Effect of Green Tea (Camellia sinensis) Extract on Healing Process of Surgical Wounds in Rat

Abstract

Green tea (Camellia sinensis) has anti-oxidant and anti-inflammatory properties and may enhance wound healing process. The present study, therefore, was aimed to examine the effect of green tea ethanolic extract on wound healing process. For this experimental study, 36 healthy male Wistar rats were randomly designated to three groups of A, B, and C which, respectively treated with, Vaseline+0.6% green tea extract, Vaseline and normal saline for 21 days. Wounds' length and area were measured by caliper every other day and specimens were taken at 3rd, 12th, and 21st day for microscopical examinations. Data were analyzed by SPSS 16 using survival analysis (Breslow test), repeated measured ANOVA, one-way ANOVA and Mann-Whitney. P < 0.05 was considered as statistically significant. The mean healing duration of surgical wounds in groups A and B was 14.66 and 20.66 (P=0.018), respectively. Decrease in healing duration in the group A was significantly higher within the first two weeks compared with control groups (P = 0.05). Microscopic examinations also indicated a significant difference in wound healing process between groups A and C throughout the whole study duration as well as groups A and B during the 3rd week of the study (P<0.05). Green tea extract could help wound healing process, probably effective on surgical wounds healing.

Full text

Original research
Effect of green tea (Camellia sinensis) extract on healing process of
surgical wounds in rat
Sayyed Yazdan Asadi
a
, Pouya Parsaei
b
, Mehrdad Karimi
c
, Sareh Ezzati
a
,
Alaleh Zamiri
d
, Fereshteh Mohammadizadeh
e
, Mahmoud Rafieian-kopaei
f
,
*
a
Medical Plants Research Center, Faculty of Medicine, Shahrekord University of Medical Sciences, Shahrekord, Iran
b
Young Researchers Club, Shahrekord Branch, Islamic Azad University, Shahrekord, Iran
c
Department of Surgery, Shahrekord University of Medical Sciences, Shahrekord, Iran
d
Faculty of Medicine, Tehran University of Medical Sciences, Tehran, Iran
e
Department of Pathology, Faculty of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran
f
Medical Plants Research Center, Shahrekord University of Medical Sciences, Shahrekord, Iran
article info
Article history:
Received 26 October 2012
Received in revised form
20 January 2013
Accepted 18 February 2013
Available online 28 February 2013
Keywords:
Green tea
Camellia sinensis
Healing process
Surgical wound
Rat
abstract
Green tea (Camellia sinensis) has anti-oxidant and anti-inflammatory properties and may enhance wound
healing process. The present study, therefore, was aimed to examine the effect of green tea ethanolic
extract on wound healing process.
For this experimental study, 36 healthy male Wistar rats were randomly designated to three groups of
A, B, and C which, respectively treated with, Vaseline þ0.6% green tea extract, Vaseline and normal saline
for 21 days. Wounds’length and area were measured by caliper every other day and specimens were
taken at 3rd, 12th, and 21st day for microscopical examinations. Data were analyzed by SPSS 16 using
survival analysis (Breslow test), repeated measured ANOVA, one-way ANOVA and ManneWhitney.
P<0.05 was considered as statistically significant.
The mean healing duration of surgical wounds in groups A and B was 14.66 and 20.66 (P¼0.018),
respectively. Decrease in healing duration in the group A was significantly higher within the first two
weeks compared with control groups (P¼0.05). Microscopic examinations also indicated a significant
difference in wound healing process between groups A and C throughout the whole study duration as
well as groups A and B during the 3rd week of the study (P<0.05).
Green tea extract could help wound healing process, probably effective on surgical wounds healing.
Ó2013 Surgical Associates Ltd. Published by Elsevier Ltd. All rights reserved.
1. Introduction
Wound healing comprises a complex pathophysiological pro-
cess including several cellular and biochemical sub processes, e.g.
inflammation, angiogenesis, and collagen deposition.
1
Inflamma-
tion maintenance and inadequate vessel formation comprise the
most noticeable causes of delayed wound healing.
2
On the other
hand wound fibrosis or abnormal accumulation of collagen in the
wound could lead to an unpleasant scar.
3
Recent research has
shown that many of the compounds that are used for wound
healing such as Acetic acid, Hydrogen peroxide, and etc., is toxic to
cells needed for healing.
4
The majority of plant extracts, e.g. Grape seed, Lemon, Rose-
mary, and Jojoba, have been employed for wound healing and
longevity increase. All of these plants have a common property, i.e.,
producing compounds with phenolic structure.
5
These phyto-
chemicals ordinarily react with some compounds such as Oxygen
free radicals and other macromolecules in order to neutralize free
radicals and/or initiate biological effects.
5
Green tea (Camellia sinensis) which is a product of dried leaves
has been consumed by East Asian people for health promotion
since 3000 B.C.
6,7
Abundantly found in Asia, green tea is also one of
the most prevalent drinks worldwide.
6,8,9
Ample evidence indicates that this plant, with anti-oxidant,
anti-cancer, anti-aging, and anti-inflammatory effects, could also
prevent exaggerate collagen production and accumulation and
induce changes in immune responses, as well
5,6,8
; the majority of
*Corresponding author.
E-mail addresses: yazdan.asadi@ymail.com (S.Y. Asadi), pouyaparsaei@
yahoo.com (P. Parsaei), karimi.mehrdad89@yahoo.com (M. Karimi), sareh_ezzati@
yahoo.com (S. Ezzati), al_zamiri@yahoo.com (A. Zamiri), mohammadizadeh@
med.mui.ac.ir (F. Mohammadizadeh), rafieian@yahoo.com (M. Rafieian-kopaei).
Contents lists available at SciVerse ScienceDirect
International Journal of Surgery
journal homepage: www.theijs.com
1743-9191/$ esee front matter Ó2013 Surgical Associates Ltd. Published by Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.ijsu.2013.02.014
International Journal of Surgery 11 (2013) 332e337
ORIGINAL RESEARCH

these properties could be attributed to the plant’s polyphenolic, in
particular, catechin compounds.
10e14
Epicatechin, Epicatechin gallate, Epigallocatechin, and Epi-
gallocatechin gallate are among the key anti-oxidant compounds of
green tea, little of which could increase collagen volume and hence
heal the wounds.
15
These compounds (e.g. epigallocatechin gallate)
have also been used as an agent for keratinocytes reproduction and
distinction.
16
Also, its anti-fibrogen effects have been confirmed in
some animal models.
17
Regarding the above mentioned and easy accessibility, we
investigated the effect of ethanolic extract of green tea on post-
surgical wound healing process.
2. Materials and methods
After obtaining approval of Shahrekord University of Medical Sciences (SKUMS)
Ethics Committee, this preclinical study was conducted in Medical Plants Research
Center of SKUMS. Throughout the experiments, we tried to follow all ethical prin-
ciples of working on laboratory animals so as to impose the lowest possible stress on
them.
2.1. Extract preparation
Maceration method was employed to prepare the extract. For this purpose,100 g
green tea (Herbarium No. 304, Medical Plants Research Center, SKUMS) was trans-
ported into an Erlenmeyer, 1 L ethanol 70% (Nasr Co. Iran) was added and the so-
lution was left at laboratory temperature. Forty eight hours later, the extract was
filtrated through a filter paper and the pulp was squeezed to discharge. Then, the
extract was concentrated by a rotary evaporator,
18
dried, and mixed with pure
Vaseline (Ehsan Chemi, Iran) to make a Vaseline-based 0.6% ointment.
5
2.2. Measurement of phenolic compounds
The phenolic compounds were evaluated equivalent to gallic acid using Folin-
Ciocalteu colorimetry method.
19
Different concentrations of standard gallic acid,
i.e., 12.5, 25, 50, 62.5, 100, and 125 ppm in methanol 60%, were prepared. Then,
0.1 ml from each sample was transferred into a test tube and 0.5 ml Folin-Ciocalteu
10% was added as reactive agent. The solutions were left for 8 min at room tem-
perature and then 0.4 ml carbonate 7.5% was added. The tubes were maintained for
30 min at room temperature and then assayed in three intervals by a spectropho-
tometer (Unico UV 2010) at 765 nm wavelength. To measure the overall phenol in
the extracts, 0.01e0.02
m
g of the extracts was solved in methanol 60%, reaching
10 ml and then, using Folin-Ciocalteu method, the overall level of phenol was
measured. However, instead of using the standard solution, 0.1 ml extract solution
was added. Finally, the overall phenol level was obtained in mg/g extract in gallic
acid equivalent.
2.3. Measurement of flavonoid compounds
Total flavonoids were evaluated equivalent to Rutin, using chloride aluminum
colorimetry.
20
First, different concentrations of standard Rutin (25, 50, 100, 250 and
500 ppm) were prepared in methanol 60%. Then, from each solution, 1 ml was
transferred into test tubes and mixed with 1 ml of chloride aluminum 2%. After-
wards, 6 ml potassium acetate 5% was added and the optical density level was read
after 40 min at 415 nm wavelength. The concentration levels of the standard solu-
tions were assayed in three intervals. In order to measure the overall level of fla-
vonoids in the extracts, 0.01e0.02 g of the extracts was dissolved in methanol 60%,
reaching 10 ml. Then, the total level of flavonoids was measured using chloride
aluminum colorimetry. However, instead of using the standard solution, 1 ml the
extract was added. The total flavonoid level was calculated in mg/g extract, equiv-
alent to Rutin.
2.4. Measurement of flavonol compounds
The total flavonol was measured using chloride aluminum colorimetry and
Rutin procedure, however the optical density level reading was obtained after 2.5 h
at 440 nm wavelength.
21
2.5. Measurement of anti-oxidant activity
b
-carotene model was employed to measure the anti-oxidant activity of the
extract.
22
Half ml chloroform, 5 ml
b
-carotene (0.2 mg), 20 ml linoleic acid (20 mg),
and 0.2 ml Tween 40 were mixed in a suitable container and incubated at 50

C for
10 min in order to removethe solvent. The solution was diluted with distilled water
and mixed with 4 ml aliquots in the following manner. The control solution was
prepared including 0.2 ml ethanol, 0.2 ml the extract sample, and 0.15 ml ethanol.
The optical density of the control group was recorded at t
0
and t
90
at 470 nm, in a
manner similar to the standard. The samples were incubated in a bain-marie at
50

C. The anti-oxidant activity was measured on the basis of the ability of the
samples in preventing
b
-carotene washing, calculated through AA ¼100[1(A
o
A
t
)/
(Ao
o
Ao
t
)]; where, A
o
is the optical density at t
0
,A
t
is optical density of the sample at
t
90
, and Ao
o
and Ao
t
are optical density values in the control samples at t
0
and t
90
respectively.
2.6. Animals and study design
36 healthy Wistar male rats weighing 200e250 g were randomly assigned to
three groups of A, B, and C and treated respectively, with Vaseline þ0.6% green tea
extract, Vaseline and normal saline for 21 days. The rats had no history of surgery
and other medical interventions, were kept in at most 3-member cages, at 23 2

C
temperature, and on nutritionally similar and standard pelleted diet (Razi Co. Karaj,
Iran).
2.7. Surgical wounds
Incisions were made by one person when the rats were anaesthetized. Rats, in all
groups, were anaesthetized by a combination of 20 mg/kg Ketamine 10% (Alfasan
Co., Netherlands) and 2 mg/kg Xylazine 2% (Alfasan Co., Netherlands), administered
intramuscularly. Then, the cases while anaesthetized were positioned prone on a
surgical table, their back skin was disinfected with Betadine 10% (Tolid Daru Co.,
Iran), and the hairs of an area of the skin preselected using a caliper were completely
shaved with a razor to make a 4 cm incision by means of a No. 24 scalpel; the depth
of incision included both dermis and hypodermis, i.e., the thickness was full. Then, 4
stitches at 1 cm intervals were made by means of 3.0 nylon string (Kamran Teb Co.,
Table 1
System for scoring the histological features of wound tissue samples.
Score Epithelial regeneration Granulation tissue thickness Angiogenesis
1 Little epithelial organization Thin granular layer Altered angiogenesis (1e2 vessels per site)
2 Moderate epithelial organization Moderate granular layer Few newly formed capillary vessels (3e6 vessels per site)
3 Complete epithelial organization Thick granular layer Newly formed capillary vessels (7e10 vessels per site)
4 Complete epithelial organization Very thick granular layer Newly formed and well-structured capillary vessels (>10 vessels per site)
Table 2
The mean decrease in wound length (cm) during 1st, 2nd, 3rd week and total.
Treatment 1st Week 2nd Week 3rd Week Total weeks Result of repeated measures
test in total weeks
Vaseline þ0.6% green tea extract (A) 2.07 0.72 1.83 0.69 0.05 0.12 3.95 0.12 <0.001
Vaseline (B) 1.50 0.35 2.13 0.39 0.30 0.09 3.90 0.08 <0.001
Normal saline (C) 1.12 0.22 2.07 0.21 0.65 0.13 3.85 0.06 0.009
Result of one-way ANOVA test <0.001 0.001 0.003 0.274
Tukey test showed statistically significant differences in these cases: between groups A and B in the third week (P¼0.005) between groups A and C in the first week
(P<0.001), second week (P¼0.001) and third week (P<0.001); between Group B and C in the first week (P¼0.001), second week (P¼0.004) and third week (P¼0.001).
S.Y. Asadi et al. / International Journal of Surgery 11 (2013) 332e337 333
ORIGINAL RESEARCH

Iran), the given area was again disinfected, and the rats were kept at suitable tem-
perature until consciousness. At 7th day, the stitches were removed.
23e26
2.8. Treatment
Treatment was performed every day at the same time by one person and
continued for 21 days, beginning from the day of making incisions. For this, 1 g
Vaseline ointment containing green tea extract was topically applied on a 2 cm
2
(4 cm 0.5 cm) area of the wound in group A; the same was performed on group B
and C using Vaseline ointment and normal saline (Daru-Pakhsh, Iran), respectively.
26
2.9. Microscopic examinations
For purpose of data collection, the animals were numbered and a checklist was
prepared to record data individually.Since the 2nd day of the study,the length of the
remaining wound was measured by a predetermined person using a caliper, every
other day; the case’s data in the given days was separately recorded.
23e26
For the purpose of comparison, photographs of all animals’wounds were taken
at the 2nd day and, since the 7th day, every other day until the study completion.
For this, 3 rats were randomly recruited from each group at 3rd, 12th, and 21st
days; on 21st day, the day of study completion, the recruited rats were euthanized
after measuring of the remaining wound length. After euthanizing them through
ether inhalation in a closed space, the wound tissue histological specimens in full
thickness accompanied with neighboring healthy skin were taken and placed into
10% Neutral Buffered Formalin fixator. Tissue processing was done by paraffin and
wax. Transverse incisions, 5
m
m thick, were made by means of Microtome fixed
blade. Lesions were stained by hematoxylin and eosin.
27,28
Histopathological ex-
aminations, in view of epithelial regeneration, granulation tissue thickness, and
angiogenesis (Table 1), were separately done by a pathologist who was blinded in
grouping.
2.10. Data analysis
Data analysis was performed by SPSS 16 using Survival analysis (Breslow test),
Repeated measures ANOVA and One-way ANOVA (for macroscopic analysis), and
ManneWhitney (for histopathology analysis). P<0.05 was considered as statisti-
cally significant.
3. Results
Totally, 5 of 6 (83%) rats in group A (Vaseline þ0.6% green tea
extract) and 3 of 6 (50%) rats in group B (Vaseline) recoveredby 21st
day. No inflammation, eczema and/or infection were observed.
Regarding Breslow test, the mean healing duration in group A
was shorter compared to group B [P¼0.018, 14.66 days (SE ¼1.94)
vs. 20.66 days (SE ¼0.373)].
The wound length in group A, based on repeated measures
ANOVA, was significantly shorter compared to group B. Fig. 1
indicates wound lengths during the study. Table 2 indicates the
0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
2 7 9 111315171921
Lenght of Wound
Da
y
of Measurement
Vaseline+0.6% Green tea extract
Vaseline
Normal saline
Fig. 1. The size of the wound incision length (cm) in the three treatment groups during
different days.
Photograph 1. Improvement process of wound healing in group A.
S.Y. Asadi et al. / International Journal of Surgery 11 (2013) 332e337334
ORIGINAL RESEARCH

mean decrease in wound length during 1st, 2nd, and 3rd week as
well as the whole study duration.
Based on one-way ANOVA, there was no significant difference in
the mean decrease in wound length between groups A and B during
1st and 2nd weeks (P¼0.094 and P¼0.536, respectively); during
the 3rd week, the difference between the two groups was signifi-
cant (P¼0.005). Also, the decrease in wound length in group A was
significantly higher compared to group B during the first 14 days
(P¼0.043). The Photograph 1 shows improving in group A in
macroscopic study.
Table 3 shows degrees of histopathological indices of wound
healing in different groups throughout the whole study. Regarding
ManneWhitney test, comparing groups A and C during the whole
duration, indicated statistically significant differences in histo-
pathological indices of epithelial regeneration, granulation tissue
thickness, and angiogenesis (P¼0.006, P¼0.007, and P¼0.016,
respectively).
In addition, this comparison at 21st day of the study indicated
statistically significant differences in histopathological indices of
epithelial regeneration, granulation tissue thickness, and angio-
genesis (P¼0.006, P¼0.007, and P¼0.016, respectively); no
significant difference in all indices was observed throughout the
whole study duration.
The Photographs 2 and 3 illustrate a significant difference be-
tween group A and the other two (control) groups.
4. Discussion
Wound healing is a complex process, which interruption could
lead to a delayed healing or excessive fibrosis.
29
Delay in wound
healing increases the possibility of getting infected, inappropriate
recovery, and unpleasant scar. Several effects of green tea and its
compounds have been already examined, indicating that this plant,
with anti-oxidant, anti-cancer, anti-aging, and anti-inflammatory
effects, could also prevent collagen production and accumula-
tion
8,14
; the majority of these properties could be attributed to the
plant’s polyphenolic compound, i.e., catechin in the leaves.
11e13
Consistent with our findings, Safari and Sadrzadeh’s study indi-
cated anti-oxidant effects of epigallocatechin, one of the green tea’s
compounds.
30
In addition, the beneficial effect of epicatechin gallate on wound
healing quality and hence leaving more pleasant scar has been
shown, which probably confirm its effect on increased level of
vascular endothelial growth factor, accelerated vessel formation,
and enhanced nitric oxide and cyclooxygenase.
31
Consistently, the
superiority of group A over the other groups in view of vessel for-
mation, in the present study, is indicative of this effect. A favorable
vascular expansion around the wound could lead to the cells’
appropriate nutrition and improved wound healing process.
32
Monitoring surgical wounds during the study demonstrated the
significant effect of Vaseline þ0.6% green tea extract in contrast to
Vaseline on the wounds’recovery acceleration. The recovery ac-
celeration could be due to the anti-oxidant effect of the present
epigalocatechin on speeding up vessel formation of the skin as well
as anti-inflammatory properties.
Also, the effects of polyphenolic compounds (e.g. epi-
galocatechin gallate) as the expressive agent for connective tissue
growth factor gene and inhibitory regulator of collagen gene
expression have been proposed and their effects on myofibroblasts’
production and distinction, connective tissue growth, and regular
classification of collagens have been proved
33
; the significant dif-
ference in granulation tissue thickness and epithelial regeneration
between group A and control groups could result from these
properties. One of the limitation of this study is lack of immuno-
histological staining method to proper assessment of angiogenesis.
Despite significant decrease in healing duration among those
treated with Vaseline þ0.6% green tea extract, no significant dif-
ference in mean wound length was seen between groups A and B
within the whole duration (P>0.05), probably because of our small
sample size. Within the first two weeks, however, decrease in
Table 3
The mean and median scores for the histological features of the wound tissue samples (mean SD and median [minmax]).
Wound healing’
scaling group
Epithelial regeneration Granulation tissue thickness Angiogenesis
Vaseline þ0.6% green
tea extract (A)
1.333 0.942 (median ¼2,
min ¼0, max ¼2)
1.444 0.831 (median ¼1,
min ¼0, max ¼3)
11.118 (median ¼1,
min ¼0, max ¼3)
Vaseline (B) 1 0.816 (median ¼1,
min ¼0, max ¼2)
0.777 0.416 (median ¼1,
min ¼0, max ¼1)
0.222 0.441 (median ¼0,
min ¼0, max ¼1)
Normal saline (C) 0.222 0.415 (median ¼0,
min ¼0, max ¼1)
0.333 0.471 (median ¼0,
min ¼0, max ¼1)
0.222 0.441 (median ¼0,
min ¼0, max ¼1)
Photograph 2. Tissue samples taken from Group A; Proper and complete angiogenesis
in different parts of the broad bands of collagen in tissue and its context is quite
apparent that the proper formation of granulation tissue (H&E staining) (100,400).
S.Y. Asadi et al. / International Journal of Surgery 11 (2013) 332e337 335
ORIGINAL RESEARCH

wound length was significantly higher in group A compared to
group B (P¼0.043).
5. Conclusion
Regarding the properties mentioned for components present in
the green tea, it could help wound healing duration decrease
considerably; therefore, further research on green tea-derived
medications could lead to ensuring its application as a treatment
for post-surgical wounds.
Ethical approval
All experiments were performed under supervision of Dr.
Shahriyar Adibi and Prof. Mahmoud Rafieian-kopaei, in accordance
to the guidelines of the Animal Ethics Committee of Shahrekord
University of Medical Sciences by the number of: 91-5-2.
Funding
This research were supported by Deputy of Research of Shah-
rekord University of Medical Sciences and Medical Plants Research
Center, Shahrekord University of Medical Sciences.
Author contribution
Sayyed Yazdan Asadi: Study design, Experimental works
(Extract Preparation and determination their components, Animal
study), Data collecting, Article writing.
Pouya Parsaei: Experimental works (Animal study), Data col-
lecting and some parts of article writing.
Mehrdad Karimi: Determination of surgical and anesthesia
procedure and its methods.
Sareh Ezzati: Data Analysing, Article writing.
Alaleh Zamiri: Data Analysing, Article writing.
Fereshteh Mohammadizadeh: Pathologic evaluation.
Mahmoud Rafieian-kopaei: Supervisor of research plan, Study
design, Determination of Antioxidant Activity, Article writing.
Conflict of interest
There are not any conflicts of interest in this manuscript.
Acknowledgments
This work was supported by the grant No. 626 by Deputy of
Research of SKUMS, Shahrekord, Iran. We also thank Medical Plants
Research Center stuff, SKUMS and all people who helped us
throughout the study.
References
1. Desmouliere A, Redard M, Darby I, Gabbiani G. Apoptosis mediates the
decrease in cellularity during the transition between granulation tissue and
scar. Am J Pathol 1995;146(1):56e66.
2. Appleton I. Wound healing: future directions. J Drugs 2003;6(11):1067e72.
3. Norrby K. Angiogenesis: new aspects relating to its initiation and control.
APMIS 1997;105(6):417e37.
4. Life extension foundation. Wound healing (surgical wound, trauma, burns)
[online]. Available from: URL, www.lef.org/protocols/abstracts/abstr-111c.
html; 2000.
5. Hsu S. Green tea and the skin. J Am Acad Dermatol 2005;52(6):1049e59.
6. Kim HR, Rajaiah R, Wu QL, Satpute SR, Tan MT, Simon JE, et al. Green tea
protects rats against autoimmune arthritis by modulating disease-related im-
mune events. J Nutr 2008;138(11):2111e6.
7. Jankun J, Selman SH, Swiercz R, Skrzypczak-Jankun E. Why drinking green tea
could prevent cancer. Nature 1997;387(6633):567.
8. Park G, Yoon BS, Moon JH, Kim B, Jun EK, Oh S, et al. Green tea polyphenol
epigallocatechin-3-gallate suppresses collagen production and proliferation in
keloid fibroblasts via inhibition of the STAT3-signaling pathway. J Invest Der-
matol 2008;128(10):2429e41.
9. Hamaishi K, Kojima R, Ito M. Anti-ulcer effect of tea catechin in rats. Biol Pharm
Bull 2006;29(11):2206e13.
10. Katiyar SK, Afaq F, Perez A, Mukhtar H. Green tea polyphenol epigallocatechin
3-gallate treatment of human skin inhibits ultraviolet radiation-induced
oxidative stress. Carcinogenesis 2001;22(2):287e94.
11. Katiyar SK, Elmets CA. Green tea polyphenolic antioxidants and skin photo-
protection (review). Int J Oncol 2001;18(6):1307e13.
12. Chan MMY, Dunne F, Ho CT, Huang HI. Inhibition of inducible nitric oxide
synthase gene expression and enzyme activity by epigallocatechin gallate, a
natural product from green tea. Biochem Pharmacol 1997;54(12):1281e6.
13. Yen GC, Chen HY. Scavenging effect of various tea extracts on superoxide
derived from the metabolism of mutagens. Biosci Biotechnol Biochem
1998;62(9):1768e70.
14. Frei B, Higdon JV. Antioxidant activity of tea polyphenols in vivo: evidence
from animal studies. J Nutr 2003;133(10):3275e84.
15. Kim H, Kawazoe T, Han DW, Matsumara K, Suzuki S, Tsutsumi S, et al.
Enhanced wound healing by an epigallocatechin gallate-incorporated. Wound
Repair Regen 2008;16(5):714e20.
16. Hsu S, Bollag WB, Lewis J, Huang Q, Singh B, Sharawy M, et al. Green tea
polyphenols induce differentiation and proliferation in epidermal keratino-
cytes. J Pharmacol Exp Ther 2003;306(1):29e34.
Photograph 3. In the first picture lack of skin epithelial tissue layers in group C, while
second one shows the complete formation of the epidermis and dermis layers is clearly
visible (H&E staining) (100).
S.Y. Asadi et al. / International Journal of Surgery 11 (2013) 332e337336
ORIGINAL RESEARCH

17. Nakamuta M, Higashi N, Kohjima M, Fukushima M, Ohta S, Kotoh K, et al.
Epigallocatechin-3-gallate, a polyphenol component of green tea, suppresses
both collagen production and collagenase activity in hepatic stellate cells. Int J
Mol Med 2005;16(4):677e81.
18. Khalaji N, Neyestani T. The inhibitory effects of black and green teas (Camellia
sinensis) on growth of pathogenic Escherichia coli, in vitro. Iran J Nutr Sci
2007;1(3):33e8.
19. Shirzad H, Shahrani M, Rafieian-Kopaei M. Comparison of morphine and tra-
madol effects on phagocytic activity of mice peritoneal phagocytes in vivo. Int
Immunopharmacol 2009;9(7e8):968e70.
20. Rafieian-Kopaei M, Nasri H, Nematbakhsh M, Baradaran A, Gheissari A,
Rouhi H, et al. Erythropoietin ameliorates genetamycin-induced renal toxicity:
a biochemical and histopathological study. J Nephropathol 2012;2(1):109e16.
21. Sharafati R, Sherafati F, Rafieian-Kopaei M. Biological characterization of Ira-
nian walnut (Juglans regia) leaves. Turk J Biol 2011;35:635e9.
22. MGh Mirzaei, Sewell RDE, Kheiri S, Rafieian-Kopaei M. A clinical trial of the
effect of St. John’s wort on migraine headaches in patients receiving sodium
valproate. J Med Plants Res 2012;6(9):1519e23.
23. Hedlund Cherly S. Surgery of the integumentary system. In: Fossum Theresa W,
Hedlund Cheryl S, editors. Small animal surgery. 3rd ed. St. Louis: Mosby; 2007.
p. 159e76. 228e32.
24. Philips Linda G. Wound healing. In: Townsend Jr Courtney M, editor. Sabiston
textbook of surgery. 18th ed. Philadelphia: Saunders; 2007. p. 131e45.
25. Malone M. Supplemental zinc in wound healing. Nutr J Clin Pract 2000;15:253e6.
26. Baumann LS, Spencer J. The effects of topical vitamin E on the cosmetic
appearance of scares. Dermatol Surg 1999;25:34e5.
27. Taghizadeh-Jahed M, Jarolmasjed SH, Mohamadnejad S, Rezaii A, Delazar A.
The effect of Echinacea purpurea aerial organ dried extract vs. zinc oxide on
skin wound healing in rat: a morphometric & histopathologic study. J Tehran
Univ Med 2008;66(9):625e32.
28. Derakhshanfar A, Oloumi M, Mirzaie M. Study on the effect of pega-
numharmala extract on experimental skin wound healing in rat:
pathological and biomechanical findings. J Comp Clin Pathol 2010;19(24):
169e72.
29. Tredget EE, Nedelec B, Scott PG, Ghahary A. Hypertropic scars, keloids, and
contractures: the cellular and molecular basis for therapy. Surg Clin North Am
1997;77(3):701e30.
30. Saffari Y, Sadrzadeh SMH. Green tea metabolite EGCG protects membranes
against oxidative damage in vitro. Life Sci 2004;74(12):1513e8.
31. Kapoor M, Howard R, Hall I, Appleton I. Effects of epicatechin gallate on wound
healing and scar formation in a full thickness incisional wound healing model
in rats. Am J Pathol 2004;165(1):299e307.
32. Hashimoto I, Nakanishi H, Shono Y, Toda M, Tsuda H, Arase S. Angiostatic ef-
fects of corticosteroid on wound healing of the rabbit ear. J Med Invest 2002;49:
61e6.
33. Klass BR, Branford OA, Grobbelaar AO, Rolfe KJ. The effect of epigallocatechin-
3-gallate, a constituent of green tea, on transforming growth factor-beta1-
stimulated wound contraction. Wound Repair Regen 2010;18(1):80e8.
S.Y. Asadi et al. / International Journal of Surgery 11 (2013) 332e337 337
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