AMELIORATIVE ROLE OF QUERCETIN AND/OR RESVERATROL ON ...
Transcript of AMELIORATIVE ROLE OF QUERCETIN AND/OR RESVERATROL ON ...
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
92
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
AMELIORATIVE ROLE OF QUERCETIN AND/OR RESVERATROL
ON ACROLEIN-INDUCED CLASTOGENESIS IN BONE MARROW
CELLS OF MALE ALBINO MICE MUS MUSCULUS
Nagla Zaky Ibrahim El-Alfy1, Mahmoud Fathy Mahmoudand
2 and Sally Ramadan
Gabr El-Ashry3*
1Professor of Cytogenetics,
2Professor of Cytology and Histology,
3PhD Researcher
Department of Biological and Geological Sciences, Faculty of Education, Ain Shams
University, PO11341, Cairo, Egypt.
ABSTRACT
Acrolein, a highly reactive unsaturated aldehyde, is considered as a
mutagenic environmental pollutant which can cause oxidative stress by
generation of reactive oxygen species. The aim of the present study is
to investigate the cytoprotective role of oral pretreatment with
quercetin (50 mg/ kg body weight) alone, resveratrol (12.5 mg/ kg
body weight) alone and the mixture of both quercetin/ resveratrol
against the clastogenesis of acrolein (10 mg/ kg body weight) in bone
marrow cells of male albino mice Mus musculus by using
chromosomal aberration assay, mitotic index and micronucleus test as
toxicological endpoints. In this study, quercetin and/ or resveratrol
were given to mice for eight days (four days prior to acrolein treatment
followed by other four days along with acrolein treatment). The results
revealed that oral administration of acrolein to mice for four consecutive days significantly (P
< 0.001) increased the incidence of aberrant metaphases, structural and numerical
chromosomal aberrations, micronuclei formation and cytotoxicity in bone marrow cells in
comparison with the control group. Also, pretreatment of mice with quercetin and/ or
resveratrol significantly (P < 0.001) reduced acrolein-induced clastogenesis and cytotoxicity
in the bone marrow cells. No considerable difference was observed between the
cytoprotective effects of quercetin alone, resveratrol alone and also the mixture of both
quercetin/ resveratrol against acrolein-induced clastogenesis and cytotoxicity. However, oral
pretreatment of quercetin alone showed the best protective effect against acrolein-toxicity.
*Corresponding Author
Sally Ramadan Gabr El-
Ashry
PhD Researcher, Department
of Biological and Geological
Sciences, Faculty of
Education, Ain
ShamsUniversity, PO11341,
Cairo, Egypt.
Article Received on
07 March 2021,
Revised on 28 March 2021,
Accepted on 18 April 2021
DOI: 10.20959/wjpps20216-18910
WORLD JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES
SJIF Impact Factor 7.632
Volume 10, Issue 5, 92-114 Research Article ISSN 2278 – 4357
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
93
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
Therefore, natural foods rich in plant polyphenols, particularly quercetin and resveratrol,
should be included in the human daily diet to protect them against the deleterious effects of
clastogenic agents like acrolein.
KEYWORDS: Clastogenesis, Bone marrow cell, Chromosomal aberration, Mitotic index,
Micronucleus test, Mice.
1. INTRODUCTION
Acrolein is a reactive α, ß unsaturated aldehyde which can react with many cellular molecules
including amino acids, proteins and nucleic acids and can arise DNA adducts, DNA damage
and inhibition of DNA repair.[1]
The sources of acrolein that are most relevant to human
exposure are grouped into dietary, endogenous and environmental sources.[2]
It is formed in
cooked foods from carbohydrates, vegetable oils, animal fats and amino acids during heating
of foods and exists naturally in some vegetables, fruits and herbs.[3]
Environmentally, the
ubiquitous presence of acrolein is attributed to incomplete combustion of petroleum fuels,
biodiesel, paper, wood and plastic, smoking of tobacco products and frying of foods in oils.[4]
In addition, acrolein can be generated endogenously by several methods during cellular
metabolism as degradation of threonine by neutrophil derived myeloperoxidase, amine
oxidase mediated catabolism of polyamines such as spermine and spermidine, metabolism of
cancer drugs such as cyclophosphamide and lipid peroxidation of polyunsaturated fatty
acids.[2]
Acrolein interferes with the tissue antioxidant defense system, produces highly
reactive oxygen free radicals[5]
and suppresses the activities of antioxidant enzymes such as
superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT).[6]
The
increased level of acrolein adducts and oxidative stress have been observed in plasma of
patients with cancer[7]
, renal failure[8]
, Alzheimer's disease[9]
and diabetes.[10]
Plant polyphenols are natural antioxidants scavenging free radicals and chelating transition
metal ions.[11]
Moreover, experimental studies have confirmed the selective cytotoxic activity
of polyphenols in cancer cells and the lack of any such action or at least only minimal
cytotoxicity in normal cells.[12,13]
Quercetin (Q) is a polyphenolic flavonoid compound present in large amounts in vegetables,
fruits and tea and it exhibits therapeutic potential including hepatoprotection and the
inhibition of liver fibrosis.[14,15]
The antioxidant activity of quercetin is related to the presence
of a phenolic hydroxyl group in its chemical stucture which is a good hydrogen donor and
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
94
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
can react with reactive oxygen and reactive nitrogen species in a termination reaction which
breaks the cycle of generation of new radicals.[16]
During cancer chemotherapy, quercetin is
suggested to play a role in reducing cytogenotoxicity induced by cyclophosphamide by
decreasing oxidative stress and inflammation.[17]
Resveratrol (RES) is a polyphenol and a powerful antioxidant which is known to have anti-
inflammatory, anti-neoplastic, anti-platelet aggregation, anti-fibrotic, anti-allergic and anti-
aging actions and occurs naturally in red grapes, mulberries and peanuts.[18]
The direct
antioxidant effects of resveratrol are probably related to the presence of hydroxyl groups that
can trap reactive oxygen species on the stilbenic skeleton.[19]
No statistically significant
difference was observed in the mean of total chromosomal aberration frequencies in mouse
bone marrow cells of the control group and resveratrol (100 mg/kg) orally administrated
group.[20]
Many reports have observed the anti-mutagenic and anti-carcinogenic activity of
resveratrol by its ability to prevent DNA damage and to increase DNA repair.[21,22]
Accordingly, the present study was designed to evaluate the effect of quercetin and/ or
resveratrol on clastogenesis induced by acrolein.
2. MATERIALS AND METHODS
2.1. Animals
Adult male mice (CD1) of nearly the same age (16-18 weeks old) with an average body
weight 22-26 g (mean 24 ± 2 g) were obtained from the closed colony of Theodor Bilharz
Research Institute, Cairo. All animals were kept under suitable laboratory conditions of
humidity, temperature (25°C ± 2) and light (12 hr light/12 hr dark) and they were fed on
standard rodent pellet diet and supplied with water, during the whole period of
experimentation. All animal procedures in the present study were conducted in accordance
with the ethical guidelines for investigations in laboratory animals and comply with the guide
for the care and use of laboratory animals.[23]
The study also approved by the research ethics
committee of Ain Shams University, Egypt.
2.2. Chemicals
The chemicals used in the present investigation were acrolein (Acr), quercetin (Q) and
resveratrol (Res). They were purchased from Sigma Aldrich (St. Louis, MO, USA). Acrolein
was obtained in the form of clear, colorless liquid, sensitive to light, heat and air. Appropriate
acrolein concentration (10 mg/ kg b. wt.) was freshly prepared directly before treatment by
dilution with distilled water and protected from light and heat. Quercetin (Q) was obtained in
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
95
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
the form of yellow crystalline powder and its dose (50 mg/ kg b. wt.) was freshly prepared
daily before treatment by suspending it in corn oil and protected from light. Resveratrol was
obtained in the form of off-white to pale yellow powder. Its stock solution was prepared by
dissolving 100 mg of powder in 2 ml of absolute ethanol and storing it in freezing
temperature (at or below 4°C) at dark condition then the required resveratrol concentration
(12.5 mg/ kg b. wt.) was freshly prepared by adding sterile distilled water. The doses were
converted from human dose to mice dose by using multiplication factors for dose conversion
between different species by Paget & Barnes.[24]
2.3. Experimental design
One hundred mice (CD-1) were acclimated for one week and randomly assigned to one of ten
groups, comprising of ten animals each as follows: Group 1: mice served as a control group
and received distilled water (the acrolein’s solvent) orally at same volume as acrolein
treatment. Group 2: mice served as quercetin-vehicle control group and received corn oil
(the quercetin’s vehicle) orally at same volume as quercetin for eight consecutive days.
Group 3: mice served as resveratrol-vehicle control group and received 2% ethyl alcohol
(the resveratrol’s vehicle) orally at same volume as resveratrol for eight consecutive days.
Group 4: mice received oral dose of acrolein (10 mg/ kg) for four consecutive days. Group
5: mice received oral dose of quercetin (50 mg/ kg) for eight consecutive days. Group 6:
mice received oral dose of quercetin (50 mg/ kg) alone for four consecutive days directly
prior to oral treatment with both quercetin (50 mg/ kg) followed by acrolein (10 mg/ kg) for
other four consecutive days. Group 7: mice received oral dose of resveratrol (12.5 mg/ kg)
for eight consecutive days. Group 8: mice received oral pretreatment with resveratrol (12.5
mg/ kg) alone for four consecutive days directly prior to oral treatment with both resveratrol
(12.5 mg/ kg) followed by acrolein (10 mg/ kg) for other four consecutive days. Group 9:
mice received both quercetin (50 mg/ kg, orally) and resveratrol (12.5 mg/ kg, orally) for
eight consecutive days. Group 10: mice received oral pretreatment with quercetin and
resveratrol (50 mg/ kg, 12.5 mg/ kg, respectively) for four consecutive days directly prior to
oral treatment with both quercetin and resveratrol followed by acrolein (10 mg/ kg) for other
four consecutive days. Oral administration of acrolein was separated from regular quercetin
and/or resveratrol treatment by one hour. All the control and treated animals were sacrificed
by cervical dislocation at 24 hr after the last treatment for collection of samples.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
96
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
2.4. Chromosomal aberration analysis
Each animal was weighed alive prior to the intraperitoneal injection with colchicine solution
(0.2 ml/ 100 g) and they were sacrificed after two hours. Bone marrow chromosome
preparations were carried out according to the method of Preston et al.[25]
Observation was
made using bright field and most photographs were taken with a 100x oil object. The best
photograph with a spread metaphase stage was chosen for making a karyotype. One hundred
well spread metaphase plates per mouse (500 metaphases for each group) were examined in
the control and all treated groups for both structural and numerical aberration.
2.5. Mitotic index
From the prepared slides, mitotic index (MI) was evaluated by counting the divided cells
among at least 1000 metaphase spreads per each group (five animals per each group) and
expressed in percentage.
2.6. Micronucleus test
In the present study Schmid’s[26]
standard procedure was followed however with slight
modification. Instead of foetal calf serum, 5% bovine albumin (from National Research
Center, Giza, Egypt) was used as suspending medium to collect the bone marrow.[27]
At the
end of experiment, Mice were sacrificed and femurs were trimmed and a blunt needle was
pushed to pierce the marrow cavity. The marrow was flushed through a syringe with 5%
bovine albumin to obtain a fine suspension which was centrifuged at 1000 rpm for 8 to 10
minutes. Then, the supernatant was discarded and fresh suspending medium was added. A
smear was prepared (3-4 slides/animals), slides were air dried overnight, fixed by methanol
for 5 minutes, dried, stained by May-Grunwald and then with a combination of May-
Grunwald and phosphate buffer at pH 6.8 for the proper color differentiation of
polychromatic erythrocytes (PCE’s) and normochromatic erythrocytes (NCE’s). Finally
slides are stained with Geimsa and buffer at pH 6.8 for micronuclei staining. After washing
with distilled water and buffer good slides were dried and mounted. Two thousand PCE’s
were screened per animal and micronucleated PCE’s (MNPCE’s) were recorded.
Consequently, identified normochromatic erythrocytes (NCE’s) were also recorded. Finally,
the percentage of MNPCE and PCE/NCE ratio were calculated for each animal.
2.7. Data analysis
Statistical analyses were carried out using the package software SPSS/PC computer program
(version 16.0). All values were expressed as mean and standard deviation (mean ± SD). The
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
97
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
independent samples T-test was used for comparison between means of two groups to
determine if the difference between means is statistically significant or due to sampling error.
A value of (P < 0.05) was considered significant, (P < 0.001) was considered highly
significant and (P > 0.05) was considered insignificant. Chart was drawn using Excel 2010.
3. RESULTS
3.1. Chromosomal aberration analysis
Current results as shown in Table 1 revealed that acrolein (10 mg/ kg) administration for four
consecutive days significantly (p < 0.001) increased the incidence of structural and numerical
chromosomal aberrations in the bone marrow cells of male mice in comparison to the
corresponding control. Table 1 and Figure 1 illustrated that acrolein administration induced
various chromosomal aberrations, which were then further classified into structural
aberrations such as centromeric attenuation (Ca), chromosome gap (Chg), chromatid gap
(Cg), centric fusion (CF), ring chromosome (R), beaded chromosomes (Bch), end to end
association (Ee), pulverized chromosomes (P), sticky chromosomes (S), deletion (D) and
fragment and numerical aberrations in the form of polyploidy. However, the mean of
centromeric attenuation, fragment, deletion and sticky chromosomes were higher as
compared to other chromosomal aberrations. On the other hand, oral pretreatment of plant
polyphenols (quercetin and/or resveratrol) prior to acrolein treatment significantly (p < 0.001)
decreased the mean of total aberrations comparable to that of acrolein-administrated group
(Group 4) which indicated the protective role of quercetin and/ or resveratrol against
clastogenicity of acrolein. In addition, this study observed that pretreatment of quercetin prior
to acrolein treatment gave more effective reduction in the total chromosomal aberrations and
abnormal metaphases than pretreatment of resveratrol or pretreatment of both quercetin/
resveratrol as shown in Figure 2. Quercetin (50 mg/kg) and resveratrol (12.5 mg/kg) were
also not observed to be clastogenic, as there were no significant changes in the frequency of
chromosomal aberrations over the corresponding quercetin-vehicle control group and the
corresponding resveratrol-vehicle control group, respectively.
3.2. Mitotic index
Mitotic index was employed to analyze potential toxicity of acrolein and potential protective
role of quercetin and/ or resveratrol in terms of cell proliferation. Table 2 and Figure 3
revealed that after the oral administration of acrolein dose (10 mg/ kg) for four consecutive
days to mice of group 4, the percentage of mitotic index was highly significant (p < 0.001)
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
98
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
decreased compared to the corresponding control group, confirming bone marrow
suppression. Moreover, a highly significant increase in the mitotic index was observed as a
result of pretreatment of quercetin and/or resveratrol prior to acrolein treatment, indicating
the cytoprotective role of them against acrolein-induced bone marrow suppression as shown
in Table 2 and Figure 3. Quercetin (50 mg/kg) and resveratrol (12.5 mg/kg) were also not
observed to be cytotoxic, as the oral pretreatment of each of them alone induced a highly
significant (P < 0.001) increase in the mitotic index over the corresponding quercetin-vehicle
control group and the corresponding resveratrol-vehicle control group, respectively.
3.3. Micronucleus test
The results of micronucleus test were summarized in Table 3. As shown in Figure 4, the
polychromatic erythrocytes were stained light blue to gray and normochromatic erythrocytes
were stained light pink to light yellow. Figure 5 showed that polychromatic erythrocytes with
micronuclei (MNPCEs) are polychromatic erythrocytes have one or more small nuclei (dark
blue in color) as a residual hereditary material remained after erythropoiesis process.
Acrolein induced a significant (P < 0.05) increase in the frequency of micronucleated
polychromatic erythrocytes over the corresponding control. On the other hand, mice treated
with quercetin and/ or resveratrol before and along with acrolein treatment showed a
significant (P < 0.05) reduction in the incidence of micronucleated polychromatic
erythrocytes (MNPCEs) as compared to the acrolein treated group. Oral pretreatment with
quercetin before acrolein administration showed more remarkable decrease in MNPCEs
frequencies than pretreatment with resveratrol alone or with both quercetin/resveratrol.
Moreover, assessment of the ratio of polychromatic erythrocytes to corresponding
normochromatic erythrocytes (PCE/NCE) of control and all treated groups was used as an
index of cytotoxicity and an indicator of the acceleration or inhibition of erythropoiesis. As
shown in Table 3, oral administration of acrolein alone resulted in a highly significant
increase in cytotoxicity of bone marrow cells by decreasing the ratio of PCE/NCE less than
the corresponding control. Also, the anti-cytotoxic potential of quercetin and/ or resveratrol
caused a significant reduction in the cytotoxicity of bone marrow cells by increasing the ratio
of PCE/NCE more than the corresponding acrolein- treated group. Treatment of mice with
quercetin or resveratrol alone without acrolein did not induce any significant variation in the
incidence of micronucleated polychromatic erythrocyte (MNPCE) as compared to the
corresponding quercetin-vehicle control group and the corresponding resveratrol-vehicle
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
99
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
control group, respectively. In addition, quercetin and resveratrol were not cytotoxic to the
bone marrow and did not cause a significant decrease in the ratio of PCEs/NCEs.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
100
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
Table 1: Average of chromosomal aberration observed in bone marrow cells of male albino mice Mus musculus of the control and all
treated groups (mean±standered deviation).
Group Treatment (mg/ kg)
Mean number (mean±SD) and types of structural and numerical chromosomal
aberrations Total aberrations
(Mean±SD) Ca Chg & Cg CF R Bch Ee P S D F Po
1 Control 12.4±
2.5
8.2±
1.05
6.06±
1.22
10.8±
1.3
1.4±
1.2
7.26±
2.8
2±
0.4
8.3±
1.5
10.1±
4.2
14.1±
.7 0
80.8±
0.220
2 Q-vehicle 17.6±
3.1
9.9±
2.5
6.4±
1.2
12.4±
2.5
0.6±
1.1
9.9±
2.5
1.6±
1
13.1±
0.8
11.2±
3.2
14.5±
1.9 0
97.6±
7.600
3 Res-vehicle 20.2±
2.2
3.4±
1.3
6.6±
1.4
8.4±
2.4
0.8±
1.3
7.2±
2.5
4.5±
0.5
6.7±
1.5
12.1±
2.5
16.1±
1.2
1.8±
0.6
88.1±
0.980
4 Acr (10) 40.4± 2.2
⁎ 17.8± 2.4
⁎ 11.2± 1.2
⁎ 16.9± 0.5
⁎⁎ 6.4± 1.3
⁎ 20.1± 2.2
⁎ 18.3± 1.5
⁎ 22.8± 3.3
⁎ 25.4± 5.06
⁎ 29.9± 2.7
⁎ 13.7± 3.4
⁎ 223.3± 0.940
⁎⁎
5 Q (50) 22.4±
2.5⁎
4.2±
2.8
5±
2.5
9.6±
2.02⁎
4.4±
0.5⁎
10.6±
2.08
7.3±
2.5⁎
13.4±
1.3
7.4±
2.5⁎
9.1±
1.8⁎
1.2±
0.7
94.9±
1.680
6 Q (50) + Acr (10) 32.4± 2.5
⁎ 6.1± 1.2
⁎ 5.4± 1.3
⁎ 11.4± 1.2
⁎ 4.1± 1.8
10± 5
7.5± 0.4
⁎ 19.2± 1.6
⁎ 8.8± 1.3
⁎ 22.5± 2.5
⁎ 2.14±
0.4 129.8± 1.880
⁎⁎
7 Res (12.5) 19.4±
2.6 8.3± 2.8
⁎
7.4± 2.7
8± 2.6
4.9± 2.5
⁎
6.7± 1.5
6.7± 0.8
⁎
12.4± 0.4
⁎⁎
11± 1.2
18.4± 1.4
⁎
0
103.4± 1.740
8 Res (12.5) + Acr
(10)
24±
3.6⁎
11.2±
3.3⁎
9.1±
1.8⁎
14.8±
2.3⁎
2.4±
2.5
13±
2.6⁎
7.6±
1.3⁎
12.4±
2.5⁎
12.4±
6.5
24.1±
6.3⁎
1.8±
0.5⁎
135.1±
1.960⁎⁎
9 Q (50) + Res (12.5) 21±
3.6⁎
7.4±
2.5
3.8±
1.3⁎
15.1±
0.9⁎
2.1±
2
14.6±
9.2⁎
3.2±
1.05⁎
20.3±
5.03⁎
11.8±
1.6
15.2±
4.1
1.4±
1.2
116.2±
1.180
10 Q (50) + Res (12.5)
+ Acr (10) 24.3± 5.1
⁎ 8.8± 3.3
⁎ 6.6± 1.5
⁎ 15.4± 1.7
⁎ 1.6± 1.44
10.7± 2.8
⁎ 4.5± 1.1
⁎ 22.1±
3 19.9± 6.6
⁎ 18.2± 3.7
⁎ 1.9± 0.9
134.5± 0.480
⁎⁎
Five hundred metaphases were scored for chromosomal aberrations per each group (five mice were examined per each group). The significances
were indicated as follow: *Significant (P < 0.05) and **Highly Significant (P < 0.001). Ca, centromeric attenuations; Cg, chromatid gaps; Chg,
chromosome gaps; CF, centric fusions; R, rings; Bch, beaded chromosomes; Ee, end to end association; P, pulverization; S, stickiness; D,
deletion; F, fragments; Po, polyploidy.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
101
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
Table 2: Effect of oral pretreatment with quercetin and/ or resveratrol on the score of
divided cells and the percentage of mitotic index (MI %) in bone marrow cells of male
albino mice treated with acrolein (mean±standered deviation).
Group Treatment (Dose mg/ kg)
No. of
examined
mice
No. of
examined
cells/ mice
Score of
divided cells (Mean± SD)
Percentage of
mitotic index (MI %)
(Mean± SD)
1 Control 5 1000 96.2± 4.86826 9.6200± 0.48683
2 Q-vehicle 5 1000 71.6± 8.5322 7.1600± 0.85323
3 Res- vehicle 5 1000 81.8± 4.43847 8.1800± 0.44385
4 Acr (10) 5 1000 53.2±
3.96232⁎⁎
5.3200± 0.39623⁎⁎
5 Q (50) 5 1000 100± 3.93700⁎⁎
10.0000±
0.39370⁎⁎
6 Q (50) + Acr (10) 5 1000 96.2±
4.60435⁎⁎
9.6200± 0.46043⁎⁎
7 Res (12.5) 5 1000 93.6±
4.82701⁎⁎
9.3600± 0.48270⁎⁎
8 Res ( 12.5) + Acr (10) 5 1000 92.4±
3.36155⁎⁎
9.2400± 0.33615⁎⁎
9 Q (50) + Res (12.5) 5 1000 120± 2.00000⁎⁎
12.0000±
0.20000⁎⁎
10 Q (50) + Res (12.5) + Acr
(10) 5 1000
112.8±
13.81304⁎⁎
11.2600±1.38130⁎⁎
The significances were indicated as follow: ⁎
Significant (P < 0.05) and ⁎⁎
Highly Significant
(P < 0.001). MI, mitotic index.
Table 3: The mean and standard deviation of micronucleated polychromatic
erythrocytes (MNPCEs) and PCEs/ NCEs ratio in 6000 polychromatic erythrocytes
(PCEs) and corresponding normochromatic erythrocytes (NCEs) scored in the bone
marrow of three male albino mice Mus musculus of the control group and treated
groups.
Group
Treatment
(Dose mg/
kg)
Total
scored cells/
No. of mice
Total
MNPCEs
Micronuclei
MNPCEs/Total PCEs %
(Mean± SD)
Total
NCEs
Cytotoxicity
PCEs/ NCEs
(Mean± SD)
1 Control 6000/3 47 0.7777± 0.25455 5988 1.0017±
0.00764
2 Q-vehicle 6000/3 119 1.9800± 0.62960
5469 1.0933±
0.01528
3 Res- vehicle 6000/3 143 2.3867± 0.19630 5790 1.0333± 0.
03512
4 Acr (10) 6000/3 373 6.2200± 0.85159⁎
6533 0.9180±
0.00700⁎⁎
5 Q (50) 6000/3 103 1.7200± 0.25534 5570 1.0733±
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
102
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
0.02082
6 Q (50) + Acr
(10) 6000/3 147 2.4400± 0.25534
⁎ 5685
1.0500±
0.04000⁎⁎
7 Res (12.5) 6000/3 117 1.9400± 0.19053 5420 1.1033±
0.01155
8 Res (12.5) +
Acr (10) 6000/3 158 2.6367± 0.17214
⁎ 5265
1.1333±
0.00577⁎⁎
9 Q (50) + Res
(12.5) 6000/3 80 1.3300± 0.17000 5920
1.0147±
0.07333
10
Q (50) + Res
(12.5) + Acr
(10)
6000/3 153 2.5533± 0.42253⁎
5930 1.0100±
0.05196⁎
The significances were indicated as follow: ⁎
Significant (P < 0.05) and ⁎⁎
Highly Significant
(P < 0.001). MNPCEs, micronucleated polychromatic erythrocytes; PCEs, polychromatic
erythrocytes; NCEs, normochromatic erythrocytes.
Figure 1: Metaphase chromosomes in mice bone marrow cells. (a) Normal chromosomes
of control group; (b, c and d) Different types of structural chromosomal aberrations in
bone marrow cells of acrolein treated mice of group 4 including Ca, centromeric
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
103
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
attenuations; Cg, chromatid gaps; Chg, chromosome gaps; CF, centric fusions; R,
rings; Bch, beaded chromosomes; Ee, end to end association; D, deletion and F,
fragments; (e) P, pulverization which is an extreme fragmentation of chromosomes as a
result of acrolein treatment; (f) Po, polyploidy was the common form of numerical
change which was observed as a result of acrolein treatment. Scale bar, 0.05 µm.
Figure 2: Different types of chromosomal aberrations in mice bone marrow cells
because of pretreatment of quercetin and/or resveratrol prior to acrolein treatment
including Ca, centromeric attenuations; Cg, chromatid gaps; CF, centric fusions; R,
rings; Ee, end to end association; S, stickiness; D, deletion; F, fragments. (a and b) Mice
of group 6 received oral pretreatment of quercetin alone prior to acrolein treatment; (c
and d) Mice of group 8 received oral pretreatment of resveratrol alone prior to acrolein
treatment; (e and f) Mice of group 10 received oral pretreatment of both quercetin and
resveratrol prior to acrolein treatment. Scale bar, 0.05 µm.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
104
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
Figure 3: Chart represents the percentages of mitotic index (MI %) of bone marrow
cells of male albino mice in the control group and all treated groups.
Figure 4: Normal bone marrow smears of male albino mice Mus musculus showing
polychromatic erythrocyte (PCE) and normochromatic erythrocyte (NCE). (a) Control
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
105
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
group; (b) Quercetin-vehicle control group; (c) Resveratrol-vehicle control group; (d)
Quercetin-treated group; (e) Resveratrol-treated group; (f) Quercetin/resveratrol-
treated group. Scale bar, 0.05 µm.
Figure 5: Bone marrow smears of male albino mice Mus musculus showing
polychromatic erythrocytes (PCE), normochromatic erythrocytes (NCE) and
micronucleated polychromatic erythrocytes (MNPCE). (a, b and c) Acrolein-treated
group; (d) Quercetin/acrolein-treated group; (e) Resveratrol/acrolein-treated group; (f)
Quercetin and resveratrol/acrolein-treated group. Scale bar, 0.05 µm.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
106
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
4. DISCUSSION
The current study investigated the protective effects of the plant polyphenols, quercetin and
resveratrol, against the genotoxicity of acrolein by measuring the frequency of chromosomal
aberrations, mitotic index and micronuclei in bone marrow cells of male albino mice Mus
musculus. Also, the previous study of[28]
investigated the genotoxicity of the widely
prescribed drugs depakine and/or epanutin in mice by using chromosomal aberration assay.
Quercetin and resveratrol were selected because they have been considered representative,
more abundant in human nutrition and more promising in terms of their positive effects in
previous studies.[29,30]
In the present study, chromosomal aberration assay indicated that the
oral administration of acrolein (10 mg/kg) highly significant (P < 0.001) increased the
incidence of structural and numerical chromosomal aberrations in bone marrow cells of male
albino mice. Also, previous in vitro experiments observed that acrolein induced chromosomal
aberrations, sister chromatid exchanges, point mutations and inhibition of DNA repair in
cultured human lymphocytes[31]
and mammalian cells[32]
due to its high binding affinity for
proteins and low molecular weight sulfhydryl compounds such as glutathione and cysteine.
Current study of bone marrow chromosomes revealed that centromeric attenuation, fragment,
deletion and sticky chromosomes occurred more frequently after acrolein administration. The
present finding coincided with the previous study of Saxena et al.[33]
which observed that
acrolein intraperitoneal administration (1 mg/kg/day) to female albino rats for 10 days
induced significant differences in term of chromosome aberrations including chromosome
break (both chromosome and chromatid type), gap and fragmentation.
The data of the present study showed that the oral administration of acrolein (10 mg/kg) for
four consecutive days significantly inhibited mitosis and cell proliferation in bone marrow
tissue of treated male albino mice when compared to that of the control. This finding can be
explained by the ability of acrolein to bind strongly with sulfhydryl groups, reduce proteins
synthesis, disrupt the function of many enzymes, deplete cellular glutathione contents,
decrease G0/G1 phase, decrease nuclear division index and centromere protein and inhibit the
formation of spindle fibers.[34]
Another possible mechanism explaining the cytotoxicity of
acrolein was involved in the study of Horton et al.[35]
which demonstrated that acrolein
inhibited the nuclear factor kappa B (NF-κB) controlling several genes, including those
involved in cell proliferation and apoptosis.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
107
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
Micronucleus test was included in many studies to detect genotoxic effect of many classes of
chemicals in mammalian system.[36-38]
Also, it was reported as the most reliable and widely
used bioassay to assess DNA damage in mammalian cells in vivo, because it could detect
genomic alterations resulting from chromosomal damage and/or damage to the mitotic
apparatus caused by clastogenic agents.[39]
In view of that, this test was used in the present
study to measure the genetic damage in bone marrow cells of treated male albino mice in
comparison with the control group. The current data of the micronucleus test showed a
significant increase (p < 0.05) in the mean of micronucleated polychromatic erythrocytes
(MNPCEs) in mice after the oral administration of acrolein (10 mg/kg) for four consecutive
days when compared to the corresponding control. The present investigation was confirmed
by the data from the previous study of Moghe et al.[40]
which revealed that acrolein induced
micronuclei and DNA damage in human, rats and mouse. In addition, Aydın et al.[41]
demonstrated that the oral administration of acrolein (5 mg/kg/day) six days per week for
30 days increased the frequency of micronuclei and decreased the ratio of polychromatic
erythrocytes (PCEs) in bone marrow of treated rats when compared to control. Also, Habibi
et al.[42]
observed that acrolein could interfere with cellular DNA causing DNA strand breaks,
DNA disintegration, structural and numerical chromosomal aberrations, formation of
micronucleated polychromatic erythrocytes and necrosis. The suppression of immature
erythrocytes (PCE) in relation to mature erythrocytes (NCE) causes decreasing in the ratio of
PCE to NCE which is considered as an important index of cytotoxicity that is routinely
included in micronucleus tests to assess the mutagenicity of chemicals to mammals.[43]
The
results of the present study revealed that the mean of PCE/NCE ratio was significantly (p <
0.001) decreased by acrolein administration (10 mg/kg/day) for four days when compared to
control indicating the cytotoxicity of acrolein in bone marrow tissue. It was reported that
spleen captured and destroyed erythrocytes with micronuclei quickly.[44]
Moreover, previous
study of Ahmed et al.[45]
considered acrolein as an eryptosis stimulating molecule and
indicated its ability to cause loss of erythrocytes, impaired formation of erythrocytes, red
blood corpuscles life span shortening and anemia.
The present investigation demonstrated that neither quercetin nor resveratrol were clastogenic
or cytotoxic at the doses tested. Also, Attia[46]
observed that quercetin at doses equivalent to
50 or 100 mg/kg failed to induce chromosomal aberrations in bone marrow cells of mice
which indicated its non-clastogenicity. The protective effects of quercetin in rodent models of
liver disease were maximal when administered at 50 mg/kg.[47]
In addition, the protective
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
108
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
effect of oral pretreatment of resveratrol dose (12.5 mg/kg body weight) was investigated
against the genotoxicity of acrolein according to Mokni et al.[48]
who confirmed that the
optimal protective effect of resveratrol on antioxidant enzyme activities and lipoperoxidation
products was obtained at this dose.
In the current study, oral pretreatment of quercetin was able to protect mice bone marrow
cells against the acrolein-induced clastogenicity by significantly (p < 0.001) decreasing the
mean of total structural and numerical chromosomal aberrations induced by acrolein
administration. This result coincided with Sekeroğlu & Sekeroğlu[49]
who observed a
decrease in the number of chromosomal aberrations induced by an anticancer drug called
methotrexate in bone marrow cells after quercetin administration to mice at a dosage of 50
mg/kg body weight.
Results of the present investigation observed that resveratrol exhibited a cytoprotective role
against the toxicity of acrolein in the bone marrow cells by significantly (p< 0.001)
decreasing the mean of total aberrations comparable to that of alone acrolein administrated
mice. This finding was confirmed previously by various investigators who examined the
antigenotoxic effect of resveratrol. Türkez & Sisman[50]
demonstrated that the high
concentrations of resveratrol were not genotoxic and could minimize the frequency of
chromosome aberrations and sister chromatid exchanges caused by aflatoxin in human
lymphocytes. Türkez & Aydin[51]
observed that the combined application of the most popular
environmental pollutant, Permethrin and the plant derived antioxidant, resveratrol,
significantly reduced the frequency of chromosomal aberrations and the formation of sister
chromatid exchanges in cultured human lymphocytes in comparison with alone Permethrin-
treated cultures.
The present data of micronucleus assay revealed that the antigenotoxic potential of the plant
polyphenols (quercetin and/or resveratrol) protected bone marrow tissue of mice against
genotoxicity induced by acrolein by significantly reducing the mean of (MNPCEs) and
significantly increasing the ratio of (PCE/NCE) when compared to acrolein-treated group.
Previously, quercetin was observed to be effective in decreasing the micronuclei frequency
and protecting cultured rat peripheral lymphocytes against nicotine-induced cellular and
DNA damage.[52]
Importantly, quercetin was able to enter erythrocytes to prevent the
oxidative damage induced by acrolein in erythrocytes because of glutathione depletion which
was followed by the release of free iron, lipid peroxidation and consequent hemolysis.[53]
The
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
109
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
role of resveratrol to prevent chromosomal abnormalities and reduce micronuclei formation
was explained by its ability to restore the levels of intracellular antioxidants such as
glutathione peroxidase, superoxide dismutase and catalase activity.[54]
Also, the stilbenic
structure of resveratrol was responsible for its strong antioxidant activity due to the presence
of a hydroxyl group which could trap reactive oxygen species.[55]
5. CONCLUSION
The findings of the present investigation demonstrated that acrolein, a toxic
cyclophosphamide metabolite and a major component in the gas phase of cigarette smoking,
automobile exhaust, over-heated oils, fried foods and forest fires, was highly clastogenic and
cytotoxic as it induced a very harmful genetic damage in the examined bone marrow cells of
male albino mice Mus musculus. Therefore, acrolein exposure should be limited by regulating
its levels in foods and in tobacco products or preventing its huge emissions into environment.
In addition, the observed protective effects of quercetin and resveratrol indicated that they
can be used as nutritional supplements or as a part of functional foods for geno protective
treatment against acrolein induced genotoxicity in vivo.
REFERENCES
1. Tang MS, Wang HT, Hu Y, Chen WS, Akao M, Feng Z, Hu W. Acrolein induced DNA
damage, mutagenicity and effect on DNA repair. Mol Nutr Food Res, 2011; 55: 1291–
1300.
2. Stevens JF, Maier CF. Acrolein: sources, metabolism and biomolecular interactions
relevant to human health and disease. Mol Nutr Food Res, 2008; 52: 7–25.
3. Abraham K, Andres S, Palavinskas R, Berg K, Appel KE, Lampen A. Toxicology and
risk assessment of acrolein in food. Mol Nutr Food Res, 2011; 55: 1277–1290.
4. Esterbauer H, Schaur RJ, Zollner H. Chemistry and biochemistry of 4-hydroxynonenal,
malonaldehyde and related aldehydes. Free Radical Biol Med, 1991; 11: 81–128.
5. Mythili Y, Sudharsan PT, Selvakumar E, Varalakshmi P. Protective effect of DL-alpha-
lipoic acid on cyclophosphamide induced oxidative cardiac injury. Chem Biol Interact,
2004; 151: 13-19.
6. Dumontet C, Drai J, Thieblemont C, Hequet O, Espinouse D, Bouafia F, Salles G,
Coiffier B. The superoxide dismutase content in erythrocytes predicts short term toxicity
of high dose cyclophosphamide. Br J Haematol, 2001; 112: 405-409.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
110
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
7. Zarkovic K, Uchida K, Kolenc D, Hlupic L, Zarkovic N. Tissue distribution of lipid
peroxidation product acrolein in human colon carcinogenesis. Free Radic Res, 2006; 40:
543-552.
8. Sakata K, Kashiwagi K, Sharmin S, Ueda S, Irie Y, Murotani N, Igarashi K. Increase in
putrescine, amine oxidase, and acrolein in plasma of renal failure patients. Biochem
Biophys Res Commun, 2003; 305: 143–149.
9. Lovell MA and Markesbery WR. Ratio of 8-hydroxyguanine in intact DNA to free 8-
hydroxyguanine is increased in Alzheimer disease ventricular cerebrospinal fluid. Arch
Neurol, 2001; 58: 392–396.
10. Daimon M, Sugiyama K, Kameda W, Saitoh T, Oizumi T, Hirata A, Yamaguchi H,
Ohnuma H, Igarashi M, Kato T. Increased urinary levels of pentosidine, pyrraline and
acrolein adduct in type 2 diabetes. Endocr J, 2003; 50: 61–67.
11. Hu JP, Calomme M, Lasure A, De Bruyne T. Pieters L, Vlietinck A, Vanden Berghe DA.
Structure activity relationships of flavonoids with superoxide scavenging ability. Boil
Trace Elem Res, 1995; 47: 327–331.
12. Shammas MA, Neri P, Koley H, Batchu RB, Bertheau RC, Munshi V, Prabhala R,
Fulciniti MF, Tai YT, Treon SP, Goyal RK, Anderson KC, Munshi NC. Specific killing
of multiple myeloma cells by (−)-epigallocatechin-3-gallate extracted from green tea:
biologic activity and therapeutic implications. Blood, 2006; 108: 2804–2810.
13. Feng R, Ni HM, Wang SY, Tourkova IL, Shurin MR, Harada H. Cyanidin-3-rutinoside, a
natural polyphenol antioxidant, selectively kills leukemia cells by induction of oxidative
stress. J Biol Chem, 2007; 282: 13468–13476.
14. Gonzalez-Gallego J, Sanchez-Campos S, Tunon MJ. Anti-inflammatory properties of
dietary flavonoids. Nutr Hosp, 2007; 22: 287–293.
15. Tieppo J, Cuevas MJ, Vercelino R, Tunon MJ, Marroni NP, Gonzalez-Gallego J.
Quercetin administration ameliorates pulmonary complications of cirrhosis in rats. J
Nutr., 2009; 139: 1339–1346.
16. Chobot V. Simultaneous detection of pro- and anti-oxidative effects in the variants of the
deoxyribose degradation assay. J Agric Food Chem., 2010; 58: 2088–2094.
17. Sekeroğlu V, Aydin B, Sekeroğlu ZA. Viscum album L. extract and quercetin reduce
cyclophosphamide induced cardiotoxicity, urotoxicity and genotoxicity in mice. Asian
Pac J Cancer Prev, 2011; 12: 2925-2931.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
111
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
18. El-Sheikh AAK, Morsy MA, Al-Taher AY. Protective mechanisms of resveratrol against
methotrexate-induced renal damage may involve BCRP/ABCG2. Fundamental & Clinical
Pharmacology, 2016; 30: 406–418.
19. Frombaum M, Le Clanche S, Bonnefont-Rousselot D, Borderie D. Antioxidant effects of
resveratrol and other stilbene derivatives on oxidative stress and NO bioavailability:
potential benefits to cardiovascular diseases. Biochimie, 2012; 94: 269–276.
20. Carsten RE, Bachand AM, Bailey SM and Ullrich RL. Resveratrol reduces radiation-
induced chromosome aberration frequencies in mouse bone marrow cells. Radiat Res,
2008; 169: 633–638.
21. Chakraborty S, Roy M, Bhattacharya RK. Prevention and repair of DNA damage by
selected phytochemicals as measured by single cell gel electrophoresis. J Environ Pathol
Toxicol Oncol, 2004; 23: 215–226.
22. Alturfan AA, Tozan-Beceren A, Sehirli AO, Demiralp E, Sener G, Omurtag GZ.
Resveratrol ameliorates oxidative DNA damage and protects against acrylamide-induced
oxidative stress in rats. Mol Biol Rep., 2012; 39: 4589–4596.
23. Institute of Laboratory Animal Resources. Guide for the care and use of laboratory
animals, Committee for The Update of The Guide and Use of Laboratory Animals,
National Research Council of The National Academies, 8th
ed., Washington, D.C.;
National Academy Press, 1996.
24. Paget GE, Barnes JM Interspecies dosage conversion scheme in evaluation of results and
quantitative application in different species, In: Evaluation of drug activities (Eds.).
Pharmacometrics, London and New York; Academic Press, 1964; 160-162.
25. Preston RJ, Brian JD, Sheila G. Mammalian in vivo cytogenetic assays, Analysis of
chromosomal aberrations in mouse bone marrow cells. Mutat Res., 1987; 189: 157-165.
26. Schmid W. The Micronucleus Test For Cytogenetic Analysis, In: Hallaender A (Eds.).
Chemical Mutagens - Principles And Methods For Their Detection, New York; Plenum
Press, 1976; 31- 53.
27. Narayan K, D’Souza UJ, Rao KPS. The genotoxic and cytotoxic effects of ribavirin in rat
bone marrow. Mutat Res., 2002; 521: 179-185.
28. El –Alfy NZI, Mahmoud MF, Alqosaibi AI, Abdullah AM. Genotoxic effects of
Depakine and/or Epanutin on male albino mice Mus musculus: cytological and molecular
study. World Journal of Pharmacy and Pharmaceutical Sciences, 2020a; 9: 352-365.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
112
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
29. Grove KA, Lambert JD. Laboratory, epidemiological, and human intervention studies
show that tea (Camellia sinensis) may be useful in the prevention of obesity. J Nutr.,
2010; 140: 446–453.
30. Ramprasath VR, Jones PJH. Anti-atherogenic effects of resveratrol. European Journal of
Clinical Nutrition, 2010; 64: 660–668.
31. Wilmer JL, Erexson GL, Kligerman AD. Effect of acrolein on phosphoramide mustard-
induced sister chromatid exchanges in cultured human lymphocytes. Cancer Research,
1990; 50: 4635-4638.
32. Storme T, Deroussent A, Mercier L, Prost E, Re M, Munier F, Martens T, Bourget P,
Vassal G, Royer J, Paci A. New ifosfamide analogs designed for lower associated
neurotoxicity and nephrotoxicity with modified alkylating kinetics leading to enhanced in
vitro anticancer activity. J Pharmacol Exp Ther., 2009; 328: 598–609.
33. Saxena AK, Singh D, Singh G. Acrolein induces unilateral hypertrophy and associated
histopathalogical changes during germ cell differentiation in mature rat ovary.
Biomedical Research, 2009; 20: 89-93.
34. Zhang S, Chen H, Zhang J, Li J, Hou H, Hu Q. The multiplex interactions and molecular
mechanism on genotoxicity induced by formaldehyde and acrolein mixtures on human
bronchial epithelial BEAS-2B cells. Environment International, 2020; 143: 1059432.
35. Horton ND, Biswal SS, Corrigan LL, Kehrer JP. Acrolein causes inhibitor κB-
independent decreases in NF-κB activation in human lung adenocarcinoma (A549) cells.
J Biol Chem., 1999; 274: 9200–9206.
36. El-Alfy NZ, Alqosaibi AI, Mahmoud MF, El-Ashry SRG. An analysis of micronuclei and
DNA damage induced by methotrexate treatment of male albino mice. The Egyptian
Journal of Hospital Medicine, 2016; 65: 504–514.
37. El-Alfy NZ, Alqosaibi AI, Mahmoud MF, Emam AA. Role of propolis against
monosodium glutamate genotoxicity by chromosomal aberration, micronucleus test and
comet assay in males. Der Pharmacia Lettre, 2020 b; 12: 13–22.
38. El-Alfy NZ, Alqosaibi AI, Mahmoud MF, Abdallah AM. Appraisal of genotoxicity and
cytotoxicity of Depakine and/or Epanutin in bone marrow erythrocytes and hepatocytes
of male albino mice by comet and micronucleus assays. Medico Legal Update, 2021; 21:
638-643.
39. Hayashi M. The micronucleus test-most widely used in vivo genotoxicity test. Genes
Environ, 2016; 38: 18.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
113
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
40. Moghe A, Ghare S, Lamoreau B, Mohammad M, Barve S, McClain, C, Joshi-Barve S.
Molecular Mechanisms of Acrolein Toxicity: Relevance to Human Disease.
Toxicological Sciences, 2015; 143: 242–255.
41. Aydın B, Atlı Şekeroğlu Z, Şekeroğlu V. Acrolein-induced oxidative stress and
genotoxicity in rats: protective effects of whey protein and conjugated linoleic acid. Drug
Chem Toxicol, 2018; 41: 225-231.
42. Habibi E, Shokrzadeh M, Ahmadi A, Chabra A, Naghshvar F, Keshavarz-Maleki R.
Genoprotective effects of Origanum vulgare ethanolic extract against cyclophosphamide-
induced genotoxicity in mouse bone marrow cells. Pharm Biol, 2015; 53: 92-97.
43. Celik A, Mazmanci B, Camlica Y, Askin A, Comelekoglu U. Cytogenetic effects of
lambda-cyhalothrin on Wistar rat bone marrow. Mutat Res, 2003; 539: 91–97.
44. Shimada K, Yamamotoa M, Takashimaa M, Wakob Y, Kawasakob K, Aokia Y, Sekia J,
Miyamaea Y, Wakata A. Repeated dose liver micronucleus assay of mitomycin C in
young adult rats. Mutat Res Gene Toxicol Environ. Mutagen, 2015; 780: 85-89.
45. Ahmed MS, Langer H, Abed M, Voelkl J, Lang F. The uremic toxin acrolein promotes
suicidal erythrocyte death. Kidney Blood Press Res, 2013; 37: 158–167.
46. Attia SM. The impact of quercetin on cisplatin induced clastogenesis and apoptosis in
murine marrow cells. Mutagenesis, 2010; 25: 281-288.
47. Moreira AJ, Fraga C, Alonso M, Collado PS, Zetller C, Marroni C, Marroni N, González-
Gallego J. Quercetin prevents oxidative stress and NF-kappa B activation in gastric
mucosa of portal hypertensive rats. Biochem Pharmacol, 2004; 68: 1939–1946.
48. Mokni M, Elkahoui S, Limam F, Amri M, Aouani E. Effect of resveratrol on antioxidant
enzyme activities in the brain of healthy rat. Neurochem Res, 2007; 32: 981-987.
49. Sekeroğlu ZA, Sekeroğlu V. Effects of Viscum album L. extract and quercetin on
methotrexate induced cytogenotoxicity in mouse bone marrow cells. Mutation Res, 2012;
746: 56–59.
50. Türkez H, Sisman T. The genoprotective activity of resveratrol on aflatoxin B₁induced
DNA damage in human lymphocytes in vitro. Toxicol Ind Health, 2012; 28: 474-480.
51. Türkez H, Aydin E. The genoprotective activity of resveratrol on permethrin induced
genotoxic damage in cultured human lymphocytes. Braz Arch Biol Technol, 2013; 56:
405-411.
52. Muthukumaran S, Sudheer AR, Nalini N, Menon VP. Effect of quercetin on nicotine-
induced biochemical changes and DNA damage in rat peripheral blood lymphocytes.
Redox Rep, 2008; 13: 217-224.
www.wjpps.com │ Vol 10, Issue 5, 2021. │ ISO 9001:2015 Certified Journal │
114
El-Alfy et al. World Journal of Pharmacy and Pharmaceutical Sciences
53. Ferrali M, Signorini C, Caciotti B, Sugherini L, Ciccoli L, Giachetti D, Comporti M.
Protection against oxidative damage of erythrocyte membrane by the flavonoid quercetin
and its relation to iron chelating activity. FEBS Letters, 1997; 416: 1873-3468.
54. Fu Y, Wang Y, Du L, Xu C, Cao J, Fan T, Liu J, Su X, Fan S, Liu Q, Fan F. Resveratrol
inhibits ionising irradiation-induced inflammation in MSCs by activating Sirt1 and
limiting NLRP-3 inflammasome activation. Int J Mol Sci, 2013; 14: 14105–14118.
55. Koohian F, Shanei A, Shahbazi-Gahrouei D, Hejazi SH, Ahmadi A, Sharifi M. The effect
of resveratrol administration in irradiated mice on the induction of micronuclei in bone
marrow. J Radiat Oncol, 2017; 6: 423–427.