This study investigated the protective effects of carvacrol on testicular damage caused by experimental testicular torsion-detorsion in rats. The study consisted of 4 groups of rats: a control group, a torsion group, a torsion-detorsion group, and a torsion-detorsion group treated with carvacrol. Histopathological analysis found increased damage in spermatogenic cells and decreased antioxidant levels in the torsion and torsion-detorsion groups compared to the control and carvacrol groups. Immunohistochemical staining showed increased endothelin-1 expression in the torsion and detorsion groups but not in the carvacrol group. The results suggest that carvacrol may prevent

1. 125
OBJECTIVE: To investigate the protective effects of
carvacrol on an experimental testicular torsion-detorsion
rat model.
STUDY DESIGN: Wistar male rats (n=48) weighing
230–250 g were assigned to 4 groups (8 per group): con-
trol, torsion, torsion-detorsion, and torsion-detorsion+
carvacrol–treated groups. Control group animals did
not undergo any surgical operation. For the torsion
group, the scrotum was opened (under general anes-
thesia) and the left testis twisted 720° clockwise and in
the last 30 minutes of 3-hour ischemia; i.p. saline was
injected. In the torsion-detorsion group, after ischemia
the left testis was reperfused for 2 hours. The torsion/
detorsion+carvacrol group protocol was similar to that
of the torsion-detorsion group but in the last 30 minutes
of 3-hour ischemia, i.p. 20 mg/kg carvacrol was admin-
istered.
RESULTS: Malondialdehyde (MDA) was highest in the
torsion-detorsion group (p<0.01). The lowest catalase
(CAT) value was found in the torsion-detorsion group.
Decrease in glutathione (GSH) levels of the torsion
and torsion-detorsion groups as compared to those of
control and carvacrol groups was significant (p<0.01).
The highest superoxide dismutase (SOD) value was
in the control and carvacrol groups. Increased apop-
tosis and degeneration of spermatogenic cells with
hyperplasic nuclei were mainly observed in the torsion
and torsion-detorsion groups. The torsion-detorsion+
carvacrol group mostly showed regular histology, but
Leydig cells were degenerated. ET-1 expression was
increased in endothelial cells in the torsion and detor-
sion groups but negative in the carvacrol group. Bax
expression was positive in luminal spermatogenic cells
in the torsion group but negative in interstitial cells
in both torsion and torsion-detorsion groups. In the
carvacrol-treated group some luminal spermatogenic
cells in seminiferous tubules showed positive Bax ex-
pression but weak in basal membrane cells and Leydig
cells.
CONCLUSION: Carvacrol influences spermatogenic
cells with strong mitotic activity in basal membranes
of seminiferous tubules and may prevent apoptotic
development and signaling of these cells. (Anal Quant
Cytopathol Histpathol 2020;42:125–132)
Analytical and Quantitative Cytopathology and Histopathology®
0884-6812/20/4204-0125/$18.00/0 © Science Printers and Publishers, Inc.
Analytical and Quantitative Cytopathology and Histopathology®
Effects of Carvacrol on Experimental Testicular
Torsion-Detorsion Model
Investigation by Immunohistochemistry
Recep Dursun, M.D., Abdullah Şen, M.D., Mahmut Yaman, M.D.,
Hasan Mansur Durgun, M.D., and Fırat Aşır, Ph.D.
From the Departments of Emergency Medicine and of Histology and Embryology, Faculty of Medicine, Dicle University, Diyarbakır,
Turkey.
Recep Dursun is Associate Professor, Department of Emergency Medicine, Faculty of Medicine, Dicle University.
Abdullah Şen is Assistant Professor, Department of Emergency Medicine, Faculty of Medicine, Dicle University.
Mahmut Yaman is Assistant Professor, Department of Emergency Medicine, Faculty of Medicine, Dicle University.
Hasan Mansur Durgun is Associate Professor, Department of Emergency Medicine, Faculty of Medicine, Dicle University.
Fırat Aşır is Research Assistant, Department of Histology and Embryology, Faculty of Medicine, Dicle University.
Address correspondence to: Fırat Aşır, Ph.D., Department of Histology and Embryology, Faculty of Medicine, Dicle University, 21280
Diyarbakır, Turkey (firatasir@gmail.com).
Financial Disclosure: The authors have no connection to any companies or products mentioned in this article.

2. Keywords: Bax protein; bcl-2-associated X pro-
tein; carvacrol; detorsion; endothelin-1; ischemia-
reperfusion injury; rats, Wistar; reperfusion injury;
spermatic cord torsion; testicular torsion; torsion
abnormality.
Testicular torsion is a urological complication lead-
ing to testicular injury and is commonly seen in
humans between the ages of 12 and 18. The sper-
matic cord twists around its axis, and blood flow
to testicular tissue is interrupted. The testis does
not receive enough blood, and ischemia, edema,
and necrosis develop.1 During this process, reac-
tive oxygen species (ROS) and apoptosis rates in-
crease. There are many articles showing a correla-
tion between reperfusion and ROS production due
to testicular oxidative stress.2 The severity of testic-
ular torsion depends on the duration and degree
of rotation. To prevent male infertility, testicular
torsion must be treated immediately.
Carvacrol [isopropyl-ortho-cresol, C6H3(OH)
(C3H7)] is a monoterpene derivative of cymene.
More technically, it is a member of phenols which
is naturally found in the subspecies of the fam-
ily Lamiaceae.3 Carvacrol has been used as a
food additive for years and is especially effective
against food-borne pathogens, including Esche-
richia coli, Salmonella, and Bacillus cereus.4 Car-
vacrol has many bioactivities such as antioxidant,
anti-inflammatory, anti-tumor activity, bactericid-
al activity, fungicidal activity, and antimicrobial
properties.1 Endothelins are 21 amino acid pep-
tides that regulate vascular homeostasis. Normally,
endothelins are vasoconstrictors and so they cause
blood pressure to elevate, but their expression is
held in balance by other mechanisms. There are
3 forms of endothelins: ET-1, ET-2, and ET-3. ET-1
is encoded by human EDN1 gene and is expressed
mainly in the endothelium. ET-1 is reported to be
associated with the cardiovascular system, uri-
nary system, nervous system, respiratory system,
immune system, and other tissues.5 Apoptosis is
regulated by functional genes and their protein
products. Bcl-2 is a protein found in the inner
membrane of the mitochondria and blocks apop­
tosis. It inhibits the passage of cytochrome c from
the mitochondria to the cytoplasm, reducing free
radical production. Therefore, it acts as an anti­
oxidant. Increased expression of Bax gene on
mitochondrial membranes induces the release of
pro-apoptotic molecules. This event triggers the
activation of effector caspases, leading to the re-
lease of various inflammatory mediators and free
radicals.6
The aim of this study was to investigate the pro­
tective effects of plant extracts on spermatogenesis
and testicular tissue damage caused by experi-
mental testicular torsion by histopathological and
immunohistochemical methods.
Materials and Methods
Study Design
All experimental protocols were conducted at Dicle
University Animal Research Center. All procedures
were approved by the Dicle University Animal
Care and Use Local Committee (ethical approval
number 2019/13). Forty-eight male Wistar rats
weighing 230–250 g were kept at 24±1°C and 12-
hour light/dark cycles with ad libitum water and
nutrition. The rats were randomly assigned to 4
groups (8 rats per group): control group, ischemia
group, ischemia-reperfusion group, and ischemia-
reperfusion+carvacrol–treated group.
Before the experiment, 40 mg/kg ketamine hy-
drochloride (Ketalar; Pfizer, Istanbul, Turkey) and
5 mg/kg xylazine (Rompun; Bayer, Istanbul, Tur-
key) were administered intramuscularly to anes-
thetize the animals. The rats were divided into 4
groups, 8 rats per group, and the following proce-
dures were applied to the groups.
Control Group. The animals did not undergo any
surgical operation and were sacrificed at the end of
the experiment.
Torsion Group. Under general anesthesia, the mid-
line of the rat scrotum was incised. The left testes
were twisted 720° clockwise, and in the last 30
minutes of 3-hour ischemia, i.p. saline was injected
into the animals. At the end of the fifth hour, all
animals were sacrificed.
Torsion/Detorsion Group. Under general anesthesia,
the midline of the rat scrotum was incised. The
left testes were twisted 720° clockwise, and in the
last 30 minutes of 3-hour ischemia, i.p. saline was
injected. At the end of the ischemia procedure the
scrotum sutures were opened and the left testes
were brought to normal position. The scrotums
were closed again, and the testes were reperfused
for 2 hours. At the end of the reperfusion period,
all animals were sacrificed.
Torsion/Detorsion+Carvacrol Group. Under general
126 Analytical and Quantitative Cytopathology and Histopathology®
Dursun et al

3. anesthesia, the midline of the rat scrotum was in­
cised. The left testes were twisted 720° clockwise,
and in the last 30 minutes of 3-hour ischemia, i.p.
20 mg/kg carvacrol was administered to the ani-
mals. The scrotum sutures were opened, and the
left testes were brought to normal position. The
scrotums were closed again, and the testes were
reperfused for 2 hours. At the end of the reperfu-
sion period, all animals were sacrificed.
Paraffin Wax Embedding Protocol and
Immunohistochemical Staining
The protocol was described by Obut and Oglak.7
Tissues were fixed with zinc-formalin solution
(catalog no. Z2902, Sigma-Aldrich, St. Louis, Mis-
souri, USA) and washed under tap water for 5
minutes. Tissues were passed through ascending
alcohol series for about 24 hours. To remove alco-
hol, tissues were washed with xylene 2×30 min-
utes and incubated in paraffin wax. Tissues were
embedded in paraffin wax, and 4–6 µm sections
were cut with a microtome (catalog no. RM2265,
Leica, Wetzlar, Germany). Sections were incu-
bated at 60°C for 3–4 hours. After deparaffinized in
xylene for 2×30 minutes, sections were brought to
distilled water. Some of the sections were stained
with routine hematoxylin and eosin, and the rest
were soaked in PBS for 3×5 minutes. Hydrogen
peroxide solution (catalog no. TA-015-HP, Thermo
Fisher Scientific, Fremont, California, USA) was
dropped on sections for 20 minutes. After washing
in PBS for 3×5 minutes, ultra V Block (catalog no.
TA-015-UB, Thermo Fisher) was applied to sec-
tions for 8 minutes. Sections were incubated with
primary antibodies Bax (catalog no. PA5-11378,
Thermo Fisher; dilution rate 1/100) and anti-
endothelin-1 (catalog no. ab117757, Abcam, Cam-
bridge, Massachusetts, USA; dilution rate 1/200)
at +4oC overnight. The next day, sections were
left at room temperature for 30–60 minutes. Follow-
ing the PBS step, sections were washed with bio­
tinylated secondary antibody (catalog no. TP-015-
BN, Thermo Fisher) for 14 minutes. Streptavidin-
peroxidase (catalog no. TS-015-HR, Thermo Fish-
er) was dropped onto sections for 15 minutes.
Clearing with PBS, DAB (catalog no. TA-001-
HCX, Thermo Fisher) was used as chromogen,
and reaction was stopped by PBS. Sections were
counter-stained with Harris hematoxylin and
mounted with Entellan (catalog no. 107961, Sig-
ma-Aldrich, St. Louis, Missouri, USA). Slides
were analyzed with Zeiss Imager A2 and photo­
micrographed.
Results
Statistical Results
Malondialdehyde (MDA), catalase (CAT), and GSH
results statistically showed normal distribution
and were evaluated by one-way ANOVA test.
Post-hoc Tukey test was used for multiple com­
parisons. Superoxide dismutase (SOD) and diam-
eters of seminiferous tubules were normally dis­
tributed and were evaluated by one-way ANOVA
test. Tamhane’s T2 test was used for multiple
comparisons between the groups. All results are
shown as mean±SEM, and p<0.05 was considered
significant (Table I).
Histopathological Results
Cross sections of the testes were stained with
hematoxylin and eosin (Figure 1). In the semini­
ferous tubules of the control group, primary
spermatocyte cells were oval with chromatin-rich
nuclei and intense mitotic activity towards the lu-
men. Sertoli cells were triangular and spermatid
cells were concentrated in their apex toward the
lumen. In the interstitial area, Leydig cells were
found to be organized in groups around small
Volume 42, Number 4/August 2020 127
Carvacrol in Testicular Torsion-Detorsion
Table I Statistical Analysis of Groups Regarding MDA, CAT, GSH, SOD, and Diameters of Seminiferous Tubules
Diameter of
seminiferous
Group MDA CAT GSH SOD tubule
Control 6.30±0.18a 0.06±0.01d 368.88±1.77h 3.21±0.13j 303.73±3.56l
Torsion 10.58±0.42b 0.03±0.00f 312.49±0.97i 1.73±0.12k 264.87±5.62m
Torsion/detorsion 13.53±0.21c 0.01±0.00g 315.95±0.27i 1.63±0.07k 262.45±5.02m
Torsion/detorsion+carvacrol 6.82±0.14a 0.04±0.00e 362.44±2.59h 3.41±0.08j 301.93±2.46l
Different superscript letters in the same row indicates significance between groups (a-b: p<0.01, a-c: p<0.01, b-c: p<0.01, d-e-f-g: p<0.05, h-i: p<0.01, j-k:
p<0.01, l-m: p<0.01).
CAT = catalase, GSH = glutathione, MDA = malondialdehyde, SOD = superoxide dismutase.

4. capillary vessels (Figure 1A). In the torsion group,
deteriorated basal membranes of the seminifer-
ous tubules, degenerated spermatogenic cells, in-
creased apoptotic cells, and significant deformation
in Sertoli cells were observed. Vascular dilation
and hemorrhage and Leydig cells with pyknotic
nuclei were observed in the interstitial area (Fig-
ure 1B). In the torsion-detorsion group, sperma­
togenic cells in the seminiferous tubules were
deformed, with hyperplastic nuclei observed. Ser-
toli cells were degenerated and detached from
the basal membrane of the seminiferous tubules.
In the intertubular area, congested blood ves-
sels and deteriorated Leydig cells with pyknotic
nuclei were observed (Figure 1C). In the torsion-
detorsion+carvacrol group, while degenerative
changes were observed in some spermatogenic
cells in some seminiferous tubules, most of the
cells were regularly localized in the basal mem-
brane with chromatin-rich nuclei. Sertoli cells
preserved their triangular structure, while Leydig
cells were degenerated (Figure 1D).
Cross sections of the testes were immunostained
with endothelin-1 antibody (Figure 2). In the con-
trol group, endothelin-1 expression was positive
in the interstitial vascular endothelial cells (Figure
2A). In the torsion group, increased endothelin-1
expression was observed in the basal membrane
of the seminiferous tubules and in interstitial in­
flammatory cells and vascular endothelial cells in
the intertubular areas (Figure 2B). In the torsion-
detorsion group, endothelin-1 expression was sig-
nificantly increased in the basal membrane of the
seminiferous tubules and in interstitial inflam-
matory cells and degenerated endothelial cells
adhering to the basal membrane in the intertu-
bular area (Figure 2C). In the torsion-detorsion+
carvacrol group, negative endothelin-1 expression
128 Analytical and Quantitative Cytopathology and Histopathology®
Dursun et al
Figure 1 Hematoxylin-eosin staining of cross sections of the testes. (A) Seminiferous tubules of the control group. (B) Torsion group.
(C) Torsion-detorsion group. (D) Torsion-detorsion+carvacrol group. H-E staining, Bar=500 µm.

5. was observed in the basal membranes of the sem-
iniferous tubules and in the interstitial cells and
vascular endothelial cells (Figure 2D).
Figure 3 shows Bax expression of cross sections
of the testes. In sections of the control group, neg-
ative Bax expression was observed in spermatoge­
nic and Sertoli cells in seminiferous tubules and
Leydig cells in the intertubular area (Figure 3A). In
the torsion group there was a significant increase
in Bax expression in luminal spermatogenic cells
and Sertoli cells, while Bax expression was nega-
tive in interstitial cells (Figure 3B). In the torsion-
detorsion group, Bax expression was increased in
spermatic cells in seminiferous tubules. Negative
Bax expression was observed in Leydig cells and
some fibroblast cells (Figure 3C). In the torsion-
detorsion+carvacrol group, Bax expression was
positive in some luminal spermatogenic cells in
seminiferous tubules, whereas it was weak in basal
membrane cells and Leydig cells (Figure 3D).
Discussion
Testicular torsion is the rotation of the spermatic
cord around its own axis, resulting in decreased
arterial blood flow and obstruction of venous
and lymphatic drainage. As a result of ischemia,
edema, necrosis, hemorrhage, and venous conges­
tion occur. Levels of lactic acid, hypoxanthine, and
lipid peroxides in tissues are increased.8 Many
studies reveal that overproduction of reactive
oxygen species are related to ischemia-reperfusion
injury in organs other than the testes, as well.9,10
Józsa et al11 showed that artificial spermatic cord
torsion for 2 hours in rats led to changes in tes-
ticular microcirculation, volume reduction, and a
slight change in hemorrhagic parameters. In the
testis, reperfusion injury is more destructive than
ischemic tissue damage. After the resumption of
blood flow in the tissues, some ischemic tissue
was recovered, and reperfusion damage was asso-
ciated with systemic shock and subendothelial
Volume 42, Number 4/August 2020 129
Carvacrol in Testicular Torsion-Detorsion
Figure 2 Cross sections of the testes were immunostained with endothelin-1 antibody. (A) Control group. (B) Torsion group.
(C) Torsion-detorsion group. (D) Torsion-detorsion+carvacrol group. Endothelin-1 immunostaining, Bar=500 µm.

6. hemorrhagic necrosis. The enzymatic antioxidant
defense system, which contains SOD, CAT, and
GSH-Px, reacts to eliminate free radicals in dam-
age caused by ischemia and reperfusion models.
It stimulates the migration of leukocytes and leads
to ischemic region. Reactive oxygen species (ROS)
production takes place. Lipid peroxidation causes
an increase in the reaction. MDA value, which is
an indicator of lipid peroxidation, is increased.12,13
Statistical analysis of the results of MDA, GSH,
SOD, and diameters of seminiferous tubules are
shown in Table I. The MDA amount was highest in
the torsion-detorsion group and was significantly
different from the control group (p<0.01). MDA
values of the torsion group were significantly dif-
ferent from those of all other groups (p<0.01). The
MDA value in the carvacrol group was close to
that of the control group (p>0.05). The difference
of the CAT value was significant in all groups
(p<0.05), and the lowest value was found in the
torsion-detorsion group. The GSH value of the
carvacrol group was similar to that of the control
group (p>0.05). The GSH value of the torsion
group was close to that of the torsion-detorsion
group (p>0.05). However, the decrease in GSH
levels of the torsion and torsion-detorsion groups
as compared to the control and carvacrol groups
was significant (p<0.01). The highest SOD value
was in the control and carvacrol groups. The
SOD levels of these groups were similar (p>0.05),
whereas the decrease in SOD levels in the tor-
sion and torsion-detorsion groups was significant
(p<0.01). Seminiferous tubule diameters were sim-
ilar in the control and carvacrol groups (p>0.05).
The decrease in seminiferous tubule diameter was
statistically significant in the torsion and torsion-
detorsion groups (p<0.01).
Karabulut et al14 stated that irregular germ cells
with coagulation necrosis, pyknotic nuclei, deteri-
orated seminiferous tubules, and vascular conges-
tion and edema in the interstitium of the contra-
lateral testicles are observed in testicular torsion.
130 Analytical and Quantitative Cytopathology and Histopathology®
Dursun et al
Figure 3 Bax expression of cross sections of testes. (A) Control group. (B) Torsion group. (C) Torsion-detorsion group. (D) Torsion-
detorsion+carvacrol group. Bax immunostaining, Bar=500 µm.

7. They also reported that these events are more
severe in testicular detorsion. In our study, testi­
cular sections of the control group showed normal
histological structure (Figure 1A). In our torsion
group, deteriorated basal membranes of the semi-
niferous tubules, degenerated spermatogenic cells,
increased apoptotic cells, and significant defor-
mation in the Sertoli cells were observed (Figure
1B). In the torsion-detorsion group, we observed
increased deformation in spermatogenic cells
with hyperplastic nuclei, and degenerated Sertoli
cells detached from the basal membrane towards
the lumen (Figure 1C). In the torsion-detorsion+
carvacrol group, most of the cells seemed to be
regular and orderly, organized in the basal mem-
brane with chromatin-rich nuclei. Sertoli cells were
apparently triangular, but Leydig cells were degen-
erated (Figure 1D).
Carvacrol is a major component of the essential
oils of certain aromatic plants and has attracted
much attention due to its biological properties in
the treatment of human diseases.15 Carvacrol ex-
hibits strong antioxidant and hydrophobic prop-
erties associated with its substituted aromatic
ring, as well as hydrophilic properties associated
with its phenolic OH group, which have pre-
viously been associated with antioxidant, anti-
inflammatory, antibacterial, antifungal, antiproto-
zoal, anticarcinogenic, antidiabetic, antinociceptive,
cardioprotective, and neuroprotective effects.1 I
·
pek
et al16 stated that carvacrol has antitumor and
anti-mutagenic activities in vitro against oxidative
damage caused by mutagenic lesions in DNA.
Luo et al17 showed that carvacrol reduced oxi-
dative stress and increased antioxidative effect
on ethanol-induced neuron damage. A study re-
vealed that apoptosis in liver ischemia-reperfusion
injury was suppressed by carvacrol treatment and
that carvacrol could reduce ischemia-reperfusion–
induced liver damage by antioxidative and anti­
apoptotic properties.18 Aristatile et al19 treated rats
with doses of 20, 40, and 80 mg/kg carvacrol to
prevent D-galactosamine–induced oxidative inju-
ry and measured oxidant/antioxidant systems in
plasma, kidney, and liver tissue. In the testicular
sections of the torsion-detorsion+carvacrol group,
although spermatogenic cells in some seminifer-
ous tubules were degenerated, most of them were
with chromatin-rich nuclei and regularly localized
throughout the lumen. The number of apoptotic
cells decreased, and Sertoli cells preserved their
triangular structure (Figure 1D).
Endothelin-1 (ET-1) constricts the vascular
structures and is expressed by endothelial cells,
which counteracts vasodilator nitric oxide.20 Bajory
et al21 stated that ET-1 has an important role in
ischemia-reperfusion–induced cystitis, and pre-
treatment with an ET-A (endothelin-1 receptor)
reduces ischemia-reperfusion–related microvascu­
lar disturbances in the bladder. Han et al22 report-
ed that their results suggest that endogenous
endothelin-1 may contribute to ischemia/reper­
fusion injury and could be attenuated by ET-1 an-
tagonists. A study showed that ET-1 elevates the
generation of reactive oxygen species and con-
tributes to the development of endothelial dys-
function.23 I
·
pek et al24 said that ET-1 could be an
effective regulator in angiogenic development by
stimulating blood flow and microcirculation of
the testicles, and the increased ET-1 expression in
endothelial cells influences angiogenic develop-
ment. In our study, in the torsion and detorsion
groups, increased ET-1 expression was observed
in endothelial cells (Figure 2B–C); however, the
expression was negative in the carvacrol group
(Figure 2D).
Increased ROS production leads to oxidative
stress, increasing apoptosis, and DNA damage.
Testicular torsion-detorsion has been shown to
cause increased apoptosis in germ cells and in-
creased expression of Bax mRNA.25 Bax expres-
sion was negative in testicular cells of the con-
trol group (Figure 3A). In the torsion group, Bax
expression was positive in spermatogenic cells
close to the lumen but negative in interstitial cells
(Figure 3B). The expression was negative in the
intertubular area of the torsion-detorsion group
(Figure 3C). In the carvacrol-treated group, some
luminal spermatogenic cells in the seminiferous
tubules showed positive Bax expression but were
weak in the basal membrane cells and Leydig cells
(Figure 3D).
In conclusion, we think that after torsion-
detorsion application, carvacrol is effective in
spermatogenic cells with strong mitotic activity in
the basal membranes of seminiferous tubules and
may prevent apoptotic development and signaling
of these cells.
References
1. DaJusta DG, Granberg CF, Villanueva C, Baker LA: Contem-
porary review of testicular torsion: New concepts, emerging
technologies and potential therapeutics. J Pediatr Urol 2013;
9(6 Pt A):723-730
2. Akgur FM, Kilinc K, Aktug T: Reperfusion injury after detor-
Volume 42, Number 4/August 2020 131
Carvacrol in Testicular Torsion-Detorsion

8. sion of unilateral testicular torsion. Urol Res 1993;21(6):395-
399
3. Kırımer N, Baser KHC, Tümen G: Carvacrol-rich plants in
Turkey. Chem Nat Compd 1995;31:37-41
4. Ultee A, Kets EP, Smid EJ: Mechanisms of action of carvacrol
on the food-borne pathogen Bacillus cereus. Appl Environ
Microbiol 1999;65(10):4606-4610
5. Samarghandian S, Farkhondeh T, Samini F, Borji A: Protec-
tive effects of carvacrol against oxidative stress induced by
chronic stress in rat’s brain, liver, and kidney. Biochem Res
Int 2016:2645237
6. Barton M, Traupe T, Haudenschild CC: Endothelin, hyper-
cholesterolemia and atherosclerosis. Coron Artery Dis 2003;
14(7):477-490
7. Obut M, Oğlak SC: Expression of CD44 and IL-10 in nor-
motensive and preeclamptic placental tissue. Ginekol Pol
2020;91(6):334-341
8. Kazaz IO, Demir S, Yulug E, Colak F, Bodur A, Yaman SO,
Karaguzel E, Mentese A: N-acetylcysteine protects testicular
tissue against ischemia/reperfusion injury via inhibiting
endoplasmic reticulum stress and apoptosis. J Pediatr Urol
2019;15:253.e1-e253.e8
9. Omar B, McCord J: Ischemia–reperfusion. In Oxidative
Stress: Oxidants and Antioxidants. Edited by H Sies. Lon-
don, Academic Press, 1991, pp 493–527
10. McCord JM: Oxygen-derived free radicals in post-ischemic
tissue injury. N Engl J Med 1985;312:159-163
11. Józsa T, Klárik Z, Kiss F, Tóth E, Mester A, Hargitai Z,
Changchien YC, Fossum M, Nemeth N: Morphological and
microcirculatory evaluation of the rat testis after detorsion
with or without a capsular release with a tunica vaginalis
flap. Asian J Androl 2016;18(3):462-466
12. Cay A, Alver A, Küçük M, Işik O, Eminağaoğlu MS, Karahan
SC, Değer O: The effects of N-acetylcysteine on antioxidant
enzyme activities in experimental testicular torsion. J Surg
Res 2006;131:199-203
13. Dokmeci D, Inan M, Basaran UN, Yalcin O, Aydogdu N,
Turan FN, Uz YH: Protective effect of L-carnitine on testi­
cular ischemia-reperfusion injury in rats. Cell Biochem Funct
2006;25:611-618
14. Karabulut Ö, Kalkanlı S, Deveci E, Isen K, Uysal E, Sevda S:
Effects of Potentilla Fulgens as a prophylactic agent in tes-
ticular ischemia reperfusion injury. Anal Quant Cytopathol
Histpathol 2016;38:306-312
15. Friedman M: Chemistry and multibeneficial bioactivities
of carvacrol (4-isopropyl-2-methylphenol), a component of
essential oils produced by aromatic plants and spices. J Agric
Food Chem 2014;62(31):7652-7670
16. I
·
pek E, Zeytinoglu H, Okay S, Tuylu BA, Kurkcuoglu M,
Baser KHC: Genotoxicity and antigenotoxicity of Origanum
oil and carvacrol evaluated by Ames Salmonella/microso-
mal test. Food Chem 2005;93:551-556
17. Luo Q, Qiao H, Ding H, Cao Y, Yu J, Liu R, Zhang Q,
Zhu H, Qu L: The neuroprotective effects of carvacrol on
ethanol-induced hippocampal neurons impairment via the
antioxidative and antiapoptotic pathways. Oxid Med Cell
Longev 2017;2017:4079425
18. Suo L, Kang K, Wang X, Cao Y, Zhao H, Sun X, Tong L,
Zhang F: Carvacrol alleviates ischemia reperfusion injury
by regulating the PI3K-Akt pathway in rats. PLoS One 2014;
9(8):e104043
19. Aristatile B, Al-Numair KS, Veeramani C, Pugalendi KV:
Effect of carvacrol on hepatic marker enzymes and antiox-
idant status in D-galactosamine-induced hepatotoxicity in
rats. Fundam Clin Pharmacol 2009;23:757-765
20. Bakhsha F, Mazandarani M, Aryaei M, Jafari SY, Bayate
H: Phytochemical and antioxidant activity of Lavandula
Angustifolia Mill. essential oil on preoperative anxiety in
patients undergoing diagnostic curettage. Int J Women’s
Health Reprod Sci 2014;2(4):268-271
21. Bajory Z, Hutter J, Krombach F, Messmer K: The role of
endothelin-1 in ischemia-reperfusion induced acute inflam-
mation of the bladder in rats. J Urol 2002;168(3):1222-1225
22. Han H, Neubauer S, Braeker B, Ertl G: Endothelin-1 con-
tributes to ischemia/reperfusion injury in isolated rat heart-
attenuation of ischemic injury by the endothelin-1 antago-
nists BQ123 and BQ610. J Mol Cell Cardiol 1995;27(2):761-
766
23. Kalani M: The importance of endothelin-1 for microvascu-
lar dysfunction in diabetes. Vasc Health Risk Manag 2008;
4(5):1061-1068
24. I
·
pek H, Dogan G, Dogan G, Deveci E: Effects of caffeic acid
phenethyl ester on testicular torsion/detorsion injury in rats.
Anal Quant Cytopathol Histpathol 2018;40:270-276
25. Shokoohi M, Shoorei H, Soltani M, Abtahi-Eivari SH, Salim-
nejad R, Moghimian M: Protective effects of the hydroal-
coholic extract of Fumaria parviflora on testicular injury
induced by torsion/detorsion in adult rats. Andrologia 2018;
50(7):e13047
132 Analytical and Quantitative Cytopathology and Histopathology®
Dursun et al