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Review Article
Regenerative medicine in dermatology: biomaterials,
tissue engineering, stem cells, gene transfer and beyond
Christina Dieckmann1*, Regina Renner2*, Linda Milkova1,2 and Jan C. Simon1,2
1
Translational Centre for Regenerative Medicine, Leipzig University, Leipzig, Germany;
Department of Dermatology, Venerology and Allergology, Leipzig University Medical Center, Leipzig, Germany
Correspondence: Regina Renner, MD, Department of Dermatology, Venerology and Allergology, Leipzig University Medical Center,
Philipp-Rosenthal-Straße 23, D-04103 Leipzig, Germany, Tel.: +49-341-9718600, Fax: +49-341-9718609,
e-mail: [email protected]
*These authors contributed equally to this work.
2
Accepted for publication 9 March 2010
Abstract: The term ‘regenerative medicine’ refers to a new and
expanding field in biomedical research that focuses on the
development of innovative therapies allowing the body to replace,
restore and regenerate damaged or diseased cells, tissues and
organs. It combines several technological approaches including the
use of soluble molecules, biomaterials, tissue engineering, gene
therapy, stem cell transplantation and the reprogramming of cell
and tissue types. Because of its easy accessibility, skin is becoming
an attractive model organ for regenerative medicine. Here, we
review recent developments in regenerative medicine and their
potential relevance for dermatology with a particular emphasis on
biomaterials, tissue engineering, skin substitutes and stem cellbased therapies for skin reconstitution in patients suffering from
chronic wounds and extensive burns.
Key words: biomaterials – gene transfer – regenerative medicine –
skin substitutes – stem cells – tissue engineering
Please cite this paper as: Regenerative medicine in dermatology: biomaterials, tissue engineering, stem cells, gene transfer and beyond. Experimental Dermatology 2010; 19: 697–706.
Introduction
Regenerative medicine is an emerging interdisciplinary field
of research and clinical application focussing on the repair,
replacement or regeneration of cells, tissue or organs to
restore impaired function because of congenital defects,
disease, trauma and ageing. It combines several technological approaches including, the use of soluble molecules, gene
therapy, stem cell transplantation, tissue engineering and
the reprogramming of cell and tissue types (Fig. 1) (1).
The earliest successful example of regenerative medicine
can be traced back to the late 1950s, when the idea of
reversing heart failure by transplanting a heart form one
individual to another had been realized, in the beginning
in animal models. In 1967, the first heart transplantation in
humans was carried out (2). Because of ethical concerns
and the lack of donor organs as well as the risk of graft
failure, scientists and clinicians now are pursuing a different strategy in regenerative medicine. Instead of replacing
whole organs, they intend to transplant biologically competent cells and engineered tissues or to stimulate tissue-resident stem cells to restore tissue or organ function. Both
adult stem cells and embryonic stem (ES) cells as well as
reprogrammed somatic cells with a multipotent, ES cell-like
ª 2010 John Wiley & Sons A/S, Experimental Dermatology, 19, 697–706
character [also referred to induced pluripotent stem (iPS)
cells], featuring a versatile growth and differentiation
potential become increasingly important for fighting severe
and yet incurable diseases. Stem cell therapies have already
been demonstrated (in clinical trials or the laboratory) to
heal ischaemic heart diseases (3–5), auto immune diseases,
like multiple sclerosis (6) and neurological diseases, like
Parkinson’s disease and stroke (7,8). Table S1 shows a representation of completed or pending human clinical trials
of stem cell therapies, indexed by clinical application and
source of stem cells used.
Skin is an attractive model organ to test novel concepts
of regenerative medicine, with a particular emphasis on
skin regeneration for acute or chronic wounds. Chronic
wounds present a worldwide growing health and economic
problem because of a steadily increasing number of
patients, high morbidity and risk of amputations, unsatisfactory results of existing therapies and heavy socioeconomic burden (9). Tissue-engineered skin substitutes
represent an innovative therapeutical option for the treatment of acute and chronic skin wounds. Bioengineered
skin replacements are not only supposed to substitute the
major physiological functions by providing a rapid and
reliable cover of the defect but also should be easily appli-
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Soluble
molecules
VEGF EGF
IGF
FGF VEGF
IGF
G
EGF
FGF
Stem cell
therapy
Biomaterials
Regenerative
medicine
Gene
therapy
Figure 1. Research areas in regenerative medicine. Regenerative
medicine is an emerging multidisciplinary field of research and clinical
applications focused on the repair, replacement or regeneration of
tissue or organs. The approaches may include the use of soluble
molecules, gene therapy, stem cell therapy and biomaterials.
cable under routine use conditions and reduce pain and
discomfort for the patient. Furthermore, they should elicit
the regeneration process from the wound bed without
causing inflammation or rejection. Skin substitutes should
be available immediately, and be non-toxic nor immunogenic. From an aesthetic point of view, skin substitutes
should be durable, elastic, with minimized scar formation,
and pigmentation should resemble natural skin. Finally, yet
importantly, the cost–benefit ratio has to be taken into
consideration (10,11).
In this review, we are surveying the existing bioengineered skin substitutes that are already in clinical applications, without claiming to be complete. Additionally, we
give an outlook on recent progress in the development
of innovative approaches in skin reconstitution, by stem
cell-based therapies.
Biomaterials as skin substitutes
Here, we will discuss the key requirements of biomaterials
for the generation of synthetically engineered skin substitutes. Biomaterials are acellular natural or synthetic substances used for creation of the backbone of skin
substitutes in clinical applications.
Skin substitutes composed of biomaterials can be used as
temporary wound cover for all thickness wounds, like
chronic ulcers or superficial and second-degree wounds
prior to autologous skin grafting. They can be used as stimulating agents for cell proliferation and angiogenesis or as
permanent dermal replacement, according to the intended
use and the individual constitution of the wound and the
patients general condition In a systematic review of randomized controlled trials, most skin equivalents are in
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favour compared to other wound dressings [reviewed by
O¢Donnell et al. and by Dini et al. (12,13)], in particular
products like Oasis (Healthpoint Ltd., Fortworth, Texas,
USA), Dermagraft (Advanced BioHealing, Westport, Connecticut, USA) and Apligraf (Organogenesis Inc., Canton,
Massachusetts, USA). Tissue engineering usually requires an
artificial extracellular matrix (ECM) allowing for infiltration
of surrounding cells. For the generation of an artificial
ECM, both naturally occurring and synthetically manufactured substances are processed. Examples of natural materials include polypeptides, hydroxyapatites, hyaluronan,
glycosaminoglycans (GAG), fibronectin, collagen, chitosan
and alginates. Because of their abundance in skin and their
recognition by cell surface receptors, these natural materials
display low toxicity and low chronic inflammatory response.
However, concerning their predominant xenogeneic origin,
they harbour an intrinsic risk for transmission of animal
viruses (9,14).
Examples of synthetic, fully degradable materials include
polyglycolide, polylactide, polylactide-coglycolide, polytetrafluoroethylene, polycaprolactone and polyethylene terephthalate, while polyurethane (PUR) represents a frequently
used non-degradable substance. New technologies like threedimensional printing and electrospinning emerged for accurate manufacture, creating scaffolds of both a defined shape
and pore sizes, facilitating infiltration of fibroblasts and
vascularization (10,14–19). A major drawback of synthetic
materials is the lack of cell-recognition signals. An approach
enabling the repopulation and regeneration of a new natural
matrix consisted in the incorporation of adhesion peptides
e.g. using RGD sequences (Arg-Gly-Asp) into biomatrices
(20,21). A so-called smart matrix is provided by biomaterials
or scaffolds capable of directing cell differentiation and
metabolism and therefore accelerating tissue regeneration
(11,22). For example, this can be accomplished via integration of polyethylene glycol (PEG) hydrogels into the scaffold.
PEG acts as an inert structural platform because of its hydrophilicity and resistance to protein adsorption (23). The gel
can be modified by addition of cell anchoring points like
RGD-containing peptides or networking functional domains
linked to a hyaluronan backbone. Furthermore, the degree of
degradability by proteases can be influenced by linkage of
protease-sensitive oligopeptides (24–27).
As the crucial role of growth factors for cell migration,
proliferation, differentiation and differentiation in the
wounded area has been recognized, efforts in tissue engineering have focused on the incorporation of growth factors like fibroblast growth factor, vascular endothelial
growth factor (VEGF), insulin-like growth factor (IGF) and
platelet-derived growth factor (PDGF) into matrix scaffolds
(28–30). Thus, the bioactivity of a polymer backbone can
be augmented and the healing process markedly accelerated
especially in chronic wounds (11,28). The challenge consists
ª 2010 John Wiley & Sons A/S, Experimental Dermatology, 19, 697–706
Skin substitution by tissue engineering
in the identification of the required factors and cytokines
according to the current condition of the wound. In the
following, acellular epidermal and dermal skin substitutes
comprising biomaterials are discussed.
A currently used double-layered biosynthetic epidermal
substitute consists of an outer silicone membrane and a knitted nylon mesh (Biobrane, Smith & Nephew Healthcare,
London, UK). Both layers enclose a chemically cross-linked
porcine type 1 collagen forming a 3D structure that allows
rapid adherence to the wound surface and initiation of the
wound healing. Indications for Biobrane include superficial and second-degree burns and large epithelial defects
(14). A new product synthetically polymerized of the three
components DL-lactidetrimethytencarbonate, trimethylencarbonate and e-caprolactone with a porous membrane is
called Suprathel. Hydrolytic degradation of this epidermal
substitute occurs about 4 weeks after application. Principal
domains of Suprathel (PolyMedics Innovations GmbH,
Denkendorf, Germany) are second-degree burns and donor
sites of skin transplantation. Promising advantages are a
rapid painless re-epithelialization and the possibility of
application next to utilized regions like joints (31).
Being engineered already since 1981, Integra (Intergralife Sciences, Plainsboro, New Jersey, USA) represents the
oldest available dermal equivalent. A double-layered synthetic skin substitute is composed of a three-dimensional
porous matrix of bovine collagen with about 10–15%
chondroitin-6-sulphate derived from shark cartilage and an
outer silicone sheet. After the infiltration of fibroblasts and
initiation of the vascularization process within 3 weeks, the
application procedure requires removement of the silicone
layer and wound covering with a sheet autograft (14,17,32–
34). Indications for Integra are treatment of full-thickness
burns and correction of scars in joint proximity. However,
healing of keratinocyte autografts on Integra is limited
(about 60%) possibly because of an increased frequency of
infections, and this material cannot be applied in persons
bearing a sensitization against bovine products (14,35).
Matriderm (Skin & Health Care AG, Suwelack, Billerbeck, Germany) is a porous and thin matrix composed of
bovine type I, II and V collagen covered with bovine elastin
hydrolysate. Within 2 weeks after application, the matrix is
degraded and replaced by the recipients own collagen.
Depending on the thickness of the sheet used for application, a one-step procedure is possible, however, sheets of
2 mm thickness and above should be used only in sufficiently vascularized wound beds during an interval of 7 days
before transplantation of split thickness autografts (36–38).
Convincing results in the treatment of patients with
chronic venous leg ulcers or superficial and second-degree
burns, respectively, have been received using porcine small
intestinal submucosa acellular collagen matrix (Oasis) or
porcine skin (Permacol, Covidien, Norwalk, Connecticut,
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USA). Despite an overall rather negligible exposure, porcine
matrices implicate the risk of transmission of prion disease
and porcine retroviruses, a concern that needs to be
addressed wherever xenogeneic material is used for skin
substitutes (9). An approach to resolve this problem consists in the use of alloplastic material. The common scaffold
of the commercially available alloplastic skin substitutes is a
foil made of PUR that is coated with various substances.
Epigard (Orthomed Medizintechnik GmbH, Wien,
Austria) possesses a teflon layer upside that renders the
membrane permeable to air, but not to bacteria or wound
secretion. If changes of dressing are performed regularly,
removal of necrotic tissue and wound exudate as well as ingrowth of fibroblasts and vascular endothelium is facilitated
(39). Syspurderm (Paul Hartmann AG, Heidenheim,
Germany) is a double-layered pad of flexible foam of PUR.
The inner layer provides an open porous matrix for tissue
granulation, whereas the outer condensated surface serves
as a barrier preventing secondary infection. Cell debris,
necrotic material and bacteria can also be easily removed
during wound dressing. In contrast, Lyomousse ⁄ Lyofoam
(Cosanum AG, Swiss) is composed of a hydrophilic porous
membrane that does not adhere to the wound ground
and an outer hydrophobic surface ensuring bacterial impermeability. The mentioned alloplastic products represent a
temporary solution for debrided wound beds until biological skin substitutes can provide a permanent wound
cover (40,41).
A temporary cover can also be achieved by cadaver skin.
Allografts function often as scaffolds and allow repopulation of the recipient’s endothelial and dermal cells. But
they are subjected to host rejection (17,42). A modification
of human cadaver skin is Alloderm (LifeCell Corporation,
Branchburg, New Jersey, USA). The immunologically inert
acellular dermal matrix facilitates the regeneration of the
underlying dermis (17,36,43).
Cellular skin substitutes
There are several cellular tissue-engineered skin substitutes
currently on the market that can be characterized and
distinguished by their origin: xenogeneic (from other species), allogeneic (from a non-genetically identical individual
of the same species), autologous (from the patient itself)
and syngeneic grafts (from a genetic identical individual
like monozygotic twins). Xenogeneic skin grafts bear the
intrinsic risk of transferring prion diseases or porcine retroviruses. In particular, the use of porcine tissue in humans
can evoke an immune response against the porcine membrane glycoprotein (GAL epitope), which can cause acute
rejection of the donated skin. It is absolutely necessary
that this membrane protein is removed in the final skin
product (44). Allogeneic transplants offer the possibility of
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large prefabrication and cryo-preservation mostly with
comparable results in regard to vitality or effectivity of
fresh allografts (45,46). Furthermore, they allow for
repeated applications. However, the keratinocytes within
these allografts are being replaced within a few weeks by
the infiltrating recipient cells and sometimes fail to produce
a satisfactory result in full-thickness wounds (47,48).
The manufactured skin substitutes can further be distinguished by the different dermal components they replace:
There are (i) epidermal equivalents, (ii) dermal equivalents
and (iii) composite substitutes.
Epidermal equivalents
Epidermal grafts consist of keratinocytes that are differentiated in vitro building a stratified epidermal layer. They can
be combined with other biocompatible substrates (bovine
collagen, hyaluronic acid), acellular natural human or
porcine materials, nylon or polyglactin meshes. The first
commercialized cultured epidermal autograft (Epicel (Genzyme Corporate Offices, Cambridge, Massachusettts, USA))
is composed of autologous keratinocytes grown in vitro
in the presence of proliferation-arrested, murine (Swiss
3T3 ⁄ J2) fibroblasts (49). It is currently used as cover for fullthickness burns in the USA and Europe. Another epidermal
product is represented by EpiDex (Euroderm GmbH, Leipzig, Germany), which is manufactured from autologous
outer root sheath (ORS) cells of plucked hair follicles for the
treatment of chronic venous leg ulcers (Fig. S1) (50–53).
The proliferation potential of the ORS derived keratinocytes
is not restricted by the age of the recipient, enabling a successful treatment also of elderly patients (54). To improve
mechanical stability some epidermal skin substitutes are
combined with a polyvinyl chloride polymer coated with a
plasma-polymerized surface (Altrika Ltd., Sheffield, UK) or
with a perforated hyaluronic acid membrane (Laserskin,
Fidia farmaceutici S.p.A., Abano Terme (PD), Italy) (55).
Another administration strategy is pursued by BioSeed-S
(not available, formerly by BioTissue Technologies GmbH,
Freiburg, Germany). Autologous keratinocytes are propagated in vitro and then suspended within a gel-like fibrin
adhesive (56,57). The gel-like skin graft is applied to the
patient’s wound using a syringe. The fibrin adhesive fixes the
cells to the wound bed and allows a better in-growth (56,57).
Although the product is approved to treat chronic leg ulcers,
it is not commercially available at present. The clinical outcome of the epidermal transplants is not always satisfying
which can be partially explained by the absence of a dermal
component and of an adequate support of the undifferentiated keratinocytes (58,59).
Dermal substitutes
The development of cutaneous substitutes by tissue engineering has evolved from simple cultured autologous epider-
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mal sheets to more complex bilayered cutaneous constructs.
Mainly, dermal constructs are created with or without a temporary synthetic epidermis. The dermal cellular component
is composed either of autologous dermal fibroblasts or of
allogeneic neonatal foreskin fibroblasts, as they are more
responsive to mitogens than adult cells (60). The scaffold for
the cells consists mostly of biocompatible and biodegradable
materials, like benzyl-esterified derivatives of hyaluronic acid
(Hyaff-11) (Hyalograft 3D, Fidia farmaceutici S.p.A.,
Abano Terme (PD), Italy), polyglycolic acid or polyglactin
(Dermagraft, Advanced BioHealing, Westport, Connecticut, USA). TransCyte (formerly Dermagraft-TC, Advanced
BioHealing, Westport, Connecticut, USA) is a comparable
product containing a nylon mesh coated with porcine dermal collagen that is seeded with newborn human fibroblasts
and fixed to an outer silicone membrane (61,62). The major
indication of application consists in temporary wound cover
in surgically excised full-thickness and partial-thickness
burns. One of the most successfully bioengineered products
is Dermagraft, an allogenic human neonatal-derived dermal fibroblast culture, grown on a biodegradable scaffold,
is able to produce several growth factors, to stimulate angiogenesis, tissue expansion and re-epithelialization from the
wound edge, even after cryopreservation and thawing
(63,64). This seems to be of particular advantage in diabetic
foot ulcers as shown in various randomized controlled trials
(63). However, Dermagraft turned out to be also as safe
and as efficacious as allograft in burns (14). Furthermore, it
can be used to provide a dermal matrix that helps facilitating
re-epithelization by the patient’s own keratinocytes (17).
In clinical trials, treatment of venous leg ulcers with Dermagraft in combination with compression therapy was
superior to compression therapy alone. Total ulcer area rate
of healing and linear rate of healing was significantly
improved in patients treated with Dermagraft, they also
had a greater increase in periulcer skin perfusion but this
was not statistically significant (65).
Composite substitutes
Composite skin equivalents are defined by epidermal cells
growing on fibroblast-containing dermal substitutes. They
have been demonstrated to provide benefit in chronic ulcers
(16). The additional dermal component reduces wound
contraction, provides better mechanical stability and
reduces the time necessary for self-assembly of host-own
granulation tissue. Furthermore, production and longdistance transport are facilitated using composites. On the
other hand, collagen-based dermal substitutes promote terminal differentiation and apoptosis of fibroblasts as well as
of keratinocytes (9,66,67), and high-density fibrin carriers
have anti-migratory and survival-compromising effects (57).
PermaDerm (previously known as Cincinnati skin
substitute, Calibrex Bioscience, Walkersville, Maryland,
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Skin substitution by tissue engineering
USA) is an example of collagen-based autologous dermoepidermal skin substitutes. The dermal component is
composed of type I bovine collagen seeded with human
fibroblasts and an overlying epidermal layer of human
keratinocytes. Similar to OrCel (Forticell Bioscience,
formerly Ortec International Inc., New York, USA), it is
applied for the permanent skin replacement in severe
burn patients (33,68).
Apligraf (formerly Graftskin) is an allogeneic dermoepidermal product consisting of cultivated keratinocytes
and a dermal layer of fibroblasts on a collagen-type-Imatrix. As it does not contain any antigen-presenting cells
such as Langerhans cells, dermal dendritic cells, endothelial
cells, melanocytes or inflammatory cells, such as leucocytes,
it is thought to be immunologically inert. It has been
approved for the treatment of venous and diabetic ulcers
(69–71). Clinical trials showed that the healing rate of the
ulcers treated with Apligraf (63% vs. 49% control) as well
as the healing time (61 vs. 181 days in the control group)
was improved (72). The mechanism of action in the promotion of wound healing is not exactly clear. First of all, it
behaves as physical wound cover but secondary, it is supposed to induce the production of a number of cytokines
and growth factors (73–75) which seems to be responsible
for stimulation of differentiation and proliferation of the
surrounding fibroblasts and keratinocytes. Healing occurs
with less fibrosis which might be because of the construction of the graft itself containing neonatal cells, stimulating
a more foetal-like scarless wound healing (76). Clinical trials demonstrated that ulcers treated with OrCel were healing better in comparison with standard care (77,78). Yet,
the major drawbacks of dermo-epidermal substitutes are
the technically challenging and time-consuming two-step
production and early clinical failures because of a delayed
vascularization of the wound bed or because of their components from xenogenic origin.
Table S2 summarizes skin substitutes that are or have
been available in the past for the treatment of skin injuries
in humans.
The potential of stem cells for skin
replacement
The use of stem cells as the basic material for skin engineering offers the potential to improve significantly the clinical
outcome, both in wound healing and in gene therapeutic
approaches. Cell-based therapies with adult stem cells, ES
cells or reprogrammed somatic cells, respectively, are attractive, particularly if used in autologous transplantation regimens, as inherent problems with rejections of the transplant
do not exist, and ethical and moral objections are avoided.
Recent studies on tissue-engineered skin indicated that epidermal stem cells might provide a superior source of multi-
ª 2010 John Wiley & Sons A/S, Experimental Dermatology, 19, 697–706
potent stem cells to replace damaged tissue in patients with
compromised wound healing (50,51,79,80). Two distinct
subpopulations of epidermal stem cells occur in the skin: a
basal keratinocyte population found in the interfollicular
epithelium and stem cells residing in the bulge region of the
hair follicle (81,82).
Epithelial keratinocytes are applied today with success in
skin-grafting technologies, e.g. burn patients routinely have
cultured skin keratinocytes engrafted (68,83–85). They are
isolated from skin biopsy and plated onto a mitotically
inactivated and lethally irradiated layer of murine fibroblasts. The feeder layer supports clonal expansion of proliferative epithelial cells. Changing the culture conditions to a
higher calcium level subsequently supports the partial differentiation and stratification of the cells (49,86,87). However, several studies have demonstrated the superiority of
progenitor cells over more differentiated keratinocytes in
the generation of tissue-engineered skin (88–90). High a6
integrin expression (a6bri) and low expression of the transferrin receptor (CD71dim) is the most accepted combination of epidermal stem cell markers (91–93). Studies
analysing the regenerative capacity of these epidermal stem
cell populations revealed that skin equivalents created from
a6bri ⁄ CD71dim keratinocytes give rise to a stratified and
thick epidermis, while skin equivalents from a6bri ⁄ CD71bri
keratinocytes produced a thin and less well-differentiated
epidermis (93). However, several other cell surface proteins
like high expression of b1 integrin (94), the ability to
exclude Hoechst 33342 dye (95–97), expression of phospho-glycoproteins (98), CD200 (99,100), frizzled homologue 1 (FZD1) (99), p63, a homologue of p53 tumor
suppressor gene) (101), Keratin 19 (102), Keratin 15 (103–
105) or b-catenin (106) have been described recently for
epidermal stem cells. However, the identification of one or
rather a combination of specific epidermal stem cells markers is mandatory, to achieve a pure cell population.
The second reservoir for skin stem cells is the bulge
region of the ORS of hair follicles (107–111). Under physiological conditions, these stem cells regenerate the hair
bulb, while after injury they can also regenerate the sebaceous gland and the epidermis (112,113). Stem cells from
hair follicles could be differentiated experimentally into
neurons, glia cells, keratinocytes, smooth muscle cells and
melanocytes, indicating that the hair follicle contains stem
cell populations of both neuroectodermal and mesodermal
origin (89,100,114–118).
Because of their multiple differentiation capacity and the
facile accessibility hair follicle stem cells offer an attractive
source of stem cells for skin substitution. However, the
issue of a reliable and specific identification and discrimination of the various stem cell populations has not been
solved yet properly. Additionally, because of the limited
number of cells within the interfollicular epithelium and
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the ORS of the hair follicle, further investigations are also
needed for determination of appropriate culture conditions
to prevent stem cell loss.
Another fascinating application for epidermal stem cells
might be their use for gene therapy approaches. Considering that the epidermis is a self-renewing tissue, any permanent genetic correction must be aimed at the stem cell
population, to achieve long-lasting therapeutical effects.
Particularly, a gene therapy approach is perfectly eligible
for the treatment of diseases involving recessive loss-offunction mutations, as it occurs in junctional or recessive
dystrophic epidermolysis bullosa (EB) and other recessive
genetic skin disorders like X-linked ichthyosis or xeroderma
pigmentosus. The re-introduction of the wild-type gene via
viral or non-viral insertion is sufficient for correction of the
phenotype. In a striking and pioneering clinical trial, a team
of researchers and clinicians demonstrated for the first time
that cutaneous gene therapy has the potential to cure a
patient with inherited EB. The treatment relied on ex vivo
transduction of autologous epidermal stem cells with a normal copy of the defective gene, here laminin beta-3 chain,
followed by generation of epithelial sheets from these genetically modified cells. The transplantation of the epidermal
grafts led to functional correction of the disease (119).
Mesenchymal stem cells (MSCs; also referred to as mesenchymal stromal cells) either from bone marrow or adipose tissue present further suitable candidates for cell
therapy. Bone marrow MSCs (BM-MSCs) are a heterogeneous group of multipotent progenitor cells with the
capacity of self-renewal and differentiation into cells with
mesodermal, ectodermal and endodermal characteristics
(120–128). They have the intrinsic capacity to leave the
bone marrow, circulate in the blood and home to injured
tissues (129–133). Not surprisingly, they have also the
capability of accelerating cutaneous wound regeneration
both in animal models and in humans (134–137). Several
reports on the clinical application of BM-MSCs in wound
therapies have clearly revealed that grafted MSCs facilitate
skin regeneration, both in acute and in chronic wounds
(138–142). For example, Badiavas et al. (138) could demonstrate that autologous BM-MSCs delivered in a fibrin
spray accelerate healing in human cutaneous wounds.
However, the underlying mechanisms of BM-MSCs on skin
healing are multiple and have not been clarified in detail
yet, but include cell differentiation, wound contraction,
release of proangiogenic factors and production and maintenance of the ECM (137,143–145).
BM-MSCs are usually isolated from the mononuclear
layer of bone marrow after separation by density gradient
centrifugation and are achieved through expansion of plastic-adherent cells.
In recent years, in analogy to BM-MSCs, researchers are
investigating whether also adipose-derived stem cells (ASC)
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can be potentially applied in skin replacement strategies.
ASCs are a population of pluripotent cells, which have
characteristics similar to bone marrow-derived MSCs (146–
148). Generally, ASCs are isolated from lipoaspirates,
obtained by suction-assisted lipectomy (147,149,150). After
extensive washing, removal of the red cells and enzymatic
break-down of the ECM, the resulting cells are known as
stromal vascular fraction (SVF). SVF contains mesenchymal
stromal cells, but also other cells, like endothelial cells,
smooth muscle cells, pericytes, fibroblasts, leucocytes and
hematopoietic stem cells (151). Finally, from this mixed
cell population, the selection for ASCs occurs simply by
their ability to adhere to plastic ware. (152,153). Some
researchers have performed additionally purification by
magnetic bead coupling to deplete cells of the hematopoietic and of the endothelial lineages. ASCs are capable of
differentiating into other mesenchymal tissue types, like
adipocytes, chondrocytes, myocytes and osteoblasts and
show angiogenic properties (147,148,154–156). They seem
also to be applicable for skin regeneration, because in
experimental wound models ASCs accelerated wound closure by improved re-epithelization and angiogenesis (157–
162). In some of these studies, the stem cells had been
delivered via a human acellular dermal matrix or a matrix
composed of atelocollagen or silk fibroin-chitosan. It could
be shown, that these ASCs persisted locally and did not
distribute systemically, and by this providing anatomically
directed support to tissue regeneration at the desired site of
surgical engraftment (158–161).
These observations are particularly promising, because
BM-MSCs can only be obtained in a limited amount and
their differentiation abilities decrease with age. Additionally, the bone marrow procurement is extremely painful for
patients. In contrast, adipose tissue is ubiquitous and cells
are easily obtainable in adequate quantities with little
patient discomfort. Therefore, ASCs may provide a superior
source of stem cells for skin regeneration purposes
(155,157,161).
Despite of the eminent potential of MSCs in tissue
engineering, unintentional side effects in connection with
MSCs engrafting, like their immunosuppressive properties,
have to be taken into consideration before these cells will
be utilized in clinical applications (163–165). And very
importantly is a distinct molecular characterization of the
cells, e.g. appropriate marker proteins to allow for (i) the
selection of cells or cell types, respectively, that are most
beneficial to wound healing and (ii) the standardization of
technique improving the clinical usefulness.
Since their discovery in 1998 (166), human ES cells have
been recognized for their regenerative properties and viewed
as a potential application in tissue regeneration. However,
their use in clinical application is limited by ethical and
moral objections and the risk of graft rejection reactions or
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teratoma formation (167–169). At least the first two problems can be prevented by the use of reprogrammed somatic
cells that can be autologously derived. Several scientists have
succeeded in generating iPS cells from adult human dermal
fibroblasts and from keratinocytes by transduction with a
combination of various transcription factors, involved in
reprogramming. These cells were phenotypically and functionally indistinguishable from ES cells (170–174). These
cells could potentially be used in the construction of tissueengineered skin, after considerations of potential tumorgenicity caused by cell regulations and genetic manipulations
have been sorted out.
Table S3 summarizes the differentiation potential of follicular epidermal stem cells, stem cells from the bulge
region, bone marrow-derived MSCs, adipose stem cells, ES
cells and iPS-cells.
Conclusion
New progress in stem cell technologies has extended greatly
our knowledge about the function and relevance of adult
stem cells for tissue development and tissue regeneration.
Additionally, new technologies in material sciences are
emerging for the accurate manufacture of degradable polymers, creating materials with defined and customized
mechanical properties for the repair or replacement of
impaired tissue or organs. Particularly in dermatology,
numerous developments in the field of tissue engineering
have been translated already into therapeutical applications,
like for the treatment of extensive chronic wounds, e.g. skin
burns, venous or diabetic leg ulcers. Initial success in treating recessive genetic skin disorders by a combined approach
using gene therapy and tissue engineering indicates clearly
that skin is an appropriate target for innovative therapeutic
strategies in regenerative medicine. Despite of all this scientific progress, the skin substitutes used at present are yet
not fully functional as they lack differentiated structures
as nerves, sweat glands, pilosebaceous structure and blood
supply. Still, intensive research has to proceed to attain a
durable and cosmetically acceptable tissue-engineered skin.
The choice of suitable cells in sufficient quantity and purity,
adequate culture conditions to maintain stem cell properties, as well as appropriate biomaterials for transient or permanent replacement are factors important for long-lasting
engraftment of a regenerated skin.
The tissue-engineered skin of the future will resemble
morphologically and functionally natural skin. Epidermal
stem cells will be utilized for generation of the epidermal
component, probably along with the addition of melanocytic stem cells. For the dermal construct, endothelial, mesenchymal and neuronal stem cells may support the
generation of the dermal component with a functional vasculature. Furthermore, improving tissue-engineered skin
ª 2010 John Wiley & Sons A/S, Experimental Dermatology, 19, 697–706
and stem cell-targeted cutaneous gene transfer will be
essential for the successful gene therapy.
In conclusion, already today regenerative medicine has
proven to have a tremendous potential in dermatological
applications that will increase in the future, as the interdisciplinary research on stem cell biology, biomaterial science,
gene therapy and tissue engineering will progress.
Acknowledgement
This work was supported by funding from the German Federal Ministry of
Education and Research (BMBF, PtJ-Bio 0313909 to C.D. and L.M.) and
by the German Research Council (DFG, TRR-SFB 67, TP B3, http://
www.trr67.de to JCS).
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Supporting Information
Additional Supporting Information may be found in the online version of
this article:
Figure S1. Scheme of a mature anagen hair follicle, plucked hair follicles
in their anagen phase, the manufacturing process of outer root sheath
(ORS) cells to an epidermal equivalent (EpiDex) and finally the clinical
application in the treatment of chronic venous leg ulcers.
Table S1. Representative human clinical trials of stem cell therapies,
both completed or pending, indexed by clinical application and source of
stem cell.
Table S2. Examples of various types of acellular and cellular skin substitutes currently commercially available without intending to be complete.
Table S3. Differentiation potential of human epidermal stem cells, bone
marrow-derived mesenchymal stromal cells, adipose stem cells, embryonic
stem cells and induced pluripotent stem cells.
Please note: Wiley-Blackwell are not responsible for the content or functionality of any supporting materials supplied by the authors. Any queries
(other than missing material) should be directed to the corresponding
author for the article.
ª 2010 John Wiley & Sons A/S, Experimental Dermatology, 19, 697–706