Basrah Journal of Veterinary Research,Vol.19,No.2,2020. Proceeding of the 17th International
Conference. College of Veterinary Medicine. University of Basrah. Iraq
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HISTOLOGICAL AND PHYSIOLOGICAL STUDY OF
DIANABOL ON THE THYROID GLAND OF FEMALE ALBINO
RATS (Rattus rattus)
Ekhlas Abid Hamza, Salim Salih Ali, Lubab Aqeel Jafer
Isam Mohamed Jaber
College of Veterinary Medicine, Al-Qasim Green University,Babylon, Iraq
Corresponding author: ekhlasalalwany@yahoo.com
Key word: dianabol, thyroid gland , TSH .
ABSTRACT
The random uses of androgenic anabolic steroids such as dianabol (Alpha pharma)
especially among young people and adolescent could have harmful effects on health
.The aim of this study is to investigate the possible harmful effects of oral
administration of dianabol on the histology and hormones levels of the thyroid gland
in female rats. The study was carried out in the animal house ( College of
veterinaryMedicine / University of Al-Qasim Green . Twenty four female rats
(Rattus rattus ) were divided into 4 groups , three groups were given dianabol orally
at the concentrations of (0.05 , 0.1 , 0.2) mg/kgvrespectively on daily basis for six
weeks , and the fourth group, served as a control group, was injected by (0.9%Nacl).
Thyroxine (T4), triiodothyronine(T3) and thyroid stimulating hormone (TSH) were
estimate, Histological study included the histopathological changes in thyroid gland
tissue. The results showed a significant increase (p< 0.05) in the levels of TSH, T4,
and T3 compared with the control group .This study also showed changes in the
thyroid tissue that included hyperplasia of thyroid follicles, colloid material in most
follicles, little or no increase in the size of follicles which suggest that increased dose
of dianabol leads to pathological (Physiological and histological) changes noticeable
in most of the studied criteria.
Basrah Journal of Veterinary Research,Vol.19,No.2,2020. Proceeding of the 17th International
Conference. College of Veterinary Medicine. University of Basrah. Iraq
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INTRODUCTION
In females, the testosterone circulating levels are about 10% of those
observed in males. This hormone is known to have an effect on the muscle growth
and build up properties. The development of the male phenotype, observed in men, is
under the influence of this hormone. Other functions such as blood production ,
plasma lipids, protein and bone metabolism are also documented (1). Due to the
anabolic effects of testosterone , it has the efficacy in the treatment of many disorders
such as renal and hepatic failure, anaemia associated with hepatic failure and
hypogonadism (2) , whereas the effects caused by anabolic androgenic drugs such as
dianabol include cardiovascular disorders, liver and kidney diseases as well as other
problems on human body (3). A reduction in HDL (High Density Lipoprotein)
cholesterol, increased inflammatory markersvand oxidative stress in AAS users (4).
Relationship between using Dianabol and hormones
Te elevated concentrations of Testosterone, DHT or Estrogens leads to the
suppression of gonadotropic hormones (5). (6) investigated the suppressing influence
of steroids on male sexual behavior after high-dose administration of (17a-alkylated)
steroids (stanozolol, oxymetholone or methyltestosterone), whereas the application of
metandienone, nandrolonedecanoate or testosterone, cypionate had little influence on
the amounts, intromissions or ejaculations of male rats. According to self-reports of
bodybuilders, boldenone, trenbolone andvnanrolone were considered as the most
unpleasant steroids due to their suppression of libido (7).
Doping in Sport
Testosterone hormone most important steroids used in sports which affects the
body in two directions is building body tissue and muscle and strength building from
intramuscularly as soon as, and this hormone gives the male characteristics to the
athlete, and adrenal hormones such as cortisol, which used to increase metabolism,
leading to a temporary increase in efficiency, but this hormone works on hair to
appear in unexpected places and the collapse of the adrenal gland, and the hormone
adrenaline and its derivative works to increase the number of heartbeat, high blood
pressure and increases the vulnerability of the body's metabolism of carbohydrates
Basrah Journal of Veterinary Research,Vol.19,No.2,2020. Proceeding of the 17th International
Conference. College of Veterinary Medicine. University of Basrah. Iraq
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which leads to degradation of glycogen in the body to modulate the simple sugar used
for energy production, but the hormone has a direct effect on the heart and use it in
larger amounts than usual may lead to stop working (8 ).
MATERIALS AND METHODS
Animals:Twenty four female albino Rats ( Rattus rattus ) were used in this study ,
their weight ranged between 200-250 gm.aged 2 months The rats were given food
and water ad libitum. In sprig section , then rats were housed in controlled conditions
of temperature ( 25± 3 Cº) and 12 hours light – dark cycles .
Experimental animals: The females rats were randomly divided into four groups,
each group consisted of 5 rats , one group served as control group and the other three
served as the experimental groups. The control group was (Nacl 0.09%) orally once a
week for six weeks. The experimental groups were given ( 0.05, 0.1) and 0.2mg/kg of
dianabol for a period of six weeks.
Blood and tissues sampling: Chloroform was used as anesthetic and female rats were
sacrificed. 5 ml of blood were obtained directly from the heart using sterile syringe.
Left to clot in plastic tubes at room temperature for 30 minutes, centrifuged at 3000
rpm for 10 minutes, and serum was taken for thyroid hormones ( TSH , T3, T4)
estimation. Thyroid gland was removed , fixed in Bouin solution, stained with Harris
hematoxylin& eosin (9, 10) for histological study .
Hormonal estimation: Enzyme Linked Immune Sorbent Assay (ELISA), which was
used to estimate the levels of hormones in serum at absorbance of 450 nm
wavelength. According to the procedure provided by Metra company (Italy) several
steps were followed to determine the levels of T3 and T4 and TSH. ELISA Reader
device, consisting of three pieces, a Washer - USA, Bio - ELISA Reader - Aspani,
Epson – LQ .
Histological Processing and Staining: Thyroid gland fixed in Bouin solution was
dehydrated with ascending concentrations of ethanol, embedded in paraffin ,
sectioned by using rotary microtome, stained with Haematoxylin & Eosin, mounted
on slides using Canada balsam (11).
Basrah Journal of Veterinary Research,Vol.19,No.2,2020. Proceeding of the 17th International
Conference. College of Veterinary Medicine. University of Basrah. Iraq
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Microscopic examination and photomicrography: The slides were examined by
using light microscope (Olympus) to detect the histopathology changes , and
photographed by using the camera of phase contrast light microscope.
Statistical analysis: SPSS software (Genstat) version (1995) was used to compare
the differences between experimental groups and control group. L.S.D, (Least
Significant Differences) was calculated for this purpose.
.
RESULTS
Estimation the levels of thyroid hormones
The results in figures(1,2,3 ) respectively , revealed that there was an
increase( P< 0.05) in TSH, T3 , T4 of female rats given dianabol in comparison
with control group.
Histological study of thyroid gland
The main histopathological changes of thyroid gland sections stained with
heamatoxylin -eosin in female rats treated by different concentrations of dianabol
showed that there was a range of changes from moderate to severe, at the
concentration 0.05 mg / kg body weight. Hyperplasia of follicle cells and a few empty
of colloid follicles (figure 5) were observed . While at the concentration of 0.1mg / kg
body weight of dianabol columnar lining epithelial , faint colloid material ,variable in
size of follicles and hyperplasia of follicle cells were noticed (figure 6). Furthermore,
severe changes at 0.2 mg / kg body weight of dianabol, such as columnar hyperplasia
of thyroid follicles , empty colloid material in most of follicles and increase in size of
follicles were observed (figure 7) in comparison with control group (figure 4) .
Basrah Journal of Veterinary Research,Vol.19,No.2,2020. Proceeding of the 17th International
Conference. College of Veterinary Medicine. University of Basrah. Iraq
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Figure (1) : TSH levels of serum rats treated with different doses of dianabol
Figure (2) :T3 levels in serum rats treated with different doses ofdianabol
0
0.1
0.2
0.3
0.4
0.5
0.6
0.05 0.1 0.2
TSH concentration (mg/dl)
dose (mg/kg)
control
sustanon
Basrah Journal of Veterinary Research,Vol.19,No.2,2020. Proceeding of the 17th International
Conference. College of Veterinary Medicine. University of Basrah. Iraq
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Figure (3):T4 levels of serum female rats treated with different doses of dianabol
Figure (4): cross section of control thyroid gland in female rats treated by different
doses of dianabol (H& E 100X )
0
0.5
1
1.5
2
2.5
3
3.5
0.05 0.1 0.2
T4 concentration (mg/dl)
dose (mg/kg)
control
Basrah Journal of Veterinary Research,Vol.19,No.2,2020. Proceeding of the 17th International
Conference. College of Veterinary Medicine. University of Basrah. Iraq
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Figure (5): cross section of thyroid gland in female rats treated by 0.05 mg/kg of
dianabol observed hyperplasia of follicle cells (green arrow) and a few follicles
showed empty (white arrow) (H& E 100X)
Figure (6): cross section of thyroid gland in female rats treated by 0.1 mg/kg of
dianabol showed faint colloid material(green arrow) ,variable in size of follicles and
hyperplasia of follicle cells (red arrow) (H& E 100X)
Basrah Journal of Veterinary Research,Vol.19,No.2,2020. Proceeding of the 17th International
Conference. College of Veterinary Medicine. University of Basrah. Iraq
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Figure (7): cross section of thyroid gland in female rats treated by 0. 2 mg/kg of
dianabol showed hyperplasia of thyroid follicles(red arrow) , empty colloid material
in most of follicles(green arrow) and increase in size of follicles (yellow arrow)
(H& E 100X )
DISCUSSION
The significant increase in TSH, T3 ,T4 (Figures 1,2,3) could be due to the hyper
androgenism which is associated with varying type of steroid used. The misuse of
steroid may result in partial pituitary atrophy thyroid hormones changes (12).
Thyroid gland is another target of testosterone when this hormone stimulates
thyrotropin secretion and increase the growth of the thyroid gland (13). In contrast,
the administration of steroids to patients with thyroid diseases causes changes in
regard to thyroid medications (14). Meanwhile, there may be indirect effect dianabol
on the pituitary-thyroid axis as a consequence of increased serum levels of thyroxin
binding globulin, and the associated changes in thyroid function as a result of
increased of metabolic rate, through uptake of anabolic steroids by athletes (15).
The histological changes of thyroid gland in female rats suggest that anabolic
androgenic steroid (dianabol) may have caused hormonal effects which led to
proliferative tissue responses, including hyperplasia and neoplasia as well as a
decrease of colloid by hyper active thyroid gland (16).
Basrah Journal of Veterinary Research,Vol.19,No.2,2020. Proceeding of the 17th International
Conference. College of Veterinary Medicine. University of Basrah. Iraq
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