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355Clinical Science (2002) 103, 355369 (Printed in Great Britain)

R E V I E W

Adult stem cell plasticity: new pathways oftissue regeneration become visible

Stuart J. FORBES*, Pamela VIG*, Richard POULSOM*, Nicholas A. WRIGHT*and Malcolm R. ALISON**Histopathology Unit, Cancer Research UK, London, U.K., Department of Medicine, Faculty of Medicine, Imperial College ofScience, Technology and Medicine (ICSTM), St Marys Hospital, London, U.K., and Department of Histopathology, ICSTM,Hammersmith Hospital, London, U.K.

A B S T R A C T

There has recently been a significant change in the way we think about organ regeneration. In

the adult, organ formation and regeneration was thought to occur through the action of organ-

or tissue-restricted stem cells (i.e. haematopoietic stem cells making blood; gut stem cells

making gut, etc.). However, there is a large body of recent work that has extended this model.

Thanks to lineage tracking techniques, we now believe that stem cells from one organ system,

for example the haematopoietic compartment, can develop into the differentiated cells within

another organ system, such as liver,brainor kidney. This cellular plasticity notonly occursunder

experimental conditions, but has also been shown to take place in humans following bone

marrow and organ transplants. This trafficking is potentially bi-directional, and even differen-

tiated cells from different organ systems can interchange, with pancreatic cells able to form

hepatocytes, for example. In this review we will detail some of these findings and attempt to

explain their biological significance.

INTRODUCTION

Each organ and tissue is perceived to possess a subpopu-

lation of cells capable of self-maintenance, indefinite

proliferative potential and the abilityto giverise to a large

family of descendants, i.e. to be clonogenic. These stem

cells usually give rise to a limited number of different cell

lineages within their normal environs, such multipoten-

tiality being a feature of tissue- and organ-specific stem

cells [1]. This review focuses on a hitherto unsuspected

property of tissue-specific stem cells, i.e. the ability to

give rise to cell types in a new location, that are not

normally present in the organ in which the stem cells are

located a property we refer to as stem cell plasticity.The stem cells that are thought to be most flexible come

from the inner cell mass of the blastocyst: these cells are

Key words: bone marrow stem cells, lineage tracking, plasticity, transdifferentiation, transplants.

Abbreviations: CNS, central nervous system; ES cells, embryonic stem cells; FAH, fumarylacetoacetatehydrolase; FGF, fibroblast

growth factor; G-CSF, granulocyte colony-stimulating factor; GFAP, glial fibrillary acidic protein; GFP, green fluorescent protein;

eGFP, enhanced GFP; HSC, haematopoietic stem cell; MSC, mesenchymal stem cell; OI, osteogenesis imperfecta; NOD,

non-obese diabetic; SCID, severe combined immunodeficient; SDF, stroma-derived factor; SP cells, side-population cells.

Correspondence: Dr S. J. Forbes, Hepatology Section, Division of Medicine, Faculty of Medicine, Imperial College London, 10th

floor QEQM Wing, South Wharf Road, London W2 1NY, U.K. (e-mail s.j.forbes!ic.ac.uk).

essentially pluripotential, being capable of giving rise tocells found in all three germ layers. However, the ethical

issues surrounding the use of embryonic stem cells (ES

cells) from early human embryos have caused concern.

There may, however, be alternatives to the use of ES

cells, as certain adult stem cells appear to be more flexible

than previously thought. Numerous papers have chal-

lenged the long-held belief that organ-specific stem cells

are lineage-restricted. In particular, haematopoietic and

neural stem cells appear to be the most versatile at cutting

across lineage boundaries (see Table 1). Of course, it is

one thing for a circulating cell to engraft in another organ

and assume some or all of the phenotypic traits of

that organ; this is known as transdifferentiation theacquisition of a new phenotype. It is quite another to

claim that the engrafted cell is a stem cell for its new

# 2002 The Biochemical Society and the Medical Research Society

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356 S . J . Forbes and other s

Table 1 Examples of adult stem cell plasticity, based on lineage tracking and phenotype determination

Abbreviations: ISH, in situ hybridization for Y chromosome; CK, cytokeratin; G-6-Pase, glucose-6-phosphatase; -Gal, -galactosidase.

Donor cells Recipient organ Cell type Proof of donor origin/proof of new phenotype References

Bone marrow Liver Oval cells, hepatocytes (rat) ISH and MHC class II antigen L21-6/morphology [74]

KTL S cells Liver Hepatocytes (mouse) -Gal/FAH+ [6]

Bone marrow Liver Hepatocyes (human) ISH/CK8 and albumin [76,77]

Pancreatic exocrine cells Liver Hepatocyte (mouse) GFP/G-6-Pase and transferrin [87]

Pancreas Liver Hepatocyte (mouse) ISH/FAH+ [88]

Bone marrow Liver Endothelium (mouse, human) ISH/factor VIII [54]

Bone marrow Kidney Tubular epithelium glomeruli (mouse, human) I SH/cytochrome P450 and CAM 5.2 [47]

Bone marrow Kidney Endothelium (human) XX chromosome and HLA typing/morphology [46]

Extra-renal Kidney Endothelium (human) Barr-body detection/morphology [44]

Bone marrow Heart Myocardium (mouse) ISH and GFP/cardiac myosin [31]

Bone marrow SP cells Heart Cardiomyocytes and endothelium (mouse) -Gal/cardiomyocytes: -actinin and endothelial cells: Flt-1 [58]

Bone marrow Lung Type 1 pneumocytes (mouse) ISH/surfactant B [65]

Neuronal Marrow Multiple haematopoietic lineages (mouse) -Gal/morphology [103]

Bone marrow CNS Neurons ISH/NeuN [107]

Bone marrow CNS Microglia and astrocytes ISH and GFP/macrophage antigen F4/80 [108]

found home. Ideally this would require the isolation and

transplantation of single cells that self-renew and produce

a large family of descendants (clonogenicity) that eventu-

ally become fully functional; these robust criteria have

been met in one or two cases. However, some commenta-

tors have added that this phenomenon should be ob-

served to occur naturally in organs not forced to

undergo organ degeneration before accepting that stem

cells jump lineage boundaries [2]. Although this does

occur to a limited extent, we will argue that it is precisely

because of severe organ damage that transdifferentiation

occurs more readily, and that the likes of haematopoieticstem cells (HSCs) can act as a back-up system when an

organs own regenerative capacity is overwhelmed. Thus

the lack of transdifferentiation in the absence of organ

damage in no way invalidates the claim that it does occur,

and it is largely in the clinical context of severe organ

damage that we would envisage exploiting the use of stem

cells with transdifferentiating potential. We will also

briefly review the evidence that some adult stem cells

may even be pluripotential, albeit in the context of

creating chimaeric animals, for example in the ability

of adult HSCs to contribute to all three germ layers

in the pre-immune foetal sheep and the NOD\SCID

(non-obese diabetic\severe combined immunodeficient)mouse after injection into the blastocyst.

STEM CELL PLASTICITY:TRANSDIFFERENTIATION OR FUSION ?

Regenerative medicine is big news in both the biomedical

and the popular press, and there has been a vigorous

debate regarding the therapeutic potential of ES cells

versus adult stem cells. Recently, doubt has been cast

upon the claims that certain adult stem cells, particularly

from the bone marrow and central nervous system

(CNS), can jump lineage boundaries to generate com-

pletely new types of cells.The evidence for adult stem cell

plasticity often relies on the appearance of Y chromo-

some-positive cells in a female recipient of a bone marrow

transplant from a male donor. Alternatively, markers

such as LacZ or green fluorescent protein (GFP) have

been used (see Figure 1), and these techniques are usually

combined with lineage markers in an attempt to show a

switch in the fate (transdifferentiation) of the trans-

planted cells. However, two publications have suggestedthat these phenomena could be due to the fusion of bone

marrow cells with the differentiated cells in the new

organ. When bone marrow from GFP transgenic mice

was mixed with ES cells, a very small proportion (211

hybrid clones\10' marrow cells) of thebone marrow cells

fused with ES cells, and these cells could subsequently

adopt some of the phenotypes typical of ES cell dif-

ferentiation [3]. A very low frequency of fusion (one

event\100000 CNS cells) was reported when mouse

CNS cells were mixed with ES cells, and here the derived

hybrid cells were able to show multilineage potential

when injected into blastocysts, most prominently into

liver [4]. While these observations do raise the possibilitythat the apparent transdifferentiation events are the result

of cell fusion (so-called heterokaryons), this speculation

is at odds with a number of observations. For example, a

recent report suggested that post-partum thyroiditis may

be due to transplacentally acquired foetal cells causing an

alloimmune disease (previously regarded as an autoim-

mune disease) [5]. In this report, one female patient had

clusters of fully differentiated thyroid follicular cells

bearing one X and one Y chromosome; of course, the

source of the transdifferentiated cells was the foetus


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357Adult stem cell plasticity

Figure 1 Methods commonly used to track the fate of transplanted bone marrow

rather than a deliberate transplant, but nevertheless no

follicular cells were XXXY, suggesting that cell fusion

was not responsible for the phenomenon.

Furthermore, in mice, the ability of bone marrow cellsto cure a metabolic liver disease has been established [6].

Female mice deficient in the enzyme fumarylacetoacetate

hydrolase (FAH/, a model of fatal hereditary tyrosi-

naemia type 1), can be biochemically rescued by 10'

unfractionated bone marrow cells that are wild type for

FAH. Moreover, as few as 50 HSCs were capable of

biochemical rescue. The very low levels of fusion

reported with ES cells also makes it unlikely that such

hybrids could be responsible for the multi-focal liver

colonization by marrow-derived hepatocytes seen in

this model. On the other hand, if fusion was responsible,

then clearly these hybrids had a selective growth ad-

vantage, turning unhealthy hepatocytes into metaboli-cally competent hepatocytes, and would not negate the

therapeutic potential of bone marrow cells in the liver.

Moreover, bone marrow stem cells are common in cord

blood and are even found in peripheral blood: if

widespread fusion exists, we would all have large num-

bers of polyploid cells in many organs. This has not been

reported outside the liver, where polyploidization does

occur on a large scale, due to binucleate cells segregating

on thesame mitotic spindle. Until experiments arecarried

out that show heterokaryon formation when adult stem

cells transdifferentiate in vivo, then extrapolations from

rare events involving ES cells are premature.

BONE MARROW

Adult bone marrow contains HSCs and mesenchymal

stem cells (MSCs), both of which may derive from a

common primitive blast-like cell precursor able to

differentiate along MSC or HSC potentials [7].

HSCsThe hierarchy of human haematolymphopoietic cells is

defined by functional assays. HSCs with extensive self-

renewal capacity are assayed in vivo for their capacity to

xenograft immunodeficient NOD\SCID mice and pre-immune sheep foetuses. These models are surrogates for

a syngeneic transplantation assay. Primitive haematolym-

phopoietic cells with limited self-renewal potential are

identified in vitro as high-proliferative-potential colony-

forming cells. Lineage-committed haematolymphopoi-

etic cells with no self-renewal activity are also defined in

vitro by clonogenic assays as colony-forming units or

burst-forming units.

Within the bone marrow, HSCs reside in niches that

support all the requisite factors and adhesive properties


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to maintain their viability and produce an appropriate

balanced output of mature progeny over the lifetime of

the organism [811]. Their survival and proliferation in

vivo is dependent on an intimate association with bone

marrow stroma, containing the progeny of mesenchymal

or marrow stromal cells (MSCs or colony-forming unit-

fibroblast), which supports and signals through soluble

and adhesive modalities [12,13] (see reviews by Quesen-

berry and Becker [14] and Chan and Watt [15]). During

development there is migration between sites capable of

supporting HSCs, although in adults a homing mech-

anism is considered to operate that causes the majority of

HSCs to return to the bone marrow within 1 day. A

number of different factors are involved in migration and

homing: the ligand for c-Kit, stem cell factor, is im-

portant, particularly the cell membrane-bound variant, as

it stimulates the adherence of HSCs to stroma. Integrin

interactions are also crucial, as 1 integrins are fun-

damental to the migration of HSCs to the foetal liver [16].

Murine knockouts of either the chemokine stroma-derived factor 1 (SDF-1) (cloned from bone marrow

stromal cells) or its receptor CXCR4 prevent haemato-

poiesis transferring from embryonic liver to marrow.

SDF-1 seems to be a chemoattractant for HSCs express-

ing CXCR4, although CXCR4 cells may also migrate

towards SDF-1, and it has been demonstrated in vivo

that CXCR4 expression by CD34+ CD38 Lin cells

confers no advantage in the rescue of irradiated NOD\

SCID mice [17]. Clinically, advantage is taken of the

ability of HSCs to migrate between stem cell compart-

ments; granulocyte colony-stimulating factor (G-CSF)

is particularly important as an agent capable of mobil-

izing HSCs, which are then harvested for transplantation[8]. Given the probable requirement for directionality

of stem cell trafficking, it is interesting to speculate

whether tissue-specific homing signals, such as selectins

[18], are invoked by damage to encourage recruitment

to the affected organ(s). In some studies of adult stem

cell plasticity, whole bone marrow aspirates have been

injected into recipients and so the compartment respon-

sible for all of the novel progeny cannot be determined,

but in others, exquisite effort has been invested to

identify the subpopulation of cells that is capable of

integrating into specific tissues.

MSCsFriedensteinand colleagues reported in 1976 that marrow

aspirates grown at low dilution formed fibroblastic

colonies [19]; it was found that they could differentiate

into bone and cartilage and were transplantable [20].

Subsequently, Owen and Friedenstein [21] proposed that

stromal cells from marrow in particular had the potential

to generate adipocyte and osteocyte progenitors. Approx.

30% of human marrow aspirate cells adhering to plastic

are considered to be marrow stromal cells [22]. They can

be expanded in vitro [23,24] and then induced to

differentiate. The fact that adult MSCs can be expanded

in vitro and stimulated to form bone, cartilage, tendon,

muscle or fat cells makes them attractive for tissue

engineering and gene therapy strategies [25].

Assessing the factors that contribute to human MSC

plasticity in vivo is complex, as there may be bias induced

during the isolation of MSCs, or it may already exist in

vivo due to the existence of regions of marrowwitha pro-

pensity to differentiate along specific pathways [26,27].

Furthermore, there is significant variation between

mouse strains in the yield of MSCs and their ability to

differentiate along selected pathways [28]. A variety of

factors might also affect MSC commitment in vitro,

including treatmentsused to subclone cells, factors within

tissue culture sera, culture plastics of different com-

position, and interactions between different colonies

expressing various growth factors and cytokines. Such

alterations are likely to have profound effects on the

differentiation repertoire of transplanted cells. In vivo

assays have been developed to assay MSC function.MSCs injected into the circulation can integrate into a

number of tissues (see below) including, importantly,

bone marrow, from which they or their descendants

might be released as part of a normal pattern of

trafficking. Skeletal and cardiac muscle phenotypes have

been reported to reside in the MSC repertoire, encour-

aged by exposure to 5-azacytidine [29], although in-

completely purified HSC populations have also been

reported to be able to contribute to the repair of mouse

tibialis anterior muscles [30], and highly purified Lin c-

Kit+ cells are extremely efficient at generatingcardiomyo-

cytes [31].

Neuronal differentiation of rat and human MSCs inculture can be induced by exposure to -mercapto-

ethanol, DMSO or butylated hydroxyanisole [32]. Fur-

thermore, MSC-derived cells are seen to integrate deep

into brain after peripheral injection as well as after direct

injection of human MSCs into rat brain; they migrate

along pathways used during the migration of neural stem

cells developmentally, become distributed widely and

start to lose markers of HSC specialization [24]. What

they become is less clear, although in related studies with

mouse MSCs, some adopted neural or astrocyte pheno-

types, with expression of glial fibrillary acidic protein

(GFAP) and neurofilament markers [33]. Mouse reci-

pients of MSCs prepared from enhanced GFP (eGFP)-transgenic mice [34] were found to have a large numberof

eGFP fluorescent cells in their brains with a variety of

morphologies that co-expressed the neuron-specific mar-

kers NeuN or NF-H or the astrocyte marker GFAP [34].

Side-population (SP) cellsA numerically minor population of cells can be isolated

from marrow and other organs of several species by

FACS on the basis of exclusion of the fluorescent dye

Hoechst 33342 [35,36]. These SP cells have considerable


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potential to differentiate and integrate into other organs

(see below); in some circumstances they lack CD34 and

thus appear uncommitted to haematopoietic lineages

[37], and it is possible that they are similar to primitive

blast-like cells that appearto be able to differentiate along

MSC or HSC lines [7]. SP cells appear to be able to

exclude xenobiotic molecules by virtue of overexpression

of a number of drug efflux membrane transporter

proteins, and this property may confer a survival ad-

vantage, a feature valuable in stem cell survival [38].

SOMATIC SITES OF STEM CELL PLASTICITY

Cardiovascular system

Blood vesselsThe endothelium of vessels in a variety of settings

experiences a degree of turnover that is detectable after

transplantation of organs or marked cells. A proportionof the endothelium derives from circulating angioblasts,

and these can be harvested during the preparation of

haematopoietic grafts from HSCs mobilized into per-

ipheral blood [39]. Endothelial cell progenitors isolated

from circulating mononuclear blood CD34+ and Flk-1+

populations [40] can differentiate into endothelial cells in

vitro, and in vivo they or their progeny contribute to

neoangiogenesis driven by ischaemic injury in mouse and

rabbit models [41]. Some progeny have been shown to

integrate into new microvessels in skin, heart, skeletal

muscle, endometrium and corpus luteum [42]. These

marrow-graft-derived angioblasts or endothelial pro-

genitor cells are mobilized following ischaemia, or afterG-CSF pretreatment [43].

Over 30 years ago, Williams and Alvarez [44] looked

for the presence of Barr bodies in a transplanted male

kidney and reported that the endothelium of a tertiary

artery (but not vein) appeared to be derived from the

female recipient. Endothelial turnover may be slow,

unless there is endothelial damage: Sinclair [45] found

that in 37 of 40 cross-gender kidney transplants no

significant endothelial repopulation had occurred, yet in

three cases where grafts were severely damaged a high

proportion (6080%) of the endothelial cells in peritu-

bular capillaries and veins were derived from the reci-

pient. It was suggested that extensive acute damagerequires repair by host cells, while less severely damaged

grafts could be restored by endothelial continuity from

surviving donor endothelial cells. The extent of replace-

ment of endothelial cells lining small renal vessels has

been reported by Lagaaij et al. [46] to be related to the

severity of vascular rejection. In this study, six of seven

grafts affected by vascular rejection showed over 33%

recipient-derived endothelial cells, whereas just two of 13

patients without evidence of rejection showed such

extensive endothelial re-colonization [46]. We have seen

occasional male endothelial cells in human renal trans-

plants in which female kidneys were grafted into male

recipients [47]. However, Andersen and colleagues [48]

studied 45 renal biopsies from 40 sex-mismatched trans-

plant patients suspected of developing acute rejection but

found no evidence of revascularization by the recipient,

even in four cases where the transplant failed.

The origin of the glomerular endothelium in trans-

planted kidneys is less clear. It might be expected that the

migration and integration of recipient endothelial pro-

genitor cells should occur in the glomerulus, yet Sinclair

[45] considered glomeruli to be unaffected, Lagaaij et al.

[46] did not comment on them and Andersen et al. [48]

found no recipient endothelium.

Williams and colleagues [44,49] studied the endothelial

repopulation of grafted segments of aorta, and found up

to 10 % of the endothelium to be host-marrow-derived.

Intriguingly, the extent of engraftment was less when

rejection was attenuated by immunosuppression. In-

jection of bone marrow cells (principally MSC-derived)into damaged rat heart muscle promoted angiogenesis,

and some of the new capillaries were MSC-derived [29].

Adult human CD34+ bone marrow cells mobilized by

G-CSF have recently been shown capable of contributing

to the repair of rat hearts following infarct induced

by ligation of the left anterior descending coronary

artery [50]. This remarkable ability was shown to

be due principally to angioblast precursors (CD34+\

CD117Bright\GATA-2Hi) generating new human capil-

laries specifically within the infarct zone that improved

the salvage of rat myocytes (not generating new myo-

cytes). Further support for marrow-derived cells con-

tributing to maintenance angiogenesis was provided byGunsilius and colleagues [51], who studied patients with

chronic myeloid leukaemia: individual endothelial cells

in the heart vessels of one patient were seen by fluores-

cence in situ hybridization to bear the chromosomal

translocation, and similarly some endothelial cells were

derived from a therapeutic HSC graft. Orlic and col-

leagues [31] found that the direct injection of highly

purified rat Lin c-Kit+ cells into infarcted rat hearts

produced substantial repair via generation of not only

new marrow-derived endothelial cells, but cardiomyo-

cytes and smooth muscle cells too. Circulating cells

derived from the recipients of heart allografts in mice

were thought to contribute substantially to the formationof neointimal hyperplasia when acute rejection was

suppressed with FK506 [52]. Whether circulating smooth

muscle progenitors are recruited in large numbers, or

proliferation occurs from just a few, was not established,

and others have suggested that marrow-derived cells do

not contribute significantly to the newly expanded

population of smooth muscle actin-expressing cells [53].

In patients that have received liver transplants, repopula-

tion of both portal and hepatic veins by endothelium of

recipient origin has been observed [54]. In the same


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report a similar observation was made in a proportion of

mice following bone marrow transplantation, suggesting

marrow origin [54].

HeartCardiomyocytes appear to have a modest capacity for

self-renewal in areas adjacent to infarcted myocardium

[55]. Non-myogenic tissue sources of cells with cardio-

myocyte differentiation potential have been identified. A

cell line (WB-F344), derived clonally from a young male

rat liver and tagged with the Escherichia coli lacZ gene,

showed cardiomyocyte differentiation 6 weeks after

direct injection into the left ventricle of nude female mice

[56]. The donor origin of the cells was confirmed by the

presence of the rat Y chromosome and the expression of

E. coli -galactosidase, and these cells expressed cardiac

troponin T and formed intercalated discs with host

myocytes. A less surprising observation, perhaps, is that

some bone marrow cells can differentiate into cardio-

myocytes [31]. In female mice, direct injection of Lin

c-Kit+ bone marrow cells from male eGFP-transgenic

donors into the contracting area bordering an infarct

induced by coronary ligation resulted in more than half

of the infarcted area being colonized by donor cells

within 9 days. Thedonororiginof thecellswas confirmed

by the presence of the Y chromosome and by eGFP

expression. These cells were seen to proliferate in situ and

expressed proteins characteristic of cardiac tissue, in-

cluding connexin 43, suggesting intercellular communi-

cation. Bone marrow cells can also contribute in a more

subtle way to the restoration of cardiac function after

myocardial infarction: labelled CD34+ human bone

marrow cells injected intravenously into athymic nuderats with an experimental myocardial infarction were

found to enhance infarct zone microvascularity and

reduce ventricular remodelling, a process which, if left

unchecked, precipitates heart failure [50]. Using the

technique of Y-chromosome detection in sex-mismat-

ched cardiac transplants, a small proportion of cardio-

myocytes, coronary arterioles and capillaries of bone

marrow origin have been identified in humans [57].

Recently, Jackson and colleagues [58] demonstrated in

mice that haematopoietic SP cells could migrate into

ischaemic cardiac muscle and blood vessels, and dif-

ferentiate into cardiomyocytes and endothelial cells.

Furthermore, studies in human heart transplant patientssupport a role for circulating stem cells (presumably from

the bone marrow) that are able to engraft into the heart

and differentiate into cardiomyocytes and endothelia

[57].

Adult mouse MSCs in culture can generate sponta-

neously beating cardiomyocytes [59], and there is evi-

dence for the generation in vivo of mature cardiac

myocytes derived from adult stem cells. Tomita and

colleagues [29] found that injection of bone marrow cells

into cryo-scarred hearts in vivo induced angiogenesis,

but only bone marrow cells cultured with 5-azacytidine

(to induce differentiation into cardiac-like muscle cells)

were able to integrate within ventricular scar tissue and

improve myocardial function. Bittner and colleagues [60]

used male wild-type bone marrow and spleen cells to

treat female mdx mice; in this model of Duchenne

muscular dystrophy, these authors observed occasional

male cardiomyocytes within the cardiac muscle syn-

cytium and a few male endothelial cells in cardiac vessels.

In contrast, Pereira andcolleagues [61] found no evidence

of MSC-derived cells in the heart or aorta of mice 2.5

months after intraperitoneal injection, and Kocher and

colleagues [50] reported that no marrow-derived cardiac

myocytes were detectable after tail vein injection of adult

human CD34+ bone marrow cells into rats with infarcts,

although it may be that the nature of the lesion promoted

incorporation and differentiation in an exclusively angio-

blast direction. The field appears to be moving rapidly

into the clinical arena, as a German team recently pub-

lished results from a series of patients suffering frommyocardial infarction who were treated by coronary an-

gioplasty and injection of autologous bone marrow into

their coronary arteries; a functional benefit in the physio-

logical function of the recipient hearts was reported [62].

Bone marrow can also contribute to pathological angio-

genesis; in a transplantable murine neuroblastoma, bone

marrow-derived cells can be partly responsible for the

tumour neovasculature [63]. This was exploited thera-

peutically: marrow cells transduced with a truncated

soluble vascular endothelial growth factor receptor-2

(tsFlk-2) slowed tumour growth and reduced tumour

vascularity.

LungThe bronchopulmonary tree is lined throughout by

epithelial cells, and indigenous multipotential stem cell

populations have been proposed to exist at several levels,

based on observations made after cell injury. In the

proximal airways (trachea and bronchi), which are lined

by pseudostratified epithelia, the so-called basal cells

appear to be the major proliferative cells. In the terminal

and respiratory bronchioles the dome-shaped Clara cells

show an enhanced proliferative rate after injury [64],

whereas in the alveoli the cuboidal-shaped Type II

pneumocyte appears to be the stem cell that proliferatesand generates progeny that can differentiate into Type I

(squamous) pneumocytes. In terms of plasticity, it has

been claimed that even a single cell from a male bone

marrow population (lineage-depleted and enriched for

CD34+ and Sca-1+ by in vivo homing to the bone

marrow) can, when injected into female recipients along

with 2i10% female supportive haematopoietic progeni-

tor cells, give rise to a variable proportion of epithelial

cells in some organs: at 11 months, a surprisingly high

proportion (20%) of cytokeratin-expressing alveolar


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pneumocytes were Y-chromosome-positive (many were

identified as Type II cells by surfactant B synthesis), with

somewhat fewer (4%) Y-positive bronchial epithelial

cells [65]. The high level of lung engraftment was

attributed to lung damage caused by either the lethal

irradiation to facilitate bone marrow transplantation or

viral infection in the temporarily immunosuppressed

animals. Somewhat different observations were made by

Kotton et al. [66], who injected plastic-adherent cultured

bone marrow cells intravenously into recipient mice 5

days after alveolar injury induced by bleomycin. The

lung was the only organ with any engraftment and, as one

might expect, injury promoted this process, but surpri-

singly observations between 1 and 30 days after injection

found only Type I and not Type II pneumocytes

(considered stem cells for Type I) of donor origin. In this

study the donor cells were found in occasional clusters,

but an absence of proliferative activity among these cells

may suggest multiple cells engrafting into particular

niches rather than clonal proliferation.

Gastrointestinal tractThe mucosa of the gastrointestinal tract has clearly

demarcated stem cell regions: in the gastricgland thisarea

is just below the foveolus, while in the intestine stem cells

arelocated close to thecryptbases. It is generally accepted

that these stem cells are multipotential, capable of giving

rise to all the indigenous lineages [67]. Adult mouse brain

neural stem cells are able to be reprogrammed and

contribute to the liver and intestine of chimaeras generat-

ed in chick or mouse embryos [68]. In adult mice, there is

some evidence supporting the integration of marrow-

derived cells into functional epithelial cells in theoesophagus, stomach, and small and large bowels. At 11

months after engraftment of a single male HSC (sup-

ported with female marrow transplantation), Krause and

colleagues [65] discovered that 0.191.81% of cells within

the gastrointestinal tract were HSC-derived and strongly

resembled, for example, absorptive villus epithelial cells

in their morphology. Despite being clearly cytokeratin-

positive and being near to the stem cell niches [69] in

colonic crypts and gastric glands, there was no evidence

that these cells were part of a local tissue-specific clonal

stem cell population, and it is possible that they had

differentiated directly from a circulating multipotential

cell. Foetal mouse liver (embryonic day 13.5) alsocontains a population of highly clonogenic cells (for

liver), but when injected into the duodenal wall these

cells canapparently form villus andcryptal epithelial cells

[70].

LiverPossibly because of its unique exposure to a host of

potentially harmful foreign compounds, the liver can call

upon indigenous populations of both functional stem

cells and potential stem cells [71]. In response to

parenchymal cell loss, the hepatocytes are the cells that

normally restore the liver mass, rapidly re-entering the

cell cycle from the G!

phase. However, even after a two-

thirds partial hepatectomy, the remaining cells only have

to cycle two or three times to restore pre-operative cell

number, and this fact led to the incorrect assumption that

hepatocytes are mere progenitor cells with only limited

division potential. However, hepatocyte transplantation

protocols, developed because of the shortage of livers for

whole-organ transplantation, have shown that the trans-

planted cells are capable of significant clonal expansion

within the diseased liver of the recipient, and so at least

some can be considered true functional stem cells. When

either massive damage is inflicted upon the liver or

regeneration after damage is compromised, a potential

stem cell compartment located within the smallest bran-

ches of the intrahepatic biliary tree is activated. This so-

called oval cell response or ductularreaction amplifies

the biliary population before these cells differentiate into

hepatocytes [72,73].One of the first demonstrations of stem cell plasticity

was in the liver. Antigens traditionally associated with

haematopoietic cells can also be expressed by oval cells,

including c-Kit, Flt-3, Thy-1 and CD34; this led to the

suggestion that perhaps bone marrow cells are at one end

of a common differentiation spectrum, with hepatocytes

at the other end. Oval cells\hepatocytes were first

discovered to be derived from circulating bone marrow

cells in the rat: Petersen and colleagues [74] followed the

fate of syngeneic male bone marrow cells transplanted

into lethally irradiated female recipient animals whose

livers were subsequently injured by a regime of 2-

acetylaminofluorene (which blocks hepatocyte regen-eration) and carbon tetrachloride (which causes hepato-

cyte necrosis) designed to cause oval cell activation. Y-

chromosome-positive oval cells were found 9 days after

liver injury, and some Y-chromosome-positive hepato-

cytes were seen after 13 days, when oval cells were

differentiating into hepatocytes. Additional evidence for

hepatic engraftment of bone marrow cells came from a rat

whole-liver transplant model. Lewis rats expressing the

MHC class II antigen L21-6 were made recipients of

livers from Brown Norway rats that were negative for

L21-6. Subsequently, ductular structures in the trans-

plants contained both L21-6-negative and L21-6-positive

cells, indicating that some biliary epithelium was of insitu derivation and some was of recipient origin, pre-

sumably from circulating bone marrow cells.

Using a similar gender-mismatch bone marrow trans-

plantation approach in mice to track the fate of bone

marrow cells, Theise and colleagues [75] reported that,

over a 6 month period, 12% of hepatocytes in the

murine liver may be derived from bone marrow in the

absence of any obvious liver damage, suggesting that

bone marrow contributes to normal wear and tear

renewal. It was thought unlikely that the bone marrow


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transplant contained a liver progenitor cell that was not

of bone marrow origin, since 200 CD34+ Lin marrow

cells produced the same degree of hepatic engraftment as

20000 unfractionated bone marrow cells.

Alison et al. [76] and Theise et al. [77] have demon-

strated that hepatocytes can also be derived from bone

marrow cell populations in humans. Two approaches

were adopted. First, the livers of female patients who had

previously received a bone marrow transplant from a

male donor were examined for cells of donor origin using

a DNA probe specific for the Y chromosome, localized

using in situ hybridization. Secondly, Y-chromosome-

positive cells were sought in female livers engrafted into

male patients, but later removed due to recurrent disease.

In both sets of patients, Y-chromosome-positive hepato-

cytes were readily identified. The degree of hepatic

engraftment of HSCs into human liver was highly

variable, most probably related to the severity of paren-

chymal damage, with up to 40% of hepatocytes and

cholangiocytes being derived from the bone marrow in aliver transplant recipient with recurrent hepatitis C.

Furthermore, a study of 27 sequential biopsies from nine

liver transplant recipients found that, whereas biliary

epithelial chimaerism was a consistent feature of most

biopsies, hepatocyte chimaerism was more prominent in

those patients suffering recurrent hepatitis, again sug-

gesting that local organ damage is necessary for signifi-

cant engraftment of circulating stem cells into the liver

[78].

Importantly, the ability of bone marrow cells to cure a

metabolic liver disease has been shown in mice (type 1

tyrosinaemia; see above) [6], thus establishing haemato-

poietic cells as a stem cell population for hepatocytes.While it seems logicalto believe that parenchymal damage

is a stimulus to hepatic engraftment by HSCs, the

molecules that mediate this homing reaction to the liver

are unknown. Petrenko and colleagues [79] speculated

that, in mice, the molecule AA4 (murine homologue of

the C1q receptor protein) may be involved in the homing

of haematopoietic progenitors to the foetal liver maybe

this receptor protein is expressed on HSCs that engraft to

the damaged liver. Clearly there may well be multiple

other signals mediating this engraftment.

PancreasThe pancreas is composed of two components, theexocrine portion organized into aciniand secretory ducts,

and the endocrine portion organized into islets of

Langerhans. Most attention has been paid to the latter,

which contain the -cells, and are responsible for plasma

glucose homoeostasis. Until recently, it had been thought

that a person is born with all the pancreatic -cells they

ever have, but it is now apparent that in adulthood low

levels of mature -cell replication and apoptosis mean

that the -cell population should be defined as a slowly

renewing population [80]. The pancreatic ducts appear to

be the site of multipotential stem cells with the potential

to generate endocrine, acinar and ductular cell pheno-

types [81], even giving rise to new islets (islet neogenesis)

when presented with a functional demand. The latter

observation has led to the belief that all mature duct cells

are potential stem cells, able to temporarily attain a less

differentiated phenotype, expand and subsequently dif-

ferentiate along any one of the pancreatic lineages [82].

Indeed, functional islet -cells have been generated in

vitro from cultured pancreatic ductal cells [83]. Recently,

Zulewski et al. [84] suggested that, in both islets and

ducts, a subpopulation of cells expressing the neuronal

stem cell marker nestin are the true stem cells; ex vivo,

these cells are highly clonogenic and can differentiate not

only into endocrine and exocrine pancreatic cells, but

also into cells with a hepatic phenotype. It is not

surprising that certain pancreatic cells can transdiffer-

entiate into hepatocytes. During development the ventral

pancreas and liver emerge from the same general area

of ventral foregut endoderm, but fibroblast growthfactor (FGF) from the cardiac mesoderm inhibits pan-

creatic development in the presumptive liver [85]. An-

other example of pancreaticliver cellular plasticity was

demonstrated by Krakowski et al. [86], who generated

insulin-promoter-regulated keratinocyte growth factor

transgenic mice. Under the influence of keratinocyte

growth factor, numerous functional hepatocytes emerged

within the islets of Langerhans. A combination of dexa-

methasone and oncostatin M (a natural hepatocyte dif-

ferentiation factor produced by haematopoietic cells in

the foetal liver) is a very effective in vitro inducer of

pancreatic exocrine cell transdifferentiation into hepato-

cytes [87]. This differentiation was associated with theinduction of the transcription factor C\EBP(CCAAT\

enhancer-binding protein ), a factor thought to ac-

celerate fatty acyl-CoA synthesis. This in turn bound to

hepatocyte nuclear factor 4, causing its translocation to

the nucleus, where it activated genes such as those

encoding -fetoprotein and transthyretin that are nor-

mally switched on during early hepatocytic differen-

tiation.

In the FAH-deficient mouse model of type 1 tyro-

sinaemia, transplantation with pancreaticcells is generally

not life-saving, but a small proportion of animals do

survive, with 5090% replacement of the diseased liver

with pancreatic-cell-derived hepatocytes [88]. Given thatanimals fed on a copper-deficient diet undergo pancreatic

exocrine cell atrophy and that refeeding induces the

surviving ducts to give rise to hepatocytes, it was

surprising that pancreatic cell suspensions enriched in

pancreatic ducts were poorer than unfractionated pan-

creatic cells at reconstituting the diseased FAH/ liver

with functional hepatocytes [89]. Moreover, we have

already noted that the pancreatic ducts appear to be the

location of mutipotential stem cells, at least for pancreatic

lineages; however, human pancreatic exocrine cells in


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vitro readily assume a ductal phenotype, and re-express

PDX-1 (pancreas\duodenal homeobox 1), a homeo-

domain protein essential for pancreatic ontogeny [90].

Thus exocrine cells can be a source of multipotential stem

cells for the pancreas, and this flexibility seems to extend

to hepatic lineages both in vitro and in vivo.

KidneyThe kidney has no recognizable stem cell zone, but

tubular cells can regenerate after injury. Adult mouse

neural stem cells injected into an early embryo contribute

to the developing kidney [68], so adult cells may be

reprogrammed to differentiate into renal cells. In human

renal transplants where female kidneys are grafted into

male recipients, we have noted male tubular cells ex-

pressing the epithelial marker CAM 5.2 [47]. Further-

more, Grimm and colleagues [91] found evidence for

circulatinghost-derived mesenchymal cells in renaltrans-

plants that were suffering chronic rejection. However,Andersen and colleagues [48] reported that tubular and

glomerular cells remained of donor origin in transplanted

kidneys even 10 months after transplantation. In the

kidney a conversion or transdifferentiation may occur

between the phenotype of epithelial cells and fibroblasts,

both being generated originally from the primitive

metanephric mesenchyme [92]. In various models, epi-

thelial cells are seen to acquire markers of fibroblasts or

myofibroblasts and adopt a fusiform morphology; in

interstitial fibrosis, cells are seen with a fibroblastic

morphology that bear epithelial markers. Transdifferen-

tiation appears to be restricted to regions where the

basement membrane is damaged, with most myofibro-blastic cells being seen where the tubular basement

membrane was extensively damaged [93].

These observations, and others on cultured cells, are

concordant with the hypothesis that the epithelium

adopts a fibroblastic morphology before proliferating,

and perhaps before helping to repair the basement

membrane. Sun and colleagues [94], examining rat kid-

neys after uranyl acetate-induced tubular necrosis, con-

sidered that the repair occurred without the movement of

cells from the interstitium into the denuded tubules, yet

they observed proliferation of flattened cells lining the

regenerating tubules that expressed vimentin, like myofi-

broblasts. So, is there in vivo an influx of cells that firstadopt a fibroblast morphology, expand and then dif-

ferentiate intoepithelium ? Kidney was one of the murine

organs studied by Krause and colleagues [65] in their

search for evidence of epithelial differentiation following

engraftment with a single male bone marrow cell.

Surprisingly, glomeruli were not commented upon, and

no donor-derived renal tubular epithelial cells were seen

in any of the five mice. Perhaps this was due to the use of

HSCs, ratherthan stromal cells, as in ourstudies of whole

bone marrow transplants we have observed marrow-

derived renal tubular epithelial cells and, within glo-

meruli, marrow-derived cells that appeared to be podo-

cytes [47]. Podocytes are central to the maintenance of

glomerular capillary permeabilityand one of the potential

therapeutic cellular targets in thekidney. Nephrin, a large

protein normally present in the podocyte plasma mem-

brane at the filtration slits, is mutated in a range of

nephrotic syndromes, including congenital nephropathy

of Finnish type. Another potential target for stem cell

therapy is Alports syndrome, in which the absence of

specific collagen IV chains in the renal basement mem-

brane leads to the progressive impairment of renal

function. Another important site of renal pathology is the

mesangium, and the bone marrow appears capable of

supplying mesangial cells. Bone marrow rescue and

transfer of a mesangial sclerosing defect has been shown

elegantly in a mouse model [95].

Nervous systemThe mammalian brain develops as a tube containing aventricular compartment filled with cerebral fluid. Dur-

ing development, the dividing cells are located in the cell

layer that lines the lumen of the neural tube (cor-

responding to the localization of the ependymal cells in

the adult). These cells show a trilineage potential, being

capable of differentiating into astrocytes, oligodendro-

cytes or neurons. In the adult, single cells isolated from

the lining of the ventricular system (ependymal cells and

cells from the subventricular zone [96], where it exists)

are capable of forming spheroids of tightly clustered cells

(neurospheres) that show the same trilineage potential

[9799]. The existence of multipotential neural stem cellsin vivo is now widely accepted, and growth factors such

as epidermal growth factor and FGF-2 are significant

players in their self-renewal [100,101]. Furthermore, cells

with considerable replication potential and the ability to

form astrocytes and neurons can be isolated from human

post-mortem tissue [102]. More intriguingly, Bjornson et

al. [103] demonstrated that single neural stem cells with

trilineage potential could transdifferentiate into several

haematopoietic lineages. Clonally derived neural stem

cells cultured from ROSA26 mice were injected into

sublethally irradiated Balb\c mice.An in vitro clonogenic

assay of the bone marrow from the transplanted mice

showed that some of the colonies were positive for -galactosidase, suggesting a neural stem cell origin. Signifi-

cantly, cultured neural stem cells neither proliferated nor

formed haematopoietic progeny in the same clonogenic

assays without prior injection into the irradiated hosts,

indicating that the appropriate microenvironment is

necessary for transdifferentiation. Likewise, clonally

derived human neurosphere cells derived from foetal

tissue and expanded in vitro by epidermal growth factor

and\or FGF-2 show no haematopoietic potential in

culture, but can establish long-term haematopoiesis in


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human bone fragmentsin SCID-hu mice [104]. However,

a recent study using a similar protocol to Bjornson et al.

rigorously tested the haematopoietic potential of murine

neurosphere cells and was unable to find any evidence of

haematopoietic differentiation in a large group of suble-

thally irradiated mice, which suggests that haemato-

poietic potential is not a general property of neural stem

cells [105]. In mousechick chimaeras, created by the

injection of neurospheres from ROSA26 mice,-galacto-

sidase-positive cells contributed to several tissues, in-

cluding liver, gastric mucosa and mesonephric tubules

[68]. These experiments demonstrate that neural stem

cells have considerable flexibility, but do not prove that

these differentiation pathways exist normally. In par-

ticular, they would not satisfy one of the criteria

stipulated by Anderson et al. [2], namely that the donor

population should be transplanted without intervening

culture manipulations, since all studies have used cells

from cultured neurospheres. Neural stem cells have other

potentialities; clonally derived murine and human adultneural stem cells can undergo apparent myogenic dif-

ferentiation in vitro when co-cultured with myoblasts,

suggesting that mature tissues can provide epigenetic

signals to neighbouring cells to undergo alternative

pathways of differentiation [106]. Moreover, a small

percentage (12%) of such cultured cells could undergo

myogenic differentiation when injected into cardiotoxin-

damaged skeletal muscle. Likewise, Clarke et al. [68]

showed that neural stem cells from ROSA26 mice co-

cultured withES cell-derived embryoid bodies are able to

undergo myogenic differentiation. Impressive as all these

data might seem, with multiple lines of evidence proffered

for myogenic conversion, they in no way provideevidence that neural stem cells can act as stem cells for

skeletal muscle, and there is no real expectation that

trafficking from the brain to skeletal muscle occurs in

vivo.

Looking for plasticity in the other direction, it is

readily apparent that cells from outside the nervous

system can differentiate into neurons and glial cells.

Mezey et al. [107], using homozygous PU.1 mutant

female mice (PU.1 is a transcription factor required for

the histogenesis of six of the haematopoietic lineages) as

recipients of a life-saving bone marrow transplant from

male wild-type donors, showed that up to 4.6% of CNS

cells were Y-chromosome-positive, and that up to 2.3%of Y-positive cells possessed the neuronal markers NeuN

and neuron-specific enolase. Similarly, Eglitis and Mezey

[108] detected significant numbers of microglia (F4\80-

positive) and astrocytes (GFAP-positive) of bone mar-

row origin in the brains of recipient female mice 6 weeks

after transplantation of male bone marrow. Marrow

stromal cells may also be able to differentiate along CNS

lines. Ventricular transplantation of myelodepleted mu-

rine stromal cells (marked by bromodeoxyuridine in

culture) resulted in their widespread distribution by 12

days post-transplantation, and some labelled cells were

either neurofilament- or GFAP-immunopositive [33].

Marrow stromal cells can also be induced to differentiate

along neuronal lines in vitro, with the cells having

neuronal morphology and being initially nestin-positive

(characteristic of neuronal precursors) before expressing

typical neuronal markers such as neuron-specific enolase

and NeuN [32]. Given the inaccessibility of conventional

neuronal stem cells, marrow stromal cells may therefore

eventually have applications in the treatment of neuro-

degenerative disease.

SkinThe epidermis and hair follicle are prime examples of

tissues under constant insult that require a highcapability

for self-renewal. In normal epidermis, proliferation is

confined to the basal layer that contains both stem cells

and more numerous transit amplifying cells. In thin

rodent epidermis the suprabasal cells are arranged in

columns (stacks) that interdigitate with neighbouringstacks; each stack is associated with a seemingly defined

group of basal cells, and a more slowly dividing cell

underneath the centre of each stack has been proposed to

be the stem cell for the so-called epidermal proliferative

unit. Human epidermis is much thicker and is generally

not stacked, and the identity of stem cells is more

controversial, although markers such as 1 integrin have

been proposed [109]. In most areas of the epidermis these

are confined to the tops of the dermal papillae [110]. The

basal cells of the interfollicular epidermis are continuous

with those of the hair follicle, and here multipotential

stem cells are tucked away in the permanent portion of

the follicle called the bulge. Bulge cells have the classicstem cell properties of low in vivo proliferation and high

in vitro clonogenic potential. Elegant experiments in-

volving creating chimaeric vibrissal (whisker) follicles by

transplanting the bulge region from ROSA26 mice into

wild-type mouse follicles have shown that bulge cells

migrate both downwards and upwards, forming all

follicular, sebaceous and epidermal lineages [111].

In terms of plasticity, the study of Krause et al. [65]

indicated that haematopoietic cells in the female mouse

could differentiate into cytokeratin-positive epidermal

cells; these authors found, using Y chromosome tracking

techniques, that approx. 2% of epidermal cells were Y-

chromosome-positive 11 months after bone marrowtransplantation. No clonal proliferation of such cells was

seen, although the authors illustrate one such cell as

possibly being located in the bulge region [65]. Epidermis

and pilosebaceous units can be generated from not-too-

dissimilar tissue, certainly indicative of plasticity. Com-

bining murine embryonic dermis with rabbit central

corneal epithelium causes the transit amplifying cells,

thought to be located here, to be reprogrammed [112].

Cells with multipotentiality have been isolated from

rodent and human skin, specifically from the dermis, and


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have been christened SKPs skin-derived precursors

[113]. These cells could undergo multiple rounds of cell

division and could be instructed to undergo differen-

tiation along neuroectodermal lines (neurons and glial

cells) or mesodermal lines (adipocytes and smooth

muscle). These cells were distinguishable in their be-

haviour from plastic-adherent bone marrow mesenchy-

mal cells, andapparently clonally derivedspheres of these

cells could generate all the above lineages. Some murine

epidermal cells may even be pluripotential; isolation of

epidermal stem cells on the basis of size and Hoechst

33342 dye exclusion from 3-day-old eGFP transgenic

mice and their injection into wild-type blastocysts results

in their incorporation into a variety of tissues in all three

germ layers [114]. Wounds created by clipping the tails of

foetal sheep at the same time that adult human MSCs

were injected intraperitoneally were found subsequently

to have human cells of fibroblastic morphology within

the dermis and dermal appendages [115]; this might

indicate that circulating MSCs have the potential to assistin skin repair processes.

Musculoskeletal system

Skeletal muscleSatellite cells are responsible for maintenance of muscle

fibres and are the local stem cells that are able to divide

and self-renew. They are mononuclear, and normally

reside between the sarcolemma and the basal lamina of

themuscle fibre. When purified from adult mouse skeletal

muscle, cultured, then injected into mice (along with

other distinguishable whole marrow), they result in a

full-range multilineage engraftment of the HSC com-partment [35]; this could be transferred to a further

mouse by bone marrow grafting. Several reports suggest

that a common haematopoietic and muscle precursor

exists in adult muscle and in marrow, and it is interesting

to speculate that the SP cell fraction of many tissues

contains a population of multipotent stem cells. The fact

that purified human muscle myoblasts, injected directly

into the muscle of patients with Duchenne muscular

dystrophy, can integrate into myotubes and express

muscle-specific transcriptswas demonstrated by Gussoni

and colleagues [116]. Subsequently, this group [36] used

the mdx mouse model of muscular dystrophy to establish

that intravenous injection of wild-type male HSCs or SPcells isolated from muscle resulted in the integration of

male nuclei into female mdx mouse myotubes, with

" 1% expressing dystrophin [117]. Ferrari and col-

leagues [30] demonstrated that whole bone marrow

contains cells able to migrate into damaged skeletal

muscleand, within weeks,contribute nuclei to myotubes.

A muscle-derived clonal cell population (mc13) express-

ing both muscle and stem cell markers was shown to

integrate at low efficiency into muscle after intravenous

injection into mdx mice; integration was greater if cells

were injected directly into the dystrophic muscle [118].

Bittner and colleagues [60] demonstrated that maledonor

marrow\spleen cells or their progeny invaded skeletal

muscleof female mdx mice and contributedto endothelial

and myotube populations; Y-chromosome-positive

nuclei wereseen withindystrophin-expressing myotubes.

Integration occurred whether or not the recipient bone

marrow was ablated by irradiation. Human skeletal

muscle cells (hybridizing to an AluI DNA probe and

expressingdystrophin, or expressinghuman#-microglo-

bulin and fast or slow myosin) were detected 5 months

after injection of adult human MSCs into foetal sheep

[115]. Neural stem cell neurospheres of human or

mouse origin are also able to contributeto skeletal muscle

fibres in vivo after transplantation into adult mice [106],

and can form myotubes after physical contact and co-

culture with C2C12 cells, which themselves showed

myotube formation. This raises a further question of

whether nuclear exchange, or the formation of hetero-

karyons, occurred in other studies that seem to supportstem cell plasticity.

BoneWhen whole male mouse marrow is injected intra-

venously into female mice that have not hadtheir marrow

ablated, donor marrow cells can contribute to the

formation of long bones at low frequency [119]. The Y-

chromosome-positive cells seen in the bones were con-

sidered to be functionally active as osteoblasts, producing

bone before being encapsulated within the bone lacunae

and terminally differentiating into osteocytes. Y-positive

flattened bone-lining cells on the periosteal bone surface

were also present. The important principle that stem cells(MSCs) can be used to direct tissue-specific gene ex-

pression wasshown clearly by Hou andcolleagues [27] in

mice. They used a reporter gene under the control of an

osteocalcin promoter; after intravenous injection, MSCs

were found throughout a wide range of tissues, but

expression of the reporter gene was found only within

bony tissues, confined to a subset of osteoblasts and

mature osteocytes within well formed lacunae. Pereira

and colleagues [120] showed that MSCs expanded from

mice transgenic for a human collagen I minigene and

injected intravenously into recipient wild-type mice were

able to infiltrate a variety of tissues; the frequency of

MSC-derived cells within the organs increased over time,and expression of mRNA from the minigene was seen in

bone, but not cartilage. The recipient lungs appeared to

contain large numbers of MSC progeny, but expressed

the mRNA at a lower level.

A variety of therapeutic protocols have been examined

using a mouse model [61] of osteogenesis imperfecta

(OI), a genetic disorder of one of the genes for collagen I

chains that form the primary protein scaffold for bone

formation, which frequently results in a generalized

osteopenia, fragile bones and short stature. These studies


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assessed the extent of engraftment of tissues with wild-

type MSCs or whole marrow cells, and sought to detect

any improvement in bone composition. The authors

hypothesized that bone cells derived from the trans-

planted marrow would have a selective advantage over

the resident OI cells, as OI MSCs are defective in

differentiation towards an osteoblast phenotype in vitro.

It was found that 3-week-old female OI mice given

several injections of male MSCs intraperitoneally showed

small but significant increases in bone collagen and

mineral content 1 month later. Male MSC-derived cells

were detected by fluorescence in situ hybridization in

primary cultures from tissues of one OI mouse at 2.5

months; male cells comprised 7% of long bone and 15%

of calvaria cells, but the phenotype of these cells was not

determined. Therapeutic intervention has been attempted

in infant patients with OI. In a proof of principle study

[121], patients were given whole bone marrowgraftsafter

ablation of their original marrow; after 3 months, all

three initial patients showed an increase in total bodybone mineral content, associated with improved growth

and less fracturing. Cultures of a trabecular bone biopsy

from a female patient receiving male marrow revealed

that "1.5% of osteoblasts were donor-derived [122],

and it is not obvious how such a low level of engraftment

could produce the substantial benefits described. Subse-

quently, additional grafts of MSCs from the original

donors have been administered to see if a greater

proportion of osteoblasts can be replaced [123].

A potential complication of studies of bone growth

and turnover in which growth or mineralization effects

are attributed to MSC grafts is that osteoclasts, the

primarycells responsible for resorption of bone normallyand under pathological conditions, arise from precursors

of the monocyte\macrophage lineage elaborated by

HSCs. Another complication is that a population of non-

adherent low-density cells exist in marrow that have the

ability to promote bone precursor development through

the release of soluble factors [124]; these cells would be

depleted in most MSC culture protocols. The balance

between osteoblast and osteoclast formation may affect

growth, and might offer an avenue for some therapeutic

interventions: the peroxisome-proliferator-activated re-

ceptor- pathway is active in the differentiation of both

HSCs to osteoclasts [125] and MSCs to osteoblasts [126].

Muscle stem cells have been isolated and clonal popula-tions produced that yield bone in vitro on exposure to

bone morphogenetic protein 2. Further, adenoviral-

transduced expression of bone morphogenetic protein 2

by these cells allows them to make ectopic bone after

intramuscular injection, or to heal skull bone damage

[118].

CartilageKey factors involved in the differentiation of MSCs to

form mature cartilage are being identified through in

vitro studies of isolated and expanded MSCs: they can

be induced by dexamethasone and transforming growth

factor 3 to secrete an extracellular matrix incorporating

type II collagen, aggrecan and anionic proteoglycans

[127].After injection of male wild-type MSCs intofemale

OI mice, 8% of cells grown from cartilage contained a Y

chromosome [61]. Injection of prelabelled MSCs intra-

peritoneally into rats at the onset of arthritis resulted in

thepresenceof labelled cells in joint cavities andsublayers

of proliferating synovial tissues, demonstrating their

targeting ability. Furthermore, human MSCs injected

intaperitoneally into foetal sheep contributed to articular

cartilage chondrocytes, based on their appropriate loca-

tion and characteristic morphology [115].

SUMMARY

There is now a large body of evidence indicating that the

concept of organ-specific stem cells could be extended toinclude populations of stem cells that are able to

contribute to the renewal of quite different lineages, even

in tissues from a separate germ layer. Perhaps a key factor

in the generation of self-renewing clones in the new

tissues is the exposure to and successful occupation

of niches emptied by damage, with the local environ-

ment of the niche defining the cell repertoire that will be

produced [128]. Extraordinary claims require extraordi-

nary proof, and some have asked for a higher standard of

evidence; requiring a clonal approach [129] or dem-

onstration of a robust, sustained multi-lineage engraft-

ment and functional activity representative of multiple

phenotypic characteristics of the converted cells to showthat full conversion has occurred [2]. Put simply,

showing partial repopulation of an organ with cells that

have come to resemble their neighbours is not the same as

showing a functional competence as diverse and broad as

that expected of the indigenous population. Yet this is

what will be needed for tissue regeneration and gene

therapy strategies relying on adult stem cell plasticity

with clonal expansion to yield all of the cell types

normally produced, and only those, together with ap-

propriate responses to the usual demands of growth,

adaptation and repair.

REFERENCES

1 Fuchs, E. and Segre, J. (2000) Stem cells: a new lease onlife. Cell 100, 143155

2 Anderson, D., Gage, F. and Weissman, I. (2001) Canstem cells cross the lineage barrier? Nat. Med. (N.Y.) 7,393395

3 Terada, N., Hamazaki, T., Oka, M. et al. (2002) Bonemarrow cells adopt the phenotype of other cells byspontaneous cell fusion. Nature (London) 416, 542545

4 Ying, Q. L., Nichols, J., Evans, E. P. and Smith, A. G.(2002) Changing potency by spontaneous fusion. Nature(London) 416, 545548


8/14/2019 1030355

13/15


5 Srivatsa, B., Srivatsa, S., Johnson, K. L., Samura, O., Lee,S. L. and Bianchi, D. W. (2001) Microchimerism ofpresumed fetal origin in thyroid specimens fromwomen: a case-control study. Lancet 358, 20342038

6 Lagasse, E., Connors, H., Al-Dhalimy, M. et al. (2000)Purified hematopoietic stem cells can differentiate intohepatocytes in vivo. Nat. Med. (N.Y.) 6, 12291234

7 Hall, F., Han, B., Kundu, R., Yee, A., Nimni, M. andGordon, E. (2001) Phenotypic differentiation of tgf-

beta1-responsive pluripotent premesenchymalprehematopoietic progenitor (p4 stem) cells from murinebone marrow. J. Hematother. Stem Cell Res. 10,261271

8 Whetton, A. and Graham, G. (1999) Homing andmobilization in the stem cell niche. Trends Cell Biol. 9,233238

9 Weissman, I. (2000) Stem cells: units of development,units of regeneration, and units in evolution. Cell 100,157168

10 Jankowska-Wieczorek, A., Majka, M., Ratajczak, J. andRatajczak, M. (2001) Autocrine\paracrine mechanismsin human hematopoiesis. Stem Cells 19, 99107

11 Chan, J. Y. and Watt, S. M. (2001) Adhesion receptorson haematopoietic progenitor cells. Br. J. Haematol. 112,541557

12 Friedenstein, A., Ivanov-Smolenski, A., Chajlakjan, R. et

al. (1978) Origin of bone marrow stromal mechanocytesin radiochimeras and heterotopic transplants. Exp.Hematol. 6, 440444

13 Clark, B. and Keating, A. (1995) Biology of bonemarrow stroma. Ann. N.Y. Acad. Sci. 770, 7078

14 Quesenberry, P. and Becker, P. (1998) Stem cellhoming: rolling, crawling, and nesting. Proc. Natl. Acad.Sci. U.S.A. 95, 1515515157

15 Chan, J.-H. and Watt, S. (2001) Adhesion receptors onhaematopoietic progenitor cells. Br. J. Haematol. 112,541557

16 Zanjani, E., Flake, A., Almeida-Porada, G., Tran, N. andPapayannopoulou, T. (1999) Homing of human cells inthe fetal sheep model: modulation by antibodiesactivating or inhibiting very late activation antigen-4-dependent function. Blood 94, 25152522

17 Rosu-Myles, M., Gallacher, L., Murdoch, B. et al. (2000)

The human hematopoietic stem cell compartment isheterogeneous for CXCR4 expression. Proc. Natl. Acad.Sci. U.S.A. 97, 1462614631

18 Mazo, I. B., Gutierrez-Ramos, J. C., Frenette, P. S.,Hynes, R. O., Wagner, D. D. and von Andrian, U. H.(1998) Hematopoietic progenitor cell rolling in bonemarrow microvessels: parallel contributions byendothelial selectins and vascular cell adhesion molecule1. J. Exp. Med. 188, 465474

19 Friedenstein, A., Gorskaja, U. and Kalugina, N. (1976)In vitro growth of bone marrow aspirate derived cellcolonies. Exp. Hematol. 4, 267274

20 Keating, A., Singer, J., Killen, P. et al. (1982) Donororigin of the in vitro haematopoietic microenvironmentafter marrow transplantation in man. Nature (London)298, 280283

21 Owen, M. and Friedenstein, A. J. (1988) Stromal stemcells: marrow-derived osteogenic precursors. CibaFound. Symp. 136, 4260

22 Prockop, D. (1997) Marrow stromal cells as stem cellsfor non-hematopoietic tissues. Science 276, 7174

23 Colter, D., Class, R., DiGirolamo, C. and Prockop, D.(2000) Rapid expansion of recycling stem cells incultures of plastic-adherent cells from human bonemarrow. Proc. Natl. Acad. Sci. U.S.A. 97, 32133218

24 Azizi, S., Stokes, D., Augelli, B., DiGirolamo, C. andProckop, D. (1998) Engraftment and migration ofhuman bone marrow stromal cells implanted in thebrains of albino rats similarities to astrocyte grafts.Proc. Natl. Acad. Sci. U.S.A. 95, 39083913

25 Koc, O. and Lazarus, H. (2001) Mesenchymal stemcells: heading into the clinic. Bone Marrow Transplant.27, 235239

26 Phinney, D., Kopen, G., Righter, W., Webster, S.,Tremain, N. and Prockop, D. (1999) Donor variation inthe growth properties and osteogenic potential of humanmarrow stromal cells. J. Cell. Biochem. 75, 424436

27 Hou, Z., Nguyen, Q., Frenkel, B. et al. (1999)Osteoblast-specific gene expression after transplantationof marrow cells: implications for skeletal gene therapy.Proc. Natl. Acad. Sci. U.S.A. 96, 72947299

28 Phinney, D., Kopen, G., Isaacson, R. and Prockop, D.

(1999) Plastic adherent stromal cells from the bonemarrow of commonly used strains of mice: variations inyield, growth and differentiation. J. Cell. Biochem. 72,570585

29 Tomita, S., Li, R., Weisel, R., Mickle, D., Kim, E., Sakai,T. and Jia, Z. (1999) Autologous transplantation of bonemarrow cells improves damaged heart function.Circulation 100, II247II256

30 Ferrari, G., Cusella-De Angelis, G., Coletta, M. et al.(1998) Muscle regeneration by bone marrow-derivedmyogenic progenitors. Science 279, 15281530

31 Orlic, D., Kajstura, J., Chimenti, S. et al. (2001) Bonemarrow cells regenerate infarcted myocardium. Nature(London) 410, 701704

32 Woodbury, D., Schwartz, E., Prockop, D. and Black, I.(2000) Adult rat and human bone marrow stromal cellsdifferentiate into neurons. J. Neurosci. Res. 61, 364370

33 Kopen, G., Prockop, D. and Phinney, D. (1999) Marrow

stromal cells migrate throughout forebrain andcerebellum, and they differentiate into astrocytes afterinjection into neonatal mouse brains. Proc. Natl. Acad.Sci. U.S.A. 96, 1071110716

34 Brazelton, T., Rossi, F., Keshet, G. and Blau, H. (2000)From marrow to brain: expression of neuronalphenotypes in adult mice. Science 290, 17751779

35 Jackson, K., Mi, T. and Goodell, M. (1999)Hematopoietic potential of stem cells isolated frommurine skeletal muscle. Proc. Natl. Acad. Sci. U.S.A. 96,1448214486

36 Gussoni, E., Soneoka, Y., Strickland, C. et al. (1999)Dystrophin expression in the mdx mouse restored bystem cell transplantation. Nature (London) 401, 390394

37 Goodell, M., Rosenzweig, M., Kim, H. et al. (1997) Dyeefflux studies suggest that hematopoietic stem cellsexpressing low or undetectable levels of CD34 antigenexist in multiple species. Nat. Med. (N.Y.) 3, 13371345

38 Bunting, K. D. (2002) ABC transporters as phenotypicmarkers and functional regulators of stem cells. StemCells 20, 1120

39 Lin, Y., Weisdorf, D., Solovey, A. and Hebbel, R. (2000)Origins of circulating endothelial cells and endothelialoutgrowth from blood. J. Clin. Invest. 105, 7177

40 Asahara, T., Murohara, T., Sullivan, A. et al. (1997)Isolation of putative progenitor endothelial cells forangiogenesis. Science 275, 964967

41 Kalka, C., Masuda, H., Takahashi, T. et al. (2000)Transplantation of ex vivo expanded endothelialprogenitor cells for therapeutic neovascularization. Proc.Natl. Acad. Sci. U.S.A. 97, 34223427

42 Asahara, T., Masuda, H., Takahashi, T. et al. (1999)Bone marrow origin of endothelial progenitor cellsresponsible for postnatal vasculogenesis in physiologicaland pathological conditions. Circ. Res. 85, 221228

43 Takahashi, T., Kalka, C., Masuda, H. et al. (1999)Ischemia- and cytokine-induced mobilization of bonemarrow-derived endothelial progenitor cells forneovascularization. Nat. Med. (N.Y.) 5, 434438

44 Williams, G. and Alvarez, C. (1969) Host repopulationof the endothelium in allografts of kidneys and aorta.Surg. Forum 20, 293294

45 Sinclair, R. (1972) Origin of endothelium in human renalallografts. Br. Med. J. 4, 1516

46 Lagaaij, E., Cramer-Knijnenburg, G., van Kemenade, F.,van Es, L., Bruijn, J. and van Krieken, J. (2001)Endothelial cell chimerism after renal transplantationand vascular rejection. Lancet 357, 3337

47 Poulsom, R., Forbes, S. J., Hodivala-Dilke, K. et al.(2001) Bone marrow contributes to renal parenchymalturnover and regeneration. J. Pathol. 195, 229235


8/14/2019 1030355

14/15


48 Andersen, C., Ladefoged, S. and Larsen, S. (1991)Cellular inflammatory infiltrates and renal cell turnoverin kidney allografts: a study using in situ hybridizationand combined in situ hybridization andimmunohistochemistry with a Y-chromosome-specificDNA probe and monoclonal antibodies. APMIS 99,645652

49 Williams, G., Krajewski, C., Dagher, F., Ter Haar, A.,Roth, J. and Santos, G. (1971) Host repopulation of

endothelium. Transplant. Proc. 3, 86987250 Kocher, A., Schuster, M., Szabolcs, M. et al. (2001)Neovascularization of ischemic myocardium by humanbone-marrow-derived angioblasts preventscardiomyocyte apoptosis, reduces remodeling andimproves cardiac function. Nat. Med. (N.Y.) 7, 430436

51 Gunsilius, E., Duba, H.-C., Petzer, A. et al. (2000)Evidence from a leukaemia model for maintenance ofvascular endothelium by bone-marrow-derivedendothelial cells. Lancet 355, 16881691

52 Saiura, A., Sata, M., Hirata, Y., Nagai, R. and Makuuchi,M. (2001) Circulating smooth muscle progenitor cellscontribute to atherosclerosis. Nat. Med. (N.Y.) 7,382383

53 Li, J., Han, X., Jiang, J. et al. (2001) Vascular smoothmuscle cells of recipient origin mediate intimalexpansion after aortic allotransplantation in mice. Am. J.Pathol. 158, 19431947

54 Gao, Z.-H., McAlister, V. and Williams, G. (2001)Repopulation of liver endothelium by bone marrow-derived cells. Lancet 357, 932933

55 Beltrami, A. P., Urbanek, K., Kajstura, J. et al. (2001)Evidence that human cardiac myocytes divide aftermyocardial infarction. N. Engl. J. Med. 344, 17501757

56 Malouf, N., Coleman, W., Grisham, J. et al. (2001)Adult-derived stem cells from the liver becomemyocytes in the heart in vivo. Am. J. Pathol. 158,19291935

57 Quaini, F., Urbanek, K., Beltrami, A. P. et al. (2002)Chimerism of the transplanted heart. N. Engl. J. Med.346, 515

58 Jackson, K. A., Majka, S. M., Wang, H. et al. (2001)Regeneration of ischemic cardiac muscle and vascularendothelium by adult stem cells. J. Clin. Invest. 107,13951402

59 Makino, S., Fukuda, K., Miyoshi, S. et al. (1999)

Cardiomyocytes can be generated from marrow stromalcells in vitro. J. Clin. Invest. 103, 69770560 Bittner, R., Schofer, C., Weipoltshammer, K. et al.

(1999) Recruitment of bone-marrow-derived cells byskeletal and cardiac muscle in adult dystrophic mdxmice. Anat. Embryol. 199, 391396

61 Pereira, R., OHara, M., Laptev, A. et al. (1998) Marrowstromal cells: a source of progenitor cells fornonhematopoietic tissues in transgeneic mice with aphenotype of osteogenesis imperfecta. Proc. Natl. Acad.Sci. U.S.A. 95, 11421147

62 Strauer, B. E., Brehm, M., Zeus, T. et al. (2001)Intracoronary, human autologous stem celltransplantation for myocardial regeneration followingmyocardial infarction. Dtsch. Med. Wochenschr. 126,932938

63 Davidoff, A. M., Ng, C. Y., Brown, P. et al. (2001) Bonemarrow-derived cells contribute to tumor

neovasculature and, when modified to express anangiogenesis inhibitor, can restrict tumor growth inmice. Clin. Cancer Res. 7, 28702879

64 Boers, J. E., Ambergen, A. W. and Thunnissen, F. B.(1999) Number and proliferation of clara cells in normalhuman airway epithelium. Am. J. Respir. Crit. CareMed. 159, 15851591

65 Krause, D., Theise, N., Collector, M. et al. (2001) Multi-organ, multi-lineage engraftment by a single bonemarrow-derived stem cell. Cell 105, 369377

66 Kotton, D. N., Ma, B. Y., Cardoso, W. V. et al. (2001)Bone marrow-derived cells as progenitors of lungalveolar epithelium. Development 128, 51815188

67 Wright, N. A. (2000) Epithelial stem cell repertoire inthe gut: clues to the origin of cell lineages, proliferativeunits and cancer. Int. J. Exp. Pathol. 81, 117143

68 Clarke, D., Johansson, C., Wilbertz, J. et al. (2000)Generalized potential of adult neural stem cells. Science288, 16601663

69 Slack, J. (2000) Stem cells in epithelial tissues. Science287, 14311433

70 Suzuki, A., Zheng Yw, Y. W., Kaneko, S. et al. (2002)Clonal identification and characterization of self-renewing pluripotent stem cells in the developing liver.

J. Cell Biol. 156, 173184

71 Alison, M., Poulsom, R. and Forbes, S. (2001) Updateon hepatic stem cells. Liver 21, 367373

72 Alison, M. R., Golding, M., Sarraf, C. E., Edwards, R. J.and Lalani, E. N. (1996) Liver damage in the rat induceshepatocyte stem cells from biliary epithelial cells.Gastroenterology 110, 11821190

73 Alison, M. (1998) Liver stem cells: a two compartmentsystem. Curr. Opin. Cell Biol. 10, 710715

74 Petersen, B., Bowen, W., Patrene, K. et al. (1999) Bonemarrow as a potential source of hepatic oval cells.Science 284, 11681170

75 Theise, N., Badve, S., Saxena, R. et al. (2000) Derivationof hepatocytes from bone marrow cells in mice afterradiation-induced myeloablation. Hepatology 31,234240

76 Alison, M. R., Poulsom, R., Jeffery, R. et al. (2000)Hepatocytes from non-hepatic adult stem cells. Nature

(London) 406, 25777 Theise, N., Nimmakalayu, M., Gardner, R. et al. (2000)

Liver from bone marrow in humans. Hepatology 32,1116

78 Kleeberger, W., Rothamel, T., Glockner, S., Flemming,P., Lehmann, U. and Kreipe, H. (2002) High frequencyof epithelial chimerism in liver transplants demonstratedby microdissection and STR-analysis. Hepatology 35,110116

79 Petrenko, O., Beavis, A., Klaine, M., Kittappa, R.,Godin, I. and Lemischka, I. R. (1999) The molecularcharacterization of the fetal stem cell marker AA4.Immunity 10, 691700

80 Bonner-Weir, S. (2001) Beta-cell turnover: itsassessment and implications. Diabetes 50 (Suppl. 1),S20S24

81 Bernard-Kargar, C. and Ktorza, A. (2001) Endocrine

pancreas plasticity under physiological and pathologicalconditions. Diabetes 50 (Suppl. 1), S30S35

82 Bonner-Weir, S., Taneja, M., Weir, G. C. et al. (2000) Invitro cultivation of human islets from expanded ductaltissue. Proc. Natl. Acad. Sci. U.S.A. 97, 79998004

83 Ramiya, V. K., Maraist, M., Arfors, K. E., Schatz, D. A.,Peck, A. B. and Cornelius, J. G. (2000) Reversal ofinsulin-dependent diabetes using islets generated in vitrofrom pancreatic stem cells. Nat. Med. (N.Y.) 6, 278282

84 Zulewski, H., Abraham, E. J., Gerlach, M. J. et al. (2001)Multipotential nestin-positive stem cells isolated fromadult pancreatic islets differentiate ex vivo intopancreatic endocrine, exocrine, and hepatic phenotypes.Diabetes 50, 521533

85 Deutsch, G., Jung, J., Zheng, M., Lora, J. and Zaret,K. S. (2001) A bipotential precursor population forpancreas and liver within the embryonic endoderm.

Development 128, 87188186 Krakowski, M. L., Kritzik, M. R., Jones, E. M. et al.

(1999) Pancreatic expression of keratinocyte growthfactor leads to differentiation of islet hepatocytes andproliferation of duct cells. Am. J. Pathol. 154, 683691

87 Shen, C. N., Slack, J. M. and Tosh, D. (2000) Molecularbasis of transdifferentiation of pancreas to liver. Nat.Cell Biol. 2, 879887

88 Wang, X., Al-Dhalimy, M., Lagasse, E., Finegold, M.and Grompe, M. (2001) Liver repopulation andcorrection of metabolic liver disease by transplantedadult mouse pancreatic cells. Am. J. Pathol. 158, 571579

89 Rao, M. S. and Reddy, J. K. (1995) Hepatictransdifferentiation in the pancreas. Semin. Cell Biol. 6,151156


8/14/2019 1030355

15/15


90 Gmyr, V., Kerr-Conte, J., Belaich, S. et al. (2000) Adulthuman cytokeratin 19-positive cells reexpress insulinpromoter factor 1 in vitro: further evidence forpluripotent pancreatic stem cells in humans. Diabetes 49,16711680

91 Grimm, P. C., Nickerson, P., Jeffery, J. et al. (2001)Neointimal and tubulointerstitial infiltration byrecipient mesenchymal cells in chronic renal-allograftrejection. N. Engl. J. Med. 345, 9397

92 Strutz, F. and Mu$ ller, G. (2000) Transdifferentiationcomes of age. Nephrol. Dial. Transplant. 15, 17291731

93 Ng, Y., Huang, T., Yang, W. et al. (1998) Tubularepithelial-myofibroblast transdifferentiation inprogressive tubulointerstitial fibrosis in 5\6nephrectomized rats. Kidney Int. 54, 864876

94 Sun, D., Fujigaki, Y., Fujimoto, T., Yonemura, T. andHishida, A. (2000) Possible involvement ofmyofibroblasts in cellular recovery of uranyl acetate-induced acute renal failure in rats. Am. J. Pathol. 157,13211335

95 Cornacchia, F., Fornoni, A., Plati, A. R. et al. (2001)Glomerulosclerosis is transmitted by bone marrow-derived mesangial cell progenitors. J. Clin. Invest. 108,16491656

96 Doetsch, F., Caille! , I., Lim, D., Garcia-Verdugo, J. andAlvarez-Buylla, A. (1999) Subventricular zone astrocytes

are neural stem cells in the adult mammalian brain. Cell97, 703716

97 McKay, R. (1997) Stem cells in the central nervoussystem. Proc. Natl. Acad. Sci. U.S.A. 276, 6671

98 Momma, S., Johansson, C. B. and Frisen, J. (2000) Getto know your stem cells. Curr. Opin. Neurobiol. 10,4549

99 Johansson, C. B., Momma, S., Clarke, D. L. et al. (1999)Identification of a neural stem cell in the adultmammalian central nervous system. Cell 96, 2534

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