https://doi.org/10.52973/rcfcv-e34365
Received: 13/12/2023 Accepted: 10/01/2024 Published: 28/02/2024
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Revista Científica, FCV-LUZ / Vol. XXXIV, rcfcv-e34365
ABSTRACT
The aim of this study was to evaluate the possible therapeutic
effect of propolis on cyclophosphamide (CP)–induced testicular
lipid peroxidation and on the associated changes in spermatological
parameters in epididymal spermatozoa and histopathological
structure of rat testis. Rats were randomly separated into 4 groups
with 7 rats in each group. Groups were formed as; 1
st
group: Control
group (untreated rats), 2
nd
group: Propolis–treated group, 3
rd
group:
CP–treated group, and 4
th
group: CP+Propolis–treated group. Propolis
was administered to the rats at dose of 200 mg·kg bw
-1
by gavage for
7 days (d). CP was administered to the rats at a single dose of 150
mg·kg bw
-1
intraperitoneally. Propolis administration was started 2 d
before CP administration and continued for 7 d. Malondialdehyde (MDA)
and reduced glutathione (GSH) levels, catalase (CAT), glutathione
peroxidase (GSH–Px), glutathione S–transferase (GST), and superoxide
dismutase (SOD) activities, spermatological parameters, the weight
of the reproductive organs, and the histopathological structure
were determined. Compared to the control group, MDA levels and
decreased, it was not found changed in GSH levels and GSH–Px
activities in CP group. In the CP–treated group, there was decreased
in sperm motility in epididymal spermatozoa, sperm density in
epididymal spermatozoa, and weight of testis, prostate, epididymis
and vesicula seminalis; while there was increased in abnormal sperm
ratio compared to the control group in epididymal spermatozoa.
Propolis normalized biochemical and spermatological parameters
in epididymal spermatozoa. The histopathological examination of
change such as cell debris, invagination and degeneration occurred
in CP group. In the pathogenesis of CP–induced testicular toxicity
may play role the deterioration in oxidant–antioxidant balance and
propolis may reduce severe side effects of CP–induced alterations.
Key words: Antioxidant; cyclophosphamide; malondialdehyde;
oxidative stress; propolis
RESUMEN
El objetivo de este estudio fue evaluar el posible efecto terapéutico
del propóleo sobre la peroxidación lipídica testicular inducida por
ciclofosfamida (CP) y sobre los cambios asociados en los parámetros
espermatológicos en los espermatozoides epididimarios y la
estructura histopatológica de los testículos de rata. Las ratas se
separaron aleatoriamente en 4 grupos con 7 ratas en cada grupo. Se
formaron grupos como; 1er grupo: grupo control (ratas no tratadas),
2do grupo: grupo tratado con propóleo, 3er grupo: grupo tratado
con CP y 4to grupo: grupo tratado con CP+propóleo. Se administró
propóleo a las ratas en una dosis de 200 mg·kg pc
-1
mediante
alimentación forzada durante 7 días (d). Se administró CP a las
ratas en una dosis única de 150 mg·kg pc
-1
por vía intraperitoneal. La
administración de propóleo se inició 2 d antes de la administración de
CP y continuó durante 7 d. Niveles de malondialdehído (MDA) y glutatión
reducido (GSH), actividades de catalasa (CAT), glutatión peroxidasa
(GSH–Px), glutatión S–transferasa (GST) y superóxido dismutasa (SOD),
parámetros espermatológicos, peso de los órganos reproductivos.
y se determinó la estructura histopatológica. En comparación con
el grupo de control, los niveles de MDA y las actividades de SOD
disminuyeron, no se encontraron cambios en los niveles de GSH y las
actividades GSH–Px en el grupo CP. En el grupo tratado con CP, hubo
una disminución en la motilidad de los espermatozoides del epidídimo,
la densidad de los espermatozoides en los espermatozoides del
epidídimo y el peso de los testículos, la próstata, el epidídimo y la
vesícula seminal; mientras que hubo un aumento en la proporción de
espermatozoides anormales en comparación con el grupo de control
en los espermatozoides epididimarios. El propóleo normalizó los
parámetros bioquímicos y espermatológicos en los espermatozoides
epididimarios. El examen histopatológico del tejido testicular mostró
celulares, invaginación y degeneración, ocurrieron en el grupo CP.
En la patogénesis de la toxicidad testicular inducida por la PC puede
desempeñar un papel el deterioro del equilibrio oxidante–antioxidante
y el propóleo puede reducir los efectos secundarios graves de las
alteraciones inducidas por la PC.
Palabras clave: Antioxidante; ciclofosfamida; malondialdehído;
estrés oxidativo; propóleos
Protective effect of propolis on the antioxidant enzymes activities,
characteristics of epididymal spermatozoa and histopathological structure
of testis from rats treated with cyclophosphamide
Efecto protector del propóleo sobre las actividades de las enzimas antioxidantes,
las características de los espermatozoides epididimarios y la estructura
histopatológica de los testículos de ratas tratadas con ciclofosfamida
Emre Kaya
1
* , Seval Yılmaz
1
, Zülal Altay
1
, Şeyma Özer Kaya
2
, Neriman Çolakoğlu
3
, Emine Sarman
4
1
Firat University, Faculty of Veterinary Medicine, Department of Biochemistry. Elazig, Türkiye.
2
Firat University, Faculty of Veterinary Medicine, Department of Reproduction and Articial Insemination. Elazig, Türkiye.
3
Firat University, Faculty of Medicine, Department of Histology and Embryology, Elazig, Türkiye.
4
Afyonkarahisar Health Sciences University, Faculty of Medicine, Department of Histology and Embryology. Afyon, Türkiye.
*Corresponding author: emrekaya@rat.edu.tr
Role of propolis in cyclophosphamide-induced testicular toxicity / Kaya et al. ______________________________________________________
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INTRODUCTION
Chemotherapy is the administration of antineoplastic drugs to slow
down, regress or stop the progression of neoplastic disease in many
cancer treatments. The most carcinogenic antineoplastic drugs are
those of the alkylating type. The most important factor limiting the use
of alkylating drugs in chemotherapy applications is that they cause
multiple organ toxicities. This prevents these types of drugs from
1]. Cyclophosphamide (CP) is
one of the most widely used antineoplastic agents today. It is used for
the treatment of chronic and acute leukemias, myelomas, lymphomas
and bone marrow transplantation. In addition, CP is thought to have
2]. Apart from its tumor–
selective activity, it also has many toxic side ef3].
Cyclophosphamide, a cytotoxic drug, undergoes hydroxylation in
the liver and turns into its metabolites phosphoramide mustard and
acrolein. CP exerts its antineoplastic effects with phosphoramide
mustard. Phosphoramide suppresses cell division by binding to
nucleic acids in the mustard organism and helps the antitumor
effects of CP. The main toxic effect of CP is known to be due to
4]. Metabolites alkylate Deoxyribonucleic
acid (DNA) and proteins, leading to cytotoxicity and the formation
5]. Their cytotoxic effects are caused by the
irreversible combination of the electrophilic alkyl radical in their
structure and the nucleophilic part of the target macromolecules
6]. Acrolein interacts with the antioxidant system in cells and tissues
and causes the formation of free radicals. These metabolites interact
with systems that produce reactive oxygen species (ROS), such
as hydrogen peroxide (H
2
O
2
), superoxide radical (O
2
–
·). In addition,
increased ROS causes DNA damage and disruption of the oxidant/
antioxidant balance, leading to cellular dysfun7].
Because testicular germinal epithelial cells have high mitotic
activity, chemotherapy drugs can cause temporary or permanent
infertility by affecting the functions of the reproductive organs
and reducing the sperm count. Chemotherapeutics cause lipid,
cholesterol and protein peroxidation and oxidative stress, causing
adverse effects such as DNA damage, apoptosis, decreased sperm
quality, infertility and sterility, especially in spermatogenic cells, in
8]. This effect is mostly caused by cytotoxic
chemotherapeutics. Chief among these is CP. Alkylating agents (CP)
are particularly harmful to the germinal epithelium and are effective
by disrupting the DNA repair mechanism in the late spermatogenesis
9]. Although the exact mechanism underlying the toxicity of
CP to testicles and other organs is unknown, a number of studies
have demonstrated that tissue redox balance can be upset by CP
exposure, raising the possibility that oxidative stress is the cause
10, 11, 12]. Long–
term azoospermia may occur in 90–100% of patients with lymphoma
treated with chemotherapeutic agents containing CP. It also takes
about 2 years for spermatogenesis to return to no13].
Propolis is a sticky, resinous substance that is collected and
deposited from the leaves and stems of different plant buds by the
honey bee (Apis mellifera L.) and protects the hive from external
14]. Numerous biological
properties of propolis have been reported, including antioxidant,
cytostatic, antimutagenic and immunomodulatory, antimicrobial,
hepatoprotective, antitumoral, and immunostimulating. These
and terpenoid contents. For these reasons, propolis is widely used
in apitherapy and medical applications15, 16].
This study was aimed to investigate the effects of propolis
application against testicular toxicity that may be caused by CP
by examining malondialdehyde (MDA), reduced glutathione (GSH)
levels, antioxidant enzyme activities, spermatological parameters
in epididymal spermatozoa and some reproductive organ weights
and the histopathological structure of the testis.
MATERIALS AND METHODS
Animals and working order
In this study, Wistar–Albino male rats (Rattus norvegicus) (3–month–
old) weighing 250–300 g, obtained from the Firat University Laboratory
Animals Breeding Unit, were used. This study was conducted with
the ethical approval of the Firat University Animal Experiments Local
Ethics Committee (Protocol No: 2014/10). The rats were kept in air–
°C and 60–65%
humidity, with a 12/12h dark/light cycle, under standard conditions.
They were fed on standard rat food (pellet) and tap water ad libitum
throughout the experimental practices. Experimental practices on rats
were performed in the Firat University Experimental Research Center.
Experimental protocol
In this study, with seven rats apiece, four groups of rats were formed:
Groups were formed as; 1
st
group: Control group (untreated rats), 2
nd
group: Propolis–treated group, 3
rd
group: CP–treated group, and 4
th
group: CP+propolis–treated group. Propolis was administered to the
rats at dose of 200 mg·kg bw
-1
16, 17]. The
CP was administered to the rats at a single dose of 150 mg·kg bw
-1
16, 18]. Propolis administration was started 2 d before
CP administration and continued for 7 d. Propolis is prepared daily by
dissolving in 40% ethanol. In the testis tissue, the levels of MDA and
GSH, as well as the activity of antioxidant enzymes like catalase (CAT),
glutathione peroxidase (GSH–Px), glutathione S–transferase (GST) and
superoxide dismutase (SOD) were measured spectrophotometrically.
Biochemical analyses
When the experiment was finalized, the rats in control and
Maxwellstraat 11, 6716 BX Ede,
Holland)
Malondialdehyde and GSH levels and the activities of CAT, GSH–Px,
GST, and SOD were analyzed in testis tissues by spectrophotometrically
The MDA level was measured by spectrophotometry as per the
method of Placer et al19]. This method was based on the reaction
of MDA with TBA, one of the aldehyde products of lipid peroxidation.
GSH level was determined by the method of Ellman et al20]. This
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method was a spectrophotometric method based on the formation of
highly stable yellow color of sulfhydryl groups when 5,5'dithiobis–2–
nitrobenzoic acid (DTNB) was added. CAT activity was evaluated by
21]. CAT activity was determined by measuring
the resolution of H
2
O
2
at 240 nm. GSH–Px activity was measured by
22]. GSH–Px catalyzes the oxidation of GSH to
oxide glutathione (GSSG) using H
2
O
2
. The rate of formation of GSSG
was measured by the GR reaction. GST activity was measured by the
method of Habig et al.23]. Enzyme activity was determined with
spectrophotometry by measuring the amount of enzyme catalyzing
1 μ
SOD activity was measured by using xanthine and xanthine oxidases
to generate O
2
–
·
,
concentration was determined based on the method of Lowry et al.25].
Sperm analyses
Epididymal spermatozoon motility
A glass slide was put on the microscope's stage, and the microscope
placed on the left cauda epididymis, and a few drops of the Tris
buffer solution —which contains tris (hydroxymethyl) aminomethane
were dropped onto the slide. By cutting a piece of the left cauda
epididymis, a tiny drop of semen was extracted and placed on the
slide. To guarantee homogeneity, the resultant sample was mixed
the motility rate (%) was 26].
Epididymal spermatozoon density
For every animal, the right cauda epididymis was taken apart and
weighed. After that, it was cut and broken up using a knife and forceps
in 1 mL of 0.9% NaCl for a 2 min period in a petri dish. To enable the
spermatozoa in the epididymal tissue to pass into the solution, the
tissue was left at room temperature for 4 h. After being collected, the
spermatozoa–containing supernatant was diluted 1:200. In order to
allow the spermatozoa in the solution to uniformly distribute over the
entire area, 10 μL of the diluted mixture was transferred to a Thoma
slide, covering both areas. The slide was then incubated for 5 min.
By counting the spermatozoa in each square in both counting areas
density was determ26].
Abnormal spermatozoon rate
A tiny drop of semen suspension extracted from the left cauda
epididymis was combined with a few drops of Tris buffer solution
froti was painted using a Diff Quik painting set. Initially, the slide was
30 s. Eosin Y solution was added to cover the slide and incubated
for 20 s after the excess solution was removed. After removing any
leftover dye, Thiazine dye was applied to the slide and it was incubated
for an additional 30 s. After being cleaned with distilled water, the
painted slide was allowed to dry. Using a light microscope (Olympus
were examined. 200 spermatozoa in all were analyzed in the smear,
and the percentages of abnormal spermatozoa in the head, tail, and
26]. For all spermatological analyses, a phase–
contrast microscope (Nikon E 200, Tokyo, Japan) was use
The absolute weight of the reproductive organs such as V. seminalis,
prostate, right and left testis, and epididymis were determined
by using a precision weighing device (Precisa BJ 410C, Precisa
Instruments AG, CH–Dietikon, TYP 160–9422–050/P, Switzerland).
Histopathological examination
ethanol and dehydrated in a succession of ethanol concentrations
that steadily increased. The tissues were cleaned in xylene and then
Haematoxylin and eosin was used to stain the tissue blocks, which
were sliced into slices that were 5 μm t27].
Statistical analyses
Using the SPSS 22 software (Version 22.0; SPSS, Chicago,
was assessed. To determine if the raw values of all the measured
parameters showed a normal distribution, the Shapiro–Wilk normality
test was employed. The test's results showed that all of the parameter
values did. One–way analysis of variance (ANOVA) was used to evaluate
group differences based on the results of this test, and the post hoc
Tukey test was employed to compare the two groups. The mean and
mean and standard error. P–values less than 0.05 were considered
istically.
RESULTS AND DISCUSSIONS
Biochemical results
In the testis tissue of the control and experimental groups,
the levels of MDA and GSH as well as the activities of antioxidant
enzymes such CAT, GSH–Px, GST, and SOD are shown in TABLE I.
The information showed that CP group levels of MDA in the testis
tissue were considerably greater than those of the control group
(P<0.001). When compared to the CP group, the MDA level appeared
to normalize during treatment with CP+propolis group. When all
groups were compared, no statistical change was detected in GSH
levels (P>0.05). No change was detected in GSH levels when both the
control and CP+propolis groups, compared to CP applied group. While
CAT activities decreased in the CP group compared to the control
group (P=0.024), no change was detected when compared to the CP
group and CP+propolis (P=0.516). When all groups were compared,
no statistical change was detected in GSH–Px levels (P=0.644). No
change was detected in GSH–Px levels when both the control and
CP+propolis groups, compared to CP applied group. While GST
activities decreased in the CP group compared to the control group
(P=0.035), no change was detected when compared to the CP group
and CP+propolis (P=0.784). A statistical increase in SOD activities
was determined in the CP group compared to both the control group
(P=0.014) and the CP+propolis group (P=0.020). There is no statistical
difference between the propolis applied group and the control group
in all biochemical parameters.
Role of propolis in cyclophosphamide-induced testicular toxicity / Kaya et al. ______________________________________________________
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Spermatological results
TABLES II and III demonstrate the differences between the control
and experimental groups in terms of spermatological characteristics
in epididymal spermatozoa and reproductive organ weight.
The total abnormal sperm ratio increased whereas epididymal sperm
concentration and sperm motility were considerably lower in the CP
group in epididymal spermatozoa (P
sperm concentration and motility as well as a decrease in the overall
rate of abnormal sperm were seen in the CP+propolis group compared
to the CP group (P<0.001). In comparison to the control group, there
concentrations, sperm motility, or overall abnormal sperm rate
between the propolis group and the CP+propolis group (TABLE II).
Histopatological results
Testicular tissue was observed in normal structure in the control
and only propolis applied groups. In the CP–administered group,
cell debris of the immature spermatogenic series in the lumen of
some seminiferous tubules (ST), invaginations in the tubule basal
membrane, and occasional interstitial edema were distinguished.
Degeneration was detected in the epithelium of some ST. In the
CP+propolis group, the epithelium of the ST was observed close to
the control group. In this group, cell debris belonging to the immature
spermatogenic series was distinguished in a few tubules, but it was
determined that propolis had a very protective effect against CP
toxicity (FIG. 1).
The basis of cancer chemotherapy; to stop the growth and proliferation
of tumor cells or, if possible, to destroy them without damaging the
patient's normal cells. Because there is not much qualitative difference
between malignant cell and normal cell; the difference is more
quantitative. Therefore, most cancer drugs have side effects on normal
cells and blood tissue. In cancer patients, the toxic side effects of cancer
drugs have been the subject of increasing stu28].
Antitumoral activity of CP, a chemotherapeutic drug with wide
clinical use, proven to be effective in the treatment of cancer and
non–malignant diseases (such as lupus erythematosus, Behçet's
disease, vasculitis, immune diseases, systemic connective tissue
disease, rheumatoid arthritis, autoimmune hemolytic anemia,
nephrotic syndrome), attributed to its use in high doses. While high–
dose alkylating (affecting DNA) agents such as CP destroy cancer
cells, they adversely affect normal tis29].
TABLE I
The eect of propolis supplementation on rat testis MDA and GSH levels and CAT, GSH–Px, GST, and SOD activities
MDA
(nmol·mL
–1
)
GSH
(µmol·mL
–1
)
CAT
(kg·g prot.
–1
)
GSH–Px
(U·g prot.
–1
)
GST
(U·mg prot.
–1
)
SOD
(U·mg prot.
–1
)
Control 0.46 ± 0.04 15.94 ± 0.20 6.58 ± 0.25 124.72 ± 2.95 59.92 ± 2.15 20.60 ± 0.51
Propolis 0.51 ± 0.10 16.59 ± 0.50 7.61 ± 0.15 134.13 ± 3.03 57.99 ± 1.55 21.54 ± 0.23
CP 0.76 ± 0.02*
a
16.22 ± 0.82 5.14 ± 0.31* 137.30 ± 3.67 51.20 ± 1.12* 23.45 ± 0.32*
a
CP+Propolis 0.52 ± 0.01 15.14 ± 0.66 5.97 ± 0.27 130.43 ± 1.24 54.51 ± 1.78 21.04 ± 0.51
*: It means that it is statistically dierent compared to the control group.
a
: It means that it is statistically dierent compared to
the CP+propolis group.
P–values less than 0.05 were considered signicant statistically. MDA: Malondialdehyde, GSH: Reduced
Glutathione, CAT: Catalase, GSH–Px: Glutathione Peroxidase, GST: Glutathione S–Transferase, SOD: Superoxide Dismutase
TABLE II
The eect of propolis supplementation on sperm motility, sperm
concentration and abnormal sperm rate in epididymal spermatozoa
Control Propolis CP CP+Propolis P
Motility (%) 82.0 ± 2.49
a
80.0 ± 0.10
a
41.43 ± 3.45
b
60.0 ± 4.95
ab
P<0.05
Concentration
(million/right
cauda epididymis)
122.0 ± 6.56
a
124.0 ± 5.77
a
91.71 ± 6.15
b
122.75 ± 6.72
a
P<0.05
Abnormal
sperm rate (%)
6.60 ± 0.34
b
5.0 ± 0.57
c
10.14 ± 0.34
a
7.50 ± 0.28
b
P<0.05
Within rows, means with dierent letters (
a, b
) are signicantly dierent (P<0.05)
in the absolute weight of the right and left testis in CP group (P<0.001).
When compared with CP group, increases were observed in the
absolute weight of the right and left testis in CP+propolis group
(P
testis weight (TABLE III).
weight of prostate in other groups compared to the control group.
in the absolute weight of the right and left epididymis and seminal
at between CP group and CP+propolis (TABLE III)
TABLE III
The eect of propolis supplementation on testis,
epididymis, v. seminalis and prostat weights
Control Propolis CP CP+Propolis P
Testis (g)
P<0.05
Right 1.99 ± 0.08
a
1.94 ± 0.08
a
1.49 ± 0.02
c
1.71 ± 0.06
ab
Left 1.91 ± 0.12
a
1.96 ± 0.05
a
1.45 ± 0.02
b
1.69 ± 0.06
ab
Epididymis Weight (g)
P<0.05Right 0.60 ± 0.03
ab
0.65 ± 0.01
a
0.52 ± 0.01
c
0.54 ± 0.01
bc
Left 0.60 ± 0.02
ab
0.63 ± 0.01
a
0.49 ± 0.01
c
0.53 ± 0.02
bc
V. Seminalis Weight (g) 1.64 ± 0.11
a
1.46 ± 0.01
ab
1.13 ± 0.06
c
1.22 ± 0.05
abc
P<0.05
Prostat Weight (g) 47.0 ± 0.03 47.0 ± 0.04 45.0 ± 0.02 35.0 ± 0.05
P>0.05
Within rows, means with dierent letters (a, b and c) are signicantly dierent (
P<0.05)
FIGURE 1. Control group (A): Seminiferous tubule epithelium (blue arrow) and interstitial Leydig cells (black arrow) are observed in normal
structure. Hematoxylin & Eosin 200×; CP group (B): Immature cells spilled into the lumen of the seminiferous tubule (red star), invaginations
of the basement membrane of the seminiferous tubule (arrow) and interstitial edema (black star). Triple painting of Masson 200×; (C):
Degeneration of the seminiferous tubule epithelium (*) in CP group. Hematoxylin & Eosin 200×; CP+Propolis group (D): Seminiferous tubule
epithelium (arrow) and interstitial Leydig cells (*) are distinguished in normal structure. Triple painting of Masson 200×
_____________________________________________________________________________Revista Cientifica, FCV-LUZ / Vol. XXXIV, rcfcv-e34365
5 of 9
30
31
32, 33
34
35
36, 37, 38, 39].
Male germ cells are highly sensitive to chemotherapeutic agents.
Many chemotherapeutic agents can cross the blood–testicular barrier
and cause permanent damage to germ cells (with hyalinization and
dose and duration. In addition, the maximum threshold doses at
which each chemotherapeutic agent can harm are also different.
Among the CP alkylators, they are the chemotherapeutics with the
40]. CP, which has a very strong toxic
effect on the germinal epithelium, can reduce the fertility potential
of children treated with this drug in the prepubertal period by up
41]. Similarly, in children treated with CP for
42, 43]
44] is claimed to be due to failures in
sperm regenera45].
In experimental rat studies with CP, decreased sperm concentration
and motility, apoptotic germ cell count, increase in the ratio of
dead and abnormal sperm, severe testicular structure disorders
32, 4646, 47] decreases in type a spermatogonia,
10] and at
48], changes in sperm
49], and double strand
50]. Many of the
Role of propolis in cyclophosphamide-induced testicular toxicity / Kaya et al. ______________________________________________________
6 of 9
reproductive side effects of CP in rats are also observed in albino
mice (Mus musculus)51, 52].
Whether the intervention of antioxidants during cancer
chemotherapy affects the efficacy of treatment or reduces
undesirable side effects is a subject of intense research. If ROS
production by a cancer chemotherapeutic agent plays an important
role in its cytotoxicity, it is likely that antioxidants interfere with
the drug's anti–neoplastic activity. However, if ROS is primarily
responsible for the drug's side effects, antioxidants can actually
reduce the severity of such effects without affecting its effectiveness.
Therefore, it is important to distinguish between the ability of a drug
to induce oxidative stress in biological systems and the role of free
radicals in the mechanism of action of the drug under investigation.
CP is an alkylating agent and its tumor cell killing activity is mainly
due to DNA alkylation. However, free radical production by acrolein
is often associated with undesirable toxic eff53].
et al.32
et al., 33
may cause.
The degree of ROS produced in the environment is directly
stress, cellular damage, and necrosis are brought on by an excess
of ROS in response to CP. These effects are mediated by a number
of processes, including protein denaturation, membrane lipid
54]. Numerous studies show that
CP exposure raises the creation of ROS inside cells, and that the
resulting physiological and biochemical abnormalities may be the
result of the emerging oxidative stress. CP has been demonstrated
to function as an oxidant precursor; by causing oxidative stress and
raising lipid peroxidation in critical organs, it lowers the activity levels
16, 32, 33, 34, 35].
Malondialdehyde is one of the often used techniques for predicting
55]. This is based on the observation that the
loss of one hydrogen atom from the unsaturated fatty acid chains
caused by ROS during the LPO process results in elevated levels of
16]. In the current investigation, the markedly elevated MDA
concentrations in the testis tissue of the CP–treated rats appear
to be the consequence of elevated ROS levels brought on by the
stress the rats' CP poisoning caused. The administration of Propolis,
however, restored the elevated MDA levels to those observed in the
control group. This suggests that Propolis may squelch free radicals,
impede the LPO process, and avert oxidative damage to the rat testis’s
membrane lipids. The persistent generation of free radicals and the
compromised defense mechanisms against antioxidants are thought
to be linked to the elevated MDA levels in CP–treated animals. One
theory for the cause could be that conditions that encourage the
generation of free radicals eventually target the cell membrane,
16]. The oxidative damage caused by CP may have OH
radicals as the initiating species. The protective effects of GSH and
interact directly as a cofactor or coenzyme with the –SH moiety,
just like ROS can. Because biological membranes are vulnerable
16]. The current study's
observed decline in GST activity may indicate that the cell is used more
GST to potentially combat ROS generation during CP metabolism.
in the CP–administered groups, this shows that the CAT enzyme is
affected by CP application by separating H
2
O
2
into water and oxygen
H
2
O
2
Cengiz et al.54], in their study aiming to determine the protective
effects of boron on CP (200 mg·kg
-1
)–induced testicular toxicity,
determined a decrease in Bcl–2, TAC and GSH levels, and an increase in
TOC, OSI, MDA, Bax and Caspase–3 levels. Accordingly, they concluded
that CP application may cause damage to the testicle. Alkhalaf et al.
56], after CP (200 mg·kg
-1
ip single dose) application, evaluated sperm
counts, motility, viability and abnormalities, testosterone, luteinizing
hormone and follicle stimulating hormone levels, as well as parameters
such as MDA, nitric oxide and total antioxidant capacity. In their study,
they showed that CP disrupts the redox balance in testicular tissues
and therefore disrupts testicular functions by negatively disrupting
sperm characteristics, hormonal levels and testicular histology.
They also concluded that CP disrupts the oxidative balance. Many
studies such as these have emphasized that CP may cause oxidative
damage in testicular tissue at different doses and cause negativities
16, 32, 33, 34, 35]. In common with all of
them, it is emphasized that CP may cause these effects by causing a
deterioration in the redox balance in the testicular tissue. In addition,
in the aforementioned studies, the effects of some substances with
high antioxidant activity against the negativities caused by the effect
of CP by showing effects in different ways were also mentioned.
CP results in irregular ST, reduced seminiferous epithelial layers,
of intertubular tissue, and histopathological reduction in the size
and number of ST. It also causes degeneration and vacuolation in
57].
et al32], in their study examining the effects of lycopene
and ellagic acid against CP toxicity, found a decrease in the diameter
of the ST and the thickness of the germinal cell layer in the testicular
tissue after CP application, as well as degeneration, necrosis,
immature germ cells, congestion and atrophy. In the current study,
conditions such as the immature cells spilled into the lumen of the
ST, invaginations of the basement membrane of the ST, interstitial
edema, and degeneration of the ST epithelium were encountered.
In the current study, it was observed that the Propolis it was used
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to biochemical, spermatological and histopathological results).
In the present literature review, which is not tissue and organ
(such as liver, kidney, heart) against CP toxicity in different doses and
16, 36, 37, 39]. The fact that no study has been found in
which the effects of propolis on the testicular tissue of CP in rats was
evaluated using biochemical, spermatological and histopathological
data increases the originality of this study.
Many studies examining the positive effects of Propolis on testicular
tissue have concluded that Propolis reduces tissue damage due to
its strong antioxidant activity. In some studies conducted on rats,
58] (Propolis 100
mg·kg
-1
59] (Propolis 100 mg·kg
-1
),
60] (Propolis 100 mg·kg
-1
), against
61] (Propolis 50 mg·kg
-1
), against the
62] (Propolis 50 mg·kg
-1
),
63] (Propolis 100 mg·kg
-1
), against the
17] (Propolis 200 mg·kg
-1
) and positive effects
were obtained. What is common in all studies is that propolis corrects
the negative effects of Propolis on sperm quality and testicular tissues
against different toxic substance exposures by increasing antioxidant
activities and/or scavenging free radicals.
in Propolis are important compounds that prevent the effects of
by interacting with the peroxy radicals of unsaturated fatty acids.
their activities in removing peroxide ions, H
2
O
2
, lipid peroxide and
antioxidant effects by inhibiting lipoxygenase and cyclooxygenase
16, 38, 64].
CONCLUSIONS
The improvement in antioxidant biomarkers with propolis
application can be explained by the ability of propolis to prevent
oxidative stress and free radicals by limiting the production of free
radicals. It can be said that Propolis, in addition to its protective
role of the cell membrane by inhibiting LPO, can reduce the toxic
side effects of cytotoxic agents such as CP, thanks to its interaction
with antioxidants.
However, in order to make a more accurate evaluation in
spermatological studies in rats or other species, spermatogenesis
times according to animal species should be taken into account. In
this study, it was tried to talk about the effects on the last stage of
spermatogenesis and/or spermatozoa in epididymal spermatozoa.
Conict of interests
declared.
Ethics approval and consent to participate
The Firat University Animal Studies Local Ethics Committee
accepted the experiments (Protocol No: 2014/10), which were carried
out strictly in compliance with the Experimental Animal Ethics
Committee's Guiding Principles.
Disclosure statement
The authors state that there are no interests at odds with one another.
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