Actas Dermosifiliogr.

2015;106(9):725---732

REVIEW

Skin Stem Cells: At the Frontier Between the

Laboratory and Clinical Practice. Part 1: Epidermal
Stem Cells夽
I. Pastushenko,a,∗ L. Prieto-Torres,b Y. Gilaberte,c,d C. Blanpaina,e

a
Institut de Recherche Interdisciplinaire en Biologie Humaine et Moléculaire (IRIBHM), Université Libre de Bruxelles (ULB),
Bruselas, Bélgica
b
Servicio de Dermatología, Hospital Clínico Lozano Blesa, Zaragoza, Spain
c
Servicio de Dermatología, Hospital San Jorge, Huesca, Spain
d
Instituto Aragonés de Ciencias de la Salud, Zaragoza, Spain
e
Walloon Excellence in Life Sciences and Biotechnology (WELBIO), Université Libre de Bruxelles (ULB), Bruselas, Bélgica

Received 1 May 2015; accepted 27 May 2015

KEYWORDS Abstract Stem cells are characterized by their ability to self-renew and differentiate into the
Adult stem cells; different cell lineages of their tissue of origin. The discovery of stem cells in adult tissues,
Epidermal stem cells; together with the description of specific markers for their isolation, has opened up new lines
Review; of investigation, expanding the horizons of biomedical research and raising new hope in the
Dermatology; treatment of many diseases.
Therapeutic In this article, we review in detail the main characteristics of the stem cells that produce the
applications specialized cells of the skin (epidermal, mesenchymal, and melanocyte stem cells) and their
potential implications and applications in diseases affecting the skin.
Part I deals with the principal characteristics and potential applications of epidermal stem
cells in dermatology.
© 2015 Elsevier España, S.L.U. and AEDV. All rights reserved.

PALABRAS CLAVE Células madre de la piel: en la frontera entre el laboratorio y la clínica. Parte I:
Células madre del células madre epidérmicas
adulto;
Resumen Las células madre son células que se caracterizan por su capacidad para autorren-
Células madre
ovarse y diferenciarse hacia células de todos los linajes que constituyen su tejido de origen. El
epidérmicas;
descubrimiento de las células madre en un organismo adulto, y la descripción de los marcadores

夽 Please cite this article as: Pastushenko I, Prieto-Torres L, Gilaberte Y, Blanpain C. Células madre de la piel: en la frontera entre el

laboratorio y la clínica. Parte I: células madre epidérmicas. Actas Dermosifiliogr. 2015;106:725---732.

∗ Corresponding author.

E-mail address: [email protected] (I. Pastushenko).

1578-2190/© 2015 Elsevier España, S.L.U. and AEDV. All rights reserved.
726 I. Pastushenko et al.

que han permitido aislar de forma específica estas células, han abierto nuevas perspectivas y
Revisión; nuevos horizontes en la investigación biomédica y también nuevas esperanzas en el tratamiento
Dermatología; de muchas enfermedades. En este artículo se revisan de forma detallada las principales car-
Aplicaciones acterísticas de las células madre que dan origen a las distintas células de la piel humana,
terapéuticas incluyendo las células madre epidérmicas, mesenquimales y melanocíticas, y sus potenciales
implicaciones y aplicaciones en las enfermedades cutáneas. La primera parte de este artículo
revisa las células madre epidérmicas, con sus principales características y sus potenciales
aplicaciones en dermatología.
© 2015 Elsevier España, S.L.U. y AEDV. Todos los derechos reservados.

Introduction on 7 differentiation programs which finally give rise to the

mature hair follicle. When the matrix cells have exhausted
Stem cells are defined by 2 fundamental characteristics: their proliferative capacity, hair growth stops and the folli-
their capacity for self-renewal and for differentiation into cle enters catagen,6 leading to degeneration of the lower
all cell lines within their tissue of origin.1 two-thirds of the follicle while the bulge region remains
In adults, stem cells have been identified in different intact.
organs, including the skin, intestine, muscle, hematopoietic The hair bulb is found in the lower part of the hair follicle.
system, and even the human brain.2 These cells are respon- This structure, which rests on the dermal papilla, is made
sible for maintaining tissue homeostasis where they reside up of differentiated progeny of stem cells from the bulge.
and also for repairing damage when it occurs. The dermal papilla contains specialized dermal fibroblasts,
The discovery of stem cells in adult organisms and char- nerve fibers, and a capillary loop. It plays a fundamental
acterization of their markers to enable isolation of specific role in the development of the hair follicle and control of
cells have opened up new perspectives and new horizons the hair cycle in adults.7 Cells in the dermal papilla can
in biomedical research, with new hopes for treatment in differentiate into neuronal and mesodermal cell lines.8,9 In
an range of diseases. Epidermal stem cells are of particular a recent study, Rahmani et al.10 eliminated stem cells from
interest as they are relatively numerous and also accessi- the dermal papilla and observed a delay in the regeneration
ble, making them easy to obtain. In the first part of this of the hair follicle, along with change in hair type, suggesting
review, we have aimed to summarize the main findings of that the dermal papilla plays a fundamental role in restoring
basic research in the field of epidermal stem cells. We then hair growth after damage, disease, or aging.
discuss their potential applications in clinical dermatology. At least 3 types of epithelial stem cells have been iden-
tified recently: these types reside in the sebaceous glands,
the infundibulum, and sweat glands. The sebaceous glands
Different Stem Cell Populations in the are maintained by unipotent Lgr6+ stem cells, which origi-
Epidermis nate from Blimp1+ progenitor cells.11 In addition, stem cells
from the isthmus express the MTS234 marker,12 and if trans-
Two types of progenitor cell are present in the basal layer planted to an immunodeficient mouse, are surprisingly able
of the interfollicular epidermis: ␣6+ CD34--- stem cells, char- to give rise to epidermal, follicular, and sebaceous cell lines,
acterized by slow division rates (4-6 times a year) and a suggesting that these might be multipotent cells.13 The stem
long lifespan, and basal layer K14+ Inv+ progenitor cells,3 as cells in the infundibulum are characterized by expression
well as possibly Axin2+ cells,4 characterized by faster divi- of the Lrig1 marker and their multipotent capacity.14 It is
sion rates (once a week) and shorter lifespan. After a certain also thought that they may contribute to the homeostasis
number of divisions these cells undergo terminal differenti- of the sebaceous glands.15 Finally, although sweat glands
ation into differentiated keratinocytes, thereby losing their have traditionally be considered as quiescent in adults,
capacity for division. Table 1 summarizes the main charac- a study published recently suggests 4 different types of
teristics of the markers. progenitor cells are present in the epithelium of these
The hair follicle has 3 phases: anagen or growth phase structures.16 Figure 1 shows a microphotograph of the hair
(lasting on average 3 years), catagen or involuting phase follicle (Fig. 1A) and a schematic of the different compart-
(which lasts several weeks), and telogen or resting phase ments in the epithelium of the skin and where the stem cells
(which lasts several months). The stem cells responsible for reside, as well as a summary of their markers (Fig. 1B).
hair follicle regeneration during anagen reside in the bulge
(the lower part of the permanent portion of the hair fol-
licle) and are characterized by expression of the markers Self-Renewal of Stem Cells
CD34, Lgr5, and K15. The cells are multipotent,5 as they
can differentiate into all cell lines present in the hair folli- As mentioned earlier, stem cells can give rise to differenti-
cle unit. During anagen, the stem cells in the bulge give rise ated cells, but they also propagate to maintain a constant
to transit-amplifying cells, which reside in the hair follicle. pool of stem cells, and can divide symmetrically or asym-
These rapidly and transiently proliferate before embarking metrically. During the process of asymmetric division, a stem
Skin Stem Cells: At the Frontier Between the Laboratory and Clinical Practice. Part 1: Epidermal Stem Cells 727

Table 1 Summary of the Main Characteristics of Markers for Epidermal Stem Cells of Different Compartments.

Marker Full Name Cells that Express the Expression Pattern

Marker
␣6 Integrin ␣6 SC of interfollicular Membrane
epidermis
Axin2 Axin-like protein Progenitors of the Cytoplasmic
interfollicular epidermis?
CD34 CD34 Bulge SC Membrane
Inv Involucrin Progenitors of the Cytoplasmic
interfollicular epidermis
K5 Keratin 5 SC of interfollicular Cytoplasmic
epidermis
K14 Keratin 14 SC and progenitors of the Cytoplasmic
interfollicular epidermis
K15 Keratin 15 Bulge SC Cytoplasmic
Lgr5 Leucine-rich Bulge SC Cytoplasmic
repeat-containing
G-protein-coupled
receptor 5
Lgr6 Leucine-rich Isthmus SC Cytoplasmic/membrane
repeat-containing
G-protein-coupled
receptor 6
Lrig5 Leucine-rich Infundibulum SC Cytoplasmic
repeats and
immunoglobulin-
like domain
protein 1
MTS24 MTS24 Isthmus SC Membrane

Abbreviation: SC, cell.

A B

Hair axis Interfollicular epidermis

Bulge Infundibular SC: Lrig1

IFE progenitors:
IFE SC: α6, K5, K14
Isthmus SC: MTS24, Lgr6 K14

Bulge SC: CD34, Lgr5, K15

Sebaceous gland
Outer root
sheath
Progenitors of the
adult sweat gland

Hair shaft
Inner root
sheath

Matrix
transit-amplifying cells
Dermal papilla

Dermal papilla

Hair follicle Sweat gland

Figure 1 Structure of the hair follicle and different types of epidermal stem cells.
The figure shows (A) microphotography of the hair follicle (hematoxylin and eosin, × 20) and (B) illustration of the different types
of stem cells and progenitor cells in the epidermis, as well as their specific markers. IFE indicates interfollicular dermis, SC, stem
cells.
728 I. Pastushenko et al.

Asymmetric cell division Symmetric cell division

Stem cell Differentiated cell Stem cell

Stem cell Stem cell Stem cell

Differentiated cell Differentiated cell Stem cell

Stratum corneum

Stratum spinosum
B C Basal layer

Figure 2 Self-renewal of stem cells.

A) Concept of symmetric and asymmetric cell division.
During the development of the embryo (B), most of the divisions are symmetric and the axis of division is parallel to the basal
membrane, thereby allowing extension of the embryo surface during growth. During stratification of the epithelium, which occurs
during morphogenesis and in adulthood (C), most of the divisions are asymmetric. During asymmetric division, the axis can be
perpendicular to the basal membrane (on division, a daughter cell on losing contact with integrins and growth factor secreted by
the basal membrane, undergoes differentiation, and the second daughter cell, on remaining in contact with the basal membrane,
maintains the characteristics of the stem cell). Division can also be parallel to the basal membrane (in this case, differentiation of
one of the daughter cells is induced by another mechanism).

cell gives rise to a daughter cell with the same phenotype, During repair of damage to the interfollicular epider-
and another daughter cell which will differentiate. Symmet- mis, massive recruitment of interfollicular stem cells to the
ric division of a stem cell gives rise to 2 identical daughter area of the wound occurs.3 Clones derived from these cells
cells, both with a differentiated or somewhat differentiated are observed migrating from the periphery of the wound
phenotype (Fig. 2 A). towards the center, and remaining there for a long time
During the development of the embryo, most basal cell after healing. However, much fewer Inv+ cells (short-lived
divisions are symmetric and parallel to the axis of the progenitors) migrate towards the damaged zone, the clones
basal membrane (Fig. 2B), thereby allowing growth of the are much smaller, and 35 days after injury, most of these
embryo surface and ensuring that the epithelium remains as have disappeared.3 That is, it seems that the short-lived pro-
a single layer. In contrast, during stratification of the epithe- genitors are responsible for maintaining homeostasis of the
lium, approximately 70% of divisions are asymmetric (Fig. epidermis in normal conditions, whereas stem cells, which
2 C),6,17 thereby allowing development of suprabasal cells are normally in a quiescent state, are activated when an
and establishment of the skin barrier during development injury occurs (wounds or drug administration).19
and epidermal homeostasis in adulthood. In addition, stem cells both in the bulge and in the
infundibulum can migrate towards the epidermis in response
to a wound stimulus, and participate in repair of the dam-
Role of Epidermal Stem Cells in Skin age. Surprisingly, and through mechanisms that are still not
well understood, when these cells migrate towards the epi-
Homeostasis and Scarring
dermis, they lose their specific hair follicle markers and
adopt a phenotype more similar to those of the stem cells
Homeostasis is a physiological process whereby the num- of the interfollicular epidermis. However, once in the epi-
ber of cells with capacity for regeneration in a given organ dermis, these cells are short-lived and disappear soon after
remains constant. In skin homeostasis, the stem cells in each repair of the damaged tissue.20,21
compartment are those responsible for replacing the dif- Another phenomenon of great interest, which was
ferentiated cells that die in that compartment. However, observed on elimination of stem cells of a specific compo-
during the evolutionary process, stem cells have acquired nent of the epidermis with laser light, was that the empty
the capacity to participate in the repair of neighboring niches were able to recruit normal differentiated cells from
compartments in the event that stem cells in those com- that compartment and induce cell proliferation and undif-
partments become damaged. ferentiation towards a state similar to that of stem cells.22,23
The form in which stem cells respond to damage varies
drastically, depending not only on the compartment where
these reside, but also how close they are to the wound.18 Epidermal Stem Cells as the Origin of
The technique of in vivo lineage tracing offers valuable Nonmelanoma Skin Cancer Cells
functional information on the behavior of stem cells in
homeostasis and during tissue repair, given that cell develop- The identification of the cells that give rise to cancer is still
ment can be followed over time in the natural environment a challenge in most malignant tumors. Theoretically, these
(Fig. 3). cells acquire the first genetic or epigenetic abnormalities
Skin Stem Cells: At the Frontier Between the Laboratory and Clinical Practice. Part 1: Epidermal Stem Cells 729

A B C

Labeling of the target cell We can determine which Stem cell Compromised
with a reporter gene cells descended from progenitor
the initial stem cell

Figure 3 Concept of lineage tracing.

The figure shows a schematic of the information obtained from lineage tracing experiments. The technique consists of introducing a
reporter gene associated with the marker of the cells of interest (A), thereby obtaining a fluorescent signal in the subgroup of cells
of interest. All daughter cells of the labelled cells also have a fluorescent signal (B), but the intensity of the signal will decrease
as the cells continue to divide. Thus, on the one hand, we can identify cells descended from the initially labelled cells, and on the
other, assess the rate of division (the cells that maintain an intense signal over time will be the quiescent cells that divide very
slowly----the stem cells----whereas the cells that lose color intensity will be the compromised progenitor cells [C]).

that culminate in the initiation of the malignant process.24 keratinocyte proliferation.32 The keratinocytes are cultured
It follows that, given their characteristics (capacity for self- until formation of a stratified epithelium that can be used to
renewal over a long period of time), stem cells are at an cover the wound. However, there are 2 main drawbacks with
increased risk of accumulating oncogenic mutations, and so this technique: the first is the long time needed for in vitro
they may be the cells that initiate the cancer.25,26 expansion of keratinocytes and the second is the high cost
Using mice modified genetically with the inducible of the procedure.33
CreER-LoxP system (CreER refers to the estrogen recep- Of note is that the skin obtained with this method does
tor [ER] Cre-recombinase enzyme, such that administration not have any appendages (hair follicles or sweat glands). In a
of tamoxifen induces recombination of Cre, with the sub- third-degree burn, the disappearance of hair follicles is due
sequent activation or deletion of the target gene), it has not only to destruction of multipotent stem cells in the fol-
been shown that activation of the SmoM2 mutation in stem licle but also to the destruction of the dermis (the papillary
cells of the bulge or in transit-amplifying cells of the dermis). To achieve regeneration of the hair follicles, cells
hair follicle did not induce tumor formation in basal cell from the papillary dermis need to be transplanted along with
carcinoma.27,28 In fact, the authors demonstrated that 90% keratinocytes.34 In a recently published study, researchers
of surface-spreading basal cell carcinomas originate from managed to transplant human cells from the papillary der-
the interfollicular epidermis and the remaining 10% originate mis to a nude mouse (immunodeficient and without fur) and
in the infundibulum. reported the regeneration of hair follicles.35 To achieve a
In the case of spindle-cell carcinoma in mice, expres- similar hair color to the original color of the patient, a group
sion of the KRas mutation in stem cells in the bulge of Swiss investigatorss added melanocytes isolated from skin
and in the interfollicular epidermis, but not in transit- biopsy to a culture of keratinocytes, achieving favorable
amplifying cells, induces the formation of benign tumors results, both in patients with clear phototypes and with dark
but combination of the KRas mutation and deletion of the phototypes.36
tumor suppressor gene p53 is necessary for progression to A culture of keratinocytes on a bed of human fibroblasts
carcinoma.29 embedded with a plasma matrix has been used as an alter-
native to transplantation of layers of keratinocytes cultured
on a bed of irradiated fibroblasts.37 This approach enables
Potential Applications of Stem Cells in Clinical restoration of both the epidermal and dermal compartment.
Dermatology After 24-26 days of culture, the authors achieved a 1000-
fold cultured-area expansion, and successfully transplanted
Burns artificial skin from 2 patients with severe burns. Researchers
from the University of Granada in Spain used fibrin-agarose
For the treatment of extensive skin burns, successful out- biomaterials as a base to generate skin substitutes from
comes have been obtained with in vitro epidermis generated human fibroblasts and keratinocytes. The artificial skin
using autologous epidermal stem cells from a biopsy of obtained was transplanted to nude mice and samples were
undamaged patient’s skin.30 The skin biopsy sample is dis- taken for histological analysis and electron microscopy after
sociated with tripsin31 and the epidermal stem cells are 10, 20, 30, and 40 days.38 The results of these analyses
isolated. These are then expanded on a base of irradiated showed that the structure of the skin obtained through tis-
fibroblasts, which secrete extracellular matrix and growth sue engineering was very similar to that of normal mouse
factors, making the environment particularly conducive to skin.
730 I. Pastushenko et al.

Two studies of murine models published recently pointed

to a possible alternative to the method used traditionally. Biopsy of a patient with
epidermolysis bullosa
Stimulation of the stem cells from the bulge of the hair
follicle in third-degree burns in mice induced with human
␣-defensin-5 derived from the intestine accelerated wound
healing and, notably, induced hair regeneration.39 Similarly, Culture of stem cells with the mutation

transplantation of Lgr6+ stem cells isolated by fluorescence-

activated cell sorting and administered by injections into
the wound promoted reepithelization, hair growth, and
angiogenesis.40 Gene transfer in patient cells with a viral
vector that expresses collagen 7

Therapeutic Correction of the Epidermal Stem Cell

Genome in Genetic Diseases Selection of collagen–7-producing cells
(correct transfection) and in vitro expansion

In recent years, the cost of genetic sequencing has

decreased significantly. As a result, the volume of data on
the human genome in different contexts and diseases has
Assessment of genome integrity and
increased exponentially. These advances in knowledge have oncogenic potential of cells obtained
generated high expectations around the potential therapeu- by transfection (in mouse model)

tic applications in genetic diseases. Although gene therapy

is not applied in everyday clinical practice by dermatolo-
gists, in recent years, large steps forward have been made
and the field shows great promise. In vitro production of epidermal layers and
transplantation of genetically corrected
To date, the 2 most powerful therapeutic genetic tech- epidermis to the patient with
epidermolysis bullosa
niques are gene therapy, which enables the reestablishment
of the lost function of a given gene through expression
of a transgene that is incorporated into the genome by
viral vectors,41 and interference RNA, which can suppress
expression of the defective gene through inhibition of mes-
Figure 4 Example of application of gene therapy in a skin
senger RNA.42 In recent years, moreover, new technologies
disease.
have been developed with a promising future potential in
The figure shows a schematic of a the study conducted by Droz-
gene therapy. These are based on the use of programmable
Georget Lathion et al. in epidermolysis bullosa, as an example
nucleases, which are able to edit the genome of cells in
of the efficacy and safety of gene therapy in a skin disease.
damaged tissues, resulting in the elimination or correc-
tion of harmful mutations or the insertion of protective
mutations.43---46
From a theoretical point of view, there are 2 possibil- Conclusions and Perspectives
ities for the treatment of genetic skin diseases (Fig. 4).
Ex vivo therapy consists of extracting epidermal stem cells The characteristics of stem cells, whereby they are able
with a genetic abnormality (such as for example, laminin to self-renew and give rise to different cells types, along
5 deficit47 or collagen 7 deficit48 in the case of patients with the startling developments in bioengineering, make for
with epidermolysis bullosa) by skin biopsy, correcting the a promising future. Epidermal stem cells are particularly
mutation in vitro by gene transfer, selecting cells in which attractive given their relatively high number proportional
the transfer has been successfully accomplished and the to the body surface and their accessibility. Although these
defect has been corrected, and transplanting keratinocytes are complex techniques with a high cost, it is likely that in
once again in the skin of the patient. In a recently pub- the coming years, knowledge of the biology of stem cells, as
lished study, before transplanting the corrected epidermal well as the safety of the techniques used, will increase, and
stem cells back into patients with epidermolysis bullosa, so allow a more widespread application in medicine and in
the authors assessed the integrity of the genome and also dermatology in particular.
the oncogenic potential of these cells.49,50 They demon-
strated that the procedure is safe (given that this is a
transfection using viral vectors, one of the main concerns Funding
and drawbacks is the possibility of mutagenesis). In vivo
therapy consists of delivery of viral or programmable nucle-
The work of I. Pastushenko was funded by the Télévie
ase vectors directly to the affected cells in their natural
grant.
environment through intravenous drug administration or
injection into the organ itself. This approach has sev-
eral technical difficulties and safety concerns, which we
will not discuss in this review but which have impeded Conflicts of Interest
the use of these techniques in patients with genetic skin
diseases. The authors declare that they have no conflicts of interest.
Skin Stem Cells: At the Frontier Between the Laboratory and Clinical Practice. Part 1: Epidermal Stem Cells 731

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