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MORPHOLOGIC AND HISTOPATHOLOGIC CHANGES ASSOCIATED WITH NANDROLONE DECAONATE (NANDURABOLIN) ON THE SEMINAL VESICLE OF RATS

ZEINAB, S.DESOUKY¹; SHAKKER, A.S.¹; THABET IBRAHIM²; SALAH H. AFIFI³ and WAFAA A. MUBARAK⁴

¹ Department of Forensic Medicine and Toxicology, Faculty of Veterinary Medicine, South Valley University, Egypt

² Department of Forensic Medicine and Toxicology, Faculty of Veterinary Medicine, Assiut University, Assiut, Egypt

³ Department of Pathology and Clinical Pathology, Assiut University, Assiut, Egypt

⁴ Department of Anatomy, Faculty of Medicine, Assiut University, Assiut, Egypt

Received: 7 March 2017;       Accepted:2 April 2017

INTRODUCTION

The Anabolic-Androgenic Steroid (AASs) is a group of synthetic derivatives of testosterone with both skeletal muscle-building (anabolic) and masculinizing (androgenic) effects. Anabolic-Androgenic Steroids (AAS) are used in high doses by athletes to improve athletic ability, physical appearance, and muscle mass. The abuse of Anabolic Androgenic Steroids (AAS) is under constant debate world-wide. A large number of young adolescents abuse AAS to improve their physical fitness and appearance (Barceloux et al., 2013). Soma et al. (2007) stated that anabolic steroids are used in equine medicine for building weight and muscle mass, and to alleviate anemia. These drugs used for growth in food producing animals like beef, cattle and sheep. 

Most of the adverse effects following the use of AASs result from the enhancement of normalphysiologic    response to testosterone by

Corresponding author: Dr. SALAH H. AFIFI

E-mail address: Afifi_s_4@hotmail.com.

Present address: Department of Pathology and Clinical Pathology, Assiut University, Assiut, Egypt

either direct receptor agonist activity or suppression of steroid biosynthesis. In general, toxic effects associated with AAS abuse involve the following: (1) anabolic side effects, (2) enhanced androgenic effects, (3) estrogenic side effects, (4) antiandrogenic effects from the suppression of the hypothalamus–pituitary– adrenal/ gonadal axes,(5) hepatotoxicity, and(6) neuropsychiatric effects (Buttner, and Thieme 2010).

Pathologic abnormalities from AAS abuse are best-documented in the cardiovascular system, reproductive system, liver, and serum lipids. Animals studies suggest that AAS can cause dysplasia of collagen fibrils and decreased tensile strength, and potentially the use of these drugs could cause disruption of connective tissue (Laseter and Russell, 1991).

Shokri et al. (2014) stated that the prostates and seminal vesicles weight were significantly different between control and group injected intramuscular with nandrolene at a dose of 10mg/kg bodyweight for 8 weeks (respectively, P<0.05 and P<0.001). The use of anabolic androgenic steroids in rats promotes structural changes in the prostate manifested by changes in the weight, volume and epithelium height of the prostate ventral lobe and a predominance of collagen fibers in rats injected intramuscular with 10mg/kg-body weight using morphometric assessment. Ferrari et al. (2013) studied testis and seminal vesicle morpho-physiology of rats treated with nandrolone decanoate (injected with 0.5mg/kg body weight) and submitted to physical training in relation to the weight of the seminal vesicles. The anabolic caused significant increases in both sedentary and trained animals.

Little is known about the alterations might occur on the seminal vesicle due to nandrolene decaonate. The objective of the present study is to identify the effect of different doses of nandrolone decanoate (2.5, 5 and 10 mg/kg body weight) on the seminal vesicle morphologically and histopathologically.

MATERIALS AND METHODS

Nandurabolin 25 mg mg/ml) as (Nandrolone Decanoate) was obtained from El-Nile Company for Pharmaceutical and Chemical Industries, Egypt.

Animals

Twenty adult male Sprague-Dawley rats 3 month old, weighting 80-190gm were obtained from the Laboratory Animal House, Faculty of Medicine, Assiut University. The animals were housed in cages, fed a standard laboratory diet and water ad libitum.

Experimental design

Rats were divided randomly into 4 groups each of 5 rats, one control (G1) and three treated groups (G2,G3,G4). The treated groups were injected intramuscularly with Nandrolene Decaonateat a dose level of 2.5, 5 and 10 mg/kg/weekly. The control group received the same volume of normal saline.

Sampling

The whole body and seminal vesicle weights of both treated and control animal were taken and recorded. Specimen from the seminal vesicle was taken from each rat after three months post-injection and fixed in 10% formalin, dehydrated, embedded in paraffin, sectioned at 4-5µ, stained with H&E and examined by light microscopy. The thickness of the smooth muscle fiber layer of the seminal vesicle was chosen for morphometeric measurements using Fiji Soft ware. The data were expressed by the mean+ SEM.

Statistical analyses

ANOVA test used for statistical comparison of data (b<0.05) was considered statistically significant. All statistical calculations were made with the SPSS computer program, version 16.0 (SPSS Inc., Chicago, IL, USA). Data are presented as the means ± standard error (SE).

RESULTS

The seminal vesicle weight in relation to the body weight of rats injected with 2.5(Group1), 5(Group2), and 10mgkg/ body weight(Group3) and collected after three months post-injection was significant increased in weight compared to the control groups as shown in Table (1).

The thickness of the smooth muscle fiber layer was 99.01+ 3.15µm in the control group (Fig.1). While, rats injected with 2.5 mg/kg body weight was 169.50+3.09µm (Fig.2). In group 2 (5mg/kg body weight) the thickness was 178.45 + 5.32 µm(Fig.3). Rats injected with 10mg/kg body weight showed thickness of 207.56+ 5.31 µm as shown in Fig.4. Variation in the thickness among different groups and control was illustrated in Fig.5.

Histopathologically, the seminal vesicle from control group had the normal appearance and consisted of mucosal folds filled with red homogenous secretion and muscular layer made of smooth muscle fibers (Fig6). The seminal vesicle of rats injected with 2.5 mg/kg BW Nandrolene Decaonate and collected after three months post-injection showed shortening of the mucosal folds, abundant secretion and fragmentation of the smooth mucle fibers (Figs.7&8). The seminal vesicle from rats injected with 5 mg/kg BW Nandrolene Decaonate after three months post-injection had or also abundant secretion, shorten folds and fragmentation of smooth muscle fibers in addition to sub-muscular hemorrhage and hyperemia of blood vessel (Figs 9,10&11). Desquamation of the epithelium lining the mucosal folds was observed in the seminal vesicle of rats injected with 10 mg/kg BW Nandrolene Decaonate after three months post-injection (Fig.12).

Table 1: The ratio (+ SEM) of the seminal vesicle weight/gm in different doses of Nandrolene Decaonatein rats to the whole body weight.

Fig. 5: Variation in the thickness of the smooth muscle fibers of the seminal vesicle.

Figure Legends

Fig.1: seminal vesicle from rat of the control group showing the smooth muscle fibers (SMF) layer with a thickness of 99.44 + 3.15um. H&E. Bar=50um.

Fig.2: Seminal vesicle of rats IM injected with 2.5 mg/kg BW Nandrolene Decaonate weekly. Note increase in the thickness of SMF (169.50+ 3.09 um, arrow) compared with the control and altered nuclei morphology H&E. Bar= 50um.

Fig.3: Seminal Vesicle of rats IM injected with 5 mg/kg BW Nandrolene Decaonate weekly showing thickness of SMF layer (178.45+ 5.32 um(arrow). H&E. bar=100um.

Fig.4: Seminal Vesicle of rats IM injected with 10 mg/kg BW Nandrolene Decaonate weekly showing thickness of the SMF = 207.56+ 5.31um, arrow.  H&E. bar=50um.

Fig.5: Variation in the thickness of the SMF of the seminal vesicle.

Fig.6: Control rat seminal vesicle IM injected with physiologic saline showing the normal appearance of seminal vesicle. Long folds of mucosa (F), SMF and homogenous red secretion (S).H&E. Bar=100um.

Fig.7: Seminal vesicle of rats IM injected with 2.5 mg/kg BW Nandrolene Decaonate weekly for three months showing abundant secretion (*), shortening of folds (arrow) and fragmentation of SMF () H&E. bar=100um.

Fig.8: Higher magnification of Fig.7 showing the SMF fragmentation (). H&E. bar= 50um.

Fig.9: Seminal vesicle of rats IM injected with 5 mg/kg BW Nandrolene Decaonate weekly for three months showing abundant secretion(*), shortening of folds (arrow) and fragmentation of SMF layer similar to 2.5mg/kg BW.  H&E. bar=100um.

Fig.10: Seminal vesicle of rats IM injected with 5 mg/kg BW Nandrolene Decaonate weekly for three months showing submuscular hemorrhage (star) and hyperemia of blood vessel (arrow).  H&E. bar=100um.

Fig.11: Seminal vesicle of rats IM injected with 5 mg/kg BW Nandrolene Decaonate weekly forthree months showing pronounced fragmentation of the SMF (star) and hyperemia of blood vessel (arrow).  H&E. bar=100um.

Fig.12: Seminal vesicle of rats IM injected with 10 mg/kg BW Nandrolene Decaonate weekly for three months showing pronounced desqumation of the epithelium lining the folds (arrows) H&E. bar=100um.

DISCUSSION

In the present study, there was an increase in the body weight by three months post-injection. The increase in body weight was also associated with the seminal vesicle weight. The increase in the body weight in animals and human due to the use of Nandrolene Decaonate was also reported by many authors (Bhasin et al., 1996, Beatriz et al., 2000). Some authors however, believed that the gain in the body weight is apparently due to the muscular exercise rather than an anabolic effect (Schürmeyer et al., 1984). Others reported that increase in body weight may be attributed to the accumulation of fluids and sodium in the body (Forbes, 1985). There is an acceptable consequence that supra-physiological doses of AASs can inhibit body growth and weight gain (Carson et al., 2002).

The seminal vesicle weight in the present study had significant increase in the weight in all treated groups (2.5, 5 and 10mg/kg body weight) compared to the control group. These results were correlated to the increase in thickness of the smooth muscle fibers layer of the seminal vesicle (169.50+3.09, 178.45 + 5.32 and 207.56+ 5.31 µm) in treated groups compared to the control group (99.01+ 3.15µm).These results may suggest that the increased weight and thickness are due to the direct effect of Nandrolene Decaonate. Ferrari et al. (2013) investigated the effect of intramuscular injections of Nandrolone Decanoate (0.5mg kg-1 body weight) on male rats sedentary control, sedentary treated, trained control and trained treated for eight weeks. The study revealed that the weight of the seminal vesicles was significantly increased in both sedentary and trained animals and demonstrating the high sensitivity of the reproductive structure to Nandrolone Decanoate. Shokri et al. (2014) reported that the relative weights of the seminal vesicle in the Nandrolone Decanoate exercise groups significantly decreased compared with the control group.

Little is known about the effect of Nandrolone Decanoateon the histopathological alterations of the seminal vesicle. In the present study, there was an increase in the secretion of the seminal vesicle at the expense of the length of the mucosal folds at a dose of 2.5 and 5mg/kg body weight as well as fragmentation of the smooth muscle fibers. Vascular changes expressed by hyperemia of blood vessels at a dose of 5mg/kg bodyweight. Nandrolone Decanoateat 10mg/kg body weight induced sloughing of the epithelial lining the mucosal folds into its lumen. These results suggested that Nandrolone Decanoate induced both degenerative and vascular changes of the seminal vesicle.

In conclusion, the present study showed that Nandrolone Decanoate affects both the weight and morphology of the seminal vesicle and suggested that the seminal vesicle can be a sensitive organ to the anabolic steroids drug.

REFERENCES

Barceloux, Donald, G.; Palmer and Robert, B. (2013): Anabolic-Androgenic Steroids. Disease-a-Month 59, 226–248.

Bhasin, S.; Storer, T.W. and Berman, N. (1996): The effects of supraphysiologic doses of testosterone on muscle size and strength in normal men. N. Engl. J. Med. 335: 1-7.

Beatriz G.; Raquel Hernando and Rafael Manso (2000): Anabolic steroid and gender- dependent modulation of cystolicHSP70s in fast-and low-twitch skeletal muscle. The Journal of Steroid Biochemistry and Molecular Biology, 74: 63-71.

Buttner, A. and Thieme, D. (2010): Side effects of anabolic androgenic steroids: Pathological findings and structure–activity relationships. Handb. Exp. Pharmacol., 195:459–484.

Carson, J.A.; Lee, WJ.; McClung, J. and Hand, GA. (2002): Steroid receptor concentration in aged rat hindlimb muscle: Effect of anabolic steroid administration. J. Appl. Physiol, 93: 242-50.

Ferrari, F.; de Paiva Foletto, M.; de Moraes, S.; Peres, S.P.; Segatelli, T.M. and da Costa, M. (2013): Testis morphophysiology of rats treated with nandrolone decanoate and submitted to physical training. Acta Scientiarum. Health Sciences, Maringá, v. 35, n. 2, 161-167.

Forbes, G.B. (1985): The effect of anabolic steroids on lean body mass. The dose response curve. Metabolism, 34,571-573.

Laseter, J.T. and Russell, J.A. (1991): Anabolicsteroid -induced tendonpathology: a review of the literature. Med Sci Sports Exerc, 23,1–3.

Schürmeyer, T.; Knuth, UA.; Belkien, L. and Nieschlag, E. (1984): Reversible azoospermia induced by the anabolic steroid 19- nortestosterone. Lancet 1: 417-20.

Shokri, S.; Aitken, Hn.; Robert Jo; Abdolvahhabi, M.; Abolhasan F.; Ghasemi, F.J.; Kashani, I.; Shahram Ejtemaeimehr, Shahin; Bagher, A.; Naraghi, M.A. and Barbarestani, M. (2014): Exercise and Supraphysiologicaldose of Nandrolone Deconoate Increase apoptosis in spermatogeniccells. Nordic Pharmacological Society. Basic & Clinical Pharmacology & Toxicology, 106, 324–330.

Soma, L.R.; Uboh, C.E.; Guan, F. and Pack, J. (2007): Pharmacokinetics of boldenone and stanozolol and the results of quantification of anabolic and androgenic steroids and in race horses and non-race horses. J. Vet. Pharmacol. Therap. 30, 101–108.