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Biodegradable soy wound dressings withcontrolled release of antibiotics: Results froma guinea pig burn model
Dana Egozi a,b, Maya Baranes-Zeevi c, Yehuda Ullmann b,d, Amos Gilhar b,Aviad Keren b,d, Elias Matanes d, Israela Berdicevsky b, Norberto Krivoy b,Meital Zilberman c,*
aDepartment of Plastic Surgery, Kaplan Medical Center, Rehovot, Israelb Faculty of Medicine, Technion – Israel Institute of Technology, Haifa 32000, IsraelcDepartment of Biomedical Engineering, Tel-Aviv University, Tel-Aviv 69978, IsraeldDepartment of Plastic Surgery and the Burn Unit, Rambam Health Care Campus, Haifa, Israel
b u r n s 4 1 ( 2 0 1 5 ) 1 4 5 9 – 1 4 6 7
a r t i c l e i n f o
Article history:
Accepted 27 March 2015
Keywords:
Gentamicin
Infection
Histology
Soy protein
Wound healing
a b s t r a c t
There is growing interest in the development of biodegradable materials from renewable
biopolymers, such as soy protein, for biomedical applications. Soy protein is a major fraction
of natural soybean and has the advantages of being economically competitive, biodegrad-
able and biocompatible. It presents good water resistance as well as storage stability. In the
current study, homogenous antibiotic-loaded soy protein films were cast from aqueous
solutions. The antibiotic drug gentamicin was incorporated into the films in order to inhibit
bacterial growth, and thus prevent or combat infection, upon its controlled release to the
surrounding tissue. The current in vivo study of the dressing material in contaminated deep
second-degree burn wounds in guinea pigs (n = 20) demonstrated its ability to accelerate
epithelialization with 71% epithelial coverage compared to an unloaded format of the soy
material (62%) and a significant improved epithelial coverage as compared to the conven-
tional dressing material (55%). Our new platform of antibiotic-eluting wound dressings is
advantageous over currently used popular dressing materials that provide controlled
release of silver ions, due to its gentamicin release profile, which is safer. Another advantage
of our novel concept is that it is based on a biodegradable natural polymer and therefore
does not require bandage changes and offers a potentially valuable and economic approach
for treating burn-related infections.
# 2015 Elsevier Ltd and ISBI. All rights reserved.
* Corresponding author at: Department of Biomedical Engineering, Faculty of Engineering, Tel.:-Aviv University, Tel-Aviv 69978, Israel.Tel.: +972 3 6405842; fax: +972 3 6407939.
E-mail address: [email protected] (M. Zilberman).
Available online at www.sciencedirect.com
ScienceDirect
journal homepage: www.elsevier.com/locate/burns
http://dx.doi.org/10.1016/j.burns.2015.03.0130305-4179/# 2015 Elsevier Ltd and ISBI. All rights reserved.
b u r n s 4 1 ( 2 0 1 5 ) 1 4 5 9 – 1 4 6 71460
1. Introduction
1.1. Soy protein
There is growing interest in the development of biodegradable
materials from renewable biopolymers, such as soy protein,
for biomedical applications [1–7]. Soy protein is a major
fraction of natural soybean and can be found in several
soybean derivatives in different quantities, depending on the
extraction method: defatted soy flour contains 50–59% protein,
soy protein concentrate contains 65–72% protein and soy
isolate contains more than 90% protein [8]. Soy protein has
been explored mainly in the polymer, food and agriculture
fields. Use of soy protein as a food source is still increasing, due
to its functional and nutritional value, availability and low
price [9]. In the materials industry, soy protein was studied as
an adhesive and as a ‘‘green’’ plastic [1]. Soy protein is an
abundant plant protein and has the advantages of being
economically competitive, biodegradable and biocompatible.
It presents good water resistance as well as storage stability
[4]. Soy protein is also very versatile and its properties can be
tailored by physical, chemical, or enzymatic treatments [10],
such that it can provide diverse requirements for different
biomedical applications. It is known that local or systemic use
of soybean or its components has an Ig E induced immuno-
genic effect more in children and less in adults, but in spite of it
the soybean is used for the preparation of an intravenous
medication widely used in surgery. As always, the public
should be informed in the package insert. The statistics says
that soybean affects approximately 0.4% of the children’s
population. Also, approximately 50% of children with soy
allergy outgrew their allergy by age of 7 years. The soy protein
extract used in the current study was provided by a company
which sells it for the food industry and therefore it is
considered as safe.
The suitability of soy protein for biomedical applications
has been investigated, due to its low cost and bioactive
properties [11]. Soy thermoplastics were found to be non-
cytotoxic, and even encouraged cell proliferation during
in vitro tests [12]. Proposed applications range from bone
cement to hydrogel and membranes for wound-dressing
applications. The most promising recent applications include
drug delivery carrier films, temporary replacement implants
and scaffolds for tissue engineering [13].
Snyders et al. [14] and Shingel et al. [15] investigated hybrid
hydrogels made of poly(ethylene glycol) and soy protein for
moist wound-dressing applications and concluded that these
hydrogels can be considered as a safe, biocompatible and
inflammatory inert wound-dressing material. Santos et al. [16]
developed chitosan/soy (cht/soy)-based membranes as
wound-dressing materials and showed that these new
membranes possess the desired features for healing/repair
stimulation, ease of handling, and final esthetic appearance,
which are valuable properties for wound dressings.
1.2. Wound dressings
Various types of wounds result in tissue damage. These
include burns, pressure ulcers, diabetic ulcers and trauma.
The main goal in wound management is to achieve rapid
healing with good functional and esthetic results. An ideal
wound dressing can restore the milieu required for the healing
process, while simultaneously protecting the wound bed
against bacteria and environmental threats. The dressing
should also be easy to apply and remove. Most modern
dressings are designed to maintain a moist healing environ-
ment and to accelerate healing by preventing cellular
dehydration and promoting collagen synthesis and angiogen-
esis [17]. However, over-restriction of water evaporation from
the wound should be avoided, since accumulation of fluid
under the dressing may cause maceration and facilitate
infection. The physical and chemical properties of the
dressing should therefore be adapted to the type of wound
as well as to the degree of wound exudation.
Bacterial contamination of a wound seriously threatens its
healing. In burns, infection is the major complication after
the initial period of shock, and it is estimated that about 75%
of the mortalities following burns are related to infections
rather than to osmotic shock and hypovolemia [18]. This has
encouraged the development of improved wound dressings
that provide an antimicrobial effect by eluting germicidal
compounds such as iodine (Iodosorb1, Smith & Nephew),
chlorohexidine (Biopatch1, J&J), or, most frequently, silver
ions (e.g., Acticoat1 by Smith & Nephew, Actisorb1 by J&J and
Aquacell1 by ConvaTec). Such dressings are designed to
provide controlled release of the active agent through a slow
but sustained release mechanism which helps avoid toxicity
yet ensures delivery of a therapeutic dose into the wound.
1.3. About the current study
It can be said that there is an increasing need to develop new
biodegradable materials for use in wound-healing applica-
tions. Soy protein is a very promising natural biodegradable
material, due to the abovementioned advantages. We there-
fore chose to develop and study soy protein films as a platform
for wound-dressing applications, in all relevant aspects.
We believe that a high quality soy protein film which is
hydrophilic but also cross-linked and therefore exhibits
desired relevant properties may provide properties that are
usually achieved using the bilayer concept.
Homogenous antibiotic-loaded yellowish films were cast
from aqueous solutions. A detailed description of the
preparation process was published elsewhere [19]. The
antibiotic drug gentamicin was incorporated into the films
in order to inhibit bacterial infection upon its controlled
release to the surrounding tissue. The mechanical and
physical properties of these films, the antibiotic release
profiles and their antimicrobial effects as well as cellular
response results, were recently published by us [19,20]. Since
these films are crosslinked and also contain plasticizer
(glycerol), they are strong and flexible and can be easily
handled. The new gentamicin-eluting soy protein wound
dressing which exhibited the best properties in the above
study was selected for the current in vivo study and compared
to a neat soy protein wound dressing without the drug and to
a standard Melolin dressing.
The guinea pig is often used as a dermatological and
infection model [21–24]. Research on guinea pigs has included
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topical antibiotic treatment [25], delivery of delayed-release
antibiotics [26], and investigation of wound-dressing mate-
rials [27,28]. A deep partial skin thickness burn is an
excellent wound model for the evaluation of wound healing,
not only for contraction and epithelialization from the
peripheral area such as in third-degree burns, but also for
evaluation of the recovery of skin appendages that serve
as the main source for the re-epithelialization which
completes the healing process. The metabolic response to
severe burn in guinea pigs is very similar to the human post-
burn metabolic response [29]. Furthermore, bacterial coloni-
zation and changes within the complement component
of the immune system in human burn victims is analogous
to guinea pigs affected by severe burns [23]. Such a model
was therefore used in the current study to evaluate the
effectiveness of our novel composite antibiotic-eluting
wound dressing.
2. Materials and methods
2.1. Materials
Non-genetically modified soy protein (Solpro 910TM, minimum
90% (w/w) protein, on a dry weight basis) was obtained as a
donation from SolbarTM (Ashdod, Israel).
Glycerol (G-7893) was used as plasticizer, glyoxal (50650)
was used as the cross-linking agent and the antibacterial drug
gentamicin sulfate (G-1264) (450–477 g/mol, Tm = 218–237 8C)
was used as the incorporated active agent (all purchased from
Sigma–Aldrich, Rehovot, Israel).
Reagent kit (8-1P31-25) and calibration kit (8-1P31-01) were
purchased from Sigma–Aldrich, Rehovot, Israel for analysis of
gentamicin concentrations.
2.2. Preparation of the soy protein films
Films of soy protein were prepared using the solvent casting
method. Soy protein solution was prepared by dissolving soy
protein powder in preheated double distilled water at 55 8C
and thoroughly stirred. Plasticizer (glycerol) and cross-linker
(glyoxal) were then added. For the gentamicin-loaded films,
gentamicin was first dissolved in 10 mL of the preheated
double distilled water and added to the soy protein solution
(following the addition of the cross-linker). All film parameters
are presented in Table 1.
The solution was constantly stirred with a magnetic stirrer
and kept at 55 8C for 30 min to allow the removal of bubbles.
The solution was then stirred at room temperature for an
additional 30 min to enable further removal of bubbles and
initial cross-linking. Finally, the solution was cast into two
Table 1 – Parameters of the studied films.
Parameter Value Relative to
Soy protein 5% (w/v) DDW
Plasticizer (glycerol) 35% (w/w) Soy protein
Cross-linker (glyoxal) 1% (w/w) Soy protein
Gentamicin 0% or 3% (w/w) Soy protein
low-density polyethylene plates (8.74 cm diameter) and left to
dry under ambient conditions for 72 h. In order to maintain
uniform thickness of the films, exactly 45 mL of solution were
cast into each plate. This amount yielded films with a
thickness of approximately 0.5 mm.
When dried, the films were carefully filed off the dishes and
cut into 45 mm discs. Each disk was then positioned between
two glass Petri dishes and subjected to thermal treatment in
an oven for 24 h at 80 8C. The specimens were kept in a
desiccator (RT, 30% RH) at room temperature and constant
humidity until use. All specimens were sterilized with
ethylene oxide and sealed in an airtight plastic bag.
2.3. In vitro gentamicin release
The soy protein films were cut into round discs (1 cm
diameter), which were individually placed in glass tubes in
triplicates and 3 mL of the release medium were added to each
tube. The tubes with the immersed discs were sealed with
plastic cups and kept in a microbiological incubator (Heraeus,
B12) at 37 8C and 100% humidity under static conditions. The
entire release medium from each tube was removed at
predetermined time points and was replaced by fresh medium
(3 mL). The removed medium was kept refrigerated at 4 8C
until analyzed.
Immunoassay analysis: Gentamicin concentrations were
determined using a high-speed immunoassay with an
ARCHITECT i2000SR (Abbott) system. Samples were diluted
with PBS (pH 7) with 0.02% sodium azide, as necessary.
2.4. In vivo burn model: procedure and treatment groups
Twenty female guinea pigs (Harlan, NL) were allocated for the
study after authorization by the Technion Animal Care and
Use Committee (authorization IL-092-09-2012). The animals
(average weight 300 g) were raised in a pathogen-free animal
facility. Following five acclimatization days, the animals were
randomly divided into three groups. All animals were
anesthetized by intramuscular injection of ketamine (40–
50 mg/kg) and xylazine (4–5 mg/kg). After shaving the animals’
skin, a depilatory cream (Orna 19, Alpha Cosmetica Israel) was
applied to complete hair removal. Two standardized deep
second-degree burns were inflicted on the back of each animal
on either side of the spine according to a validated method
described by Kaufman et al. (1990). Iron templates (circle,
D = 40 mm) were immersed in water preheated to 75 8C and
then placed in perfect contact with the animal’s skin for
exactly 5 s by applying light pressure. The burn area was
traced onto a transparent paper as a reference for later follow-
up. After infliction of the burns, each animal was seeded with
0.5 mL broth containing 1 � 108 CFU/mL Pseudomonas aerugi-
nosa using a micropipette. Each group was then treated with
the relevant treatment option, as follows:
Group 1 (6 animals, 12 wounds) was treated with a neutral
non-adherent dressing material (Melolin1, Smith & Nephew).
Melolin1 consists of three layers: a low adherent perforated
film, a highly absorbent cotton/acrylic pad and a hydrophobic
backing layer. According to the manufacturer, it enables rapid
drainage of wound exudate, thus reducing trauma to the
healing tissue. The dressing was placed directly on the burn,
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and was secured by an elastic adhesive bandage (Tenso-
plastTM, Smith & Nephew).
Group 2 (7 animals, 14 wounds) was treated with our soy
dressing, without antibiotics. A round disk slightly larger than the
burn area (D �45 mm) was placed directly on the burn, covered
with Melolin1, and secured as described above. This dressing
material was tested in order to evaluate the effect of the dressing’s
texture,materials,anddegradationonthewound-healingprocess.
Group 3 (7 animals, 14 wounds) was treated with the soy
dressing which released gentamicin. The dressing materials
were placed and secured as described above. These dressing
materials were tested in order to evaluate the effect of
antibiotic release kinetics on the wound-healing process.
Each of the animals was placed in an individual cage with
food ad libitum and allowed to recover. The animals received
analgesic treatment prior to the procedure and for 5
consecutive days (buprenorphine, 5 mg/kg per os).
2.5. Wound closure
Twelve days following the burn, wound areas were traced on a
transparent paper and scale bar photographs were taken. The
burn wounds were evaluated macroscopically by two quanti-
tative parameters: (i) percentage of the original area subjected
to burn which was still an open wound, and (ii) wound
contraction as depicted by the total wound area (epithelialized
and non-epithelialized) as a percentage of the original area
subjected to burn. The measurements of the two burns within
one animal were averaged to produce a single data, from
which the total group average was calculated.
2.5.1. Post mortem examinationThe animals were sacrificed and 1 cm2 biopsies were taken
from the center of the wound and immediately fixed in
phosphate buffered formalin.
2.5.2. Biopsy evaluationsFollowing the fixation stage, biopsies were embedded in
paraffin using a standard embedding protocol, and 5 mm
sections were prepared using a Leica microtome. Sections
were stained with hematoxylin and eosin (H&E), observed and
photographed under �20 and �100 magnification using an
Olympus upright light microscope. Wound-healing analysis
was conducted in a blinded manner by two separate
evaluators using a semi-quantitative grading system. The
data obtained from the observers was analyzed and based on
the average of the two burns within one animal to produce a
single observation per animal. The sections were evaluated
based on structure and content. The wound-healing criteria
included epithelial coverage, epidermal thickness, epidermis-
dermis attachment, neoangiogenesis, number of follicles
(adnexa), dermis thickness, white blood cells (WBC) and
collagen. Grading was between 0 and 3: 0 absence, 1 minimal
presence, 2 moderate presence and 3 extensive presence,
except for WBC, where 0-extensive presence and 3-absence.
2.6. Statistical analysis
Means and standard deviation of sample (SD) were calculated
for each data set. Differences between means were analyzed
for statistical significance using a one-way ANOVA with the
Tukey-Kramer multiple comparisons posttest (SPSS version
17.0). p values �0.05 were considered significant.
3. Results
3.1. In vitro gentamicin release profile
Cumulative release of gentamicin from the soy protein wound
dressings presented an initial burst release (6 h) of approxi-
mately 54% followed by two phases of release. The first phase
exhibited a decreasing release rate for a period of 12 days,
followed by a second phase of a constant slow release rate of
0.15% drug/day (R2 = 0.9685) for more than 40 days (Fig. 1).
3.2. Wound closure
Second-degree burns in guinea pigs were used as a wound-
healing model for testing our novel soy wound dressing, which
is a membrane that covers the wound and allows slow release
of gentamycin. Pseudomonas was applied topically immedi-
ately after the creation of the burns in order to bypass the
sterile conditions of the pathogen-free room and to mimic
burn contamination that typically occurs in patients with
burns.
Twenty guinea pigs were included in the study and were
divided into three groups: Melolin-control group, soy protein
dressing without antibiotic, and soy protein dressing with
controlled release of gentamicin (Fig. 2). The closed (epithe-
lialized) wound area and the open (non-epithelialized) wound
area were traced on a transparent paper 12 days after creation
of the burns. The degree of healing was calculated by the
percentage of the epithelialized (closed) area on the 12th day
compared to the total burn area on the 12th day. On the 12th
day post-burn, epithelialization of the wounds that were
dressed with Melolin covered 55% � 15% of the burn area, the
average closure of the soy protein group was 62% � 13%,
whereas the closure area of the soy protein group with
gentamicin release reached 71% � 11% (Fig. 3A). The differ-
ence in epithelialization and wound closure between the soy
gentamicin group and the Melolin group was statistically
significant ( p = 0.04), while no significant difference was noted
between the soy group and the Melolin group.
3.3. Wound contraction
Wound contraction was calculated as 1 minus the total wound
area (epithelialized and non-epithelialized) divided by the
original area subjected to burn. A significant reduction was
found in the contraction rate between the soy treated groups
(36% � 8%, p < 0.02 for soy only, and 31% � 10%, p < 0.005 for
soy with gentamicin release) as compared to the Melolin group
(51% � 12%) (Fig. 3B).
3.4. Histological evaluation
Animals were sacrificed after 12 days, and 1 cm2 biopsies
were taken from the center of the wound and immediately
fixed in phosphate buffered formalin. Two different observers
Fig. 1 – In vitro cumulative release (mean W SD) of gentamicin during (A) 56 days and (B) the first 12 h of release. Second
phase of continuous release (14–56 days) was fitted to a linear line (C) to demonstrate a constant release rate.
Fig. 2 – Representative photographs of wounds 7 and 12
days post-treatment with MelolinW (A1 and A2,
respectively), soy dressing without antibiotics (B1 and B2,
respectively) and soy dressing with controlled gentamicin
release (C1 and C2, respectively). Bars = 1 cm.
b u r n s 4 1 ( 2 0 1 5 ) 1 4 5 9 – 1 4 6 7 1463
examined the biopsies that were stained with H&E (Fig. 4A–C),
according to the abovementioned criteria. The gentamicin
soy group was significantly superior to the other two groups
( p < 0.05) (Fig. 4D): better epithelialization and neoangiogen-
esis, thicker dermis, fewer WBC, more follicles, and greater
collagen content. The general scores differed significantly for
the gentamicin release soy dressing (10.8 � 1.4) versus the soy
and the Melolin treatment groups (7.2 � 1.7; 7.2 � 1.2, respec-
tively, p < 0.05). Wounds treated with controlled release of
gentamicin obtained significantly higher histological scores
than both controls after 12 days. This was mainly due to
significant re-epithelialization of the wound ( p = 0.02), better
epidermis-to-dermis adherence (p = 0.02), amount of collagen
(p = 0.01) and more skin appendages ( p = 0.03). The differ-
ences in the amount of neoangiogenesis, WBC and the
thickness of the dermis were not significant.
Furthermore, the soy dressing material was found to create
good contact with the wound, and the soy itself did not cause
any irritation to the animal’s skin.
4. Discussion
4.1. Gentamicin release profile
Commercial wound dressings that incorporate iodine (Iodo-
sorb1 by Smith & Nephew), chlorohexidime (Biopatch1 by J&J)
Fig. 3 – (A) Percentage of wound healing at 12 days post-
burn creation. (B) Wound contraction as percentage of total
measured wound area (epithelialized and non-
epithelialized) at 12 days post-burn creation relative to the
wound area at the time of wound creation. Each dot
represents the average of two burns on each animal
(mean W SD). Square (&) represents measurements of
superimposed dots from two different animals. Triangle
(~) represents measurements of superimposed dots from
three different animals. *Statistically significant as
compared to Melolin treated group.
b u r n s 4 1 ( 2 0 1 5 ) 1 4 5 9 – 1 4 6 71464
or most frequently silver ions (e.g., Acticoat1 by Smith &
Nephew, Actisorb1 by J&J and Aquacel1 by ConvaTec) as
active agents are available in the market. Such dressings are
designed to provide controlled release of the active agent
through a slow but sustained release mechanism which helps
avoid toxicity yet ensures delivery of a therapeutic dose to
the wound. Despite frequent usage, there is growing evidence
that silver is highly toxic to keratinocytes and fibroblasts and
may delay burn wound healing if applied indiscriminately
to healing tissue areas [30–32]. Furthermore, most of these
dressing materials undergo frequent changes. Bioresorbable
dressings may successfully address this shortcoming, since
they do not need to be removed from the wound surface once
they have fulfilled their role.
In our previous study of the soy protein wound dressings
loaded with gentamicin we in vitro studied the antimicrobial
effect of gentamicin release profiles on bacterial inhibition
and also evaluated the cytotoxicity on fibroblast cells. Our
main conclusions were that the gentamicin release from our
soy films demonstrated great efficiency toward the relevant
bacteria, which are abundant on human skin [20]. The films
could effectively inhibit S. aureus and S. albus infections for at
least two weeks and P. aeruginosa for three days. Filtered film
extracts released during the first 24 h and from the 24–48 h
showed no cytotoxic effect on the fibroblast cells, although the
gentamicin quantities released were much higher than these
of silver ion that are used in commercial wound dressings [20].
Based on these results, a selected soy protein film was used
in the current animal study.
The in vitro gentamicin release profile from our selected
film showed a moderate burst effect (54%) during the first 6 h,
followed by a continuous decrease in release rates during the
first two weeks. This was followed by a third stage of zero-
order release (constant release rate) that lasted until the end
of the experiment (8 weeks). It should be mentioned that the
specimens maintained their integrity throughout the entire
test period. The burst effect and release profile during the first
two weeks were typical of diffusion-controlled systems. The
third phase of constant release rate presumably involved
some degradation of the soy protein matrix combined with
diffusion of the remaining drug that was more firmly attached
to the protein chains.
The obtained release profile could be beneficial for
application as antibiotic-eluting wound dressings. During
the first hours after being wounded, a relatively high drug
release is essential to eliminate infections that may not have
been eliminated during wound cleansing. Later, a continuous
low release rate can keep the wound ‘‘infection free’’ for more
than 2 weeks, which is the time usually taken for wound
healing.
It should be noted that most of the gentamicin (75%) was
released within the first week of the study, due to the
hydrophilic nature of the antibiotic and the soy protein matrix.
The hydrophilic nature of soy protein enables relatively rapid
water intake, leading to full swelling of the matrix within a few
hours of immersion. Unfavorable more rapid drug release
rates have been reported in the literature for other antibiotic-
eluting systems [33]. Controlling the release of antibiotics from
these systems is challenging due to the hydrophilic nature of
both the drug and the host polymer. In most cases the drug
reservoir is depleted in less than 2 days, resulting in a very
short antibacterial effect. Thus, our new antibiotic-eluting soy
protein wound dressing is advantageous over other systems.
The obtained gentamicin release profile can be explained by
the fact that many antibiotic drugs bind proteins via van der
Waals or ionic interactions. The bonded portion may act as a
reservoir and be released more slowly than the unbound form.
0–30% of the adsorbed gentamicin binds to albumin [34].
Furthermore, gentamicin is a highly charged polycation (+3.5,
pH 7.4) [35], whereas soy protein has many negatively charged
Fig. 4 – Representative histological sections of wound areas taken from the center of the wound 12 days post-burn creation.
(A) Melolin treated group. (B) Soy dressing without antibiotics. (C) Soy dressing with controlled gentamicin release. (D)
Scoring of the histological sections of the wound area taken from the center of the wound 12 days post-burn creation. The
examined wound healing criteria included epithelial coverage, epidermal thickness, epidermis-dermis attachment,
neoangiogenesis, number of follicles (adnexa), dermis thickness, WBC and collagen. Grading was between 0 and 3: 0
absence, 1 minimal presence, 2 moderate presence and 3 extensive presence, except for WBC, where 0-extensive presence
and 3-absence (mean W SD).
b u r n s 4 1 ( 2 0 1 5 ) 1 4 5 9 – 1 4 6 7 1465
carboxyl groups. It is thus probable that some ionic bonding
took place. Such a bonding mechanism has been shown before
between deprotonated carboxyl groups of succinylated colla-
gen and positive anion groups of the gentamicin molecule [36].
4.2. In vivo study
The post-burn end-point was chosen to be twelve days. At this
stage, the wounds in all groups demonstrated epithelialization
of more than half of the original wound area. The 3 study
groups, with or without controlled release of gentamicin, were
found to have 29–45% non-epithelialized areas and 31–50%
contraction. A better epithelialization rate was observed for
the soy with gentamycin dressing compared to soy without
gentamycin and compared to the standard of care treatment
of Melolin. Furthermore, a significant decrease in contraction
rate was observed in the soy dressing with and without
controlled release of gentamicin as compared to the Melolin
dressing group (31% � 10% (p < 0.005), 36% � 8% ( p < 0.02) and
51% � 12, respectively). Wound contraction although it is an
inherent part of a normal wound healing we would like to keep
it to a minimum, since in humans it may lead to disfigurement
of the skin and poor esthetic results. It may also lead to loss of
the normal flexibility of the skin – a fixed deformity that entails
a functional disability. The reduced effect of the contraction,
which becomes a dominant mechanism when healing is
delayed, is achieved by the significantly accelerated epitheli-
alization as observed in the controlled release of gentamicin
from the wound dressing group. Furthermore, it should be
stressed that the soy dressing did not evoke a negative
response due to its chemical, structural or physical properties.
The controlled release of gentamicin dressing may have a
systemic effect, and thus each animal received the same
dressing from each treatment group to prevent cross
contamination.
A clear benefit for the soy with gentamycin dressing was
also found in the histological parameters compared to the
control groups, meaning that the quality of healing was better
when using the gentamicin dressing. Histological evaluation
of the wounds showed superior organization of the epithelium
and dermis in the gentamicin soy dressing compared to the
control groups. The general scores differed significantly
(10.8 � 1.4, for the gentamicin soy dressing compared to the
soy group, 7.2 � 1.7, and the Melolin group, 7.2 � 1.2, p < 0.05).
The quality of the epidermis-dermis junction was better and
there were more collagen fibers in the gentamicin dressing.
b u r n s 4 1 ( 2 0 1 5 ) 1 4 5 9 – 1 4 6 71466
Another important finding in groups treated with controlled
release of gentamicin was the dominance and good quality of
the hair follicles and adnexa which are the source for re-
epithelialization from within the burn area.
In conclusion, in vivo evaluation of the gentamicin-eluting
soy protein dressing material in a contaminated wound
demonstrated its advantage over the other investigated dres-
sings. We consider the better qualities of the healed woundinthe
gentamycin soy dressing to be one of the most important
advantages of this new dressing. The gold standard of local
treatment with topical antibacterial agents, e.g. Silverol1, is
changed daily or twice-daily, which is time consuming and
painful to the patient as well as less economical. Several of the
currently used dressing materials that provide controlled release
of silver ions as an antibacterial agent might have toxic effect on
cells, which can delay wound healing. The current dressing
material shows promising results, although it still needs
evaluation in a clinical trial.
Conflict of interest statement
All the authors state no conflict of interest.
Acknowledgements
The authors are grateful to the Israel Science Foundation (ISF,
grant no. 1055/11) for supporting this research. We would like
to thank SolbarTM (Ashdod, Israel) for kindly providing the soy
protein isolate used in this study.
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