ADVANCED THERAPIES OF SKIN INJURIES

FORTSCHRITTLICHE THERAPIEN DER HAUTSCHADEN

Tina MAVERa, Uroš MAVERb,*, Karin STANA KLEINSCHEKa, Irena Mlinarič RAŠČANc and

Dragica Maja SMRKEd a University of Maribor, Faculty of Mechanical Engineering, Laboratory for Characterisation and

Processing of Polymers, Smetanova 17, SI-2000 Maribor, Slovenia

b University of Maribor, Faculty of Medicine, Taborska ulica 8, SI-2000 Maribor, Slovenia

c University of Ljubljana, Faculty of Pharmacy, Aškerčeva 7, SI-1000 Ljubljana, Slovenia

d University Medical Centre Ljubljana, SI-Ljubljana, Slovenia

Acknowledgements: The paper was co-produced within the framework of the operation entitled

“Centre of Open innovation and ResEarch UM (CORE@UM)”. The operation is co-funded by the

European Regional Development Fund and conducted within the framework of the Operational

Programme for Strengthening Regional Development Potentials for the period 2007 – 2013,

development priority 1: “Competitiveness of companies and research excellence”, priority axis 1.1:

“Encouraging competitive potential of enterprises and research excellence”, contact No. 3330-13-

500032. The authors acknowledge also the financial support from the Ministry of Higher Education,

Science and Technology of the Republic of Slovenia.

*Corresponding author

Assist. Prof. dr. Uroš Maver, University of Maribor, Department of Pharmacology and Experimental Toxicology, Faculty of Medicine, Taborska ulica 8, SI-2000 Maribor, Slovenia Phone: +386 2 234 58 23 Fax: +386 2 234 59 23 uros.maver@um.si  

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ADVANCED THERAPIES OF SKIN INJURIES

FORTSCHRITTLICHE THERAPIEN DER HAUTSCHADEN

Abstract

The loss of a tissue is still one of the most challenging problems in healthcare. Efficient laboratory

expansion of skin tissue to reproduce the skins barrier function can make the difference between life

and death for patients with extensive full-thickness burns, chronic wounds or genetic disorders such as

bullous conditions. This engineering has been initiated based on the acute need in the 1980s and today,

tissue-engineered skin is the reality. The human skin equivalents are available not only as models for

permeation and toxicity screening, but are frequently applied in vivo as clinical skin substitutes. This

review aims to introduce the most important recent development in the extensive field of tissue

engineering and to describe already approved, commercially available skin substitutes in clinical use.

Die Zusammenfassung

Der Verlust eines Gewebes ist nach wie vor eine der größten Herausforderungen im

Gesundheitswesen. Effiziente Labor Expansion des Hautgewebes, um die die Haut Barriere-funktion

zu reproduzieren kann den Unterschied zwischen Leben und Tod bei Patienten mit dicken

Verbrennungen, chronischen Wunden oder genetischen Störungen wie bullösen Bedingungen

bedeuten. Die Gewebetechnik wurde auf der Grundlage des akuten Bedarfs in den 1980er Jahren

gestartet und Heute ist Haut, produziert durch diese Technik, die Realität. Die menschlichen

Hautäquivalente sind nicht mehr nur Modelle für die Permeation und Screening der Toxizität, aber

sind immer mehr auch als klinische Hautersatze verwendet. Dieser Übersichtsartikel beschreibt die

wichtigsten neuen Entwicklungen in dem umfangreichen Gebiet des Tissue-Engineering unter dessen

auch die bereits genehmigten, im Handel erhältlichen Hautersatze in der klinischen Anwendung.

Keywords: allograft, autograft, scaffold, tissue engineered skin, tissue engineering

Die Schlűsselwőrte: allograft, autograft, scaffold, tissue engineered skin, tissue engineering

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1. Introduction

Skin is the largest organ of the human body. Its main role is to protect the body from toxins and

microorganisms from the environment and to prevent dehydration. Other crucial skin functions are

immune surveillance, sensory detection, and self-healing. Loss of its integrity due to the injury or

illness may acutely result in substantial physiologic imbalance and ultimately in significant disability

or even death.

Among the most common causes to skin damage are acute trauma, chronic wounds, infections [1],

surgical interventions and genetic disorders. Major skin loss with substantial damaged areas is often

caused by thermal trauma.

According to the extent of skin damage, we can divide injuries of the skin into:

a) Epidermal injuries, which represent minor lesions that affect only the epidermis and therefore do

not require any specific surgical treatment. Such injuries regenerate rapidly without significant

scarring.

b) Superficial partial-thickness wounds, which affect the epidermis and superficial parts of the

dermis. Such wounds heal by epithelialization, which originates from the wounds edges, where

basal keratinocytes transform into a proliferating migratory cell type and cover the damaged area.

Cells migrate either from the wound edge, hair follicle or from sweat gland remnants that lie in the

deeper dermis, which has been preserved in the injury [2, 3].

c) Deep partial-thickness injuries, which involve greater dermal damage resulting in fewer skin

appendages and take longer to heal. For this type of wounds, scarring is more pronounced when

compared with superficial partial-thickness wounds.

d) Full thickness injuries are characterized by the complete destruction of epithelial-regenerative

elements. This type of injury heals by contraction, with epithelialization from only the edge of the

wound, leading to cosmetic and functional defects [2].

According to the type of injury, different wound healing approaches are used.

2. The existing approaches in the field of wound healing

Silver wound dressings

3

Silver is an established antibacterial agent and has been applied in wound dressings in the treatment of

chronic wounds and burns to manage infections for many years. Dressings containing silver

sulfadiazine (Ag-SD) or silver nanomaterials are most commonly used to treat external infections and

their use is increasing in importance due to the recent increase of antibiotic-resistant bacteria [4].

Moist wound healing

Moist wound healing has been shown to accelerate re-epithelialization of acute wounds [5], and these

observations have led to the development of many moisture retentive dressings. For chronic wounds,

moist wound healing has not been shown to significantly improve epidermal healing; nevertheless it

was proven that it helps in the formation of granulation tissue and in relieving pain. Painless

debridement is another important property of moisture-retentive dressings. Proposed advantages of

moist wound healing include retention of cytokines within the wound, facilitation of keratinocyte

migration and prevention of bacterial entry [6].

Negative pressure wound therapy

Negative-pressure wound therapy includes fluid removal, wound contraction, microdeformation and

moist wound healing. The success of this treatment is the consequence of an increased wound blood

flow, the increased granulation tissue formation, decreased bacterial counts and the stimulation

of wound healing pathways through shear stress mechanisms. Over the last decade, numerous uses of

this wound management approach have been reported, ranging from the treatment of acute and

chronic wounds, to the closure of open sternal and abdominal wounds, to the assistance with skin

grafts [7].

Factor-based approach

Identification of putative wound-healing factors has led to several attempts to speed wound healing by

local application of one or more factors that promote cell attachment and migration. Most factor-based

approaches have had marginal success. Candidates for this purpose have been: transforming growth

factor-β (TGF-β), epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), and

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platelet-derived growth factor (PDGF). Only PDGF has shown efficacy in clinical trials and is

approved for clinical use (Regranex®). It was shown that the use of factors is not enough in situations

of severe or massive loss of skin tissue [8].

Skin grafting with Autografts

Despite the fact that skin grafting with an “autograft” is the treatment of choice, there are situations in

which it is necessary to replace large areas of epidermis or in which normal regeneration is deficient.

Wounds that extend deep into the dermis tend to heal very poorly and slowly because no keratinocytes

remain to reform the epithelium. The scar tissue that forms in the absence of dermis lacks elasticity,

flexibility, and strength of the normal dermis. Consequently, scar tissue limits movement, causes pain,

and is cosmetically undesirable [9]. Skin grafting with autografts is extremely well established and

evolved technique [10]. Surgeons “shave” a thin layer of skin with the dermatome (a surgical

instrument that holds a razor-sharp blade parallel to the skin surface) from the donor site (most

commonly a non-conspicuous area such as inner thighs and buttocks) that includes the full epidermis

and portion of the dermis, or what is commonly known a split thickness graft [11]. The skin graft is

then placed on the wound site. Generally, the thicker the split skin graft is, the less contraction there

will be at the site of application but the longer it will take to heal the donor site [12]. Autologous skin

grafts avoid issues of immunogenicity, but they have significant limitations: the donor site creates

another wound, and in severe-burn patients there may be no appropriate donor site.

Skin grafting with Allografts and Xenografts

When autologous skin grafts are not possible, human cadaver allograft skin has been used. Xenografts

made of different animal skin have also been used for the same purpose. Problems associated with all

allografts and xenografts include the possibility of an immune rejection reaction [13], possible

infection, and problems of supply and variability in the quality of the material [14].

Skin replacement products

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Since suitable autografts for transplantation are very rarely present and the demand for allograft tissues

exceeds the supply, not to mention the possible rejection of foreign tissue, there has been a significant

need for tissue engineering [15]. In the following chapters we will further expand this topic.

3. Tissue engineered skin

The importance and demand for skin-replacement products encouraged many research groups

worldwide to focus on creating biomaterials for skin substitution. Engineered tissues that not only

close wounds but also stimulate the regeneration of dermis would provide a significant benefit in

human wound healing.

The main demand for tissue-engineered skin substitute bio-constructs is safety for the patient. These

have to be non-toxic, non-immunogenic, without any transmissible disease risk and should not cause

excessive inflammation. Their clinical effectiveness has to be proven; the biomaterial for skin

reconstruction has to support the reconstruction of normal tissue, with similar physical and mechanical

properties of the skin it replaces, and be biodegradable. Application of such materials supports pain

relief, prevents fluid and heat loss from the wound surface and protects the wound from infection. Last

but not least, tissue-engineered skin substitutes have to be convenient in handling and application,

which includes their availability, user-friendliness and a long shelf life [16].

In general, an engineered skin graft should incorporate as many naturally occurring skin components

as possible to imitate structural and biological characteristics of the skin: the extracellular matrix,

dermal fibroblasts, the epidermis, and a semipermeable membrane. It is very important, that these

components act synergistically as a part of a fully integrated tissue to protect the underlying tissues of

a wound bed to direct the healing process [17].

Various biological and synthetic materials have been combined with in vitro cells cultures to generate

functional tissues. The basic principle of skin tissue engineering is shown in Fig. 1 [18].

*Figure 1*

We will herein review some of the currently marketed and clinically available tissue-engineered skin-

substitutes and list them according to the following anatomical structure classification: 1. epidermal, 2.

6

dermal and 3. dermo-epidermal skin substitutes. Their availability in the European Union is addressed

in the last column of the respective tables.

3.1. Epidermal skin substitutes

Re-epithelialization of the wound is the paramount concern, since these substitutes provide limited

defense against contamination of the exposed underlying tissue or against extensive fluid loss, since

they do not provide full coverage of the exposed area. The approaches to re-establishing epidermis are

numerous, ranging from the use of cell suspensions to full-thickness skin equivalents possessing a

differentiated epidermis [19]. This process of growing complexity is shown in Fig. 2.

*Figure 2*

A key step in the designing and production of epidermal substitutes is the isolation of keratinocytes

from a donor and the subsequent in vitro cultivation of these cells to obtain the necessary number of

keratinocytes for therapeutic needs [20]. Regardless of approach, living epidermal keratinocytes are

necessary to achieve permanent, biologic wound closure.

Cultured epithelial autograft sheets

Growing human epidermal keratinocytes from small patient-biopsy samples (usually 2 – 5 cm2) using

co-culture methods is today possible with some well-established techniques [21]. Commonly, the

epidermis is separated from the dermis and single keratinocytes are released from the sheet through

exposure to enzymes [22]. The mouse 3T3 fibroblast feeder cell system allows substantial expansion

of epidermal keratinocytes and can be used to generate a sufficiently thin, multilayered epidermal

sheet to resurface the body of a severely burned patient [23]. Once transplanted, the epidermal sheets

quickly form the epidermis and re-establish epidermal coverage. In vitro keratinocyte expansion

techniques involve production of the cultured epithelial autograft (CEA) sheets, which are large

enough to cover the entire surface of the body in three to four weeks from an only 3 cm2 big skin

biopsy sample [20]. With time, the CEA stimulates formation of new connective tissue immediately

beneath the epidermis. Scarring and wound contractions remain a significant problem of this type of

treatment. Studies have shown that grafting of CEA onto pregrafted cadaver dermis greatly improves

7

graft take [24]. The oldest approach to deliver keratinocytes into the wound are the cultured epithelial

autografts (Epicel®, EPIBASE®, EpiDex®), which are manufactured using the patient’s own

keratinocytes, grown to confluency within 15 days to form CEA sheets. Disadvantages of CEA sheets

are a long culturing time, complicated handling and application procedures and a non-predictable

clinical success related to the confluent, layered cell culture system [25].

Sub-confluent keratinocytes

Studying and evaluating the disadvantages of cultured epithelial autograft sheets mentioned above,

resulted in the investigation of the use of sub-confluent keratinocytes (CellSpray®), which have a

greater in vivo proliferative activity, are harvested earlier, after 5–7 days of growth, and exhibit

flexibility in the coordination of cell propagation, harvesting and clinical application processes. They

can be applied to the wound bed via an aerosolized cell suspension [26]. Sub-confluent keratinocyte

suspensions contributed to an earlier basement membrane formation with a mature dermal–epidermal

junction region when compared with CEA sheets [27]. Although this method allows a more

convenient way of delivering keratinocytes to the wound bed at earlier stages of post-wounding, such

an application is limited to partial-thickness and graft donor site wounds, while full-thickness wounds

still require a dermal element to achieve functional permanent skin restoration [28].

Cultured sub-confluent keratinocytes on delivery membrane

Another approach to deliver sub-confluent keratinocytes to the wound bed is to culture a sub-confluent

keratinocytes on delivery membranes which can be either mechanically peeled off the culture vessel

[29] or can be applied with the cultured cells directly to the wounded site [30]. Delivery membranes

can be made from synthetic materials such as a silicone support membrane with a specially formulated

surface coating (MySkin®); polyurethane; or based on biological materials such as collagen, fibrin

glue, hyaluronic acid (Laserskin® (Vivoderm®)) or decellularized dermis [20]. The benefits of the use

of these delivery systems include earlier clinical cell application [16].

Some of the already approved or clinically tested epidermal skin substitutes are gathered in table 1.

8

Table 1: Currently commercially available epidermal bioengineered constructs [16, 20, 31, 32]. The

figures in the table were reproduced with permission of The Royal Society from 15.

The listed epidermal substitute products enable a permanent wound closure. They are effective in

treating chronic ulcers and improving the quality of life of patients. Their efficiency and long-term

outcomes for burn treatment are still questioned by many. In order to achieve effective full-thickness

wound healing, combination with dermal substitute is necessary.

3.2. Dermal skin substitutes

Wound bed preparation is very important for an effective graft take [19]. Cultured epithelial autograft

take is reportedly successful in only 15 % of cases when grafted onto chronic granulation tissue, in

28–47 % of cases if grafted onto early granulation tissue or a freshly debrided wound, but would have

a 45–75 % chance of integration when applied to the wound with a dermal or neodermal bed [33]. Pre-

grafting is also important for the successful take of cultured autologous keratinocytes and for

successful full-thickness wound epithelialization by sprayed keratinocytes. The majority of products

for dermal substitutions are acellular, based either on allogeneic, xenogeneic or synthetic materials

[34].

The main approaches for re-establishing dermis are shown in Fig. 3.

*Figure 3*

Allogeneic human acellular dermis

Decellularized cadaver dermal tissue has been used to recapitulate as much of the normal skin

architecture as possible, while simultaneously providing a natural scaffold for re-epithelialization [14].

Cadaver allograft dermis can be processed to make an immunologically inert, acellular dermal matrix

with an intact base membrane to aid the take and healing of ultrathin autografts (AlloDerm®) [35].

Initially intended as a dermal replacement material, allogeneic human acellular dermis showed

uncertain rates of vascularization [36] and has therefore recently gained more popularity for the

procedures, where immediate revascularization is of less importance. It is still used for acute thermal

injury treatment with very promising results [37].

9

Xenogeneic acellular dermis

The benefits of decellularized dermal products of animal origin (Permacol Surgical Implant®,

Matriderm®, OASIS Wound Matrix®) are reduced risk associated with transferable human viral

diseases (HIV, HepB), an easier and cheaper production due to the availability of raw materials. On

the other side, the weaknesses involve slow bio-integration and vascularization [38], which is why its

use for dermal reconstruction is limited.

Different xenogeneic/synthetic scaffolds for dermal extracellular matrix

The mentioned approach uses the possibility of redirecting granulation-tissue formation through the

use of scaffolds and livings cells. In one of the earliest tissue engineering approaches for improving

dermal healing, Yannas et al. [39] designed a collagen–glycosaminoglycan sponge to serve as a

scaffold/template for dermal extracellular matrix. The goal has been to promote fibroblast repopulation

in a controlled way to decrease scarring and wound contraction [39]. A commercial version of such a

material composed of bovine type I collagen, shark chondroitin-6-sulfate glycosaminoglycan and a

silicone membrane covering (Integra Dermal Regeneration Template®) is currently approved for use

in burns [40]. The exogenical grafted dermal layer is slowly resorbed and the silicone membrane is

eventually removed and replaced by a thin autograft. Among the advantages of this treatment approach

are the long shelf life, simple handling, low risks of immunogenic response and disease transmission

and good cosmetic outcomes with reduced rates of contraction and scarring [41]. A meticulous

surgical preparation of the wound bed that is required to guarantee a good take of these products is the

main disadvantage. Additional limitation are its non-applicability on infected wounds, a relatively long

time (10–14 days) for vascularization and the necessity of a second operative procedure to achieve

permanent wound closure [16].

Several variations of the collagen sponge have been studied and more products have been put on the

market (Terudermis®, Pelnac Standard Type®/Pelnac Fortified With Mesh Type®). Efforts have been

made to improve the fibroblast infiltration and collagen persistence by collagen cross-linking [42], by

inclusion of other matrix proteins and by modifying the porosity of the scaffold [43]. Another

10

variation is in the use of hyaluronic acid (Hyalomatrix PA®, Hyalograft 3D®), one of the main

polysaccharide components in the dermal extracellular matrix (ECM) that promotes migration and

proliferation of skin fibroblasts and keratinocytes [42]. Products based on hyaluronic acid serve as a

temporary dressing to stimulate wound regeneration after dermabrasion and are reported to be a good

and feasible approach for such wound treatments. Because they do not contain any animal or

allogeneic human-derived components, these are also appealing from the safety point of view [44].

Although matrix scaffolds have shown some improvement in scar morphology, no acellular matrix has

yet been shown to lead to true dermal regeneration. This may be due to limitations in the cell

repopulation, the type of fibroblast repopulating the graft, and due to lack of the inflammatory and

remodeling processes [19].

Cultured neonatal fibroblasts grown on different scaffolds

Recently several advances have been made in the design of artificially grown dermal tissues using

human neonatal fibroblasts grown on rectangular sheets of biodegradable polygalactin mesh

(Dermagraft®). The scaffold degrades by hydrolysis in 20–30 days while fibroblasts produce growth

factors and ECM components (vitronectin, tenascin, collagens and glycosaminoglycans), helping to

reconstitute a dermal layer. The disadvantages of this product include a necessity for multiple

applications, higher cost and safety issues owing to allogeneic cells incorporated into this bio construct

[45].

Related products are nonviable temporary coverings for burns. In this case a nylon mesh coated with

porcine collagen, layered with a pseudo-epidermal semipermeable silicone membrane (Biobrane®)

serves as a platform for deposition of human matrix proteins and associated factors by the human

dermal fibroblasts [19]. TransCyte® exhibits additional viable cultured neonatal allogeneic fibroblasts,

which are incorporated into the scaffold. These bioengineered constructs provide matrix proteins,

growth factors and cytokines necessary for wound healing improvement; they are effective for vapor

loss control, pain relief and are reported to reduce healing time when compared to conventional

dressings [46]. Being non-degradable and synthetic, these materials must be removed after 7–14 days

11

and are intended for temporary wound coverage until permanent closure with either autologous skin

graft or cultured epithelial cells are achieved [16].

Currently commercially available dermal bioengineered constructs are listed in table 2.

Table 2: Currently commercially available dermal bioengineered constructs [16, 20, 31, 32]. The

figures in the table were reproduced with permission of The Royal Society from 15.

3.3. Dermo-epidermal composite skin substitutes

Dermo-epidermal or composite skin substitutes aim to mimic the histological structure of normal skin,

where both epidermal and dermal layers are present. These are not only the most advanced and

sophisticated products, when compared to epidermal and dermal substitutes, but are also the most

expensive tissue-engineered biological constructs for tissue repair [34].

The main approaches for re-establishing epidermis and dermis are shown in Fig. 4.

*Figure 4*

Allogeneic skin cells incorporated into a dermal scaffold

This approach allows the production of large quantities of uniform product batches, which act like a

temporary biologically active wound dressing [47], providing growth factors, cytokines and ECM for

host cells while initiating and regulating wound healing. Normal cell populations seem to have an

intrinsic ability to re-express their differentiated program in vitro [48].

One of the first commercially available dermo-epidermal skin substitutes has been composed of viable

allogeneic neonatal fibroblasts, grown in a bovine type I collagen gel matrix, combined with viable

allogeneic neonatal keratinocytes (Apligraf®). Although this product does not cause immunological

rejection, allogeneic cells of the construct do not survive after one to two months in vivo [49].

Therefore, it can only be considered as a temporary bioactive dressing. Another tissue-engineered skin

construct OrCell® includes cultured allogeneic fibroblasts and keratinocytes obtained from the same

neonatal foreskin. This artificial skin substitute product showed reduced scarring, and a shorter healing

time when compared to the acellular bioactive wound dressing Biobrane® [50]. Because it is

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composed of allogeneic cells, it resorbs in 7 – 14 days. To deliver a definitive wound closure in full-

thickness injuries, co-grafting with an autologous epithelial source it needed.

Autologous skin cells incorporated into different matrices

The next step in the development of bioengineered skin has been to include autologous cultured

keratinocytes and fibroblasts into the matrix. PolyActive® is such a product, and is built of a soft

polyethylene oxide terephthalate component and a hard polybutylene terephthalate component with

incorporated autologous skin cells [51]. Advantage of this product is almost no risk of immune

rejection or cross-contamination by infective agents, but the use of autologous cells may limit the

product’s ‘off-the-shelf’ availability and increase its costs when compared with competitive

allogeneic-based products.

Combination of dermal replacement constructs and epidermal substitutes in one product

The TissueTech Autograft System® combines two tissue-engineered biomaterials; the dermal

replacement construct Hyalograft 3D® and the epidermal substitute Laserskin® [52]. These are based

on autologous keratinocytes and fibroblasts, grown on microperforated hyaluronic acid membranes.

Although this system may allow a definitive wound closure, it is not a ‘true’ bilayered skin substitute

where both dermal and epidermal layers are present, as it requires grafting of two products, and may

be complicated to use in a clinical setting. The preceding literature therefore suggests that no

commercially available true bilayered ‘skin substitute’ for permanent deep wound closure is yet

available [16].

”True skin substitute”

Some attempts have been made to combine commercially available dermal substitutes with either

cultured or non-cultured autologous cells in pre-clinical studies [53, 54] with promising results.

Unfortunately no follow-up clinical trial results are yet available. There is only one clinically used

three-dimensional reconstructed skin substitute, the Cincinnati Shriners Skin Substitute or

PermaDerm®, which has been designed by Boyce et al. [47]. It is based on the collagen sponge,

13

seeded with autologous fibroblasts and keratinocytes and therefore enables permanent wound closure

and can be viewed as a ‘true skin substitute’ [16].

The current marketed dermo-epidermal (composite) skin substitutes are listed in table 3.

Table 3: Currently marketed dermo-epidermal bioengineered constructs [16, 20, 31, 32]. The figures

in the table were reproduced with permission of The Royal Society from 15.

4. Future directions in development of bioengineered skin substitutes

Different skin and dermal substitute constructs are currently under investigation, either in in vitro

studies or have entered stage II–III clinical trials and could therefore possibly hit the market in the near

future.

The future challenges can be divided into four directions: improvement of preparation procedures,

inclusion of “new” biomaterials, functionalization of skin substitutes and the use of stem cells.

Improvement of preparation procedures

Among the challenges facing bioengineers supplying live cell products are long, complicated and

expensive cultivation procedures, specific and expensive transport and storage conditions, limited

product shelf-lifes, the friable nature of cell-containing biomaterials, especially for products based on

living cells and the need for precise coordination between the tissue culture facility and the clinic if

autologous cells are used. All above mentioned reasons as well as the always important cost-

effectiveness of cell-based biomaterials, which are mostly only partially effective at fulfilling skin

functions, make it also very difficult for any cell-based skin substitute product to reach the clinical use

let alone commercialization [26].

“New” biomaterials

Novel potential skin substitute biomaterials and scaffolds include human hair keratin-collagen sponge,

hyaluronan coupled with fibronectin functional domains, poly(lactic-co-glycolic acid)/chitosan hybrid

nanofibrous membrane, biodegradable polyurethane microfibres, polycaprolactone (PCL) collagen

nanofibrous membranes, silk fibroin and alginates, polyvinyl alcohol/chitosan/fibroin blended

14

sponges, Tegaderm-nanofibre constructs, bacterial cellulose, ICX-SKN skin graft replacements,

porcine collagen paste, bovine collagen cross-linked with microbial transglutaminase, collagen–

glycosaminoglycan– chitosan dermal matrix seeded with fibroblasts, composite nano-titanium oxide–

chitosan artificial skin, keratinocytes and fibroblasts grown on Collatamp, deacetylated chitin or plant

cellulose transfer membranes and many others [16, 55, 56]. New materials could also be found and

evaluated by novel computational methods [57].

Search for functional skin replacements

Currently available products for permanent skin substitution can partially replace its protective barrier

function, while other functions, including touch and temperature sensation, excretion, perspiration,

thermoregulation, protection from ultraviolet rays, synthetic function, not to mention the aesthetic

function, cannot be restored by the existing skin tissue-engineered products [26]. The combination of

different skin cell types including keratinocytes, melanocytes, fibroblasts and endothelial cells derived

from postnatal skin is aiming to create a functional skin replacement [58].

Use of stem cells

A new approach, where either embryonic or adult stem cells are used, both true skin regeneration and

avoidance of scarring may be successfully achieved. Stem cells are undifferentiated cells that renew

themselves for the entire life and they can develop from a common precursor into multiple cell types

with specialized functions. Due to their ability to differentiate into various tissue types by asymmetric

replication, may help create those skin components that are not found in the tissue engineered skin

substitutes. Among the sources of cells that might be used for regeneration of injured skin are adult

stem cells and pluripotent stem cells [59]. The number of investigations in the use of adult stem cells

is constantly growing [60, 61].

Adult stem cell research exhibits potential for tissue-engineering approaches for skin regeneration and

treatment of extensively burned patients and other acute and chronic skin defects. The already present

huge scientific interest will certainly lead to a significant progress in this field in the coming years.

15

The authors report no conflicts of interest.

Acknowledgements

The authors acknowledge the financial support from the Ministry of Higher Education, Science and

Technology of the Republic of Slovenia, as well as the financial contributions from the mnt-era.net

funded project WoundSens with the grant number 3211-12-00002.

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20

TABLES

Table 1:

Commercial

name/

Manufacturer

Schematic representation

Incorporated

human cells/cell

source

Scaffold material

/ scaffold source

Duration

of the

cover

Available

in EU?

Epicel/

Genzyme

Biosurgery,

USA

cultured

keratinocytes

(confluent cell

sheet)/autologous

-/- permanent

EpiDex/

Modex

Therapeutiques,

Switzerland

cultured

keratinocytes

from outer root

sheath of scalp

hair follicles

(confluent cell

sheet)/autologous

-/- permanent

EPIBASE/

Laboratories

Genevrier,

France

cultured

keratinocytes

(confluent cell

sheet)/autologous

-/- permanent

MySkin/

CellTran Ltd,

UK

cultured

keratinocytes

(subconfluent cell

sheet)/autologous

silicone support

layer with a

specially

formulated surface

coating/synthetic

permanent

Laserskin

(Vivoderm)/

Fidia Advanced

Biopolymers,

Italy

cultured

keratinocytes

(confluent cell

sheet)/autologous

hyaluronic acid

membrane

(microperforated)/

recombinant

permanent

CellSpray/

Clinical Cell

Culture,

Australia

non-/cultured

keratinocytes

(subconfluent cell

sheet)/autologous

-/- permanent

21

Table 2:

Commercial name/

Manufacturer Schematic representation

Incorporated

human

cells/cell

source

Scaffold material

/scaffold source

Duration

of the

cover

Available

in EU?

AlloDerm/

LifeCell

Corporation, USA

-/-

human acellular

lyophilized dermis/

allogeneic

permanent

Karoderm/

Karocell Tissue

Engineering AB,

Sweden

-/- human acellular

dermis/allogeneic permanent

SureDerm/

HANS BIOMED

Corporation, Korea

-/-

human acellular

lyophilized dermis/

allogeneic

permanent

GraftJacket/

Wright

MedicalTechnology,

USA

-/-

human acellular pre-

meshed dermis/

allogeneic

permanent

Matriderm/

Dr Suwelack Skin

and HealthCare AG,

Germany

-/-

bovine non-cross-

linked lyophilized

dermis, coated with α-

elastin hydrolysate/

xenogeneic

permanent

Permacol Surgical

Implant/

Tissue Science

Laboratories plc,

UK

-/-

porcine acellular

diisocyanite cross-

linked dermis/

xenogeneic

permanent

OASIS Wound

Matrix/

Cook Biotech Inc.,

USA

-/-

porcine acellular

lyophilized small

intestine submucosa/

xenogeneic

permanent

Integra Dermal

Regeneration

Template/

Integra

NeuroSciences, USA

-/-

polysiloxane, bovine

cross-linked tendon

collagen,

glycosaminoglycan /

xenogeneic+synthetic

semi-

permanent

Terudermis/

Olympus Terumo

Biomaterial Corp.,

Japan

-/-

silicone, bovine

lyophilized cross-

linked collagen

sponge made of heat-

denatured collagen/

xenogeneic+synthetic

semi-

permanent

Pelnac

Standard/Pelnac

Fortified/

Gunze Ltd, Medical

Materials Center,

Japan

-/-

silicone/silicone

fortified with silicone

gauze (TREX),

atelocollagen derived

from pig tendon/

xenogeneic+synthetic

semi-

permanent

22

Biobrane/

UDL Laboratories,

Inc., USA

-/-

silicon film, nylon

fabric, porcine

collagen/

xenogeneic+synthetic

temporary

TransCyte /

Advanced

BioHealing, Inc.,

USA

cultured

neonatal

fibroblasts/

allogenic

silicon film, nylon

mesh, porcine dermal

collagen/

xenogeneic+synthetic

temporary

Dermagraft/

Advanced

BioHealing, Inc.,

USA

cultured

neonatal

fibroblasts/

allogenic

polyglycolic acid /

polylactic acid,

extracellular matrix /

allogeneic+synthetic

temporary

Hyalomatrix PA/

Fidia Advanced

Biopolymers, Italy

-/-

a derivative of

hyaluronan layered on

silicone membrane/

allogeneic+synthetic

semi-

permanent

Hyalograft 3D/

Fidia Advanced

Biopolymers, Italy

cultured

fibroblasts/

autogenic

hyaluronic acid

membrane

(microperforated) /

allogeneic

permanent

23

Table 3:

Commercial

name/

Manufacturer

Schematic representation Incorporated human

cells/cell source

Scaffold material

/scaffold source

Duration

of the

cover

Available

in EU?

Allograft/

From - not for

profit skin

banks

native/allogeneic

native human skin

with dermal

and epidermal

cells/allogeneic

permanent NA

Karoskin/

Karocell

Tissue

Engineering

AB, Sweden

native/allogeneic

native human

cadaver skin with

dermal and

epidermal cells/

allogeneic

permanent

Apligraf/

Organogenesis

Inc., USA

cultured keratinocytes

and

fibroblasts/allogeneic

bovine collagen /

xenogeneic permanent

OrCel/

Ortec

International,

Inc., USA

cultured keratinocytes

and

fibroblasts/allogeneic

bovine collagen

sponge/xenogeneic permanent

PolyActive/

HC Implants

BV, The

Netherlands

cultured keratinocytes

and fibroblasts/

autologous

polyethylene oxide

terephthalate/

polybutylene

terephthalate/

synthetic

permanent

TissueTech

Autograft

System

(Laserskin and

Hyalograft

3D)/

Fidia

Advanced

Biopolymers,

Italy

cultured keratinocytes

and fibroblasts/

autologous

hyaluronic acid

membrane

(microperforated)/

recombinant

permanent