Journal of Pathology

J Pathol 2009; 217: 144160

Published online 13 November 2008 in Wiley InterScience
(www.interscience.wiley.com) DOI: 10.1002/path.2498

Invited Review

Attributes of adult stem cells

MR Alison* and S Islam
Centre for Diabetes and Metabolic Medicine, St. Bartholomews and the London School of Medicine and Dentistry, London, UK

*Correspondence to: Abstract

MR Alison, Centre for Diabetes
and Metabolic Medicine, ICMS, 4
Newark Street, London E1
2AT, UK.
E-mail: m.alison@qmul.ac.uk
No conflicts of interest were
declared.
While cultured embryonic stem (ES) cells can be harvested in abundance and appear to be
the most versatile of cells for regenerative medicine, adult stem cells also hold promise, but
the identity and subsequent isolation of these comparatively rare cells remains problematic
in most tissues, perhaps with the notable exception of the bone marrow. The ability to
continuously self-renew and produce the differentiated progeny of the tissue of their location
are their defining properties. Identifying surface molecules (markers) that would aid in stem
cell isolation is a major goal. Considerable overlap exists between different putative organ-
specific stem cells in their repertoire of gene expression, often related to self-renewal, cell
survival and cell adhesion. More robust tests of stemness are now being employed, using
lineage-specific genetic marking and tracking to show production of long-lived clones and
multipotentiality in vivo. Moreover, the characterization of normal stem cells in specific
tissues may provide a dividend for the treatment of cancer. The successful treatment of
neoplastic disease may well require the specific targeting of neoplastic stem cells, cells that
may well have many of the characteristics of their normal counterparts.
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John
Wiley & Sons, Ltd.
Keywords: asymmetrical cell division; label-retaining cells; multipotential stem cells; self-
renewal; side population; stem cells

Introduction specific cell type. Examples include stem cells in the

Stem cells have varying potential: only the fertilized
oocyte (the zygote) and the descendants of the first two
divisions are totipotent, able to form the embryo and
the trophoblasts of the placenta. After about 4 days
these totipotent cells begin to specialize, forming a
hollow ball of cells, the blastocyst, and a cluster of
cells called the inner cell mass (ICM), from which
the embryo develops. The ICM cells are considered
to be pluripotent, able to differentiate into almost all
cells that arise from the three germ layers but not the
embryo, because they are unable to give rise to the
placenta and supporting tissues: ES cells are derived
from the ICM. Most adult tissues have multipotential
stem cells, cells capable of producing a limited range
of differentiated cell lineages appropriate to their loca-
tion, eg small intestinal stem cells can produce all
four indigenous lineages (Paneth, goblet, absorptive
columnar and enteroendocrine), while CNS stem cells
have trilineage potential, capable of generating neu-
rons, oligodendrocytes and astrocytes. This said, we
should perhaps be a little cautious with our strict defi-
nitions because of the recent flurry of papers indicating
that pluripotency can be reinstated in ordinary somatic
cells through the introduction of ES genes so-called
induced pluripotent stem (iPS) cells. We also recognize
unipotential stem cells, cells capable of generating one
basal layer of the interfollicular epidermis that produce
only keratinized squames, and certain adult hepato-
cytes that have long-term repopulating ability [1].
The key properties of stem cells, viz. indefinite
self-renewal and multilineage potential, were first
recognized in the bone marrow. They were first
described experimentally back in 1961, by Till and
McCulloch [2], as cells that gave rise to multilineage
haematopoietic colonies in the spleen [colony forming
units spleen (CFU-S)]. All tissues appear to have
stem cells, even the central nervous system (CNS) and
the heart, although in these two tissues their activation
after damage seems too little, too late. Across tissues,
many attributes of stem cells are broadly similar, often
related to self-renewal pathways, cell adhesion to the
niche and cell survival, and these are discussed below.
Stem cell properties: some common
themes
A hierarchy of potential
When cells are continually being produced to replace
worn-out or exfoliated cells, then there is often a
unidirectional flow of cells, with stem cells at the
beginning of the flux. For example, in the epidermis,
stem cells are at one end of the cell escalator in

Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
www.pathsoc.org.uk
Attributes of adult stem cells
the basal layer, with cells being shed at the surface.
Likewise, in the colon stem cells are at the very base of
the crypts; this makes sense if the stem cell population
is not to be lost in the migratory flow toward the
luminal surface. Stem cells are a small percentage of
the total cellularity. In the small intestine there are
perhaps up to 10 stem cells near the bottom of the
crypt out of a total crypt population of <300 cells
[3]. In skeletal muscle, satellite (stem) cells comprise
about 5% of all nuclei, but in the bone marrow the
multipotential haematopoietic stem cell is much rarer,
with a frequency of perhaps 1 in 10 000 or more
amongst all bone marrow cells.
It was widely held that stem cells are slowly cycling
but highly clonogenic. Teleologically, it would seem
prudent to restrict stem cell division because DNA
synthesis can be error-prone. Thus, in many tissues
we see that stem cells divide less frequently than
transit-amplifying cells (TACs). In the mouse small
intestine, the stem cells located four or five cell
positions up from the crypt base cycle less often
than the more luminally located TACs [3]; on the
other hand, another population of stem cells has
been identified, located at the crypt base (so-called
crypt base columnar cells), that are invariably (??)
in the cell cycle [4]. In the haematopoietic system as
well, stem cells with long-term repopulating activity
might also be as cell cycle-active as their immediate
descendants [5]. On the other hand, in murine whisker
follicles, the hair shaft and its surrounding sheaths
are derived from TACs in the hair bulb, which is
itself replenished by the bulge stem cells. As formerly
believed of all stem cells, the bulge cells divide
less frequently but are more clonogenic than the
TACs of the hair bulb [6]. A simple hierarchical
system is depicted in Figure 1, and the continually
renewing populations of the gastrointestinal tract and
Figure 1. In most renewing systems a cell hierarchy can
be recognized in which a few self-renewing stem cells
(p = probability of self-renewal) give rise to a limited number
of committed progenitors, also called transit-amplifying cells
(TACs). TACs have limited proliferation potential and eventually
give rise to reproductively sterile terminally differentiated cells.
In vitro, stem cells, early TACs and late TACs are thought to
generate holoclones, meroclones and paraclones, respectively
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
145
haematopoietic systems can be broadly equated with
such an arrangement. When studied in vitro, colonies
formed of tightly packed undifferentiated cells are
considered to represent stem cell colonies, named
holoclones, first described for human epidermal cells
[7]. Other colony types, meroclones and paraclones,
are considered to represent early and late TACs, with
correspondingly reduced replicative capacity.
As a consequence of the perceived slow-cycling
nature of many stem cells, a common strategy for
supposedly identifying them has relied on finding cells
that have retained a label, either tritiated thymidine
or bromodeoxyuridine (BrdU), to which all cells had
been briefly exposed. Hence, stem cells would be
label-retaining cells (LRCs), since the more rapidly
cycling TACs would dilute their label more rapidly
as a consequence of faster cycling clearly this
technique requires re-evaluation in the light of finding
that some stem cells are not slow-cycling.
Selective DNA strand segregation
Some stem cells appear to have devised a strategy for
maintaining genome integrity; another cause of label
retention. Termed the immortal strand hypothesis by
John Cairns [8], newly-forming stem cells designate
one of the two strands of DNA in each chromo-
some as a template strand, such that in each round
of DNA synthesis, while both strands of DNA are
copied, only the template strand and its copy is allo-
cated to the daughter cell that remains a stem cell
(Figure 2). Thus, any errors in replication are read-
ily transferred (within one generation) to TACs that
are soon lost from the population. Such a mecha-
nism will produce LRCs after injection of DNA labels
when stem cells are being formed, but validation of
Figure 2. The immortal strand hypothesis. Genomic integrity
of stem cells can be preserved if newly synthesized strands
of DNA are always allocated to TACs, and stem cells retain
the immortal (template) strand. The template strand of each
chromosome is established when stem cells are being created
J Pathol 2009; 217: 144160 DOI: 10.1002/path
146 MR Alison et al

the hypothesis requires the observation that LRCs go

through further rounds of DNA synthesis, yet still seg-
regating the immortal DNA strands from sister chro-
matids into the daughter cells that remain as stem
cells; this has been demonstrated in the small intes-
tine and breast [9,10], and in some muscle satellite
cells [11]. The existence of LRCs during normal stem
cell renewal is based upon the random segregation of
sister chromatids; however, this is somewhat incom-
patible with the immortal strand hypothesis, which
predicts loss of a newly incorporated DNA synthesis
marker within two cycles [12]. An alternative epige-
netic explanation for asymmetrical cell division has
been proffered, termed the silent sister hypothesis,
proposing that the daughter cell remaining as a stem
cell selectively retains chromatids with active stem cell
genes, with loss of stem cell properties in the cell that
inherits the opposite silent sister chromatids [13].
In studying embryonic fibroblasts in vitro, immortal
strand co-segregation has been found to require both
p53 and down-regulation of inosine monophosphate
dehydrogenase (IMPDH), the rate-limiting enzyme for
guanine nucleotide biosynthesis [14]. Not all studies
concur with the hypothesis; haematopoietic stem cells
(HSCs) with long-term repopulating activity have cer-
tainly been shown not to retain older DNA strands
during division [5]. Figure 3. Mechanisms for the maintenance of stem cell
numbers. (A) Stem cells can all divide asymmetrically. (B) Equal
numbers of symmetrical divisions producing either stem cells
Self-renewal and the stem cell niche or TACs. (C) A combination of asymmetrical and symmetrical
divisions. (D) Both intrinsic and extrinsic signals determine cell
Stem cells are defined by their ability to produce more fate (EI and EII). In the Drosophila testis, correct orientation to
stem cells and cells that differentiate. By undergoing the hub niche cells is important for self-renewal; misorientation
asymmetrical cell division (Figure 3A), both tasks can can result in supernumerary stem cells or cell cycle arrest.
(EIII) In stratified epithelia a mitotic axis perpendicular to the
be accomplished in a single step. Equally well, stem basement membrane can maintain the correct number of stem
cell numbers would remain constant if only symmetri- cells in the basal layer. Stem cells, yellow; TACs, blue
cal divisions occurred, provided that each time a stem
cell gave rise to two daughter TACs, another stem
cell gave rise to two daughter stem cells (Figure 3B).
A further possibility is a mixture of symmetrical and
asymmetrical cell divisions (Figure 3C). Symmetrical
stem cell divisions provide a mechanism to increase
the stem cell population after stem cell loss, but sym-
metrical divisions can also lead to a dangerous escala-
tion of stem cell numbers, as seen with disruption of
asymmetrical neuroblast cell division in Drosophila
melanogaster that leads to lethal, expansive, tumour-
like lesions when such cells are transplanted into adult
hosts [15]. Alternatively, lack of appropriate signalling
can lead to stem cell exhaustion, as seen in mice
lacking Tcf-4 and so unable to form -cateninTcf- Figure 4. Possible consequences of asymmetrical and
4 complexes essential for Wnt signalling in the small symmetrical divisions. Stem cells, yellow; TACs, blue
intestine (Figure 4) [16].
Two main mechanisms appear to govern asymmet- Par6 and Bazooka (Par3 in other species) serves to ori-
ric cell division (Figure 3D); (a) asymmetrical por- entate the mitotic spindle along the basalapical axis
tioning of intracellular components that determine the [18, in this issue], or cell fate determinants are asym-
cell fate termed intrinsic; and (b) asymmetrical metrically partitioned to daughter cells that are des-
assignment of daughter cells relative to external cues tined to differentiate (Figure 3DII). Examples include
from the stem cell niche termed extrinsic [17]. In the distribution of inhibitors of self-renewal, such
Drosophila neuroblasts, a conserved polarity complex as Brat and Mei-P26, to daughter cells destined to
(the Par complex) of atypical protein kinase C (aPKC), become TACs in the Drosophila CNS and ovaries,

J Pathol 2009; 217: 144160 DOI: 10.1002/path

Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Attributes of adult stem cells
respectively [19,20], and the similar destination of
Numb, an inhibitor of Notch signalling, that has a
conserved role of controlling lineage specification in
a variety of cell types [21]. The relationship between
these two intrinsic mechanisms is becoming clearer
[22]; in the Drosophila CNS, Aurora A phosphorylates
Par6, a regulatory subunit of aPKC that becomes acti-
vated and phosphorylates Lethal (2) giant larvae (Lgl),
an interphase inhibitor of aPKC. Phosphorylated Lgl
is released from aPKC, allowing Baz to join the com-
plex. This changes the substrate specificity of aPKC,
allowing it to phosphorylate Numb, causing its release
from one side of the cortex and its asymmetrical distri-
bution. Thus, cell polarity (the Par complex) is coupled
to cell fate decisions.
Extrinsic signals that govern asymmetrical cell divi-
sion can emanate from the stem cell niche. All stem
cells are thought to exist in specialized microenviron-
ments known as niches (Figure 5), and the cells of
the niche regulate stem cell behaviour through direct
physical contact and paracrine signalling (reviewed in
[2327]). Specialized junctions may anchor stem cells
to the niche cells, and orientation of the centroso-
mal components via this anchorage can determine the
plane of direction of the mitotic spindle and thus the
location of daughter cells and their access to extrin-
sic signals from the niche (Figure 3EI). Misorientated
centrosomes in male germ cells are a feature of age-
ing in the Drosophila testis (Figure 3EII), leading to
cell cycle arrest and a decline in spermatogenesis [28].
In stratified epithelia, a division plane perpendicular to
the basement membrane would also maintain the status
quo (Figure 3EIII).
A paradigm of nichestem cell interactions can be
found in the Drosophila ovary and testis (Figure 6A,
B). In the ovary, blind-ending tubes called ovarioles
house a stem cell niche known as the germarium, and
here germline stem cell (GSC) number is maintained
by the close apposition of GSCs with niche (cap) cells;
Armadillo (fly -catenin) and decapentaplegic (Dpp;
Figure 5. The essentials of a stem cell niche. The niche is a
vascular microenvironment of cells and ECM components. Stem
cells make intimate contact with niche stromal cells, and signals
from these cells regulate stem cell fate decisions. In Drosophila,
extrinsic signals from niche cells help maintain the stemness of
the daughter cell still in contact with the niche
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
147
a member of the bone morphogenetic protein ligand
family) signalling are involved. Dpp signalling from
cap cells silences bag of marbles (bam) transcription
in the stem cell remaining attached to the niche,
inhibiting its differentiation [29]. In the Drosophila
testis, GSCs and somatic stem cells (SSCs) are also
attached to niche (hub) cells, and the hub cells secrete
Unpaired (Upd), which ensures self-renewal of both
types of stem cell [30]. In both gonads, disruption of
DE-cadherin-catenincentrosome interactions can
result in supernumerary GSCs, due to alterations in
the orientation of the mitotic axes [31].
In mammals too, cadherin/catenins and BMP sig-
nalling are also involved in the maintenance of
haematopoietic stem cell (HSC) number through inter-
actions with osteoblasts [32] (Figure 6C), although
a more recent study has questioned the role of
osteoblasts as niche cells, suggesting that HSCs are
more numerous in the vascular sinusoids [33]. A pro-
tein known as Merlin, encoded by the Neurofibromato-
sis 2 (Nf2 ) gene, also plays a role in nicheHSC
interactions; mice deficient in NF2 show a rise in
HSC numbers and their disengagement from the niche
and relocation to the circulation [34]. Other well-
characterized stem cell niches include the bulge region
of the hair follicles (Figure 6D), where highly clono-
genic multipotential stem cells are located, capable of
forming all the cell lineages of the hair follicle and
sebaceous gland [35], and the base of small intestinal
crypts, where Wnt signalling (amongst others) from
intestinal subepithelial myofibroblasts helps regulate
stem cell homeostasis (Figure 6E).
As might be expected, putative stem cells in
many tissues are highly expressive of cell adhesion
molecules (CAMs). We have noted the roles of cad-
herins in attachment of stem cells to niche-maintaining
cells, but also involved are integrins, cell surface het-
erodimeric receptors for extracellular matrix (ECM)
components, such as collagen IV and laminin, found
in basement membranes. Integrins provide a physical
linkage between the ECM and cytoskeletal proteins
and help to regulate cell behaviour through discrete
regulatory cues. In stratified epithelia, cells sorted on
the basis of high levels of either 1 or 6 integrins
appear to have superior clonogenic ability (see below).
The transcription factor p63, which transactivates
Perp, a protein involved in the assembly of desmo-
somes, is another factor maintaining the integrity of
stratified epithelia [36,37]; Perp-deficient mice suffer
severe blistering. p63 has two major isoforms (termed
TA and N) that drive at least three transcripts encod-
ing transactivating (TA) and nominally transactivating
(N) isoforms, producing up to six splice variants
[38], and it is the TAp63 isoforms that appear to be
involved in maintaining cells in an immature state.
MicroRNAs (miRs) are small, non-coding RNAs that
regulate gene expression post-transcriptionally, and
p63 is a target of miR-203, thus miR-203 induces cell
J Pathol 2009; 217: 144160 DOI: 10.1002/path
148 MR Alison et al

Figure 6. Examples of stem cell niches. (A) In the Drosophila germarium, terminal filament cells and cap cells make up the niche,
and the TAC derived from the asymmetrical division of the germ stem cell (GSC) undergoes four successive divisions to produce
cysts of 16 cells (cystoblasts) that subsequently undergo further maturation. Somatic stem cells (SSCs) provide the cyst-lining
cells. (B). In the Drosophila testis a broadly similar arrangement produces cysts composed of 16 gonialblasts. (C) The niche for
mammalian HSCs may involve osteoblasts attached to trabecular bone and stem cell differentiation may be inhibited by bone
morphogenetic protein (BMP) ligands. (D) The stem cells of the hair follicle bulge region can provide cells for the complete follicle
and sebaceous gland, and also contribute to the interfollicular epidermis (IFE), which may itself contain stem cells. (E) In the small
intestine, there may be two stem cell populations, the crypt base columnar cells (CBCs) located amongst Paneth cells (PC), and the
so-called label-retaining cells (LRCs) slightly higher up the crypt. Subepithelial myofibroblasts make up the niche, secreting Wnts
that regulate stem cell self-renewal; in turn, Wnt signalling is inhibited by BMPs, and BMP signalling is inhibited by the likes of Noggin
and Gremlin, produced by other mesenchymal cells. Stem cells, yellow; TACS, blue. DP, dermal papilla; Dpp, decapentaplegic; IRS,
inner root sheath; ORS, outer root sheath; TA, transit amplifying compartment; TD, terminally differentiation; Upd, unpaired

cycle exit and promotes differentiation in the epider-
mis, and K14-miR-203 transgenic mice are depleted
of basal stem cells [39].
Cytoprotective strategies
Many putative stem cells have acquired the ability
to withstand cytotoxic insults through either efficient
enzyme-based detoxification systems or by the abil-
ity to rapidly export potentially harmful xenobiotics.
In 1996, Goodell et al [40] reported a new method
for the isolation of HSCs, based on the ability of
HSCs to efflux a fluorescent dye, an ability that was
inhibited by verapamil. Cells are subjected to Hoechst
33 342 dye staining and fluorescence-activated cell
sorting (FACS) analysis; those that actively efflux the
Hoechst dye appear as a distinct population of cells
on the side of the profile, hence the name side pop-
ulation (SP). The SP phenotype of HSCs in mice
and humans is largely determined by the expres-
sion of a protein known as the ABCG2 transporter
(ATP-binding cassette [ABC] subfamily G member 2,
also known as BCRP1) [41,42]. A SP is now rec-
ognized in many solid organs, a population enriched
for clonogenic activity [43]. The ABC superfamily
of membrane transporters is one of the largest pro-
tein classes known, and is characterized by expres-
sion of an ATP-binding cassette region functioning
to hydrolyse ATP to support energy-dependent sub-
strate exportation against steep concentration gradi-
ents across membranes, principally from the intra-
cellular cytoplasm to the extracellular space [44].
Various websites detail the 49 human ABC trans-
porters, which are organized into seven families, AG
(http://www.nutrigene.4t.com/humanabc.htm,
http://www.humanabc.bio.titech.ac.jp/).
ABC transporters play a role in the transport of
drugs (xenobiotics) and drug conjugates. Their role

J Pathol 2009; 217: 144160 DOI: 10.1002/path

Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Attributes of adult stem cells
Figure 6. Continued
is exemplified by MDR1 (ABCB1 or P-glycoprotein),
MRP1 (the multidrug resistance protein 1, ABCC1)
and BCRP1 (breast cancer resistance protein 1,
ABCG2), whose expression is associated with mul-
tidrug resistance (MDR) in cancer cells [45]. Drug
resistance results from the ability of the transporters to
extrude several classes of anticancer drugs, lowering
effective concentrations within the cell. For example,
MDR1 is able to cause the greatest resistance to bulky
amphipathic drugs such as paclitaxel (taxol), anthra-
cyclines and Vinca alkaloids. SP cells can be isolated
from human tumours, and probably contribute signif-
icantly to tumour drug resistance [46]. It is important
to recognize that high expression of an ABC trans-
porter(s) is not the sole provenance of stem cells; for
example, small intestinal villus cells express high lev-
els, as do hepatocytes that transport bile acids into the
biliary canalicular system.
ABC transporters have emerged as an important
field of investigation in the regulation of stem cell biol-
ogy [47] and, significantly, ABCG2, an ABC trans-
porter found in many stem cells, is up-regulated in
hypoxic environments, eg stem cell niches, medi-
ated by HIF1- [44]. The detection of SP cells can
also facilitate the enrichment of stem/progenitor cells
from tissues otherwise lacking in cellular markers for
prospective stem cell isolation; in bovine synovial
membrane, 2% of cells were ABCG2-positive, and
these could be differentiated into either bone or carti-
lage [48].
ABC transporters are not the only cytoprotective
molecules present in adult stem cells; the aldehyde
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
149
dehydrogenase (ALDH) gene superfamily encode
detoxifying enzymes for many pharmaceuticals and
environmental pollutants [49]. Using ALDEFLUOR
staining of live cells, combined with low side scat-
ter (an indication of undifferentiated cells with few
organelles and protrusions), clonogenic, multipoten-
tial stem/progenitor cells have been isolated, eg
from mouse CNS [50], human bone marrow [51]
and cord blood [52]. As expected, high expression
of ALDH can be detrimental to tumour eradica-
tion; cyclophosphamide treatment of human colonic
xenografts enriches for CD44+ ALDH+ cells, and
these double-positive cells are more tumourigenic than
cells selected by solely CD44-positivity [53]. Likewise
in human hepatocellular carcinoma cell lines, cells
selected on the basis of CD133+ ALDH+ are more
tumourigenic than cells selected on CD133 expression
alone [54]. A further mechanism that stem cells appear
to employ to reduce susceptibility to potential toxins
is through low expression of certain cytochrome P450
enzymes, a superfamily of haemoproteins involved in
oxidative (phase I) metabolism; in the pulmonary air-
ways there are so-called pollutant-resistant stem cells
and in the liver, stem/progenitor cells clonally expand
after hepatocyte loss in the face of toxic insult by
virtue of low cytochrome P450 enzymes (see below).
Markers related to stem cell function
The transcription factor Oct-4, important for the main-
tenance of pluripotency in ES cells, is considered to
be expressed by a variety of adult stem cells [55],
J Pathol 2009; 217: 144160 DOI: 10.1002/path
150 MR Alison et al

but the identity of the cells illustrated is unclear.
There is no doubt that the Wnt signalling pathway
is involved in stem cell renewal in HSCs and many
other stem cells, but can we exploit this fact to
identify stem cells? A consequence of active Wnt
signalling is the nuclear localization of -catenin,
and indeed this has been demonstrated for murine
colonic basal cells [56], however, nuclear -catenin
is also seen in small intestinal Paneth cells, Wnt sig-
nals being required for their maturation [57]. On the
other hand, Wnt target genes have been exploited
for stem cell identification Lgr5-positive cells in
the mouse small intestine giving rise to long-lived
clones containing all cell lineages [4]. A compre-
hensive survey of Wnt signalling can be found at
http://www.stanford.edu/%7ernusse/wntwindow.html
Bmi1 is a member of the Polycomb group family of
transcriptional repressors, required for the self-renewal
of HSCs [5860]. Loss of Bmi1 leads to a depletion
of neural stem cells, associated with up-regulation of
p16Ink 4a [61] (Figure 7).
The premature senescence of murine neural stem
cells is prevented by Bmi1, suppressing transcrip-
tion at the Ink4aArf locus that encodes p16Ink 4a
and p19Arf [62]. In the mouse small intestine, Bmi1-
expressing cells can be localized by in situ hybridiza-
tion as being located at cell position 45, counting
from the bottom of the crypt, and lineage tracing from
Bmi1-expressing cells suggests that these cells can also
generate long-lived clones containing all the intestinal
cell lineages [63].
Musashi-1 (Msi-1) is an mRNA-binding protein that
blocks the translation of m-Numb mRNA, so having an
enhancing effect on Notch signalling (Figure 7c). Msi-
1 is expressed by neural progenitors, enhancing the
effects of Notch signalling, maintaining the undiffer-
entiated state [64,65]. Msi-1 has also been advocated
as a stem cell marker in the gut; in the rat stomach Msi-
1-expressing cells are located just beneath the foveolar
region (the putative niche), although there was some
expression in parietal cells [66]. Msi-1 also marks
putative stem cells in the mouse intestine [67]; inter-
estingly, in the small intestine, not only were the cells
at cell positions 45 immunolabelled, but also very
slender cells intermingled with Paneth cells the so-
called crypt base columnar cells, now known to
express Lgr5 and capable of multilineage differenti-
ation [4]. Although Msi-1 has not been considered
as a marker of colorectal cancer stem cells, a quite
remarkable suppression of tumour growth occurred
when siRNA-knockdown of Msi-1 was achieved in
a xenografted colorectal cancer cell line [68].
Prominin-1 (CD133) was the first-identified mem-
ber of the rapidly growing prominin family of pen-
taspan membrane proteins [69]. The specific func-
tions and ligands of the prominins are still relatively
unclear, but they are distinct in their restricted expres-
sion within plasma membrane protrusions, such as

Figure 7. (a) Simplified cartoon of Hedgehog signalling that can help maintain the stem cell state through up-regulation of Bmi1
(see text for further details). (b, c) Simplified cartoon of the Notch signalling system that can maintain stem cell self-renewal. (b) In
the absence of Notch ligands the DNAbinding protein CSL acts as a transcriptional repressor. (c) Upon ligand binding, proteolytic
cleavage of the intracellular domain of Notch (icN) occurs, followed by its translocation to the nucleus, where it converts CSL
to a transcriptional activator. Downstream targets include the repressor Hes1, which in turn represses differentiation-inducing
genes such as Math1. The m-Numb protein binds icN, probably targeting it for degradation. The RNA-binding protein Musashi1
(putative stem cell marker) represses the translation of m-Numb RNA, thus potentiating Notch signalling

J Pathol 2009; 217: 144160 DOI: 10.1002/path

Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Attributes of adult stem cells
epithelial microvilli [70]. Two antibodies, CD133/1
(also known as AC133) and CD133/2 (also known
as AC141) recognize different glycosylated epitopes,
and most studies use CD133/1. In combination with
other markers, CD133 has been used to isolate HSCs
and endothelial precursor cells (EPCs; see below), but
its greatest utility has been in the enrichment of cells
with tumour-initiating ability (so-called cancer stem
cells) in immune-deficient mice from a variety of
human solid tumours, including brain, prostate, liver
and colon [71]. However, a recent study has found
that most normal and malignant colonic epithelial cells
express CD133, and moreover CD133-negative cells
isolated from colorectal liver metastases were at least
as tumourigenic as their CD133-positive counterparts
[72].
Epigenetic modifications
Levels of compaction of chromatin dictate the acces-
sibility of genomic DNA and, together with epige-
netic mechanisms such as histone modifications and
methylation of DNA at cytosine residues in CpG din-
ucleotides, will largely confer the unique properties
we call stemness and be pivotal in the maintenance
of stem cell identity and the subsequent fate of differ-
entiating progeny (reviewed in [73,74]). For example,
histone acetylation, mediated by histone acetyltrans-
ferases, removes positive charges, thereby reducing the
affinity of histones with DNA, and is associated with
increased transcription. On the other hand, trimethyla-
tion of lysine 9 and lysine 27 of histone 3 (H3K9Me3,
H3K27Me3) are so-called repressive histone marks
associated with inactive regions of chromatin, whereas
H3K4Me3 and acetylation of H3 and H4 are associ-
ated with active transcription. H3K27Me3 serves as
a docking site for the bulky Bmi1-containing poly-
comb repressive complex-1 (PRC-1), whose presence
at this site blocks the accessibility to many gene loci,
including the Ink4a/Arf locus [75]. DNA methylation
is expected to be associated with the stable repres-
sion of pluripotency genes, such as Oct4 and Nanog,
observed in differentiating ES cells [76].
Organ-specific stem cells at a glance
Bone marrow
Adult marrow contains at least three ostensibly dis-
crete stem cell populations; haematopoietic stem cells
(HSCs), mesenchymal stem/stromal cells (MSCs) and
endothelial progenitor cells (EPCs).
HSCs are rare cells, with a frequency of 1 in 104
to 1 in 105 amongst bone marrow cells. Selection of
human HSCs based on expression of the cell surface
sialomucin CD34 is still the preferred method. Alter-
natives to CD34 may include the expression of CD133
(CD133/1 = AC133) [77] and ALDH. HSCs with
long-term repopulating ability have been identified in
bone marrow and cord blood, based on the combined
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
151
expression of CD34 and ALDH [51,52]. High ALDH
expression has also been combined with CD133 to
select for human HSCs from bone marrow [78].
Sca-1 (stem cell antigen 1, Ly-6A/E) is an 18 kDa
phosphatidylinositol-anchored protein expressed on
murine multipotent HSCs, and is frequently used
in combination with negative-selection for a num-
ber of cell surface markers characteristic of commit-
ment toward haematolymphoid lineages (Lin ). Mem-
bers of the signal lymphocyte activation molecule
(SLAM) family help define murine HSCs; adding
CD150+ CD48 to the Sca-1+ Lin c-kit+ selection cri-
teria greatly improved long-term multilineage reconsti-
tution in irradiated mice [79]. Recent advances in HSC
self-renewal have been reviewed elsewhere [80,81].
In comparison to HSCs, MSCs, also known as mar-
row stromal cells, are not well characterized antigeni-
cally. In vitro, single cell suspensions of bone mar-
row generate colonies of adherent stromal cells that
can readily differentiate into either bone, cartilage or
fat this trilineage potency being a gold standard
for MSC identity. Each colony is derived from a single
cell called a colony-forming unit fibroblast (CFU-F).
MSCs are rare, declining in frequency with age, being
present in bone marrow aspirates at a frequency of
0.15/105 cells in rodents and 120/105 in humans
[82]. To the properties of plastic adherence and tri-
lineage potential, a working party has suggested that
human MSCs be further defined as expressing CD105,
CD73 and CD90, but lacking expression of CD45,
CD34, CD14 or CD11b, CD79 or CD19 and HLA-
DR surface markers [83]. Added to this list could be
the expression of the 1-integrin (CD49a) [84].
In the mouse, MSCs express Sca-1 and CD44, but
not CD45 or CD11b, and it has been suggested that
positive selection of CD34 cells prior to plastic adher-
ence can provide additional enrichment for MSCs [85].
Current concepts regarding MSCs have recently been
discussed [86].
EPCs constitute a unique population of peripheral
blood mononuclear cells derived from bone marrow
that are involved in postnatal neovascularization dur-
ing wound healing, limb ischaemia, post-myocardial
infarction, atherosclerosis and tumour development.
HSCs and EPCs are seemingly derived from a com-
mon precursor called a haemangioblast. In humans,
EPCs are prospectively isolated on the basis of the
expression of VEGFR2, c-kit, CD34 and CD133, but
negative for CD45; in mice, Sca-1 is an additional
positive marker [87].
Musculoskeletal stem cells
Sources of stem cells for musculoskeletal repair
include bone marrow, peritrabecular tissues in can-
cellous bone, cartilage, muscle, fat and pericytes
collectively known as connective tissue stem cells,
most of which seem multipotent. Articular cartilage
is a unique avascular load-bearing live tissue that
J Pathol 2009; 217: 144160 DOI: 10.1002/path
152
does not normally self-repair. In the bovine, Dowth-
waite et al [88] have suggested that stem/progenitor
cells on the articular surfaces achieve appositional
cartilage growth; these cells expressed the 51 inte-
grin, the classical fibronectin receptor and Notch 1,
which appeared to identify the most clonogenic cells.
Osteoblastic progenitors can be recognized in vitro as
cells that make colonies that express alkaline phos-
phatase (CFU-APs), and in humans there are on aver-
age 55 CFU-APs/106 nucleated bone marrow cells
[89]; in both sexes there is a decline in frequency
with age. The exact nature of osteogenic stem cells
is unclear; Muschler et al [90] advocate the term
connective tissue progenitors (CTPs) to encompass
a heterogeneous group of multipotential stem cells
responsible for musculoskeletal renewal and repair.
This would include pericytes, present outside the base-
ment membrane of small blood vessels and fibroblastic
cells in the bone marrow known as WestinBainton
cells; it is uncertain whether these cells are true
stem cells or TACs, but progenitors eventually give
rise to secretory osteoblasts with limited self-renewal
capacity (short-lived, 40 days) that produce bone
matrix before long-lived osteocytes are finally pro-
duced. Cells with the characteristics of MSCs can
be found in regions of intervertebral discs in humans
[91], and it is likely that MSCs from various loca-
tions can show osteogenic and chondrogenic differen-
tiation.
Unipotential satellite cells, comprising 5% of the
nuclei present within muscle fibres, located outside
the myofibre but beneath the basement membrane, are
the major source of myogenic cells for growth and
repair of postnatal skeletal muscle. Markers include M-
cadherin [92], the transcription factor Pax7, but once
activated by injury the myogenic genes Myf-5 and
MyoD are switched on [93]. Transplantation of single
satellite cells expressing CD34, Sca-1, Pax7 and the 7
integrin in mouse muscle results in their considerable
expansion and, moreover, these cells could be re-
isolated, providing evidence of self-renewal [94].
Digestive tract
The digestive tract is lined by region-specific epithelial
coverings, beginning with stratified squamous epithe-
lia lining the oral cavity and oesophagus, progressing
to the gastric glands of the stomach, moving distally to
the small intestine, where the simple epithelial lining is
thrown into invaginations (crypts) and finger-like pro-
jections (villi), and finally to the colorectum, where
crypts are still present but villi are absent.
The surface of the human oesophagus is relatively
flat, but invaginations of the basement membrane
produce tall papillary structures within the stratified
epithelium. Cell proliferation is confined to the basal
and immediately epibasal layers, with cell division
more common in the papillary basal layer (PBL) than
in the flat interpapillary basal layer (IBL) [95]. In
the PBL, the mitotic axes tend to be parallel to the
J Pathol 2009; 217: 144160 DOI: 10.1002/path
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
MR Alison et al
basement membrane, so both daughter cells remain
in the basal layer, whereas in the IBL, the mitotic
axes tend to be perpendicular to the basement mem-
brane; thus, one daughter cell appears to remain as
a stem cell and the other daughter cell becomes a
TAC in the more proliferatively active epibasal layers
(Figure 3EIII). Okumura et al [96] isolated a fraction
of human oesophageal cells that expressed a relatively
low level of both CK13 and involucrin, and observed
that these cells were additionally characterized by high
expression of the low-affinity neurotrophin receptor
p75NTR but low 1 integrin expression; such cells
behaved as stem cells in vitro. The expression of
p75NTR may also characterize unipotent stem cells of
other squamous epithelia, such as uterine cervix. In the
mouse, a quiescent LRC cell population in vivo, but
highly proliferative in vitro, is characterized as being
highly expressive of the 6 integrin but low in expres-
sion of the transferrin receptor (6bright CD71low [97].
In the stomach, the epithelial lining is folded to
form gastric glands; stem cells are thought to be pri-
marily located just below the foveolus (pit). Cell flux
in the glands is bidirectional, cells descending down-
wards toward the gland base and upwards toward the
surface via the gastric pit. In the mouse, RTPCR
analysis of cells retrieved from the niche by laser-
capture microdissection reveals a gene expression pro-
file closely matching that of HSCs [98]. There are
few markers described for gastric stem cells; however,
Bjerknes and Cheng [99] established that multipoten-
tial progenitor cells exist in the mouse stomach; using
a mutagenicity strategy with ethylnitrosourea, they
created mutant clones in adult hemizygous ROSA26
mice in vivo that did not express -galactosidase,
and all cell lineages could be found within a sin-
gle clone persuasive evidence for the existence of
multipotential cells. From ultrastructural observations
in the isthmic region of human stomach, Karam et al
[100] have suggested that perhaps the stem cells are
so-called mini-granule cells, cells with minute, dense
or cored, secretory granules, somewhat akin to the
granule-free cells found in the mouse. In the isthmic
region of mouse antral glands, a population of LRCs
have been found expressing a protein more character-
istic of the small intestine, villin [101]. Lineage trac-
ing from villin-positive cells using Cre recombinase
suggested these cells were multipotential, but gener-
ally they were proliferatively very quiescent unless
exposed to inflammatory cytokines. Msi-1+ cells have
been found in the putative niche region of the rat
stomach, although Msi-1+ was also expressed in some
parietal cells [66].
In the small intestine the stem cells are located in
a postulated stem cell niche around the crypt base,
in the large intestine stem cells occupy the crypt bot-
tom. The intestinal subepithelial myofibroblasts and
their secreted basement membrane factors (in particu-
lar Wnts) are believed to form and maintain the stem
cell niche, and thereby regulate epithelial cell func-
tion. Nuclear -catenin is found in putative colonic
Attributes of adult stem cells
stem cells [56] but it is also located in small intesti-
nal Paneth cells [57]. Msi-1 positively regulates the
transcriptional repressor molecule, Hes-1 (Figure 7c).
In the mouse small intestine, Msi-1 and Hes-1 are
co-expressed in the cells just above the Paneth cell
zone, although Hes-1 expression additionally extends
to proliferative cells higher up the crypt [102]. Msi-
1 also co-localizes with another putative stem cell
marker, DCAMKL-1, at cell position 4 [103]. Pot-
ten et al [67] observed Msi-1 expression, not only
at cell positions 45, where LRCs exist, but also in
crypt base columnar cells (CBCs, Figure 6E). CBCs
are actively cycling cells nevertheless, they appear
to function as stem cells; using a tamoxifen-activatable
Cre recombinase knocked into the Lgr5 locus, it has
been possible to find long-lived clones derived from
Lgr5-positive cells that contained all small intesti-
nal lineages, likewise in the large intestine [4]. A
similar strategy based on Bmi1-positive cells, also
generated multilineage clones, but the Bmi1-positive
cells were located at cell position 45 [63]. Thus,
in the mouse small intestine at least, we appear to
have two distinct stem cell populations, rapidly cycling
Lgr5+ CBC stem cells and a more slowly cycling
Bmi1+ stem cell population at positions 45. It is
a widely held view that cancer arises from stem
cells, so it is worth noting that tumour development
in the ApcMin+/ mouse can be greatly suppressed
by elimination of Wip1 phosphatase, an enzyme that
functions to turn off the DNA damage response, and
both Wip1- and p53-dependent apoptosis occur at cell
positions 45, not in CBCs [104]. Several recent
reviews on renewal of the gut epithelium are available
[105,106].
The liver is normally proliferatively quiescent, but
partial hepatectomy (PH) stimulates a rapid regenera-
tive response from all cell types in the liver to restore
the organ to its pristine state [1, in this issue]. Hepa-
tocytes carry out normal wear-and-tear renewal, but
chronic damage, often associated with cell senescence,
results in the activation of a facultative stem cell com-
partment located within the intrahepatic biliary tree,
that gives rise to cords of bipotential TACs (oval
cells/hepatic progenitor cells), that ultimately differ-
entiate into hepatocytes and biliary epithelial cells.
If all surviving hepatocytes can regenerate in
response to a large hepatocyte loss, are there any
markers that identify a particular subset that may be
more clonogenic? When rats are treated with retror-
sine, this compound is metabolized by the hepatocytes
cytochrome P450 (CYP) mixed function oxidase sys-
tem to a compound that forms DNA adducts, blocking
the cells ability to proliferate; then, the response to
PH regeneration is activation, expansion and differ-
entiation of so-called small hepatocyte-like progen-
itors (SHPCs) [107]. These cell clusters lack CYP
enzymes that are usually readily induced by retror-
sine, and this probably accounts for their resistance
to the anti-proliferative effects of retrorsine. Ferry and
colleagues [108] have provided persuasive evidence
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
153
that these SHPCs are clonally-derived, seemingly at
random from amongst all hepatocytes, but we have
no further clues as to the identity of these seemingly
relatively undifferentiated hepatocytes.
In mouse liver, the location of LRCs has suggested
a parenchymal stem cell niche close to the portal area
[109], and in human liver, rare putative stem cells
strongly expressing STAT3 and the embryonic stem
cell-associated pluripotency-associated factors Oct4
and Nanog are also located near portal tracts [110].
The pancreas is essentially two different tissues, the
exocrine pancreas, organized into acini with a branch-
ing duct system that produces digestive enzymes, and
the endocrine tissue, the islets of Langerhans, that pro-
duce the hormones glucagon, insulin, somatostatin and
pancreatic polypeptide from four separate cell types,
, , and PP cells, respectively. There are about
1 million islets in a human pancreas, each composed
of 3000 cells of which 75% are insulin-producing
cells. It is the renewal of cells that is of primary
interest for the treatment of diabetes, a disease that
currently afflicts 200 million people worldwide. There
is no doubt that insulin-producing cells can and do
divide throughout life, but there is as yet no clearly
identifiable adult pancreatic stem cell. Heroic efforts
have been made to find stem cells in the mouse pancre-
atic islets, all to no avail. It was suggested, after using
a genetic labelling strategy, that cells arose from
self-duplication, with no cells derived from undiffer-
entiated (insulin-negative) stem cells, despite almost a
doubling of cell mass during the first year of life
[111]. With a similar motive, Teta et al [112] adopted a
simple labelling approach, sequentially administering
two different DNA labels, whereby each label could
separately be identified within any cell that had incor-
porated them. It was reasoned that a DNA-synthesizing
cell (having taken up the first label which would mark
both its daughter cells), would yield double-labelled
cells if either daughter entered the cell cycle again dur-
ing the second labelling period. Double labelled cells
were very rare, even after massive pancreatic dam-
age, leading to the conclusion that no cell hierarchy
exists.
There is, however, evidence of cells derived
from pancreatic ducts in the mouse. Direct lineage
tracing, in which Cre recombinase was driven by a
duct cell-specific promoter (carbonic anhydrase II),
has shown that after pancreatic duct ligation (PDL),
a population of less differentiated ductal cells emerges
that expresses Pdx1 and contributes to islets [113].
PDL also causes up-regulation of the embryonic
pancreas transcription factor Ngn3 in the ductal cells,
and direct lineage tracing has also found that these
cells can become insulin-positive cells [114].
Stratified squamous epithelia
The skin is lined by a stratified keratinizing squa-
mous epithelium, the interfollicular epidermis (IFE).
Normally, cell proliferation is confined to the basal
J Pathol 2009; 217: 144160 DOI: 10.1002/path
154
layer of cells that make contact with the underly-
ing basement membrane. The basal layer is contigu-
ous with cells of the outer root sheath that form
the outermost layer of hair follicles. Until relatively
recently, epidermal stem cell identity was inferred
either by the presence of LRCs or by their growth
potential in vitro [7]. Of course, neither label reten-
tion nor in vitro clonogenicity allow easy isolation of
epidermal stem cells, but a number of studies have
highlighted that selection of basal keratinocytes with
the highest expression of the 1 integrin (integrinbri
cells) enriches for cells with a high colony-forming
efficiency. These integrinbri cells tended to be clus-
tered over the tips of dermal papillae in most skin
sites (with the notable exception of the plantar and
palmar surfaces), had a low level of cell prolifera-
tion, and it was suggested that TACs migrated lat-
erally away from the stem cell clusters [115]. This
notion was supported by the fact that integrindull
cells were more mobile on type IV collagen than
integrinbri cells. However, following the fate of genet-
ically marked cells in human epidermis has sug-
gested instead that stem cells are scattered all along
the basal layer, giving rise to cells that migrate
solely upwards, forming columns of cells reminiscent
of epidermal proliferative unit (EPU)-like structures
[116].
In mouse epidermis, where there is a relatively flat
interface with the underlying dermis, individual stem
cells are thought to give rise to TACs and their dif-
ferentiated progeny within spatially-distinct EPUs that
extend from the basal layer to the surface. A LRC
occupies a central position in the basal layer of each
EPU, along with up to a dozen other cells, most of
which are TACs [117]. Rather than selecting for 1
integrin-expressing cells, Kaur and colleagues [118]
found blast-like 6bri CD71dim cells comprising about
25% of CK14+ basal cells in human epidermis, that
were highly clonogenic and could regenerate a full-
thickness epidermis in organotypic culture. Not all
agree that mouse skin has both a stem cell and a
TAC compartment; from genetic marking of occa-
sional mouse basal cells in the tail and subsequent
analysis of clone size, it was deduced that epidermal
homeostasis could be maintained by a single popula-
tion of progenitor cells [119].
In the hair follicle, the bulge region is clearly the
stem cell niche (Figure 6D) [120], and very elegant
transplantation studies in rodents have demonstrated
that bulge stem cells can regenerate the whole fol-
licle [6,35]. Transcriptional profiling of bulge cells
has shown that they express many mRNAs typical
of stem cells, including SCF, Eph receptors, CD34
and tenascin [120], while Morris et al [121] have
also noted that bulge cells have a gene expression
profile consistent with maintaining proliferative qui-
escence and an undifferentiated state. Putative bulge
stem cells in mouse hair follicles express K15, and
lineage tracing based on the K15 promoter show
that these cells generate all the epithelial cells in
J Pathol 2009; 217: 144160 DOI: 10.1002/path
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
MR Alison et al
the hair follicle and sebaceous gland [121,122]. In
the human hair bulge, cell selection based upon
CD200 enriches for in vitro colony-forming efficiency
[123].
Female reproductive tract
The uterus is lined principally by the endometrium,
an epithelial lining thrown into deep folds that
vary in depth, dependent on the menstrual cycle.
Endometrium gives way to the endocervical canal
of the uterine cervix, lined by a simple colum-
nar epithelium, again with contiguous glands, the
endocervical crypts. At the neck of the uterus, the
simple columnar epithelium gives way, at the so-
called transformation zone, to the stratified squa-
mous epithelium of the ectocervix. The acidic vagi-
nal environment results in squamous metaplasia of
the transformation zone, preceded by the appearance
of subcolumnar reserve cells that undergo hyper-
plasia before forming metaplastic squamous epithe-
lium indistinguishable from the squamous epithelia
of the ectocervix. In mouse endometrium, rare stro-
mal LRCs that are oestrogen receptor -positive,
have been proposed as niche cells [124], and LRCs
(stem cells?) are also found in surface epithelial
cells that are not menstruated in the mouse [125].
SP cells are found in the stratum basalis of human
endometrial glands [125] and, based upon DNA
methylation patterns, individual glands, like intestinal
crypts, appear to be populated by multiple stem cells
[126].
Mammary glands
Stem cells may reside in the terminal ductal lobulo-
alveolar units (TDLUs) as small undifferentiated cells
in the luminal cell layer that do not make contact with
the lumen. These cells are bipotential, giving rise to
luminal and myoepithelial cells [127,128]. In murine
mammary glands the SP fraction is enriched for stem
cells, determined by the proportion of LRCs and the
ability to repopulate cleared (indigenous epithelial
cells removed to allow engraftment of transplanted
cells) mammary fat pads in vivo with both myoepithe-
lial and luminal lineages [129]. In fact, single cells
expressing either CD49f (6-integrin) or CD29 (1-
integrin) can accomplish this [130,131].
Based on a number of studies in humans, three spe-
cific phenotypes appear to be emerging; myoepithelial
cells are common acute lymphoblastic leukaemia anti-
gen-positive (CALLA+ ), CK14+ SMA+ vimentin+ ,
luminal cells are MUC1+ CK18+ epithelial surface
antigen-positive (ESA+ ), while stem cells have an
6+ CK19+ ESA+ MUC1 CALLA phenotype [128].
Using DNA radiolabelling of human breast tissue
implanted into nude mice, LRCs can be found that
often express oestrogen and progesterone receptors;
moreover, in common with many other putative stem
cells, these cells were enriched for the expression
Attributes of adult stem cells
of p21CIP 1 and Msi-1 [132]. Using the previously
described gates for mouse progenitor cells, a popula-
tion of human ductal cells, highly proficient in mam-
mosphere production, has been isolated [133]; these
cells were lin CD49+ EpCamhigh , with a dual K14/19
phenotype.
Male gonadal and accessory sex tissue
In the mammalian testis the mitotic divisions occur
amongst the various generations of so-called type A
spermatogonia that are located on the basement mem-
brane of the seminiferous tubule; the meiotic reduction
divisions and further differentiation towards terminally
differentiated spermatozoa occurs centripetally. In the
mouse testis, putative stem cells have been selected
on the basis of expression of c-Kit and the 1 and
6 integrins [134]. The expression of the transcrip-
tion factors Oct-4 and Plfz has also been used to
define murine testicular stem cells [135], and an SP
population (dependent on ABCG2 activity) was found
highly-enriched for in vivo cell repopulation ability,
with these cells expressing 6 integrin and Stra8,
a premeiotic germ cell-specific cytoplasmic protein
[136]. In man, as in many primates, there are two
types of A spermatogonia; Adark , likely to be fac-
ultative stem cells, and Apale , being progenitor cells
[137].
Anatomically, the prostate gland varies greatly
between species, but can essentially be described as
an exocrine gland enveloping the urethra at the base
of the bladder, composed of blind-ending tubules that
open into the urethra. In the mouse, based on the
location of LRCs, prostatic epithelial stem cells could
be in the proximal regions of prostatic ducts, close
to the urethra [138]. Bipotential cells have also been
isolated based on p63 expression [139]. Remarkably,
single murine prostatic cells can form complete func-
tional acini when transplanted under the renal capsule,
these cells expressing CD117, CD44, CD133 and Sca-
1 [140].
Human prostatic stem cells are thought to be multi-
potential, able to generate secretory, basal and neu-
roendocrine cells, located amongst basal cells that
form a continuous layer between the luminal secre-
tory cells and the basement membrane [141]. Based
on colony growth in vitro, a cytokeratin-based differ-
entiation pathway has been proposed in which stem
cells are CK5/CK14-positive, giving rise to TACs
that lose expression of CK14 but gain CK17 and
19 [141]. Richardson et al [142] found expression of
CD133 in 1% of human prostate basal cells; these cells
uniquely co-expressed 21 integrins and possessed
two important attributes of putative stem cells a
high level of proliferative potential in vitro, coupled
with the ability to reconstitute acini in immunocom-
promised mice. A SP has also been identified, con-
stituting 1% of basal cells, associated with a stem
cell gene expression profile including TERT, Bmi1 and
nestin [143].
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
155
Heart
Cardiomyocytes can proliferate; up to 4% of human
myocyte nuclei adjacent to infarcts are found to
express Ki-67 [144]. In the mouse heart, cardiac
stem cells (about 1 for every 30 00040 000 car-
diomyocytes) are thought to be located in distinct
niches, along with TACs expressing transcription fac-
tors indicative of either cardiomyocyte commitment
(Nk 2.5 and GATA4) or endothelial or smooth mus-
cle specification [145,146]. These stem cells express c-
Kit, Sca-1 and MDR1 and are clonogenic in vitro, able
to produce cardiomyocytes, endothelia and smooth
muscle cells from single cells. A contribution of undif-
ferentiated stem/progenitor cells to the repair of the
murine heart after myocardial infarction has been ele-
gantly shown [147].
Putative stem cells have also been isolated from
human heart [148]; these cells expressed CD34, c-
kit, KDR (VEGFR2) and CD31, formed large colonies
(cardiospheres) in vitro, and could differentiate into
cardiomyocytes, endothelia and smooth muscle cells
after orthotopic transplantation into SCID/beige mice
with myocardial infarction. Stem cell selection in the
human heart based solely on c-kit has also been
suggested [149].
Kidney
The kidney has a low rate of cell turnover under
steady-state conditions, but can regenerate tubular
epithelium after injury. The identity of any renal stem
cells has not been readily forthcoming. The renal
papilla has been proposed as a niche area, based on
the distribution of BrdU-labelled cells after a period
of postnatal labelling in rodents, but the frequency
of LRCs (40%) was far too high for them all to be
stem cells [150]: nevertheless, LRCs disappeared after
ischaemic injury and displayed clonal growth in vitro.
Cells expressing Pax-2 and vimentin, with extensive
clonogenic capacity in vitro and tubulogenic capacity
in vivo, have been isolated from rat kidney; in one
case they were CK-negative [151] and in another study
came from the proximal tubule S3 segment [152].
A putative kidney stem cell has been isolated from
the cortical interstitium of human kidney, making
up 0.8% of all cortical cells and expressing Pax-2
and CD133 along with mesenchymal markers such
as CD73, CD29 and CD44 [153]. These cells were
capable of in vitro expansion and differentiation into
epithelia and endothelia, and formed tubular structures
and endothelia in the damaged kidneys of SCID mice.
This somewhat contentious issue has recently been
reviewed [154 in this issue,155].
Lung
Progress in identifying lung stem cells has been
impeded by the slow turnover of airway and alveo-
lar epithelium, but the consensus view is that there
is no single multipotential stem cell for the lung,
J Pathol 2009; 217: 144160 DOI: 10.1002/path
156
but rather there are regiospecific stem cell zones in
the proximal and distal lung [156, in this issue].
The mouse bronchial tree is lined by a number of
cell types, including basal, ciliated and non-ciliated
cells [serous, goblet and Clara cell secretory protein
(CCSP)-expressing (CE) cells]. Here, both CE cells
and basal cells appear to be able to act as multi-
potential stem cells, for when CE cells are selec-
tively ablated, a major subset of basal cells (basal
cells identified by the binding of the lectin Griffo-
nia simplicifolia isolectin B4 [GSI-B4 ]) up-regulate
CK14 and are able to regenerate the entire epithelium
[157]. In the bronchiolar epithelium, stem cell function
appears to be the property of rare pollutant-resistant
CE cells, co-localized with pulmonary neuroendocrine
cells (PNECs), normally located in cell clusters termed
neuroepithelial bodies (NEBs); PNECs can only act
as progenitors for more PNECs [158]. Pollutant resis-
tance of CE cells is related to a deficiency in the phase
I drug-metabolizing enzyme CYP450 2F2. At the
bronchiolar alveolar duct junction (BADJ), the branch
point between terminal bronchioles lined mostly by
Clara cells and the alveolar spaces lined by pneumo-
cytes, stem cells expressing both CCSP and surfactant
protein C have been identified [159]. In the alve-
oli, type II pneumocytes are widely believed to be
the stem/progenitor cells, undergoing hyperplasia in
response to the loss of the squamous type I pneu-
mocytes, giving rise to type I cells as well as self-
renewing. Type II cells are characteristically recog-
nized by the presence of intracellular lamellar bodies
and the production of surfactant proteins.
Central nervous system
During development, the neuroepithelial cells in the
embryonic ventricular layer generate most of the
neurons and glia (astrocytes and oligodendrocytes);
in the adult human brain the consensus view is that
astrocytes are the main stem cell population, with
small numbers of neural stem cells in other regions
[160]. In the mouse, the subventricular zone (SVZ)
of the lateral ventricles and the dentate gyrus of the
hippocampus are the main sites of adult neurogenesis.
The SVZ is separated from the lateral ventricle by a
single layer of multiciliated ependymal cells; the stem
cells are GFAP+ astrocytes, sometimes extending a
single cilium between the overlying ependymal cells
[161]. In the hippocampus, these astrocytes are located
in the subgranular zone (SGZ). Putative murine neural
stem cells, capable of neurosphere formation and
with trilineage potential, have been isolated based on
either CD133 expression [162] or high ALDH activity
combined with low side scatter [163]. A transcriptional
regulator, Hmga2, functions to suppress p16Ink 4a and
p19Arf in the neural stem cells of young mice, but its
down-regulation in older mice is thought to contribute
to loss of stem cell renewal and a consequent reduction
in stem cell numbers [164].
J Pathol 2009; 217: 144160 DOI: 10.1002/path
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
MR Alison et al
In human brain, Sanai et al. [165] have identified
ribbons of astrocytes in the SVZ lining the lateral ven-
tricles; a Ki-67 labelling index of 0.7% was observed
in these cells in vivo and neurospheres displaying tri-
lineage differentiation could be derived from single
cells of this region.
Eye
Previously it was thought that the corneal epithelium
was renewed from the limbus, a narrow ring of tissue
between the cornea and conjunctiva, and that newly
generated cells migrated centripetally to the centre of
the cornea. This process does occur after extensive
injury, but not in normal renewal in vivo [166]; in a
number of species, cells right across the cornea are
capable of forming holoclones expressing p63 and
CK3. The limbus may be enriched in stem cells,
and in man a SP based on ABCG2 activity has
been identified that appeared to be highly clonogenic
in vitro; these cells also expressed high levels of p63
and the 9 integrin [167,168]. In the mouse retina,
about 0.2% of pigmented cells in the ciliary margin
have stem/progenitor cell properties; they express
transcription factors such as Pax6 and Hes1, nestin
and p27kip1 , and are able to proliferate clonally and
show multipotential differentiation [169,170].
Conclusions
Stem cells in different tissues have common attributes
that enable their self-renewal, survival and mainte-
nance of genomic integrity. In all tissues, stem cells
are located in a specialized vascular microenviron-
ment, the niche; intrinsic and extrinsic signals from
the niche regulate self-renewal and cell fate. Stem cell
location is often inferred by the presence of LRCs,
commonly taken as indicative of a slowly cycling habi-
tus; whether all stem cells additionally operate a mech-
anism to preserve immortal DNA strands remains to
be established. Recently, more robust functional stud-
ies of stemness have been forthcoming, involving
marking putative stem cells to identify the niche, and
then performing lineage tracing to demonstrate that
the proposed stem cell has multipotentiality, and this
approach should be the gold standard of stem cell
identification.
Teaching Materials
Power Point slides of the figures from this Review
may be found in the supporting information.
References
1. Alison MR, Islam S, Lim S. Stem cells in liver regeneration,
fibrosis and cancer: the good, the bad and the ugly. J Pathol
2009; 217:282298.
Attributes of adult stem cells
2. Till JE, McCulloch EA. A direct measurement of the radiation
sensitivity of normal mouse bone marrow cells. Rad Res
1961;14:213222.
3. Marshman E, Booth C, Potten CS. The intestinal epithelial stem
cell. Bioessays 2002;24:9198.
4. Barker N, van Es JH, Kuipers J, Kujala P, van den Born M,
Cozijnsen M, et al. Identification of stem cells in small intestine
and colon by marker gene Lgr5 . Nature 2007;449:10031007.
5. Kiel MJ, He S, Ashkenazi R, Gentry SN, Teta M, Kushner JA,
et al. Haematopoietic stem cells do not asymmetrically segregate
chromosomes or retain BrdU. Nature 2007;449:238242.
6. Oshima H, Rochat A, Kedzia C, Kobayashi K, Barrandon Y.
Morphogenesis and renewal of hair follicles from adult
multipotent stem cells. Cell 2001;104:233245.
7. Barrandon Y, Green H. Three clonal types of keratinocyte with
different capacities for multiplication. Proc Natl Acad Sci USA
1987;84:23022306.
8. Cairns J. Mutation selection and the natural history of cancer.
Nature 1975;255:1972000.
9. Potten CS, Owen G, Booth D. Intestinal stem cells protect their
genome by selective segregation of template DNA strands. J Cell
Sci 2002;115:23812388.
10. Smith GH. Label-retaining epithelial cells in mouse mammary
gland divide asymmetrically and retain their template DNA
strands. Development 2005;132:681687.
11. Shinin V, Gayraud-Morel B, Gom`es D, Tajbakhsh S. Asymmet-
ric division and cosegregation of template DNA strands in adult
muscle satellite cells. Nat Cell Biol 2006;8:677687.
12. Rando TA. The immortal strand hypothesis: segregation and
reconstruction. Cell 2007;129:12391243.
13. Lansdorp PM. Immortal strands? Give me a break. Cell
2007;129:12441247.
14. Rambhatla L, Ram-Mohan S, Cheng JJ, Sherley JL. Immortal
DNA strand cosegregation requires p53/IMPDH-dependent
asymmetric self-renewal associated with adult stem cell. Cancer
Res 2005;65:31553161.
15. Caussinus E, Gonzalez C. Induction of tumor growth by altered
stem cell asymmetric division in Drosophila melanogaster. Nat
Genet 2005;37:11251129.
16. Korinek V, Barker N, Moerer P, van Donselaar E, Huls G,
Peters PJ, et al. Depletion of epithelial stem-cell compartments
in the small intestine of mice lacking Tcf-4. Nat Genet
1998;19:379383.
17. Morrison SJ, Kimble J. Asymmetric and symmetric stem-
cell divisions in development and cancer. Nature 2006;
441:10681074.
18. Yamashita Y. Asymmetric stem cell division and pathology:
insights from Drosophila stem cell systems. J Pathol
2009;217:181185.
19. Betschinger J, Mechtler K, Knoblich JA. Asymmetric segrega-
tion of the tumor suppressor brat regulates self-renewal in
Drosophila neural stem cells. Cell 2006;124:12411253.
20. Neumuller RA, Betschinger J, Fischer A, Bushati N, Poern-
bacher I, Mechtler K, et al. Mei-P26 regulates microRNAs and
cell growth in the Drosophila ovarian stem cell lineage. Nature
2008;454:241245.
21. Knoblich JA. Mechanisms of asymmetric stem cell division. Cell
2008;132:583597.
22. Wirtz-Peitz F, Nishimura T, Knoblich JA. Linking cell cycle to
asymmetric division: Aurora-A phosphorylates the Par complex
to regulate Numb localization. Cell 2008;135:161173.
23. Fuchs E, Tumbar T, Guasch G. Socializing with the neighbors:
stem cells and their niche. Cell 2004;116:769778.
24. Niemann C. Controlling the stem cell niche: right time, right
place, right strength. Bioessays 2006;28:15.
25. Wilson A, Trumpp A. Bone-marrow haematopoietic stem-cell
niches. Nat Rev Immunol 2006;6:93106.
26. Morrison SJ, Spradling AC. Stem cells and niches: mechanisms
that promote stem cell maintenance throughout life. Cell
2008;132:598611.
27. Jones DL, Wagers AJ. No place like home: anatomy and function
of the stem cell niche. Nat Rev Mol Cell Biol 2008;9:1121.
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
157
28. Cheng J, Turkel N, Hemati N, Fuller MT, Hunt AJ, Yamashita
YM. Centrosome misorientation reduces stem cell division during
ageing. Nature 15 October 2008;(Epub ahead of print).
29. Chen D, McKearin D. Dpp signaling silences bam transcription
directly to establish asymmetric divisions of germline stem cells.
Curr Biol 2003;13:17861791.
30. Voog J, DAlterio C, Jones DL. Multipotent somatic stem cells
contribute to the stem cell niche in the Drosophila testis. Nature
2008;454:11321136.
31. Yamashita YM, Jones DL, Fuller MT. Orientation of asymmetric
stem cell division by the APC tumor suppressor and centrosome.
Science 2003;301:15471550.
32. Zhang J, Niu C, Ye L, Huang H, He X, Tong WG, et al.
Identification of the haematopoietic stem cell niche and control
of the niche size. Nature 2003;425:836841.
33. Kiel MJ, Radice GL, Morrison SJ. Lack of evidence that
hematopoietic stem cells depend on N-cadherin-mediated
adhesion to osteoblasts for their maintenance. Cell Stem Cell
2007;1:204217.
34. Larsson J, Ohishi M, Garrison B, Aspling M, Janzen V, Adams
GB, et al. Nf2/merlin regulates hematopoietic stem cell behavior
by altering microenvironmental architecture. Cell Stem Cell
2008;3:221227.
35. Claudinot S, Nicolas M, Oshima H, Rochat A, Barrandon Y.
Long-term renewal of hair follicles from clonogenic multipotent
stem cells. Proc Natl Acad Sci USA 2005;102:1467714682.
36. Ihrie RA, Marques MR, Nguyen BT, Horner JS, Papazoglu C,
Bronson RT, et al. Perp is a p63-regulated gene essential for
epithelial integrity. Cell 2005;120:843856.
37. Carroll DK, Carroll JS, Leong CO, Cheng F, Brown M, Mills AA,
et al. p63 regulates an adhesion programme and cell survival in
epithelial cells. Nat Cell Biol 2006;8:551561.
38. McKeon F. p63 and the epithelial stem cell: more than status
quo? Genes Dev 2004;18:465469.
39. Yi R, Poy MN, Stoffel M, Fuchs E. A skin microRNA
promotes differentiation by repressing stemness. Nature
2008;452:225229.
40. Goodell MA, Brose K, Paradis G, Conner A, Mulligan R. Isola-
tion and functional properties of murine hematopoietic stem cells
that are replicating in vivo. J Exp Med 1996;183:17971806.
41. Zhou S, Morris JJ, Barnes Y, Lan L, Schuetz JD, Sorrentino BP.
Bcrp1 gene expression is required for normal numbers of side
population stem cells in mice, and confers relative protection to
mitoxantrone in hematopoietic cells in vivo. Proc Natl Acad Sci
USA 2002;99:1233912344.
42. Guo Y, Lubbert M, Engelhardt M. CD34 Hematopoietic
stem cells: current concepts and controversies. Stem Cells
2003;21:1520.
43. Challen GA, Bertoncello I, Deane JA, Ricardo SD, Little MH.
Kidney side population reveals multilineage potential and renal
functional capacity but also cellular heterogeneity. J Am Soc
Nephrol 2006;17:18961912.
44. Krishnamurthy P, Schuetz JD. Role of ABCG2/BCRP in biology
and medicine. Annu Rev Pharmacol Toxicol 2006;46:381410.
45. Litman T, Brangi M, Hudson E, Fetsch P, Abati A, Ross DD,
et al. The multidrug-resistant phenotype associated with
overexpression of the new ABC half-transporter, MXR (ABCG2).
J Cell Sci 2000;113:20112021.
46. Hirschmann-Jax C, Foster AE, Wulf GG, Goodell MA, Bren-
ner MK. A distinct side population of cells in human tumor
cells: implications for tumor biology and therapy. Cell Cycle
2005;4:203205.
47. Bunting KD. ABC transporters as phenotypic markers and
functional regulators of stem cells. Stem Cells 2002;20:1120.
48. Teramura T, Fukuda K, Kurashimo S, Hosoi Y, Miki Y, Asada S,
et al. Isolation and characterization of side population stem cells
in articular synovial tissue. BMC Musculoskel Disord 2008;9:86.
49. Vasiliou V, Nebert DW. Analysis and update of the human
aldehyde dehydrogenase (ALDH ) gene family. Hum Genom
2005;2:138143.
50. Corti S, Locatelli F, Papadimitriou D, Donadoni C, Salani S, Del
Bo R, et al. Identification of a primitive brain-derived neural stem
J Pathol 2009; 217: 144160 DOI: 10.1002/path
158
cell population based on aldehyde dehydrogenase activity. Stem
Cells 2006;24:975985.
51. Mirabelli P, Di Noto R, Lo Pardo C, Morabito P, Abate G,
Gorrese M, et al. Extended flow cytometry characterization of
normal bone marrow progenitor cells by simultaneous detection
of aldehyde dehydrogenase and early hematopoietic antigens:
implication for erythroid differentiation studies. BMC Physiol
2008;8:13.
52. Christ O, Lucke K, Imren S, Leung K, Hamilton M, Eaves A,
et al. Improved purification of hematopoietic stem cells based on
their elevated aldehyde dehydrogenase activity. Haematologica
2007;92:11651172.
53. Dylla SJ, Beviglia L, Park IK, Chartier C, Raval J, Ngan L,
et al. Colorectal cancer stem cells are enriched in xenogeneic
tumors following chemotherapy. PLoS ONE 2008;3:e2428.
54. Ma S, Chan KW, Lee TK, Tang KH, Wo JY, Zheng BJ, et al.
Aldehyde dehydrogenase discriminates the CD133 liver cancer
stem cell populations. Mol Cancer Res 2008;6:11461153.
55. Tai MH, Chang CC, Olson LK, Trosko JE. Oct4 expression in
adult human stem cells: evidence in support of the stem cell
theory of carcinogenesis. Carcinogenesis 2005;26:495502.
56. van de Wetering M, Sancho E, Verweij C, de Lau W, Oving I,
Hurlstone A, et al. The -catenin/TCF-4 complex imposes a
crypt progenitor phenotype on colorectal cancer cells. Cell
2002;111:241250.
57. van Es JH, Jay P, Gregorieff A, van Gijn ME, Jonkheer S,
Hatzis P, et al. Wnt signalling induces maturation of Paneth cells
in intestinal crypts. Nat Cell Biol 2005;7:381386.
58. Park IK, Qian D, Kiel M, Becker MW, Pihalja M, Weissman IL,
et al. Bmi-1 is required for maintenance of adult self-renewing
haematopoietic stem cells. Nature 2003;423:302305.
59. Park IK, Morrison SJ, Clarke MF. Bmi1, stem cells, and
senescence regulation. J Clin Invest 2004;113:175179.
60. Rizo A, Dontje B, Vellenga E, de Haan G, Schuringa JJ. Long-
term maintenance of human hematopoietic stem/progenitor cells
by expression of BMI1. Blood 2008;111:26212630.
61. Molofsky AV, Pardal R, Iwashita T, Park IK, Clarke MF, Mor-
rison SJ. Bmi-1 dependence distinguishes neural stem cell self-
renewal from progenitor proliferation. Nature 2003;425:962967.
62. Molofsky AV, He S, Bydon M, Morrison SJ, Pardal R. Bmi-1
promotes neural stem cell self-renewal and neural development
but not mouse growth and survival by repressing the
p16Ink4a and p19Arf senescence pathways. Genes Dev
2005;19:14321437.
63. Sangiorgi E, Capecchi MR. Bmi1 is expressed in vivo in
intestinal stem cells. Nat Genet 2008;40:915920.
64. Imai T, Tokunaga A, Yoshida T, Hashimoto M, Mikoshiba K,
Weinmaster G, et al. The neural RNA-binding protein Musashi1
translationally regulates mammalian numb gene expression by
interacting with its mRNA. Mol Cell Biol 2001;21:38883900.
65. Okano H, Kawahara H, Toriya M, Nakao K, Shibata S, Imai T.
Function of RNA-binding protein Musashi-1 in stem cells. Exp
Cell Res 2005;306:349356.
66. Nagata H, Akiba Y, Suzuki H, Okano H, Hibi T. Expression of
Musashi-1 in the rat stomach and changes during mucosal injury
and restitution. FEBS Lett 2006;580:2733.
67. Potten CS, Booth C, Tudor GL, Booth D, Brady G, Hurley P,
et al. Identification of a putative intestinal stem cell and early
lineage marker; musashi-1. Differentiation 2003;71:2841.
68. Sureban SM, May R, George RJ, Dieckgraefe BK, McLeod HL,
Ramalingam S, et al. Knockdown of RNA binding protein
musashi-1 leads to tumor regression in vivo. Gastroenterology
2008;134:14481458.
69. Mizrak D, Brittan M, Alison MR. CD133: molecule of the
moment. J Pathol 2008;214:39.
70. Bauer N, Fonseca AV, Florek M, Freund D, Jaszai J, Bornhauser
M, et al. New insights into the cell biology of hematopoietic
progenitors by studying prominin-1 (CD133). Cells Tissues
Organs 2008;188:127138.
71. Burkert J, Wright NA, Alison MR. Stem cells and cancer: an
intimate relationship. J Pathol 2006;209:287297.
J Pathol 2009; 217: 144160 DOI: 10.1002/path
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
MR Alison et al
72. Shmelkov SV, Butler JM, Hooper AT, Hormigo A, Kushner J,
Milde T, et al. CD133 expression is not restricted to stem cells,
and both CD133+ and CD133 metastatic colon cancer cells
initiate tumors. J Clin Invest 2008;118:21112120.
73. Shukla V, Vaissi`ere T, Herceg Z. Histone acetylation and
chromatin signature in stem cell identity and cancer. Mutat Res
2008;637:115.
74. Chen L, Daley GQ. Molecular basis of pluripotency. Hum Mol
Genet 2008;17:R2327.
75. Bracken AP, Kleine-Kohlbrecher D, Dietrich N, Pasini D,
Gargiulo G, Beekman C, et al. The Polycomb group proteins
bind throughout the INK4AARF locus and are disassociated in
senescent cells. Genes Dev 2007;21:525530.
76. Gopalakrishnan S, Van Emburgh BO, Robertson KD. DNA
methylation in development and human disease. Mutat Res
2008;647:3038.
77. Bhatia M. AC133 expression in human stem cells. Leukemia
2003;15:16851688.
78. Hess DA, Wirthlin L, Craft TP, Herrbrich PE, Hohm SA,
Lahey R, et al. Selection based on CD133 and high aldehyde
dehydrogenase activity isolates long-term reconstituting human
hematopoietic stem cells. Blood 2006;107:21622169.
79. Yilmaz OH, Kiel MJ, Morrison SJ. SLAM family markers are
conserved among hematopoietic stem cells from old and
reconstituted mice and markedly increase their purity. Blood
2006;107:924930.
80. Zon LI. Intrinsic and extrinsic control of haematopoietic stem-cell
self-renewal. Nature 2008;453:306313.
81. Orkin SH, Zon LI. Hematopoiesis: an evolving paradigm for
stem cell biology. Cell 2008;132:631644.
82. Short B, Brouard N, Occhiodoro-Scott T, Ramakrishnan A,
Simmons PJ. Mesenchymal stem cells. Arch Med Res 2003;34:
565571.
83. Dominici M, Le Blanc K, Mueller I, Slaper-Cortenbach I,
Marini F, Krause D, et al. Minimal criteria for defining
multipotent mesenchymal stromal cells. The International
Society for Cellular Therapy position statement. Cytotherapy
2006;8:315317.
84. Rider DA, Nalathamby T, Nurcombe V, Cool SM. Selection
using the 1 integrin (CD49a) enhances the multipotentiality of
the mesenchymal stem cell population from heterogeneous bone
marrow stromal cells. J Mol Histol 2007;38:449458.
85. Nadri S, Soleimani M. Isolation murine mesenchymal stem
cells by positive selection. In Vitro Cell Dev Biol Anim
2007;43:276282.
86. Bianco P, Robey PG, Simmons PJ. Mesenchymal stem cells:
revisiting history, concepts, and assays. Cell Stem Cell
2008;2:313319.
87. Khoo CP, Pozzilli P, Alison MR. Endothelial progenitor cells
and their potential therapeutic applications. Regen Med
2008;3:863876.
88. Dowthwaite GP, Bishop JC, Redman SN, Khan IM, Rooney P,
Evans DJ, et al. The surface of articular cartilage contains a
progenitor cell population. J Cell Sci 2004;117:889897.
89. Muschler GF, Nitto H, Boehm CA, Easley KA. Age- and
gender-related changes in the cellularity of human bone marrow
and the prevalence of osteoblastic progenitors. J Orthop Res
2001;19:117125.
90. Muschler GF, Midura RJ, Nakamoto C. Practical modeling
concepts for connective tissue stem cell and progenitor
compartment kinetics. J Biomed Biotechnol 2003;3:170193.
91. Risbud MV, Guttapalli A, Tsai TT, Lee JY, Danielson KG,
Vaccaro AR, et al. Evidence for skeletal progenitor cells in the
degenerate human intervertebral disc. Spine 2007;32:25372544.
92. Goldring K, Partridge T, Watt D. Muscle stem cells. J Pathol
2002;197:457467.
93. Chen JC, Goldhamer DJ. Skeletal muscle stem cells. Reprod Biol
Endocrinol 2003;1:101.
94. Sacco A, Doyonnas R, Kraft P, Vitorovic S, Blau HM. Self-
renewal and expansion of single transplanted muscle stem cells.
Nature 17 September 2008;[Epub ahead of print].
Attributes of adult stem cells
95. Seery JP. Stem cells of the oesophageal epithelium. J Cell Sci
2002;115:17831789.
96. Okumura T, Shimada Y, Imamura M, Yasumoto S. Neurotrophin
receptor p75(NTR) characterizes human esophageal keratinocyte
stem cells in vitro. Oncogene 2003;22:40174026.
97. Croagh D, Thomas RJ, Phillips WA, Kaur P. Esophageal stem
cells a review of their identification and characterization. Stem
Cell Rev 2008;4:261268.
98. Mills JC, Andersson N, Hong CV, Stappenbeck TS, Gordon JI.
Molecular characterization of mouse gastric epithelial progenitor
cells. Proc Natl Acad Sci USA 2002;99:1481914824.
99. Bjerknes M, Cheng H. Multipotential stem cells in adult mouse
gastric epithelium. Am J Physiol Gastrointest Liver Physiol
2002;283:G767777.
100. Karam SM, Straiton T, Hassan WM, Leblond CP. Defining
epithelial cell progenitors in the human oxyntic mucosa. Stem
Cells 2003;21:322336.
101. Qiao XT, Ziel JW, McKimpson W, Madison BB, Todisco A,
Merchant JL, et al. Prospective identification of a multilineage
progenitor in murine stomach epithelium. Gastroenterology
2007;133:19891998.
102. Kayahara T, Sawada M, Takaishi S, Fukui H, Seno H, Fukuzawa
H, et al. Candidate markers for stem and early progenitor cells,
Musashi-1 and Hes1, are expressed in crypt base columnar cells
of mouse small intestine. FEBS Lett 2003;535:131135.
103. May R, Riehl TE, Hunt C, Sureban SM, Anant S, Houchen CW.
Identification of a novel putative gastrointestinal stem cell
and adenoma stem cell marker, doublecortin and CaM
kinase-like-1, following radiation injury and in adenomatous
polyposis coli/multiple intestinal neoplasia mice. Stem Cells
2008;26:630637.
104. Demidov ON, Timofeev O, Lwin HN, Kek C, Appella E,
Bulavin DV. Wip1 phosphatase regulates p53-dependent apop-
tosis of stem cells and tumorigenesis in the mouse intestine. Cell
Stem Cell 2007;1:180190.
105. Scoville DH, Sato T, He XC, Li L. Current view: intestinal stem
cells and signaling. Gastroenterology 2008;134:849864.
106. Montgomery RK, Breault DT. Small intestinal stem cell markers.
J Anat 2008;213:5258.
107. Gordon GJ, Coleman WB, Hixson DC, Grisham JW. Liver
regeneration in rats with retrorsine-induced hepatocellular injury
proceeds through a novel cellular response. Am J Pathol
2000;156:607619.
108. Avril A, Pichard V, Bralet MP, Ferry N. Mature hepatocytes
are the source of small hepatocyte-like progenitor cells in the
retrorsine model of liver injury. J Hepatol 2004;41:737743.
109. Kuwahara R, Kofman AV, Landis CS, Swenson ES,
Barendswaard E, Theise ND. The hepatic stem cell niche:
identification by label-retaining cell assay. Hepatology 2008;
47:19942002.
110. Tang Y, Kitisin K, Jogunoori W, Li C, Deng CX, Mueller SC,
et al. Progenitor/stem cells give rise to liver cancer due to
aberrant TGF and IL-6 signaling. Proc Natl Acad Sci USA
2008;105:24452450.
111. Dor Y, Brown J, Martinez OI, Melton DA. Adult pancreatic
cells are formed by self-duplication rather than stem-cell
differentiation. Nature 2004;429:4146.
112. Teta M, Rankin MM, Long SY, Stein GM, Kushner JA. Growth
and regeneration of adult cells does not involve specialized
progenitors. Dev Cell 2007;12:817826.
113. Bonner-Weir S, Inada A, Yatoh S, Li WC, Aye T, Toschi E,
et al. Transdifferentiation of pancreatic ductal cells to endocrine
cells. Biochem Soc Trans 2008;36:353356.
114. Xu X, DHoker J, Stange G, Bonne S, De Leu N, Xiao X, et al.
cells can be generated from endogenous progenitors in injured
adult mouse pancreas. Cell 2008;132:197207.
115. Owens DM, Watt FM. Contribution of stem cells and dif-
ferentiated cells to epidermal tumours. Nat Rev Cancer
2003;3:444451.
116. Ghazizadeh S, Taichman LB. Organization of stem cells
and their progeny in human epidermis. J Invest Dermatol
2005;124:367372.
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
159
117. Wu WY, Morris RJ. In vivo labeling and analysis of epidermal
stem cells. Methods Mol Biol 2005;289:7378.
118. Li A, Kaur P. FACS enrichment of human keratinocyte stem
cells. Methods Mol Biol 2005;289:8796.
119. Clayton E, Doupe DP, Klein AM, Winton DJ, Simons BD,
Jones PH. A single type of progenitor cell maintains normal
epidermis. Nature 2007;446:185189.
120. Tumbar T, Guasch G, Greco V, Blanpain C, Lowry WE, Rendl
M, et al. Defining the epithelial stem cell niche in skin. Science
303:359363.
121. Morris RJ, Liu Y, Marles L, Yang Z, Trempus C, Li S, et al.
Capturing and profiling adult hair follicle stem cells. Nat
Biotechnol 2004;22:411417.
122. Cotsarelis G. Epithelial stem cells: a folliculocentric view. J
Invest Dermatol 2006;126:14591468.
123. Ohyama M, Terunuma A, Tock CL, Radonovich MF, Pise-
Masison CA, Hopping SB, et al. Characterization and isolation
of stem cell-enriched human hair follicle bulge cells. J Clin Invest
2006;116:249260.
124. Gargett CE, Chan RW, Schwab KE. Hormone and growth factor
signaling in endometrial renewal: role of stem/progenitor cells.
Mol Cell Endocrinol 2008;288:2229.
125. Gargett CE, Chan RW, Schwab KE. Endometrial stem cells. Curr
Opin Obstet Gynecol 2007;19:377383.
126. Kim JY, Tavare S, Shibata D. Counting human somatic cell
replications: methylation mirrors endometrial stem cell divisions.
Proc Natl Acad Sci USA 2005;102:1773917744.
127. Smalley M, Ashworth A. Stem cells and breast cancer: a field in
transit. Nat Rev Cancer 2003;3:832844.
128. Clarke RB, Anderson E, Howell A, Potten CS. Regulation of
human breast epithelial stem cells. Cell Prolif 2003;36(suppl
1):4558.
129. Alvi AJ, Clayton H, Joshi C, Enver T, Ashworth A, Vivanco MM,
et al. Functional and molecular characterisation of mammary side
population cells. Breast Cancer Res 2003;5:R18.
130. Stingl J, Eirew P, Ricketson I, Shackleton M, Vaillant F, Choi D,
et al. Purification and unique properties of mammary epithelial
stem cells. Nature 2006;439:993997.
131. Shackleton M, Vaillant F, Simpson KJ, Stingl J, Smyth GK,
Asselin-Labat ML, et al. Generation of a functional mammary
gland from a single stem cell. Nature 2006;439:8488.
132. Clarke RB, Spence K, Anderson E, Howell A, Okano H, Pot-
ten CS. A putative human breast stem cell population is enriched
for steroid receptor-positive cells. Dev Biol 2005;277:443456.
133. Villadsen R, Fridriksdottir AJ, Rnnov-Jessen L, Gudjonsson T,
Rank F, LaBarge MA, et al. Evidence for a stem cell hierarchy
in the adult human breast. J Cell Biol 2007;177:87101.
134. Shinohara T, Avarbock MR, Brinster RL. 1- and 6-integrin are
surface markers on mouse spermatogonial stem cells. Proc Natl
Acad Sci USA 1999;96:55045509.
135. Buaas FW, Kirsh AL, Sharma M, McLean DJ, Morris JL,
Griswold MD, et al. Plzf is required in adult male germ cells
for stem cell self-renewal. Nat Genet 2004;36:647652.
136. Lassalle B, Bastos H, Louis JP, Riou L, Testart J, Dutrillaux B,
et al. Side population cells in adult mouse testis express Bcrp1
gene and are enriched in spermatogonia and germinal stem cells.
Development 2004;131:479487.
137. Ehmcke J, Wistuba J, Schlatt S. Spermatogonial stem cells:
questions, models and perspectives. Hum Reprod Update
2006;12:275282.
138. Tsujimura A, Koikawa Y, Salm S, Takao T, Coetzee S,
Moscatelli D, et al. Proximal location of mouse prostate epithelial
stem cells: a model of prostatic homeostasis. J Cell Biol
2002;157:12571265.
139. Signoretti S, Loda M. Defining cell lineages in the prostate
epithelium. Cell Cycle 2006;5:138141.
140. Leong KG, Wang BE, Johnson L, Gao WQ. Generation of a
prostate from a single adult stem cell. Nature 22 October
2008;[Epub ahead of print].
141. Hudson DL. Epithelial stem cells in human prostate growth and
disease. Prostate Cancer Prostatic Dis 2004;7:188194.
J Pathol 2009; 217: 144160 DOI: 10.1002/path
160
142. Richardson GD, Robson CN, Lang SH, Neal DE, Maitland NJ,
Collins AT. CD133, a novel marker for human prostatic epithelial
stem cells. J Cell Sci 2004;117:35393545.
143. Pascal LE, Oudes AJ, Petersen TW, Goo YA, Walashek LS,
True LD, et al. Molecular and cellular characterization of
ABCG2 in the prostate. BMC Urol 2007;7:6.
144. Beltrami AP, Urbanek K, Kajstura J, Yan SM, Finato N, Bus-
sani R, et al. Evidence that human cardiac myocytes divide after
myocardial infarction. N Engl J Med 2001;344:17501757.
145. Barile L, Messina E, Giacomello A, Marban E. Endogenous
cardiac stem cells. Prog Cardiovasc Dis 2007;50:3148.
146. Leri A, Kajstura J, Anversa P, Frishman WH. Myocardial
regeneration and stem cell repair. Curr Probl Cardiol
2008;33:91153.
147. Hsieh PC, Segers VF, Davis ME, MacGillivray C, Gannon J,
Molkentin JD, et al. Evidence from a genetic fate-mapping study
that stem cells refresh adult mammalian cardiomyocytes after
injury. Nat Med 2007;13:970974.
148. Messina E, De Angelis L, Frati G, Morrone S, Chimenti S,
Fiordaliso F, et al. Isolation and expansion of adult cardiac stem
cells from human and murine heart. Circ Res 2004;95:911921.
149. Bearzi C, Rota M, Hosoda T, Tillmanns J, Nascimbene A, De
Angelis A, et al. Human cardiac stem cells. Proc Natl Acad Sci
USA 2007;104:1406814073.
150. Oliver JA, Maarouf O, Cheema FH, Martens TP, Al-Awqati Q.
The renal papilla is a niche for adult kidney stem cells. J Clin
Invest 2004;114:795804.
151. Gupta S, Verfaillie C, Chmielewski D, Kren S, Eidman K,
Connaire J, et al. Isolation and characterization of kidney-derived
stem cells. J Am Soc Nephrol 2006;17:30283040.
152. Kitamura S, Yamasaki Y, Kinomura M, Sugaya T, Sugiyama H,
Maeshima Y, et al. Establishment and characterization of renal
progenitor like cells from S3 segment of nephron in rat adult
kidney. FASEB J 2005;19:17891797.
153. Bussolati B, Bruno S, Grange C, Buttiglieri S, Deregibus MC,
Cantino D, et al. Isolation of renal progenitor cells from adult
human kidney. Am J Pathol 2005;166:545555.
154. Hopkins C, Li J, Rae F, Little MH. Stem cell options for kidney
disease. J Pathol 2009;217:265281.
155. Gupta S, Rosenberg ME. Do stem cells exist in the adult kidney?
Am J Nephrol 2008;28:607613.
156. Snyder JC, Teisanu RM, Stripp BR. Endogenous lung stem cells
and contribution to disease. J Pathol 2009;217:254264.
J Pathol 2009; 217: 144160 DOI: 10.1002/path
Copyright 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
MR Alison et al
157. Hong KU, Reynolds SD, Watkins S, Fuchs E, Stripp BR. Basal
cells are a multipotent progenitor capable of renewing the
bronchial epithelium. Am J Pathol 2004;164:577588.
158. Reynolds SD, Giangreco A, Power JHT, Stripp BR. Neuroep-
ithelial bodies of pulmonary airways serve as a reservoir of
progenitor cells capable of epithelial regeneration. Am J Pathol
2000;156:269278.
159. Kim CF, Jackson EL, Woolfenden AE, Lawrence S, Babar I,
Vogel S, et al. Identification of bronchioalveolar stem cells in
normal lung and lung cancer. Cell 2005;121:823835.
160. Dietrich J, Imitola J, Kesari S. Mechanisms of disease: the role
of stem cells in the biology and treatment of gliomas. Nat Clin
Pract Oncol 2008;5:393404.
161. Doetsch F. The glial identity of neural stem cells. Nat Neurosci
2003;6:11271134.
162. Lee A, Kessler JD, Read TA, Kaiser C, Corbeil D, Huttner WB,
et al. Isolation of neural stem cells from the postnatal cerebellum.
Nat Neurosci 2005;8:723729.
163. Corti S, Locatelli F, Papadimitriou D, Donadoni C, Salani S, Del
Bo R, et al. Identification of a primitive brain-derived neural stem
cell population based on aldehyde dehydrogenase activity. Stem
Cells 2006;24:975985.
164. Nishino J, Kim I, Chada K, Morrison SJ. Hmga2 promotes neu-
ral stem cell self-renewal in young but not old mice by reducing
p16Ink4a and p19Arf expression. Cell 2008;135:227239.
165. Sanai N, Tramontin AD, Quinones-Hinojosa A, Barbaro NM,
Gupta N, Kunwar S, et al. Unique astrocyte ribbon in adult
human brain contains neural stem cells but lacks chain migration.
Nature 2004;427:740744.
166. Majo F, Rochat A, Nicolas M, Jaoude GA, Barrandon Y.
Oligopotent stem cells are distributed throughout the mammalian
ocular surface. Nature 2008;456:250254.
167. Chen Z, de Paiva CS, Luo L, Kretzer FL, Pflugfelder SC, Li DQ.
Characterization of putative stem cell phenotype in human limbal
epithelia. Stem Cells 2004;22:355366.
168. de Paiva CS, Chen Z, Corrales RM, Pflugfelder SC, Li DQ.
ABCG2 transporter identifies a population of clonogenic human
limbal epithelial cells. Stem Cells 2005;23:6373.
169. Boulton M, Albon J. Stem cells in the eye. Int J Biochem Cell
Biol 2004;36:643657.
170. Coles BL, Horsford DJ, McInnes RR, van der Kooy D. Loss of
retinal progenitor cells leads to an increase in the retinal stem
cell population in vivo. Eur J Neurosci 2006;23:7582.