoli et al. 2001). The
number of circulating HSCs increases from the first tri-
mester to peak in the second trimester in utero,
probably because of cells migrating from the foetal
liver to establish haematopoiesis in the foetal bone
marrow (Clapp et al. 1995). Some HSCs remain in the
umbilical cord at delivery, where they can be collected
for allogeneic or occasionally autologous cell transplan-
tation (see §2.2.1.).
In the second trimester, CD34þ cells constitute 4 per
cent of cells in blood, 16.5 per cent in bone marrow,
6 per cent in liver, 5 per cent in spleen and 1.1 per
cent in the thymus (Lim et al. 2005). This frequency
of CD34þ cells in the blood gradually diminishes
during the third trimester, probably reflecting establish-
ment of the marrow as the primary site of
haematopoiesis and the declining role of the foetal
liver in that regard ( Jones et al. 1994; Wagers et al.
2002). The number of cells negative for CD38 within
the CD34þ population is also higher in early foetal
J. R. Soc. Interface
blood, suggesting that these cells are more primitive
and have greater potential than HSCs circulating later
in ontogeny (O’Donoghue & Fisk 2004). Even relatively
differentiated erythroblasts are more primitive in first
compared with later trimester foetal blood. Foetal
blood HSCs proliferate more rapidly than those in
cord blood or adult bone marrow, and produce all
haematopoietic lineages (Campagnoli et al. 2001).
Foetal liver is also a source of HSCs, and, despite
their relatively small number, it is possible to generate
sufficient numbers of cells for transplantation in four
weeks (Rollini et al. 2004), although, to our knowledge,
it has not been proved that foetal liver HSCs can suc-
cessfully transplant a human recipient. The phenotype
of foetal liver HSCs changes with gestation age. For
example, their differentiation potential decreases mark-
edly with gestational age, and cells that expressed both
CD34 and c-kit (CD117) were identified in the first-
trimester foetal liver whereas the cells expressing more
committed hepatic markers only appeared during the
second trimester (Nava et al. 2005).
2.2. Extraembryonic tissue
2.2.1. Umbilical cord blood. Umbilical cord blood is now
an established source of transplantable HSCs that have
a greater proliferative capacity, lower immunological
reactivity and lower risk of graft-versus-host disease
(GVHD) than those derived from adult bone marrow
(Broxmeyer et al. 1989; Yu et al. 2001; Ballen 2005;
Schoemans et al. 2006; Brunstein et al. 2007; Hwang
et al. 2007; Broxmeyer 2010). These cells are capable
of repopulating bone following intra-bone injection of
severe combined immunodeficiency mice (Mazurier
et al. 2003; Wang et al. 2003) and are used clinically
as an alternative to adult bone marrow HSCs in some
cases (Delaney et al. 2010). The cells are CD34þ and
CD382 and their frequency is greater than that of
bone marrow or peripheral blood following cytokine
mobilization (Pappa & Anagnou 2009). It has also
been demonstrated that cord blood contains MSCs
(Weiss & Troyer 2006; Secco et al. 2008) that can sup-
port the in vivo expansion of HSCs and function as an
accessory cell population for engraftment ( Javazon
et al. 2004; Wang, J. et al. 2004). The frequency of
MSCs in umbilical cord blood is low however, with
MSCs successfully isolated from only a third of all
samples collected (Bieback et al. 2004).
Cord blood stem cells expressing baseline levels of ES
cell markers such as Oct-4, Nanog, SSEA-3 and SSEA-4
have also been described (Zhao et al. 2006). Using a
two-stage isolation approach whereby the erythrocytes
in cord blood are lysed and the remaining cells are
sorted using flow cytometry, it has been possible to iso-
late and enrich for a subpopulation of cells that are
CXCR4þ, CD133þ, CD34þ, Lin2 and CD452.
These cells have been described as very small
embryonic-like (VSEL) stem cells because they are
only 3 – 5 mm in diameter and express Oct-4, Nanog
and SSEA-4 (Kucia et al. 2007; Zuba-Surma et al.
2010). Compared with MSCs, VSEL cells are smaller,
have a large nucleus to cytoplasm ratio and open-type
chromatin. As a similar population has also been
 Downloaded from rsif.royalsocietypublishing.org on August 28, 2010
Review. Characteristics of stem cells
found in adult bone marrow, where they also express the
same primitive markers, it has been hypothesized that
VSEL cells are related to a population of early PGCs
that are retained during development (Kucia et al.
2006).
2.2.2. Wharton’s jelly. Wharton’s jelly is the connective
tissue surrounding the umbilical vessels and includes
the perivascular, intervascular and subamnion regions
(Troyer & Weiss 2008). MSCs have been isolated from
all three regions and it remains unclear whether they
represent distinct cell populations (Karahuseyinoglu
et al. 2007). The cells have been given different names
by different groups (Troyer & Weiss 2008). MSCs iso-
lated from Wharton’s jelly share similar properties
with other cord blood MSCs as well as adult bone
marrow MSCs on the expression of markers, differen-
tiation potential and cytokine production (McElreavey
et al. 1991; Wang, H.-S. et al. 2004; Anzalone et al.
2010). Wharton’s jelly MSCs can also be induced to
differentiate into cells similar to neural, expressing
neuron-specific enolase and other neural markers
(Mitchell et al. 2003; Anzalone et al. 2010).
2.2.3. Amniotic fluid. In recent years, amniotic fluid has
emerged as a major source of putative pluripotent stem
cells that avoid many of the problems associated with
ES cells such as their non-suitability for autologous
use, their capacity for tumour formation and the ethical
concerns they raise.
In humans, the amniotic fluid starts to appear at the
beginning of week 2 of gestation as a small film of
liquid between the cells of the epiblast. The fluid expands
separating the epiblast (i.e. the future embryo) from the
amnioblasts (i.e. the future amnion), thus forming the
amniotic cavity (Miki & Strom 2006). The origin of
amniotic fluid cells is still very much debatable
(Kunisaki et al. 2007; Cananzi et al. 2009). However,
what is known is that the majority of cells present are
terminally differentiated and have limited proliferative
capacity (Gosden & Brock 1978; Siegel et al. 2007). How-
ever, a number of studies have demonstrated the presence
of a subset of cells with a proliferative and differentiation
potential (Torricelli et al. 1993; Streubel et al. 1996).
A variety of different types of stem cells have been iso-
lated and characterized from amniotic fluid. These
include cells found in mid-gestation expressing the haema-
topoietic marker CD34 (Da Sacco et al. 2010) as well as
cells with mesenchymal features, able to proliferate in
vitro more rapidly than comparable foetal and adult
cells (Kaviani et al. 2001; in‘t Anker et al. 2003a,b;
Nadri & Soleimani 2007; Roubelakis et al. 2007; Sessarego
et al. 2008). Amniotic fluid MSCs are negative for
haematopoietic markers such as CD45, CD34 and CD14
(Prusa & Hengstschlager 2002; in‘t Anker et al. 2003a,b;
Prusa et al. 2003; Tsai et al. 2004). Despite their high pro-
liferation rate, these cells display a normal karyotype
when expanded in vitro and do not form tumours in
vivo (Sessarego et al. 2008). They exhibit a broad differen-
tiation potential towards mesenchymal lineages (in‘t
Anker et al. 2003a,b; Kolambkar et al. 2007; Nadri &
Soleimani 2007; Tsai et al. 2007).
J. R. Soc. Interface
H. Abdulrazzak et al. 5
De Coppi et al. isolated c-kit-positive (CD117) cells
that represent about 1 per cent of cells present in
second-trimester amniotic fluid. These cells were
named amniotic fluid stem (AFS) cells. They can be
cultured without feeders, double in 36 h, are not
tumorigenic, have long telomeres and retain a normal
karyotype for over 250 population doublings (De
Coppi et al. 2007a). Cultured human AFS cells are posi-
tive for ES cell (e.g. Oct-4, Nanog and SSEA-4) and
mesenchymal cell markers such as CD90, CD105
(SH2), CD73 (SH3/4) and several adhesion molecules
(e.g. CD29 and CD44; Tsai et al. 2006; Chambers et al.
2007; De Coppi et al. 2007a). Furthermore, it was poss-
ible to generate clonal human lines from these cells,
verified by retroviral marking, which were capable of dif-
ferentiating into lineages representative of all three EG
layers. Almost all clonal AFS cell lines express Oct-4
and Nanog, markers of a pluripotent undifferentiated
state (De Coppi et al. 2007a). c-kitþ Lin2 cells from
human and mouse amniotic fluid display a multi-lineage
haematopoietic potential in vitro and in vivo, despite
having low or negative CD34 expression (Ditadi et al.
2009). c-kit-positive human AFS cell lines can also
form embryoid bodies (EB) with an incidence of
18–82%. EB formation was accompanied by a decrease
in Oct-4 and Nodal and the induction of differentiation
markers such as Pax 6, Nestin (ectodermal), GATA 4
and HBE1 (mesodermal) (Valli et al. 2010). The poten-
tial for a wide range of regenerative medicine and tissue
engineering applications as well as their relatively few
ethical concerns makes them an attractive source
for cell therapy, particularly considering that a bank of
100 000 amniotic fluid specimens could potentially
supply 99 per cent of the US population with a perfect
match for transplantation (Atala 2009).
2.2.4. Placenta. The placenta is a fetomaternal organ
involved in maintaining foetal tolerance and allows
nutrient uptake and gas exchange with the mother,
but also contains a high number of progenitor cells or
stem cells (Parolini et al. 2010). It has two sides: one
foetal (amnion and chorion) and one maternal (decid-
uas). The amnion membrane contains two cell types:
the amniotic epithelial cells (AECs) derived from the
epiblast; and amniotic mesenchymal cells derived from
the hypoblast. The chorion layer consists of cytotropho-
blasts and syncytiotrophoblasts that are derived from
the outer layer of the blastocyst (trophectoderm)
(Ilancheran et al. 2009; Insausti et al. 2010).
The availability, phenotypic plasticity and immuno-
modulatory properties of placenta-derived progenitor/
stem cells are useful characteristics for cell therapy
and tissue engineering. Cells can be isolated during
ongoing pregnancy using minimally invasive techniques
such as chorionic villus sampling (CVS) and placental
tissues are readily available at delivery for allogeneic
or autologous use. Cells that have been isolated from
placenta include the human AECs, human amnion
mesenchymal stromal stem cells (AMSCs), human
chorionic mesenchymal/stromal stem cells (CMSCs),
human chorionic trophoblastic cells and HSCs (Parolini
et al. 2008).
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6 Review. Characteristics of stem cells H. Abdulrazzak et al.
Mesenchymal placenta cells are plastic adherent,
share a similar immunophenotype and have lineage
differentiation potential. They express stromal markers
such as CD166, CD105, CD73, CD90 and others, while
they are negative for the haematopoietic markers CD14,
CD34 and CD45 (Igura et al. 2004; Sudo et al. 2007).
Additionally, they express pluripotency markers such as
SSEA3, SSEA4, Oct-4, Nanog, Tra-1-60 and Tra-1-81
(Yen et al. 2005; Battula et al. 2007). They lack or have
very low expression of MHC class I antigens and do not
express MHC class II antigens or the T-cell co-stimulatory
molecule B7, giving the cells immunomodulatory
properties (Bailo et al. 2004; Miao et al. 2006).
2.2.4.1. Amniotic epithelial cells. AECs are commonly
isolated from the amniotic membrane using digestive
enzymes. The cells are plastic adherent and grow under
MSC conditions, i.e. Dulbecco’s modified Eagle’s
medium (DMEM) supplemented with 10–20% FBS,
with the addition of growth factors such as epidermal
growth factor (EGF) (Miki et al. 2010). c-Kit and CD90
(Thy-1) expression is either negative or at a low level
(Miki et al. 2005; Miki & Strom 2006). However, evidence
suggests that they express pluripotency markers and have
the ability to form xenogeneic chimera with mouse ES
cells in vitro (Tamagawa et al. 2004). The cells have sub-
sequently been differentiated in vitro into cell types from
all three germ layers (Miki et al. 2005; Miki & Strom
2006; Ilancheran et al. 2007; Parolini et al. 2008).
However, in contrast with ES cells, Miki et al. (2005)
demonstrated that human AE cells did not form gross
or histological tumours up to seven months post-
transplantation in SCID/Beige mice.
2.2.4.2. Amnion-derived mesenchymal stromal cells.
Both amnion and chorion MSCs have been extensively
characterized and can be isolated throughout gestation
from first trimester to delivery. Both cells share
common characteristics, such as plastic adherence,
and, despite limited proliferation capacity, show in
vitro differentiation down the osteogenic, adipogenic,
chrondrogenic and neurogenic lineages, although some
report and show in vivo multi-organ engraftment
capacity. In addition, the volume of term placenta
makes it an attractive source of stem cells, as on average
human term placenta weighs more than 590 g
(Bolisetty et al. 2002). Human placenta contains var-
ious types of cells of different developmental origin.
Cells of the chorion and decidua are derived from tro-
phectoderm, while amnion is derived from the epiblast
of the developing embryo (Crane & Cheung 1988).
MSCs have been successfully isolated from first-,
second- and third-trimester placental compartments,
including the amnion, chorion, decidua parietalis and
decidua basalis (in‘t Anker et al. 2004; Portmann-
Lanz et al. 2006; Soncini et al. 2007; Poloni et al.
2008), and represent less than 1 per cent of cells present
in the human placenta (Zhang et al. 2004; Alviano et al.
2007). First-trimester placenta stem cells generally grow
faster than those found in the third trimester
(Portmann-Lanz et al. 2006), and the majority of
J. R. Soc. Interface
term placenta-derived stem cells are found at the
quiescent G0/G1 phase of the cell cycle.
2.2.4.3. Chorion-derived stem cells. CMSCs have been
isolated during pregnancy (CVS) or from the term pla-
centa at delivery. The immunophenotype of term
placenta cells is similar to that of adult bone marrow
MSCs, although renin and flt-1 have been shown to
be expressed uniquely in placenta MSCs (Fukuchi
et al. 2004). Similarly to human amnion isolated from
term placenta, term chorion MSCs successfully engraft
in neonatal swine and rats and failed to induce a xeno-
geneic response (Bailo et al. 2004), indicating that these
cells may have an immunoprivileged status consistent
with their low level of HLA I and absence of HLA II
expression (Kubo et al. 2001). Of relevance for pre-
natal autologous and allogeneic therapy, chorion
MSCs can be successfully isolated from chorionic villi
during first-trimester gestation. The chorionic villi do
not express typical MSC cell surface antigens and
have the capacity to differentiate into lineages of the
three germ layers, with a subset of cells expressing the
pluripotency markers Oct-4, ALP, Nanog and Sox2
(Spitalieri et al. 2009). In addition to differentiating
into adipogenic, chondrogenic and osteogenic cells in
vitro (Portmann-Lanz et al. 2006; Ilancheran et al.
2007), they can also differentiate into cells with some
characteristics of hepatocytes in vitro and have the abil-
ity to store glycogen (Chien et al. 2006; Huang 2007;
Tamagawa et al. 2007). Differentiation into other cell
types such as endothelial cells (Alviano et al. 2007),
cardiomyocytes (Zhao et al. 2005), neurons, oligoden-
drocytes and glial cells (Miao et al. 2006; Yen et al.
2008; Portmann-Lanz et al. 2010) has also been
reported, but these findings have mostly relied on upre-
gulation of tissue-specific markers without providing
robust functionality tests.
3. THERAPUTIC APPLICATIONS OF
FOETAL STEM CELLS
Regenerative medicine aims to restore diseased or
damaged tissue using the body’s own cells in order to
overcome the shortage of donated organs and the risk
associated with their rejection (Atala 2009). The use
of stem cells from foetal, cord blood and extraembryonic
tissues for regenerative medicine applications has
increased over the past few years. However, the surpris-
ing picture that has emerged is that cell replacement is
probably not the major mechanism by which cell
therapy confers functional benefit. Rather, stem cells
act beneficially by exerting trophic effects on host tis-
sues. Moreover, because they can be expanded to very
large numbers compared with adult stem cells, they
are ideal candidates for seeding scaffolds and matrices
used for tissue engineering applications such as the
repair of hollow and solid organs. In this section, we
will consider some of the therapeutic applications
FSCs have been put to in pre-clinical studies for repair-
ing diseased or damaged tissue as well as in cancer
treatment (summarized examples are listed in table 2).
 Table 2. Examples of therapeutic applications of foetal stem cells.

regenerated
cell type cell origin treated recipient tissue method of cell delivery disease treated outcome references

J. R. Soc. Interface
foetal tissues
blood and bone human oim mice bone intrauterine osteogenesis imperfecta transplantation resulted in decrease in Guillot et al. (2008a)
marrow transplantation (OI) fractures, increased bone strength, and unpublished
thickness and length. Transplanted data
cells were found clustered in areas of
active bone formation
human postnatal scid/ muscle intra-arterial delivery into Duchenne muscular galectin-1-exposed foetal MSCs formed Chan et al. (2006,
mdx dystrophic post-natal mice/ dystrophy (DMD) fourfold more human muscle fibres in 2007)
mouse model/ intrauterine post-natal scid/mdx mice than non-
prenatal Mdx transplantation using stimulated cells. In pre-natal
dystrophic mice intramuscular, immunocompetant mdx mice,
intravascular and intravascular delivery led to demise of
intraperitoneal delivery mice, intraperitoneal delivery led to
of cells into pre-natal systemic spread of cells and
mice intramuscular injections resulted in
local engraftment. The cells
differentiated at low level into
skeletal and myocardial muscle but
not at a curative level
liver human human foetus bone intrauterine OI cell engraftment of bone despite the Le Blanc et al. (2005)
diagnosed with transplantation donor cells being HLA-mismatched
severe OI with recipient. Bone histology showed
regularly arranged and configured
bone trabeculae
pancreas human sheep pancreas intrauterine type II diabetes the cells functionally engrafted in the Ersek et al. (2010)
transplantation pancreas in 50% of sheep. The
engrafted cells differentiated and were
able to secrete insulin
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extraembryonic
Review. Characteristics of stem cells

cord blood human human bone systematic administration malignant and non- used in the treatment of leukaemia/ Broxmeyer (2010)
marrow malignant blood cancer, Fanconi anaemia, severe
disorders aplastic anaemia, myelodysplastic
syndrome and other disorders
human mouse pancreas intra peritoneal type II diabetes cord blood stem cells introduced into Zhao et al. (2006)
streptozotocin-induced diabetic
mouse model differentiated into
functional insulin-producing cells and
eliminated hyperglycaemia
(Continued.)
H. Abdulrazzak et al. 7
 Table 2. (Continued.)

regenerated

J. R. Soc. Interface
cell type cell origin treated recipient tissue method of cell delivery disease treated outcome references

Wharton’s human rat brain injection into the brain Parkinson’s disease the cells ameliorated apomorphine- Weiss et al. (2006)
Jelly striatum induced rotations in a rat model of
Parkinson’s disease
amniotic fluid sheep sheep muscle cells seeded on collagen partial diaphragmatic diaphragmatic hernia recurrence was Fuchs et al. (2004)
hydrogel and placed on replacement significantly higher in animals that
intestinal cellular graft did not receive the cell grafts
rat rat smooth cells injected into site of wound healing of cells prevented cryo-injury-induced De Coppi et al.
muscle injury injured bladder hypertrophy of surviving bladder (2007a)
8 Review. Characteristics of stem cells

smooth muscle cells but failed to

differentiate specifically to smooth
muscle
mouse mouse brain intra ventricular injection focal cerebral the cells significantly reversed Rehni et al. (2007)
ischaemia neurological deficits in the treated
reperfusion injury animals
human mouse lung systematic administration lung injury AFS cells exhibited a strong tissue Carraro et al. (2008)
engraftment in a mouse lung injury
model and expressed specific alveolar
and bronchiolar epithelial markers
H. Abdulrazzak et al.

human rat heart intramyocardial injection myocardial infarction treated animals showed a preservation Yeh et al. (2010)
of the infarcted thickness,
attenuation of left ventricle
remodelling, a higher vascular density
and general improvement of cardiac
function
placenta human amnion mouse brain intracranial injection Parkinson’s disease cells transplanted into a mouse model of Kong et al. (2008)
AEC Parkinson’s disease differentiated into
neuron, astrocyte and
Downloaded from rsif.royalsocietypublishing.org on August 28, 2010

oligodendrocyte and promoted

endogenous neurogenesis
human amnion rat heart intramyocardial injection myocardial infarction cells integrated into normal and Zhao et al. (2005),
MSC infarcted rat cardiac tissue where Ventura et al.
they differentiated into (2007)
cardiomyocyte-like cells and resulted
in considerable improvements to
ventricular function, capillary density
and scar tissue
 Downloaded from rsif.royalsocietypublishing.org on August 28, 2010

Review. Characteristics of stem cells H. Abdulrazzak et al. 9

3.1. Bone Human AFS cells, transplanted into an immune-

competent and ciclosporin immune-suppressed rat
Osteogenesis imperfecta (OI) is characterized by osteo-
model of myocardial infarction, were rejected probably
penia and bone fragility due to abnormal collagen
because of the expression of B7 co-stimulatory
production caused by mutations in the a chains of col-
molecules as well as macrophage marker CD68.
lagen type I. First-trimester allogeneic HLA-
Furthermore, chondro-osteogenic cell masses were
mismatched male foetal liver MSCs were transplanted
observed in the infarcted hearts of some of the trans-
in utero into a female foetus diagnosed with severe
planted animals (Chiavegato et al. 2007). However, in
OI. Bone biopsy showed regularly arranged and config-
another study it was found that these masses occurred
ured bone trabeculae and no adverse immune reaction
independently of AFS cell injection and that the cells
was observed (Le Blanc et al. 2005). Using a mouse
did not increase the presence of these masses (Delo
model of OI, our group has shown that in utero trans-
et al. 2010).
plantation of foetal blood MSCs ameliorated the
Human AMSCs are able not only to express cardiac-
disease phenotype, producing a clinically relevant
specific genes under specific culture conditions, but also
two-thirds reduction in fracture incidence along with
to integrate into normal and infarcted rat cardiac tissue
an improvement in bone structure and mechanical
where they differentiate into cardiomyocyte-like cells
properties (Guillot et al. 2008a). Foetal bone
(Zhao et al. 2005). The cells result in considerable
marrow MSCs loaded on a highly porous scaffold were
improvements to ventricular function, capillary density
able to reach confluence within the scaffold more
and scar tissue (Ventura et al. 2007) and similar find-
quickly than umbilical cord and adult MSCs. Further-
ings have been reported using native rat AMSCs
more, they demonstrated higher in vitro and in vivo
(Fujimoto et al. 2009). Stem cells derived from pre-
osteogenic differentiation capacity, highlighting their
natal chorionic villi were also successfully engineered
suitability for bone tissue engineering applications
into a living autologous heart valve, which could have
(Zhang et al. 2009).
postnatal applications for repairing congenital cardiac
malformations (Schmidt et al. 2006).
3.2. Muscle
Duchenne muscular dystrophy (DMD) is an X-linked 3.3. Kidney
myopathy affecting one in 3500 boys. The main genetic
In a mouse model of collagen deficiency characterized
defect leads to absence of dystrophin, resulting in
by abnormal collagen deposition in renal glomeruli
muscle damage and wasting. The affected boys
(Phillips et al. 2002; Brodeur et al. 2007), we recently
become wheelchair-bound by 12 years of age and suc-
showed that intrauterine transplantation of foetal
cumb to respiratory failure or cardiopmyopathy by
blood MSCs led to a reduction of abnormal homotri-
the third or forth decade of life (Manzur & Muntoni
meric collagen type I deposition in the glomeruli of
2009). The main animal model for studying DMD is
4 – 12 week old col1a2-deficient mice. Furthermore, we
the mdx mouse that has a premature stop codon result-
showed that the damaged kidneys preferentially
ing in a termination in exon 23 of the dystrophin gene
recruited donor cells in the glomeruli. The study sup-
(Grounds et al. 2008). Galectin-1-treated foetal blood
ports the feasibility of pre-natal treatment for renal
and bone marrow MSCs, transplanted into postnatal
diseases such as Alport syndrome and the polycystic
severe combined immunodeficiency/dystrophic mice
kidney diseases (Guillot et al. 2008b).
(scid/mdx) as well immunodeficient mice whose
Acute tubular necrosis could also be potentially trea-
muscle was induced to regenerate by cryodamage,
ted with foetal cells. Human AFS cells injected into an
formed fourfold more human muscle fibres than non-
immunodeficient mouse model of the disease decreased
stimulated MSCs (Chan et al. 2006). Intrauterine
the number of damaged tubules and reduced apoptosis.
transplantation of foetal MSCs into immunocompetent
The cells also promoted the proliferation of tubular epi-
E14-16 mdx mice resulted in widespread long-term
thelial cells and appeared to have a beneficial
engraftment in multiple organs with a predilection for
immunomodulatory effect (Perin et al. 2010).
muscle compared with non-muscle tissues. However,
the engraftment level observed (0.5 – 1%) falls signifi-
cantly below the levels required for functional
3.4. Lung
improvement in DMD. The low engraftment might be
linked to an absence of muscle pathology at the time Human AFS cells injected into the tail vein of nude
of transplantation (Chan et al. 2007). mice following hyperoxia injury show the capacity to
AFS cells have been used in an ovine model of home to the lung and engraft. They also expressed
diaphragmatic hernia: repair of the muscle deficit specific alveolar and bronchiolar epithelial markers
using grafts engineered with autologous mesenchymal (e.g. TFF1, SPC and CC10; Carraro et al. 2008).
amniocytes leads to better structural and functional AECs and AMSCs from either human or mouse have
results than equivalent foetal myoblast-based and acel- been used in a mouse model of bleomycin-induced lung
lular grafts (Fuchs et al. 2004; Kunisaki et al. 2006). In injury and shown to cause a reduction in severity of
a rat model of bladder cryo-injury, AFS cells also show lung fibrosis. This occurs regardless of cell source (allo-
the ability to differentiate into smooth muscle and to geneic or xenogeneic) or administration route (systemic,
prevent the compensatory hypertrophy of surviving intravenous or intraperitoneal; local, intratracheal;
smooth muscle cells (De Coppi et al. 2007b). Cargnoni et al. 2009).

J. R. Soc. Interface
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10 Review. Characteristics of stem cells H. Abdulrazzak et al.
3.5. Liver
Second-trimester foetal liver cells, enriched for CD326
(hepatocyte progenitor marker) and labelled with Tc-
d,l-hexamethyl-propylene-amine oxime (Tc-HMPAO)
were infused into the hepatic artery of 25 patients
with end stage liver cirrhosis. The cells restored the
lost liver function in the patients without provoking
an immune reaction, probably because of the absence
of HLA class II expression of the infused cells (Khan
et al. 2010).
Human AECs can secrete albumin in culture and,
when b-galactosidase-tagged AECs were transplanted
into immunodeficient mice, the cells integrated into
the liver parenchyma and could be detected until day
7 post transplant (duration of study; Sakuragawa
et al. 2000). In another study, the engrafted cells
resulted in the detection of human a-1 antitrypsin in
the serum of recipient animals, confirming that AECs
are capable of performing this important hepatic
function in vivo (Miki & Strom 2006).
3.6. Brain
Human Wharton’s jelly MSCs ameliorate apomorphine-
induced behavioural deficits in a hemiparkinsonian rat
model (Weiss et al. 2006). There was a significant
decrease in apomorphine-induced rotations at four
weeks continuing up to 12 weeks post transplantation
in Parkinson’s disease (PD) rats that received human
Wharton’s jelly MSCs transplants compared with the
PD rats that received a sham transplant. The behav-
ioural findings correlated with the numbers of tyrosine
hydroxylase-positive cell bodies observed in the mid-
brain following sacrifice at 12 weeks, indicating a
‘rescue from a distance’ phenomenon. One explanation
for this effect may be that Wharton’s jelly MSCs
synthesize glial cell-derived neurotrophic factor
(GDNF), a potent survival factor for dopaminergic
neurons, as well as other trophic factors such as
vascular endothelial growth factor (VEGF) and ciliary
neurotrophic factor. In another report, Wharton’s
jelly MSCs were first induced towards dopaminergic
neurons using neuron-conditioned media, sonic hedge-
hog and fibroblast growth factor 8, and then
transplanted into hemiparkinsonian rats (Fu et al.
2006). Despite the fact that the rats were not immune
suppressed, Wharton’s jelly MSCs were identified five
months later and prevented the progressive degener-
ation/behavioural deterioration seen in control rats
with unilateral lesions. Similarly, rat Wharton’s jelly
cells transplanted into the brains of rats with global cer-
ebral ischaemia caused by cardiac arrest and
resuscitation significantly reduced neuronal loss, appar-
ently owing to a rescue phenomenon (Jomura et al.
2007). Lund et al. administered Wharton’s jelly MSCs
into the eyes of a rodent model of retinal disease.
Here, Wharton’s jelly MSCs were compared with bone
marrow-derived mesenchymal stem cells (BMSCs) and
placental stem cells. They reported that the Wharton’s
jelly MSCs exhibited the best histological evidence of
photoreceptor rescue with increased production of
trophic factors such as brain-derived neurotrophic
J. R. Soc. Interface
factor and fibroblast growth factor 2 (Lund et al.
2007). Interestingly, Wharton’s jelly MSCs trans-
planted into the vitreous demonstrated a rescue effect,
indicating that they could enhance the survival of
photoreceptor cells without being in close proximity
to them. This effect was presumably the result of
diffusible growth factors.
Very similar findings have also been reported for
AECs that confer neuroprotection and functional recov-
ery in animal models of Parkinson’s disease (Bankiewicz
et al. 1994; Kakishita et al. 2000, 2003; Kong et al.
2008) and ischaemia (Liu et al. 2008). The observed
therapeutic effects are probably mediated by secretion
of diffusible factors, including neurotransmitters
(Elwan & Sakuragawa 1997; Sakuragawa et al. 1997,
2001) and neurotrophic and growth factors (Koizumi
et al. 2000; Uchida et al. 2000).
AFS cells can harbour trophic and protective effects
in the central and the peripheral nervous systems. Pan
et al. (2006, 2007, 2009) showed that AFS cells facilitate
peripheral nerve regeneration after injury, particularly
if accompanied with suppressors of inflammatory cyto-
kines such as fermented soya bean extracts. After
transplantation into the striatum, AFS cells are capable
of surviving and integrating in the rat adult brain and
can migrate towards areas of ischaemic damage
(Cipriani et al. 2007). Moreover, the intra-ventricular
administration of AFS cells in mice with focal cerebral
ischaemia – reperfusion injuries significantly reverses
neurological deficits in the treated animals (Rehni
et al. 2007).
3.7. Cancer therapy
HSCs from bone marrow or umbilical cord blood have
long been used to re-establish the haematopoietic
system following radiation and/or chemotherapy
(Osawa et al. 1996; Bhatia et al. 1997). More recently,
research has shown that MSCs are attracted to tumours
because the latter often act as unresolved wounds pro-
ducing a continuous source of chemoattractant
inflammatory mediators. This property is being
exploited by using exogenously delivered MSCs as
vehicles for delivering anti-cancer molecules to tumours
(Loebinger & Janes 2010). Wharton’s jelly cells, like
neural stem cells and MSCs (Aboody et al. 2000;
Studeny et al. 2004), appear to migrate to areas of
tumour growth. Human breast carcinoma cells were
intravenously injected into SCID mice, followed by
intravenous transplantation of fluorescently labelled
Wharton’s jelly MSCs. One week after transplant
Wharton’s jelly MSCs were found near or within lung
tumours and not in other tissues. They were engineered
to express human interferon beta and were administered
intravenously into SCID mice bearing tumours. This
treatment significantly reduced the tumour burden
(Rachakatla et al. 2007). Similarly, human umbilical
cord blood-derived MSCs (UCB – MSCs) have been
used to deliver a secretable trimeric form of tumour
necrosis factor-related apoptosis-inducing ligand
(stTRAIL), via adenoviral transduction mediated by
cell-permeable peptides, to human glioma cells in
nude mice. The genetically modified cells showed
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Review. Characteristics of stem cells
greater efficacy in terms of inhibiting tumour growth
and prolonging the survival of glioma-bearing mice
compared with direct injection of adenovirus encoding
the stTRAIL gene into the tumour mass (Kim et al.
2008).
4. USE OF FSCs FOR REPROGRAMMING
The major advantage of human ES cells over other stem
cell types is their capacity to differentiate into lineages
from the three germ layers. However, there are two
major objections to their use. The moral objection is
that their derivation requires the destruction of
embryos. The practical objection is that they have a
limited clinical application as they can only be used in
an allogeneic fashion. This is also true for some foetal
tissues such as liver and bone marrow. However, extra-
embryonic tissues can be used autologously.
Thus, it was seen as advantageous to develop a
method of creating ES-like cells from somatic cells.
Early attempts at achieving this were nuclear trans-
plantation and fusion of somatic cells and ES cells
( Jaenisch 2009). However, a considerable step forward
was the generation of the induced pluripotent stem
(iPS) cells. The production of iPS cells with quasi-
identical genetic and functional properties offers the
possibility to bypass both moral conflicts and different
genetic background inherent to the technologies
mentioned above. iPS are developed from a non-
pluripotent cell, usually an adult somatic cell, by
causing a forced expression of several genetic sequences
and were first produced in 2006 by Takahashi &
Yamanaka from mouse somatic cells. The key genes
Oct-4, the transcription factor Sox2, c-Myc protoonco-
gene protein and Klf4 (Krueppel-like factor 4) were
sufficient to reprogramme fibroblasts to cells closely
resembling ES cells (Takahashi & Yamanaka 2006;
Takahashi et al. 2007).
Despite the high similarity between iPS and ES cells,
tumour formation in iPS cell chimeric mice was high,
presumably because of the expression of c-Myc in iPS
cell-derived somatic cells (Maherali et al. 2007).
Thomson et al. showed that c-Myc was not necessary
as they were able to generate iPS cells using Oct-4,
Sox2, Nanog and Lin28 using a lentiviral system (Yu
et al. 2007). More recently, it was shown that human
fibroblasts cells can be used to generate iPS cells if
transduced with Oct-4 and Sox2 only in the presence
of valproic acid (Huangfu et al. 2008).
Terminally differentiated cells might not be the ideal
candidate for iPS cell generation because a greater
number of steps could be required to ‘reprogramme’
their genome than somatic stem cells. Kim et al.
(2009a,b) have shown that it is possible to generate
iPS cells by transducing adult mouse neural stem cells
(NSCs) and human foetal NSCs with Oct-4 only. Fur-
thermore, when human adipose MSCs were used, they
were reprogrammed more efficiently than fibroblasts
(Sun et al. 2009).
Few reports have also emerged on the use of cord and
extra ES cells for iPS generation. Galende et al. used
terminally differentiated amniotic fluid cells as
J. R. Soc. Interface
H. Abdulrazzak et al. 11
candidates for reprogramming and found that the iPS
cell colonies were generated twice as fast, yielding
nearly a 200 per cent increase in number compared
with cultured adult skin cells. However, they did not
provide absolute efficiency data (Galende et al. 2009).
Li et al. reprogrammed human amniotic fluid-derived
cells (hAFDCs) with efficiencies varying from 0.059 to
1.525 per cent. The iPS colonies generated expressed
pluripotency markers such as Oct-4, Sox2 and SSEA 4
and they maintained a normal karyotype (Li et al.
2009). Wharton’s jelly cells and term placenta cells
have also been used, giving reprogramming efficiencies
of 0.4 per cent and 0.1 per cent, respectively, which com-
pare rather well with efficiencies reported for adult
fibroblasts (less than 0.01%). The generated iPS cell
colonies expressed several pluripotency markers, formed
EB and, when injected into nude mice, they generated
teratoma-containing derivates of the three germ layers
(Cai et al. 2010). Giorgetti et al. selected CD133þ (hae-
matopoietic stem and progenitor cell marker) cord blood
cells and managed to reprogramme them by retroviral
transduction with Oct-4 and Sox2 only without needing
additional chemical compounds. iPS cell colonies derived
from CD133þ cord blood cells (expressing pluripotency
markers and capable of forming EB and teratomas)
were generated in two weeks whereas no colonies were
obtained with control keratinocytes or fibroblasts using
the two factors only (Giorgetti et al. 2009, 2010).
CD133þ cord blood cells and foetal NSCs already
have a baseline level expression of Oct-4 and/or Sox2
and that, along with a generally more permissive chro-
matin organization, might be the key to their greater
reprogramming efficiency compared with adult cells.
5. CONCLUSION
Human FSCs can be isolated from foetal organs or
extraembryonic sources. They have the advantage of
rapid proliferation, stable karyotype and low or negli-
gible immunogenicity. Unlike ES cells, they do not
form teratomas in vivo and have far fewer ethical con-
cerns as they are mostly obtained from tissues that
would otherwise be discarded. Many of these cells
seem to express some of the same pluipotency markers
found in ES cells, a feature largely absent from most
adult-derived stem cells. They also have the further
advantage over adult stem cells of senescing much
later and being more readily amenable to genetic
modification.
All of these features make them valuable for poten-
tial therapy applications. Thus far they have been
used in pre-clinical settings to treat a variety of diseases
such as osteogenesis imperfecta, congenital diaphrag-
matic hernia, Parkinson’s disease and cancer with
encouraging results. Finally, their usefulness for iPS
generation is very likely to expand their future clinical
use even further.
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