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Oguji et al Journal of Drug Delivery & Therapeutics. 2021; 11(3-s):19-26
ISSN: 2250-1177 [19] CODEN (USA): JDDTAO
Available online on 15.06.2021 at http://jddtonline.info
Journal of Drug Delivery and Therapeutics
Open Access to Pharmaceutical and Medical Research
© 2011-21, publisher and licensee JDDT, This is an Open Access article which permits unrestricted
non-commercial use(CC By-NC), provided the original work is properly cited
Open Access Full Text Article Research Article
Protection of histomorphology of vital organs by methanolic seed extract
of Nigella sativa against cadmium-induced tissue injuries in rats
1Emeka Cyprian Oguji , 1Chibueze Joseph Obigeorge , 1Johnson Obiechina Omeh , 1Amechi Jnr. Odeku ,
2Tachia Jaclyn Wanger , 1Chukwuebuka Abel Udeh , 1Chidubem John Ochi , 1Kosisochukwu Doris Ogbonna
, 1Amarachi Chioma Ikekpeazu , 1Chinecherem Maudlyne Nnam , 1Ikenna Kingsley Uchendu*
1 Department of Medical Laboratory Science, University of Nigeria, Enugu, Nigeria
2 Department of Physics, University of Agriculture, Markurdi, Benue, Nigeria
Article Info:
_________________________________________
Article History:
Received 17 April 2021
Reviewed 21 May 2021
Accepted 27 May 2021
Published 15 June 2021
_________________________________________
Cite this article as:
Oguji EC, Obigeorge CJ, Omeh JO, Odeku AJ, Wanger
JT, Udeh AC, Ochi CJ, Ogbonna KD, Ikekpeazu AC,
Nnam CM, Uchendu IK, Protection of
histomorphology of vital organs by methanolic
seed extract of Nigella sativa against cadmium-
induced tissue injuries in rats, Journ al of Drug
Delivery and Therapeutics. 2021; 11( 3-S):19-26
DOI:http://dx.doi.org/10.22270/jddt.v11i3-S.4 822
_________________________________________
*Address for Correspondence:
Ikenna Kingsley Uchendu, Department of Medical
Laboratory Science, University of Nigeria, Enugu
State, Nigeria. ORCID ID: https://orcid.org/0000-
0002-1503-3759
Abstract
______________________________________________________________________________________________________
Background: Chemical-induced organ injuries have been on a fast rise for decades and these
injuries have become common causes of mortality and morbidity in the society. Edible plant
materials with medicinal properties have been used for treating various diseases for many
centuries in folk medicine. Recently, the role of food or medicinal plants in human health has
received considerable attention. Traditional uses of N. sativa seed range from soothing
wounds to remedying cough, eczema, diabetes, inflammation of the bronchi and tooth aches;
and these point to substantial tissue effects.
Objective: We investigated the protective effects of methanolic seed extract of Nigella sativa
(MENS) against cadmium-induced histomorphological alterations in heart, kidney and liver
tissues of albino rats.
Methods: Twenty five (25) male albino rats, weighing (200±20g), were randomly grouped
into five groups: A, B, C, D, and E. Group B (Negative Control) received intraperitoneal
administration of cadmium chloride (CdCl2, 5mg/kg) only, group C received CdCl2 and low
dose MENS (300mg/kg, oral), group D received CdCl2 and high dose MENS (600mg/kg, oral),
and group E (Positive control) received CdCl2 and Vitamin C (200mg/kg, oral), for 14 days.
Group A (Normal control) received no administration. Heart, kidney and liver were harvested
for histopathological analyses.
Results: Cadmium (CdCl2) induced significant histomorphological changes in the studied
organs, and the heart was the most damaged of all the organs studied; however a
significantly ameliorative effect by methanolic seed extracts was observed.
Conclusion: Nigella sativa seed extract is potentially tissue-protective against harmful
chemical toxins like cadmium.
Keywords: anticardiotoxic, cadmium, Nigella sativa, medicinal food, antinephrotoxic,
ethnopharmacology, antihepatotoxic
1. INTRODUCTION
Nigella sativa (N. sativa) (Family Ranunculaceae), commonly
known as black seed or black cumin or seed of blessing in
different languages, is a grassy plant, and has green to blue
flowers with small black seeds.1 A few foods of plant origin
and plants species have been thoroughly evaluated for their
medicinal properties.2 The medicinal properties of Nigella
sativa has been reported to be due to its composition of
stable and volatile oils which contain good amounts of
unsaturated fatty acids, arachidonic acid and eicosenoic
acids in little amount.1,3 Traditional uses of N. sativa seed
range from soothing wounds to remedying cough, eczema,
diabetes, inflammation of the bronchi and tooth aches.3 One
of the active ingredients of N. sativa is thymoquinone, a
flavanoid with proven anti-inflammatory, anti-cancerous,
anti-bacterial along with anthelmintic properties.4 With the
rapid increase in developments, environmental pollutants,
heavy metals (e.g. cadmium and lead inclusive), now top the
list of toxicants which pose serious risk or threat to human
health and wellbeing.
Cadmium is of a great use especially in the utilization of its
conductor properties and thus found to be of great
importance in the manufacture of batteries and reactors.
However, this element is found to be very toxic, even in very
little amount and could cause abnormalities in man after
exposure.5 Cigarette smoking is considered to be the most
significant source of human cadmium exposure. When
ingested, cadmium accumulates in the body with age and has
an extremely long biological half-life.6-8
Although cases of cadmium-induced nephropathy have been
described in chronic exposures, recent studies suggest that
Oguji et al Journal of Drug Delivery & Therapeutics. 2021; 11(3-s):19-26
ISSN: 2250-1177 [20] CODEN (USA): JDDTAO
cadmium at acute or low environmental exposures in
industrialized countries can also cause subtle renal effects
leading to a modest increase in the urinary excretion of low
weight micro proteins.9 Exposure to cadmium could produce
serious adverse effects in humans and one of the ways
through which this happens is through induction of
oxidative stress. Cadmium chloride (CdCl2), recognized as an
experimental toxin, is known to be highly cardiotoxic, 11,12
and may cause severe liver damage.12
Despite all studies performed to date, therapy choices for
cardiac, liver and kidney injuries are very few. Foods or
plants with medicinal value have proven to be foremost or
firstline therapy choice in the treatment of diseases in most
of the developing countries, and they provide important
sources of most of the world’s pharmaceutical; thus they
have served a valuable starting material in drug discovery or
drug development.13 To date, no study has holistically
evaluated the protective effect of black seed extracts against
cadmium-induced histomorphological alterations of heart,
kidney and liver in rat.
2. MATERIALS AND METHODS
2.1 Plant Material
Fresh samples of Nigella sativa seeds were obtained from
local market in Enugu, Nigeria. The plant material was
authenticated by a consultant taxonomist at the herbarium
section of the Department of Plant Science and
Biotechnology, University of Nigeria, and a voucher
specimen was deposited at the herbarium with reference
number UNH No 662 for future reference.
2.1.1 Preparation of Methanolic extracts of Nigella sativa
(MENS)
Nigella sativa seeds were dried and shaded from sun light,
then powdered with a grinder. Extraction was done using
Babaei et al.14 method with minor modifications. Five
hundred gram (500g) of N.sativa powder was macerated
with 2 litres of absolute (100%) methanol (as methanolic
extract) for seventy two (72) hours. The mixture was stirred
in an Erlenmeyer flask for twenty four (24) hours using a
laboratory shaker. At the end of the extraction, the extract
was filtered through a Whatman filter (Whatman Clifton, NJ,
USA). Finally, using a water bath set at 30°C, the solvent
evaporated, and 4g of dried methanolic extracts was
obtained. This was reconstituted in distilled water, used to
prepare the required concentration, and stored at 4°C until
when needed for use.
2.1.2. Acute toxicity test (LD50)
The median lethal dose (LD50) of methanolic extracts of
N.sativa (MENS) was performed on mice and Lorke
procedure of LD50 determination was used.15
2.1.3 Phytochemical analysis of Nigella sativa seeds
Preliminary phytochemical screening of N.sativa was carried
out at Department of Pharmacognosy, Faculty of
Pharmaceutical Science, University of Nigeria. Procedures
outlined by Trease and Evans16 were employed for the
analyses.
2.2 Drug and Chemicals used
The chemicals used in the study include analytical grade of
absolute methanol, for plant extraction, and Cadmium
chloride salt for induction of selected organs toxicity. The
chemicals were purchased from Ogbete main market, Enugu.
Drug used includes vitamin C (Alpha Pharmaceuticals,
Enugu, Nigeria).
2.2.1 Preparation of vitamin C solution
Stock concentrations (20mg/ml) of vitamin C were prepared
and used for the research.
2.2.2 Preparation of cadmium chloride solution
Eighty milligram (80mg) of cadmium salt (CdCl2) was
dissolved in distilled water and made up to 100ml in a
measuring cylinder to give a stock concentration of
0.8mg/ml.
2.3 Induction of selected organs toxicity
Sub-acute cardiac, kidney and liver injuries were induced in
each animal by intraperitoneal injection with cadmium
chloride solution (5mg/kg), daily for 14 days.
2.4 Animals and maintenance
Twenty five (25) adult albino rats, weighing (200±20g),
were obtained from the animal house of the College of
Veterinary Medicine, University of Nigeria. The animals were
housed in standard condition and properly fed with
commercial rat pellets and water ad libitum. The animals
were kept under observation for 14 days before the onset of
the experiment for acclimatization. The experimental
protocol was approved by the institution animal ethics
committee of the University of Nigeria Teaching Hospital
(UNTH/CSA. 452/VOL. 19).
2.5 Experimental Design
The twenty five (25) male rats were grouped into (A-E) and
received the following treatments daily and within 2 hours.
Group A: (normal Control): No treatment was administered
to this group.
Group B: (Negative Control): received CdCl2 (5mg/kg, i.p)
only for 14 days.
Group C: received CdCl2 and low dose (300mg/kg, oral) of
methanolic extract of N. sativa MENS for 14 days.
Group D: received CdCl2 and high dose (600mg/kg, oral) of
methanolic extract of N. sativa (MENS) (600mg/kg, oral) for
14 days.
Group E (Positive control): received CdCl2 and Vitamin C
(200mg/kg, oral) for 14 days.
2.6 Sample collection
After 14 days, the animals were sacrificed via cervical
dislocation under chloroform anesthesia. The heart, kidney
and liver were harvested for histopathological analysis.
2.7 Histopathological analysis
The excised heart, kidney and liver tissues were processed
using the paraffin wax embedding technique, sectioned at 5
microns and stained using the Haematoxylin and Eosin [H
and E] staining procedure.17 The histological sections were
examined using an Olympus TM light microscope.
2.7.1 Histopathological image analysis
Interpretation was done following standard guidelines as
described by Gurcan et al.,18.
Oguji et al Journal of Drug Delivery & Therapeutics. 2021; 11(3-s):19-26
ISSN: 2250-1177 [21] CODEN (USA): JDDTAO
3. RESULTS
3.1 Acute toxicity studies result.
Median lethal dose (LD50) value of the extract was 2400
mg/kg which indicates that MENS is safe and is not toxic to
mice, at the doses used in the experiment (Table 1).
3.2 Phytochemical results.
The result of the preliminary phytochemical analysis of
Nigella sativa revealed abundant presence of alkaloids and
flavonoids (+++); moderate presence of tannins and phenols
(++). However glycosides, saponins and steroids were
absent (table 2).
3.3 Histopathological results
In Figure 1, the heart of normal control rats appeared
functionally and structurally normal. The cardiac fibres
showed a well conserved morphology (1A). The heart of
CdCl2-treated group (negative control) showed abnormal
changes; there was evidence of fibrosis and mild infiltration
by inflammatory cells. Fibres appear wavy showing signs of
significant degeneration (1B). However, the cardiac fibres of
test group rats (low dose MENS at 300mg/kg) appeared
normal with very mild infiltration by inflammatory cells
(1C). While in the other test group rats (high dose MENS at
600mg/kg), the myocardial fibres appear wavy; some fibres
are necrotic, with presence of leucocyte infiltration (1D).
Furthermore, photomicrograph of heart section from CdCl2 +
Vitamin C (200mg/kg), showed normal appearance of
cardiac fibres (1E).
In Figure 2, the kidney of normal control rats appeared
functionally and structurally normal. The glomeruli and
tubule showed a well conserved morphology (1A). The
kidney of CdCl2-treated group (negative control) showed
abnormal changes; there was severe tubular degeneration;
the glomeruli are all enlarged (1B). However, the kidney
tubules of test group rats (low dose MENS at 300mg/kg)
showed mild signs of autolytic degeneration and erosions;
most glomeruli were normal while some were enlarged (1C).
While in the other test group rats (high dose MENS at
600mg/kg), the kidney tubules showed moderate signs of
autolytic degeneration, mild infiltration by inflammatory
cells; and the glomeruli were enlarged and hyper cellular
(1D). Furthermore, photomicrograph of kidney section from
CdCl2 + Vitamin C (200mg/kg), showed normal appearance
of tubules; glomeruli also appear normal with a few eroded
Bowman’s capsule (1E).
In Figure 3, the liver of normal control rats appeared
functionally and structurally normal. The hepatocytes
showed a well conserved morphology (1A). The Liver of
CCl4-treated group (negative control) showed abnormal
changes; there was extensive vacuolation of the hepatocytes
with mild infiltration by inflammatory cells (1B). However,
the liver of test group rats (low dose MENS at 300mg/kg)
showed mild pericentral vacuolation of hepatocytes with
mild infiltration (1C). In the other test group rats (high dose
MENS at 600mg/kg), the liver showed evidence of fatty
degeneration (giving a foamy appearance) and vacuolation
of some hepatocytes. Furthermore, photomicrograph of
liver section from CdCl2 + Vitamin C (200mg/kg), showed
normal appearance of hepatocytes (1E).
Table 1: The median lethal dose (LD50) of methanolic extracts of N.sativa (MENS)
Phase
Dose
Death
Observation
1
10
100
1000
0/3
0/3
0/3
Nil
Nil
Nil
2
1200
2500
3500
5000
0/4
2/4
2/4
4/4
Calm and no death occurred
Died within 48hours
Died within 48hours
Died within 24hours
Two doses, 1200mg/kg and 5000mg.kg were used to calculate the LD50 of the plant extract.
LD50 = √ A x B
A = Maximum dose with 0% mortality (1200mg/kg)
B = Minimum dosed with 100% mortality (5000mg/kg)
LD50 of methanolic extract of N.sativa seed = √1200 X 5000 = 2449.48mg/kg
LD50 of methanolic extract of N.sativa seed ≈ 2400mg/kg
Table 2: Preliminary qualitative phytochemical results of methanol extract of N.sativa (MENS)
Test
Result
Alkaloid
+ + +
Flavonoid
+ + +
Tannins
+ +
Glycoside
-
Phenol
+ +
Saponin
-
Terpenoid
-
Steroid
-
Key: +++ = present (in abundance); ++ = present (in moderate amount); − = absent
Oguji et al Journal of Drug Delivery & Therapeutics. 2021; 11(3-s):19-26
ISSN: 2250-1177 [22] CODEN (USA): JDDTAO
Figure 1: Photomicrograph of heart section. (A) Cardiac fibres (black arrows) appear normal with no degenerative changes.
(B) Evidence of fibrosis (#) and mild infiltration by inflammatory cells (arrows). Fibres appear significantly wavy and damaged
(*). (C) A section of the cardiac fibres appear necrotic and inflamed with infiltration by inflammatory cells (arrows).. (D)
Evidence of fibrosis (#) and mild infiltration by inflammatory cells (arrows). (E) Cardiac fibres (red arrow) appear normal with
very mild infiltration by inflammatory cells (black arrows) [Stain: H and E; ×400].
Oguji et al Journal of Drug Delivery & Therapeutics. 2021; 11(3-s):19-26
ISSN: 2250-1177 [23] CODEN (USA): JDDTAO
Figure 2: Photomicrograph of kidney section. (A) There is normal appearance of glomeruli (black arrows) and renal tubule
(blue arrow). (B) There is severe tubular degeneration (red arrows); the glomeruli are all enlarged (*). (C) Most glomeruli are
enlarged and hypercellular (#); few are still normal (*). (D) The tubules show signs of autolytic degeneration (red arrows); the
glomeruli are enlarged (*) and there is infiltration by inflammatory cells (blue arrows). (E) Most tubules appear normal (red
arrows); glomeruli also appear normal (*) with a few eroded Bowman’s capsule (#). [Stain: H and E; ×100].
Oguji et al Journal of Drug Delivery & Therapeutics. 2021; 11(3-s):19-26
ISSN: 2250-1177 [24] CODEN (USA): JDDTAO
Figure 3: Photomicrograph of liver section. (A) Hepatocytes (arrows) are normal with no signs of degenerative
lesions/changes. (B) There is extensive vacuolation of the hepatocytes (red arrows) with mild infiltration by inflammatory cells
(arrow head). (C) Hepatocytes are normal; there is mild infiltration of inflammatory cells (arrow head) around the central ve in.
(D) Hepatocytes appear normal with mild vacuolations (arrow). (E) Liver section appears normal. Hepatocytes (arrow) and
central vein appear normal. CV- central vein [Stain: H and E; ×400].
4. DISCUSSION
The heart is a muscular organ; made up of cardiac muscles
which constantly pump blood through the blood vessels of
circulatory system in human body and animals; while the
kidneys are very effective in the excretion of metabolic
wastes.19 Kidney disease is both a cause and a consequence
of cardiovascular disease, hence the term cardiorenal
syndrome.19 Cardiorenal syndrome is a medical term for
disorders involving both the heart and kidneys whereby
acute or chronic dysfunction in one organ may induce acute
or chronic dysfunction in the other organ.20 The liver is the
main metabolic organ in the body especially in
lipometabolism and glycometabolism.21
Accumulation of toxic substances such as drugs, heavy
metals, poisons etc can cause significant damages to the
histomorphology of vital organs such as the heart muscles,
kidney and liver tissues. Exposure to cadmium could
produce serious adverse effects in humans and one of the
ways through which this happens is through induction of
oxidative stress; whereby it induces early hyperproduction
Oguji et al Journal of Drug Delivery & Therapeutics. 2021; 11(3-s):19-26
ISSN: 2250-1177 [25] CODEN (USA): JDDTAO
of Reactive Oxygen Species (ROS) that impair antioxidant
defense system leading to oxidative stress in target organs
and tissues.22
In this study, the LD50 value of N.sativa seed extract was
established at 2400mg/kg which indicates that the
methanolic extract of N. sativa (MENS) was safe and is not
toxic to mice, at the doses used in the experiment (table 1).
The preliminary phytochemical analysis of N.sativa revealed
abundant presence of alkaloids (+++) and flavonoids (+++);
moderate presence of tannins (++) and phenols (++).
However glycosides (-), terpenoids (-) saponins (-) and
steroids (-) were absent (table 2). In the histopathological
analyses, cadmium significantly induced histomorphological
changes in the heart fibres, kidney and liver tissues.
However, treatment with low and high dose methanolic seed
extracts of Nigella sativa (MENS) separately, revealed a
marked ameliorative effect, thereby protecting the
histomorphological architecture of the vital organs studied.
Although this present study was not aimed to evaluating the
mechanism through which the seed extract showed
ameliorative effects; we however observed that the extract,
particularly the low dose, acted in similar way as the
standard drug (Vitamin C), a known antioxidant, which was
used (Figures 1-3). Interestingly, we did not observe dose-
dependent protection by the seed extract; and these
observations once against support the world wide claim that
N.sativa is a potent medicinal plant. These findings could be
as a result of the singular or combined actions of one or
more of these bioactive phytochemical constituents present
in the Nigella sativa. Thymoquinone, an active ingredient of
Nigella sativa (black seed), is a constituent of flavonoids.4
Several other studies have reported the ameliorative effects-
and potential pharmacotherapeutic effects - of crude
extracts of N.sativa and thymoquinone separately. N.sativa
has been reported to offer protections against cadiotoxic
drug such as cyclosporine A; and against toxic heavy metals
such as lead (Pb) and cadmium (Cd).23.24 Crude extracts of
the seeds of N. sativa have been evaluated for
hepatoprotective activity in Wistar rats against various
hepatotoxicants, which are widely known for their ability to
induce hepatotoxicity in experimental animals.25,26 N. sativa
has not only been studied for its protective effects against
tissue injuries, it has also been evaluated for antimicrobial
activities. Different crude extracts of N. sativa exhibited
antimicrobial efficacy against different bacterial strains
which comprised either gram negative or gram positive
bacteria; and these have been reported.27,28
Thymoquinone (TQ) is chemically known as 2-methyl-5-
isopropyl-1, 4-benzoquinone.4 Nemmar et al.29 reported that
TQ showed strong anti-inflammatory effects against diesel
exhaust particles-induced cardiopulmonary injury in mice.
Due to potent anti-oxidant and free radical scavenging
action, TQ has been shown to normalize the adverse effect of
various environmental toxins or xenobiotics causing
oxidative damage and organ dysfunctions leading to
pathogenesis of various diseases.30 Alkaloids and Phenols,
found to be significantly present in the methanol extracts,
also act as natural antioxidants, scavenge free radicals and
inhibit their production, stimulate the synthesis of
antioxidant enzymes thereby prevent oxidative stress.31
CONCLUSION
The findings of this work show that cadmium induced
histomorphological alterations in heart, kidneys and liver in
the experimental rats; however, methanolic seed extract of
Nigella sativa ameliorated the effects in the test groups,
although not in a dose-dependent manner. Thus, the result
suggests that methanolic seed extract of Nigella sativa
(MENS) has anti-cardiotoxic, antinephrotoxic and
hepatoprotective properties and could therefore offer
significant amelioration against cadmium- induced organ
injuries.
LIST OF ABBREVIATIONS
MENS = Methanolic Extract of Nigella sativa
ROS = Reactive Oxygen Species
CdCl2 = Cadmium Chloride
Cd = Cadmium
Pb = Lead
TQ = Thymoquinone
H and E = Haematoxylin and Eosin
ANOVA = Analysis Of Variance
SEM = Standard Error of Mean
AUTHORS’ CONTRIBUTION
I.K.U and E..C.O- Conceptualized and designed the
experiment, performed the experiment, performed the
histopathological image analysis, and prepared the
manuscript
C.J.O, J.O.O, A.J.O, T.J.W and C.A.U- Assisted in literature
search, and copy-edited the manuscript
C.J.O, K.D.O, A.C.I and C.M.N- Assisted in literature search
ETHICS APPROVAL AND CONSENT TO
PARTICIPATE
The experimental protocol was approved by the institution
animal ethics committee of the University of Nigeria
Teaching Hospital (UNTH/CSA. 452/VOL. 19), Nigeria.
Experiments were conducted according to the Guidelines on
the Care and Use of Laboratory Animals (National Institutes
of Health, Bethesda, MD, USA).
HUMAN AND ANIMAL RIGHTS
No humans were used in this research. The procedures
involving the use of animals were in accordance with the
standards set forth in the eighth edition of “Guide for the
Care and Use of Laboratory Animals”
(grants.nih.gov/grants/olaw/guide-for-the-care-and-
use-of-laboratory animals_prepub.pdf published by the
National Academy of Sciences, The National Academies
Press, Washington, D.C.).
AVAILABILITY OF DATA AND MATERIALS
The datasets used and/or analyzed during the current study
are available from the corresponding author on reasonable
request.
FUNDING
This study did not receive any fund from any grant agency or
organization
COMPETING INTERESTS STATEMENT
The authors declare no conflicts of interest.
ACKNOWLEDGEMENTS
Declared none.
Oguji et al Journal of Drug Delivery & Therapeutics. 2021; 11(3-s):19-26
ISSN: 2250-1177 [26] CODEN (USA): JDDTAO
REFERENCES
1. Hosseini M, Mohammadpour T, Karami R, Rajaei Z, Sadeghnia H R,
Soukhtanloo M. Effects of the hydro-alcoholic extract of Nigella
sativa on scopolamine-induced spatial memory impairment in
rats and its possible mechanism. Chinese Journal of Integrative
Medicine 2015; 21(6):438-444.
https://doi.org/10.1007/s11655-014-1742-5
2. Ahmad A, Husain A, Mujeeb M, Khan SA, Najmi AK, Siddique NA,
Damanhouri ZA, Anwar F. A review on therapeutic potential of
Nigella sativa: A miracle herb. Asian Pacific Journal of Tropical
Biomedicine 2013; 3(5):337-352.
https://doi.org/10.1016/S2221-1691(13)60075-1
3. Amin, B, Hosseinzadeh H. Black cumin (Nigella sativa) and its
active constituent, thymoquinone: an overview on the analgesic
and anti-inflammatory effects. Planta Medica 2015; 82: 8-16.
https://doi.org/10.1055/s-0035-1557838
4. Darakhshan S, Pour AB, Colagar AH, Sisakhtnezhad S.
Thymoquinone and its therapeutic potentials. Pharmacoogical
Reseasrch 2015; 95-96: 138-158.
https://doi.org/10.1016/j.phrs.2015.03.011
5. Rafati R, Kazemi S, Moghadamnia A. Cadmium toxicity and
treatment: An update. Caspian Journal internal medicine 2017;
8(3):135-145.
6. Yu C,, Ling Q, Yan S, Jing L, Zhongda C, Zhenyu P. Cadmium
contamination in various environmental materials in an
industrial area, Hangzhou, China. Chemical Speciation and
Bioavailability 2010; 22(1):35-42.
https://doi.org/10.3184/095422910X12631439471494
7. Sharma H, Blessy NR, Mathew B. The characteristics, toxicity and
effects of cadmium. International Journal of Nanoscience and
Nanotechnology 2015; 3:1-9.
8. Bernhoft RA. Cadmium toxicity and treatment. The Scientific
World Journal 2013; 2013: 394652.
https://doi.org/10.1155/2013/394652
9. Wallin M, Sallstern G, Lundh T, Barregard L. Low level Cadmium
exposure and effects on kidney function. Occupational and
environmental medicine 2014; 71(12):848-854.
https://doi.org/10.1136/oemed-2014-102279
10. Shen J, Wang X, Zhon D, Li T, Tang L, Gong T, Su J, Liang P.
Modelling cadmium-induced cardiotoxicity using human
pluripotent stem cell-derived cardiomyocetes. Journal of
Cellular and Molecular Medicine 2018; 22(9):4221- 4235.
https://doi.org/10.1111/jcmm.13702
11. Borné Y, Barregard L, Persson M, Hedblad B, Fagerberg B,
Engstrom G. Cadmium exposure and incidence of heart failure
and atrial fibrillation: a population-based prospective cohort
study. British Medical Journal open 2015; 5(6):e007366.
https://doi.org/10.1136/bmjopen-2014-007366
12. Hyder O, Chung M, Cosgrove D, Herman JM, Li Z, Firoozmand A,
Gurakar A, Koteish A, Pawlik TM. Cadmium exposure and liver
disease among US adults. Journal of Gastrointestinal Surgery
2013; 17(7):1265-1273. https://doi.org/10.1007/s11605-013-
2210-9
13. Ajibesin KK. Dacryodes edulis (G. Don) HJ. Lam: A review on its
medicinal, phytochemical and economic properties. Research
Journal of Medicinal Plant 2011; 5(1):32-41.
https://doi.org/10.3923/rjmp.2011.32.41
14. Babaei HA, Motamedifar M, Khalifat S, Mohammadi A, Khosrow
Z, Motamedifar A. In vitro study of antibacterial property and
cytotoxic effects of aqueous, ethanolic, methanolic, and
hydroalcoholic extracts of fenugreek seed. Pakistan Journal of
Medical and Health Sciences 2018; 12(2):906-910.
15. Lorke D. A new approach to practical acute toxicity testing.
Archives of Toxicology 1983; 54(4):275-287.
https://doi.org/10.1007/BF01234480
16. Trease G, Evans SM. Pharmacognosy: (15th Edition). English
Language Book Society. Bailliere Tindall, London, 2002. p. 23-
67.
17. Baker FJ, Silverton RE, Pallister CJ Baker and Silverton's
Introduction to Laboratory Technology. 7th Edition,
Butterworth-Heinemann, Wobrun, MA, USA, 1998; page 448.
18. Gurcan MN, Boucheron L, Can A, Madabhushi A, Rajpoot N, Yener
B. Histopathological image analysis: A review. IEEE Review
Biomedical Engineering 2009; 2:147-171.
https://doi.org/10.1109/RBME.2009.2034865
19. Okamoto R, Ali Y, Hashizume R, Suzuki N, Ito M. BNP as a major
player in the heart-kidney connection. International Journal of
Molecular Sciences 2019; 20(14):e3581.
https://doi.org/10.3390/ijms20143581
20. Rangaswami J, Bhalla V, Blair EA, Chang TI, Costa S, Mezue K. et
al. Cardiorenal syndrome: Classification, pathophysiology,
Diagnosis, and Treatment strategies: A scientific statement from
the American Heart Association. Circulation. 2019; 139:e840-
878. https://doi.org/10.1161/CIR.0000000000000664
21. Ding HR, Wang JL, Ren HZ, Shi XL. Lipometabolism and
glycometabolism in liver diseases. BioMed Res Int. 2018; 2018:
ID 1287127, 7 pages. https://doi.org/10.1155/2018/1287127
22. Bücker-Neto L, Paiva ALS, Machado RD, Arenhart RA, Margis-
Pinheiro M. Interaction between plant hormones and heavy
metals responses. Genetics and Molecular Biology 2017;
40(1):373-386. https://doi.org/10.1590/1678-4685-gmb-
2016-0087
23. Ebru U, Burak U, Yusuf S, Reyhan B, Arif K, Faruk TH, Emin M,
Aydın K, Lhan Atilla I, Semsettin S, Kemal E. Cardioprotective
effects of Nigella sativa oil on cyclosporine A‐induced
cardiotoxicity in rats. Basic and Clinical Pharmacology and
Toxicology 2008; 103(6):574-580.
https://doi.org/10.1111/j.1742-7843.2008.00313.x
24. Massadeh AM, Al-Safi SA, Momani IF, Al-Mahmoud M, Alkofahi
AS. Analysis of cadmium and lead in mice organs. Biological
Trace Element Research 2007; 115(2):157-167.
https://doi.org/10.1007/BF02686027
25. Adam GO, Rahman MM, Lee SJ, Kim GB, Kang HS, Kim JS, Kim SJ.
Hepatoprotective effects of Nigella sativa seed extract against
acetaminophen-induced oxidative stress. Asian Pacific Journal
of Tropical Biomedicine 2016; 9(3):221-227.
https://doi.org/10.1016/j.apjtm.2016.01.039
26. Abdel-Daim MM, Ghazy EW. Effects of Nigella sativa oil and
ascorbic acid against oxytetracycline-induced hepato-renal
toxicity in rabbits. Iranian Journal of Basic Medical Sciences
2015; 18(3): 221-227.
27. Hannan A, Saleem S, Chaudhary S, Barka M, Arshad MU. Anti-
bacterial activity of Nigella sativa against clinicalisolates of
methicillin resistant Staphylococcus aureus. Journal of Ayub
Medical College, Abbottabad 2008; 20(3):72-74.
28. Morsi NM. Antimicrobial effect of crude extracts of Nigella sativa
on multiple antibiotics-resistant bacteria. Acta Microbiologica
Polonica 2000; 49(1):63-74
29. Nemmar A1, Al-Salam S, Zia S, Marzouqi F, Al-Dhaheri A,
Subramaniyan D, Dhanasekaran S, Yasin J, Ali BH, Kazzam EE.
Contrasting actions of diesel exhaust particles on the pulmonary
and cardiovascular systems and the effects of thymoquinone.
British Journal of Pharmacology 2011; 164(7):1871-1882.
https://doi.org/10.1111/j.1476-5381.2011.01442.x
30. Alkharfy KM, Ahmad A, Khan RM, Al-Asmari M. High-
performance liquid chromatography of thymoquinone in rabbit
plasma and its application to pharmacokinetics. Journal of
Liquid Chromatography & Related Technologies 2013;
36(16):2242-2250.
https://doi.org/10.1080/10826076.2012.717062
31. Liaudanskas M, Viškelis P, Raudonis R, Kviklys D, Uselis N,
Janulis, V. Phenolic composition and antioxidant activity of
malus domestica leaves. The Scientific World Journal 2014,
2014: ID 306217, 10 pages.
https://doi.org/10.1155/2014/306217
