Non-Animal Models of Wound Healing in Cutaneous Repair:
In Silico, In Vitro, Ex Vivo And In Vivo Models Of Wounds And Scars In Human Skin
Sara Ud-Din, MSc1, Ardeshir Bayat, MBBS, PhD1,2
1Plastic and Reconstructive Surgery Research, Centre for Dermatology Research, University of Manchester,
Manchester, UK.
2Bioengineering Research Group, School of Materials, Faculty of Engineering & Physical Sciences, The
University of Manchester, Manchester, UK.
Corresponding Author: Dr Ardeshir Bayat (MBBS, PhD) Associate Professor Plastic and Reconstructive Surgery Research Centre for Dermatology Research University of Manchester Stopford BuildingOxford Road Manchester M13 9PT England, UK. Tel: 0161 306 0607Email: [email protected]
Running title: Models of Wound Repair in Human Skin
Keywords: Models, wound repair, wound healing, tissue repair, in vivo, ex vivo, in vitro, in silico,
experimental, human
1
ABSTRACT
Tissue repair models are essential in order to explore the pathogenesis of wound healing and
scar formation, identify new drug targets/biomarkers and to test new therapeutics. However,
no animal model is an exact replicate of the clinical situation in man as in addition to
differences in the healing of animal skin; the response to novel therapeutics can be variable
when compared to human skin. The aim of this review is to evaluate currently available non-
animal wound repair models in human skin, including: in silico, in vitro, ex vivo, and in vivo.
The appropriate use of these models is extremely relevant to wound-healing research as it
enables improved understanding of the basic mechanisms present in the wound-healing
cascade and aid in discovering better means to regulate them for enhanced healing or
prevention of abnormal scarring. The advantage of in silico models is that they can be used as
a first in virtue screening tool to predict the effect of a drug/stimulus on cells/tissues and help
plan experimental research/clinical trial studies but remain theoretical until validated. In vitro
models allow direct quantitative examination of an effect on specific cell types alone without
incorporating other tissue-matrix components, which limits their utility. Ex vivo models
enable immediate and short-term evaluation of a particular effect on cells and its surrounding
tissue components compared with in vivo models that provide direct analysis of a stimulus in
the living human subject before/during/after exposure to a stimulus. Despite clear advantages,
there remains a lack of standardisation in design, evaluation and follow-up, for acute/chronic
wounds and scars in all models. In conclusion, ideal models of wound-healing research are
desirable and should mimic not only the structure plus cellular and molecular interactions, of
wound types in human skin. Future models may also include organ/skin-on-a-chip with
potential application in wound-healing research.
2
INTRODUCTION
Many types of models exist that help to investigate the processes involved in normal
cutaneous wound healing in order to optimize the ultimate results of tissue repair. 1 These
models are also essential to explore the pathogenesis of chronic wounds and abnormal scar
formation, identify new drug targets and to test new therapeutics.2 One factor in choosing the
ideal model is to determine which type of wound and phase of healing is to be investigated.3
Thus, tissue repair models can be evaluated with respect to the following distinct stages of
wound healing: wound granulation tissue formation, contraction, re-epithelialization and scar
remodelling.3
Animal models, in vitro cell culture and tissue-engineered models have been used with
varying degrees of success to represent human skin.1 However, animal models are not perfect
as they do not always accurately represent the structure of human skin or develop similar
scars, which are comparable to abnormal scars observed in humans such as keloid scars.4,5 In
addition, the potential cost, ethical, and moral issues associated with their use can be
challenging and problematic.6-8 The best current animal model in terms of dermal structure
and underlying mechanisms is considered to be the pig.9-11 Nevertheless, no animal model is
shown to be an exact replicate of the clinical wound healing scenario seen in man.
Numerous models have been developed to study wound repair in humans which aim to
identify key underlying mechanisms and better understanding of the process of healing.
These include in silico, in vitro, ex vivo and in vivo models (Figure 1). In silico models can be
useful but lack the biophysical characteristics of human skin.7 In vitro models provide
valuable information about one or few components of the skin at a time, but do not correlate
with their in vivo counterparts due to their lack of complexity.6,12 Additionally, ex vivo models
enable immediate and short-term evaluation of a particular effect on cells as well as its
surrounding tissue components although there is a lack of standardisation and the in vivo
3
environment in toto.11 Furthermore, patient and volunteer studies remain essential and pivotal
to testing of emerging novel therapeutics but the appropriate methodology should be chosen
for the type of wound or scar being studied.6 In view of this, the aim of this review is to
evaluate currently available models of wound repair in human skin and offer a detailed
overview of their clinical and scientific relevance and utility.
MATERIALS AND METHODS
The available literature was reviewed regarding non-animal models of wound repair. The
PubMed and Scopus databases were used to search the recent literature to 2016. Different
combinations of terms were used, including models, wound repair, wound healing, tissue
repair, in vivo, ex vivo, in vitro, in silico, experimental, human. Articles not written in English
and those that did not include relevant information were excluded. Relevant articles
referenced within the articles identified with the above search were also included. All
retrieved articles were reviewed for their relevance to the specific topic.
4
RESULTS
The results will now be presented under the headings in silico, in vitro, ex vivo and in vivo
models.
In silico models
The complexity of the biochemical and biophysical processes involved in tissue regeneration
highlight the need for computational models in order to understand cell growth and to design
efficient scaffolds and tissue substitutes.13 Multiphase models have been used to describe
time-dependent processes in vitro in a perfusion bioreactor, with particular focus on cell
growth, access to nutrients, and scaffold degradation.14 Other computational methods have
modelled cell spreading and tissue regeneration in vitro using porous scaffolds. They looked
at transport and nutrient intake, cell population, ECM deposition, cell attachment and
migration.13,15,16 Agent based models have been used to investigate the role of growth
factors,17 organisation of keratinocytes,18 presence of fibroblasts,19 and the importance of stem
cells in long-term skin epithelium regeneration.20 In silico models (Figure 1) may be useful
theoretically and in combination with other models but are lacking the physical
characteristics of human skin.6
A number of models have been designed using differential equations.21-24 These models have
been developed to analyse strategies for improved healing, such as wound VACs,
commercially engineered skin substitutes and hyperbaric oxygen therapy. Mathematical
models using ordinary differential equations have been developed to investigate the effects of
commercially engineered skin substitutes. Waugh and Sherratt suggest that the model can be
used to explore the effects on interpatient variability or the gradual release of treatment
5
components.22 Models using partial differential equations have also been designed to analyse
the effects of hyperbaric oxygen on the promotion of wound angiogenesis.23,24 Both models
used the ‘snail-trail’ model of angiogenesis.25
A mechanical model using a finite-element method has also been used to analyse VAC
therapy and the simulation results are consistent with the in vitro strain levels shown to
promote cell proliferation.26 This approach may be used to evaluate treatment strategies that
promote cell proliferation, growth-factor production and wound angiogenesis by applying
micromechanical forces.
Computational approaches for deterministic systems in wound healing have been tested.
There have been analyses of changes in wound morphology using either planar or
axisymmetric wounds.27 A two-dimensional epidermal wound was used to assess the wound
boundary using a level-set method. Additionally, a further study used a similar method
analysing wound contraction using the finite-element method.28 Agent-based models have
been developed to analyse different treatment strategies with wound debridement and topical
administration of growth factor.21 Many studies have used agent-based models for a multi-
scale approach to wound healing.29-32
A multi-scale approach has also been developed for deterministic systems by using a hybrid
continuous-discrete model when studying the flow through a vascular network.33,34 Recently,
a continuum hypothesis-based model has also been used for the simulation of the formation
and regression of hypertrophic scar tissue after dermal wounding.35
6
In vitro models
There are several in vitro models of tissue repair that can assist with answering specific
mechanistic questions related to wound repair.36,37 Many of these in vitro models are used to
answer basic questions related to cell signalling in response to cell stress or injury.37 In vitro
cell culture models have been used for many years to gain insight into different aspects of
scar pathogenesis.36,37 In vitro models can be categorised as single cell, co-culture and
organotypic (Figure 2, Figure S1).
Single cell models work by the relevant cells being grown in vitro after which a ‘wound’ is
made in the cell layer by scraping the dish surface with either rubber devices or the tip of a
pipette, thus removing cells from the scraped area, creating a scratch and trauma to the cells
next to the wound.37 Subsequently, the rate at which cells divide and migrate into the ‘wound’
area is determined microscopically.37 The basic physiological processes are then analysed
including proliferation, protein production and secretion, viability, migration, gene
expression and differentiation.38 Single cell models are often used with dermal fibroblasts and
keratinocytes39-41 testing various agents for the enhancement of fibroblast migration or re-
epithelialisation.
In order to overcome the limitations of single cell models, several three-dimensional in vitro
models have been utilised.42.43 Fibroblasts are seeded in a type I collagen solution which is
solidified to gel, resulting in cells being embedded in a three-dimensional collagen lattice that
is then transferred to a culture dish and covered with medium.44 This model enables the
evaluation of cell contraction and matrix compaction. Another wound healing model45
embedded fibroblasts in a 3D collagen construct and the cell migration is followed from the
denser collagen matrix into a surrounding matrix. The limitations of this model include the
use of only one type of ECM protein and one cell type.
7
A number of recent quantitative studies have analysed in vitro wounding assays to investigate
aspects of cell migration for various cell types.46-50 Human in vitro models were used in a
study to investigate human skin integrity, wound closure and scar formation.51 These methods
included, cell culture, fibroblast and endothelial cell migration scratch assay, fibroblast
chemotaxis assay, endothelial cell in vitro tube formation assay and skin equivalent. The
authors showed that these models provided a platform for testing the mode of action of novel
compounds for enhanced and scar free wound healing. Two dimensional cell culture models
do not represent the interactions and mechanisms present in whole skin, such as the effect of
the extracellular matrix (ECM) in terms of structure, as well as cell signalling mechanisms
and the processes of metabolism.52
Indirect co-cultures of keratinocytes and fibroblast monolayers using transwell systems has
also enabled the study of keratinocyte–fibroblast interactions.53,54 The transwell assay or
Boyden chamber assay has been used to analyse the chemotactic responses of leukocytes.55
The principle of this assay is based on two mediums containing chambers separated by a
porous membrane.54 The transwell migration assay can be used for many different cell types
including epithelial,56 mesenchymal57 and brain cancer cell lines58 as well as many primary
cells. However, it is difficult to obtain the 3D macroscopic fibrotic tissue structure of a scar.
The expression of biomarkers from gene and protein studies is affected by the 3D structure of
a scar.37 The use of a more 3D environment such as collagen and mechanical load can
positively influence the behaviour of fibroblasts by creating a more realistic condition.59,60
One study cultured human cells derived from patients with chronic venous ulcers using
biopsies and demonstrated that they maintain their in vivo phenotype.61 Fibroblasts derived
from different wound locations were tested for their migration capacities and another portion
of the patient biopsy was used to develop primary fibroblast cultures after rigorous passage.
8
Another study also used chronic wounds for in vtiro assessment by taking punch biopsies
from patients with venous ulcers and evaluated their histology, biological response to
wounding (migration) and gene expression profile.62
Scratch assays are commonly used to observe the migration and proliferation of cells.63-65
Cells are placed on a culture dish and incubated, eventually forming a monolayer.66 An
artificial wound is created and images of the cell spreading by migration and proliferation are
captured over 12–24 h.67-69 Various cell types have been analysed for migration with this
assay including epithelial and mesenchymal cancer cells, keratinocytes, normal epithelial
cells, endothelial cells and fibroblasts.70-75 The majority of scratch assays falls into one of two
categories; wound assay or scrape assay. The wound assay involves the creation of a thin
wound,64,66 which produces two opposing directed cell fronts which merge.67 The scrape
assay, involves a monolayer that has been scratched further so there is only one cell type.67
Scratch assays have been used to investigate the influence of various chemical compounds
and potential drug treatments on the rates of cell motility and proliferation.63,65
When keratinocytes are cultured separately and then placed on top of the collagen gel
containing fibroblasts, an organotypic skin-like three-dimensional culture is created.76 This
model enables analysis of the interaction between these two types of cells in a three-
dimensional manner. Organotypic skin equivalents have been used to investigate scar
pathogenesis.77 3D skin equivalent models have used keloid fibroblasts in combination with
normal skin-derived keratinocytes.53,78 Using a similar method, a fully differentiated
epidermis constructed from keratinocytes isolated from hypertrophic scars on a fibroblast
populated dermal matrix showed characteristics of an abnormal scar including dermal
thickness, epidermal thickness and collagen I and illustrated the role of keratinocytes in
hypertrophic scar formation.79 Use of these models for testing therapies is limited due to the
lack of validated biomarkers and the dependence on excised scar tissue. Whilst in vitro
9
models have many benefits, they also have some limitations. The majority of cell culture
models employ serum to support and enhance the growth of cells in vitro.47,48 Serum is a
pathologic, inflammatory fluid compared to plasma and interstitial fluid that normal, resting
cells experience. Therefore, any observations made of cells in a serum environment should be
made with caution when assuming normal cellular function and behaviour.
Ex vivo models
Ex vivo human skin tissue is more appropriate for certain types of research. The use of whole
skin biopsies in culture allows the effect of individual ingredients and formulations to be
tested in an environment more closely mimicking normal skin.80 Full-thickness skin organ
culture is already an established tool used for understanding human skin pathophysiology and
evaluation of novel treatments.81-84 A number of wound healing organ culture models have
been previously reported in studying cutaneous wound healing processes.76,85,86 Therapies
including anti-inflammatory interleukin 10, anti-fibrotic streptolysin O and dermal substitutes
have been evaluated using this model.81,82,86 Despite promising results, there is a lack of
standardisation and conformity in these studies. Variations in the methodologies include the
use of different wound types such as a partial or full thickness wound, different culture
conditions, and a range of support and growth mediums.83,87 Ex vivo model types can be
divided into normal skin models and pathological skin models (Figure 3, Figure 4).
A study looking at the effects of topical formulations on human skin demonstrated that
altered expression of key gene and protein markers could be quantified in an optimised whole
tissue biopsy culture model.1 This model could be adapted to study a range of compounds or
10
disease mechanisms, negating the requirement for animal models, prior to study in a clinical
trial environment.
The ex vivo donut-shaped wound healing model has been used to investigate human
cutaneous repair.88,89 In a study by Sebastian et al, human tissue was collected and wounded
using a standardised punch biopsy (full thickness wound) and the effect of electrical
stimulation was evaluated.88 Another study also used this model for studying the effect of
dermal substitutes on human dermal fibroblasts and keratinocytes.89 Full thickness lower
human patient abdominal skin was collected and 8mm punch biopsies taken. In the centre of
these cylindrical skin biopsies, a 4mm full thickness artificial wound was created by a further
punch biopsy. Models were placed into 24-well plate inserts and prepared dermal substitutes
were inserted into the wound area. This model has been shown to be useful in studying
cutaneous wound healing.
Ex vivo models have been used when analysing stretch marks and scars including
hypertrophic and keloid scars.90-92 Multipotent keloid-derived mesenchymal-like stem cells,
found in the scar, have been implicated in keloid formation.91,92 Therefore, keloid explant
models are interesting as they allow these cells to remain in their location. Ex vivo biopsies
have been cultured at air–liquid interface, embedded in collagen gels.93,94 The functionality of
this model was confirmed by the reduced epidermal thickness and scar volume after
treatment with the dexamethasone. A further study evaluated the effect of two promising
candidate anti-fibrotic therapies: (-)-epigallocatechin-3-gallate and plasminogen activator
inhibitor-1 (PAI-1) silencing in a long-term human keloid organ culture (OC).95 This model
has several advantages; the keloid explants remain viable in culture for up to 6 weeks without
tissue deterioration, the maintenance of the keloid cellular and microenvironments enable
examination of keloid pathophysiology in situ and anti-fibrotic agents and gene silencing
using RNA interference technology can be used to investigate the cellular and molecular
11
interactions in the epidermis and dermis.94 The results showed that these therapies inhibited
growth and induced shrinkage of human keloid tissue and found this to be an effective
model.95 Another study assessed the extent to which differences relating to cell type influence
PAI-1 gene regulation with regard to growth state-associated mechanisms of expression
control using keloid fibroblasts and normal dermal fibroblasts for cell culture.96
Another ex vivo study utilised an organ culture model of dermal fibrosis to investigate the
effect of photodynamic therapy on skin scars and stretch marks in humans.90 This unique ex
vivo model showed the morphological and cellular effect of photodynamic therapy and
correlated this with the degree and severity of dermal fibrosis.
In vivo models
Human in vivo models are useful as the pathology and physiology of healing (in particular in
acute wounds) is identical to that found in the patient. There are a number of wound models
that can be inflicted upon a human patient or volunteer to provide accurate and representative
research tools (Figure 5, Figure 6). Due to the difficulty in obtaining suitable volunteers with
chronic wounds and incorporating them into a study, human acute wound models provide the
best opportunity to understand the wound healing process and the performance of different
products that aid and promote wound healing. Both patient and volunteer models have their
advantages and limitations. Patient models allow assessment in a variety of wound or scar
12
types, whilst volunteer models only enable evaluation of one type of wound/scar. This is
significant when when interpreting and analysing the results of these studies.
Patient model
Patient studies remain essential and are always necessary to validate potential novel wound
and scar therapies identified in other wound and scar models. Patient models are useful as any
wound type can be assessed using a variety of methods including incisions, excisions and
sequential punch biopsies (in non-keloid formers). However, chronic wound studies have
limitations due to the heterogeneity of the population and lack of uniformity of the wound.
Patients with scars can be used to explore the pathogenesis of scar formation, however, keloid
formers are rarely included due to concerns for recurrence and worsening of existing scars.
Incisional wound models are useful as they can be accurately reproduced and can be used for
multiple investigations. This model can evaluate the wound healing process and the influence
of different local compounds/dressings, response to energy based therapies, as well as
systemic effects of drugs. This model has been used extensively following cutaneous surgery
in order to evaluate quantitative outcome measures including rate of wound closure, quality
of healing as well as wound strength.97,98 Biomechanical analysis of soft tissue and incisional
wounds in different types of tissues have also been described.99
Acute wounds such as split-thickness donor sites have the benefit in that two wounds can be
created in a controlled fashion in the same patient. In a randomized, blinded fashion, one
wound can be treated with the therapy and the other, the control which is the optimal study
design.100,101 However, these wounds heal fast, therefore, it is difficult to demonstrate a
clinically significant increase in healing rate.
13
Volunteer models
Human volunteer models enable assessment of a homogeneous population and the volunteers
can act as their own controls. The limitations of these models are the inability to create other
wound or scar types such as chronic wounds or keloid scars or study different conditions. A
number of volunteer models have been utilised including excisional, incisional, scratch,
blister, tape stripping and burns.
Biopsy models have been used which enable a standardised approach as the subjects can act
as their own controls. These wounds are useful from an experimental perspective in that their
size and depth can be precisely controlled. As they involve all the components of the healing
process, they provide an excellent research tool and have been used to investigate
angiogenesis, wound contraction, and wound closure.102 Numerous studies have been
conducted evaluating various treatments for both wounds and scars using a sequential skin
punch biopsy model to assess various characteristics and correlated by clinical
measurements.102-107 Furthermore, studies have also monitored treatment efficacy in raised
dermal scarring including hypertrophic and keloid scars using objective non-invasive
measures.108-110 However, difficulties arise when attempting to experimentally monitor the
tissue in particular in keloid scars as we would be unable to biopsy the tissue as this would
lead to further abnormal scarring. For these in vivo studies, it is important to clarify the
definition of healing in the human wound. The endpoint should be identified (when the
wound has healed), and the determination of scar formation. The advantages of this model are
that it uses a wound with loss of tissue as found in clinical practice. These wounds enable the
evaluation of different therapies with the ability to create the same wounds each time and
measure different healing parameters quantitatively. Early healing can be monitored followed
14
by scarring and surrounding wound area can be observed. The disadvantage is that the wound
heals from the edges as well as by contraction.
A study by Dunkin et al investigated the association between scarring and depth of dermal
injury. They developed a novel device, which produced a wound that was deep dermal at one
and superficial at the other to healthy volunteer hips. They showed that this dermal scratch
model provided a standardised and reproducible model for investigating the healing response
to dermal injury of different depths.121
The blister model is used for the evaluation of epidermal regeneration and the influence of
various therapies. Blisters are produced by suction using different types of devices which
produce standardised, small, epidermal bulges.111,112 Normal human skin tends to blister after
35-55minutes and a scalpel can then remove the blister roofs, therefore, creating identical
superficial wounds of similar dimensions.113 Absoprtion effect can also be evaluated under
occlusive and semi-occlusive conditions and the trans-epidermal water loss can be measured
for the skin barrier function.114 The advantages of this model are that the wounds are identical
and can be in the same anatomical location on the body. The disadvantages are that
evaluation of such superficial wounds may have no relevance to much deeper dermal wound
types.
The tape stripping model involved successive stripping of the epidermis using adhesive tape
in order to disintegrate the skin barrier in the lowest part of the stratum corneum.115 This can
then be measure by a trans-epidermal water loss measurement device.114 This model is less
damaging to the epidermis than the blister model. A study used the tape stripping model to
assess the penetration of quantum dots.116 Adhesive tape was applied and pressed onto the
skin using a roller. The skin was stripped with 15 pieces of adhesive tape. Negative control
15
strips were obtained from a non-treated area of the upper arms. Tape strips removed from
skin were analyzed using confocal microscopy.
There have been studies that have created a burn in healthy volunteers to assess various
therapies. A randomised double-blind cross-over study in six volunteers investigated the
effects of intravenous lidocaine infusion on partial thickness skin burns.117 A thermoprobe
was used to induce a standardised thermal injury (1cm2) on the flexor side of one forearm and
was repeated on the opposite side 1 week later. Another study directly compared UVB
induced inflammation and the more established thermal burn and topical capsaicin pain
models.118 They delivered UVB to small areas of volar forearm skin in healthy volunteers and
found that the degree of inflammation and concomitant increase in sensitivity to cutaneous
stimuli were UVB dose and time dependent.
DISCUSSION
This review has evaluated currently available wound repair models in human skin, including:
in silico, in vitro, ex vivo and in vivo. The appropriate use of tissue repair models is extremely
relevant to wound healing research as it enables improved understanding of the basic
mechanisms behind the wound healing cascade and aid in discovering better means for
enhanced healing or prevention of abnormal scarring.6
16
The advantage of in silico models is that they can be used as a first in virtue screening tool to
predict the effect of a drug/stimulus on cells/tissues and help plan experimental
research/clinical trial studies but remain theoretical until validated.7 Additional limitations of
the in silico mathematical model is the lack of standardisation which is highlighted by the
arbitrary choice of specific healing parameter values use in these models.7,8 If the aim of the
model is to provide a quantitative prediction of a physiological process, the model parameters
require better quantification.13 In vitro models allow direct quantitative examination of a
particular effect on specific cell types alone without incorporating other tissue matrix
components, which limits their utility.12 Additionally, in vitro models have had a significant
role in the study of wound healing and scar formation.36,37 This had led to a better
understanding of biomarker expression, the role of keratinocytes and fibroblast activity in
wound and scar development.46-51 Although the disadvantage is that, it can be difficult to
extrapolate the findings of in vitro research to the in vivo wound scenario. Three dimensional
models including extracellular matrix, human skin constructed from keloid-derived tissue and
mesenchymal cells, have been recently made to mimic the conditions upon which these scars
form.91,92 Ex vivo models enable immediate and short-term evaluation of a particular effect on
cells and its surrounding tissue components compared with in vivo models that provide direct
analysis in the live human subject before, during and after exposure to a stimulus.12 Human in
vivo models are useful as the pathology and physiology of healing is identical to that found in
the patient.102 Both patient and volunteer models have their advantages and limitations.
Patient models allow assessment in a variety of wound or scar types, whilst volunteer models
only enable evaluation of one type of wound or similarly induced scar.102,108-110 Despite clear
advantages, there remains a lack of standardisation and conformity in design, evaluation and
follow-up, for all wound types including acute, chronic wounds and scars.
17
Many of the human wound and scar models have a limited duration of days or weeks,
whereas human scars develop over a period of months or years.52,53 Therefore, these models
enable us to look at the early stages of wound healing and scar formation but may not
accurately reflect the later stages of wound contraction or scar formation. Another limitation
is that a number of cell types that are involved in the healing process have not yet been
incorporated into the relevant human cell culture models.119,120
An emerging model is the organ-on-a-chip or skin-on-a-chip which involves engineered
tissues, which are similar to the in vivo equivalents and consist of multiple different cell types
interacting with each other under closely controlled conditions and are grown in a
microfluidic chip.121-123 These controlled conditions will make it possible to mimic the
environment of the skin (humidity, temperature, pH, oxygen levels), the elasticity of the skin,
and the complex structures and cellular interactions within and between the different cell
types.122
In summary, understanding the mechanism and pathophysiology of how scars form and
wounds heal, is essential if we are to develop optimal strategies in the future to prevent
adverse scar formation and prolonged healing. Ideal models of wound healing research
should mimic not only the structure but also the cellular and molecular interactions, of wound
types in the human skin.
Sources of funding: No sources of funding to declare.
Conflict of interest: No conflict of interest to declare.
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Figure Legends
Figure 1: Diagram summarising the four main categories for human models of wound repair
including; in silico, in vitro, ex vivo, in vivo. Within each of these domains the assays are
shown and the types of wound models.
Figure 2: In vitro models. Flowchart depicting the three types of in vitro models; single cell,
co-culture and organotypic.
Figure 3: Ex vivo models. Flowchart depicting the two categories of ex vivo models; normal
skin (full-thickness and partial thickness) and pathological skin (stretch marks, scars,
hypertrophic scars and keloid scars).
Figure 4: Ex vivo assays. A diagram to demonstrate the assays including angiogenesis,
apoptotic, proliferation, fibrotic and rate of epithelialisation.
Figure 5: In vivo models. Flowchart depicting the two categories of ex vivo models and the
types of wounds used; patients (surgical wounds, incisional wounds, excisional wounds,
donor site wounds) and human volunteers (Incisional wounds, excisional biopsy wounds,
scratch, blister tape stripping and burns).
Figure 6: In vivo assays. A diagram to demonstrate the assays including invasive (gene and
protein studies) and non-invasive devices and those used for specific healing parameters.
Supplementary Figures
Supplementary Figure S1: In vitro assays. A diagram to demonstrate the different assays for
migration, invasion proliferation and senescence.
38
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