Ameliorative Potentials of a Combination of Fenugreek and α-tocopherol on Cadmium induced Testicular
Ameliorative Potentials of a Combination of Fenugreek and Α-Tocopherol on Cadmium Induced...
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Ameliorative Potentials of a Combination of Fenugreek and α-tocopherol on Cadmium induced Testicular toxicity: An Ultrastructural Study Adel M. Hussein, Hesham N. Mustafa, Mohamed H. Badawoud Anatomy Department, Faculty of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia Introduction Cadmium is an ecological pollutant which clas- sified amongst the 10 most poisonous elements for human beings. Due to its long biological half-life, exposure to cadmium causes toxic effects after long periods of a cessation of exposure due to its residual effects. Perox- idation induced by Cd is the cause of the release of free oxygen radicals. The oxygen radicals lead to the destruction of sensitive molecules and tissues [1]. α-tocopherol causes the elimination of the free oxygen radicals and the inhibition of the peroxidation [2]. Fenu- greek seed demonstrate an antioxidant impact due to presence of different ingredients like flavonoids, alkaloids and amino acids [3]. Objectives The purpose of this study is to evaluate the cytoprotective effcts of fenugreek and α-toco- pherol on cadmium chloride induced testicular damage. Materials & Methods Forty male adult Wistar rats weighed 175-200 gm, randomly placed into four groups (N=10). Group I served as the control group. Group II received 5 mg/kg/day cadmium chloride (1/15 LD50) [4]. Group III received 5 mg/kg/day cad- mium chloride and 150 mg/kg/day fenugreek and 100 mg/kg/day of α-tocopherol [5]. The prophylactic combination started one week before CdCl2 administration to acquire the benefis of both curative and preventive regi- mens. All groups were treated by oral gavage for 60 consecutive days. Testis where subject- ed to histology using H&E. Immunohistochem- istry (IHC) using Anti-CD68+ for macrophages, Anti CD3+ for T-lymphocytes, and Anti CD20+ for B-lymphocytes. Morphometry and Ultra- structural examination. Results Graph (1) Mean testes weight and body weight by grams Fig. (1A): Control. Fig. (1B): CdCl2 showed disin- tegration of the testicular architecture, vacuola- tion (V), a separation of the spermatogonia from the basal lamina (↑), and a widening of the inter- stitial spaces (W). Fig. (1C): Higher magnification showed multinucleated giant cells (G) and con- gestion of the capillaries (C). Fig. (1D): CdCl2 and combined fenugreek and α-tocopherol showed preservation of the architecture of the testicular tissues. Fig. (2A). Control. Fig. (2B) CdCl2 showed a no- ticeable reduction in epithelial cells. Vacuolations (V) and intercellular areas (sp) among the germ cells can be observed. Fig. (2C) CdCl2 showed depletion, and separation of the spermatogonia layer, vacuolization (V) of the germinal epitheli- um. (2D) Combined fenugreek and α-tocopherol with CdCl2 shows an apparent improvement of the testicular architecture with a partial restora- tion of a low germinal epithelium. Graph (2): Mean Diameter of Seminiferous Tubules and Germinal Epithelial Height. Graph (3): Mean area percentage of CD3, CD20, and CD68 immunostaining. Fig. (3A). CdCl2 group showed a Sertoli cell with an irregular euchromatic nucleus (St) with the characteristic indentation of the nucleus. Prima- ry spermatocyte (Sc) is separated by intercellu- lar spaces (white star) and the vacuolated distort- ed mitochondria (black arrows). Vacuoles (V) all over the cytoplasm. Note the apparently thick- ened basal lamina (BL) containing collage (black star) and a myoid cell (curved arrow). Fig. (3B). A mid piece of the sperm (MP) with distorted and swollen mitochondrial sheaths (black arrow) sur- round the central axoneme (white arrow). End (EP) pieces can be observed. Fig. (3C). A Leydig cell (L) with an oval, indented nucleus. An elec- tron dense bodies, vacuoles (V), an empty space (black star), lymph vessels (Ly), and the ruptured vacuolated mitochondria can be observed (white arrow). Spermatogonia (Sg) and Sertoli cell (St) are resting on an apparently thickened basal lam- ina (BL) with a myoid cell (star). Fig. (3D). Sper- matids (Sd) with rounded or oval nuclei that have disturbed chromatin and peripherally arranged the mitochondria with lost cristae (black arrow) and the characteristic acrosomal cap (Ac) can be observed. Fig. (4A). Combined fenugreek and α-tocopherol and CdCl2 showed a Sertoli cell (St) with a eu- chromatic-indented nucleus and apparent nu- cleolus (N) and the spermatogonia (Sg) resting on the thickened basal lamina (BL) with an ap- parent myoid cell (black star). Some mitochon- dria are affected, and others are healthy (arrow). Fig. (4B). Reconstruction of the structure of the cross-sections of the mid pieces (MP) with ax- oneme (white arrow) surrounded by a mitochon- drial sheath (black arrow). Fig.(4C). Leydig cells with oval euchromatic nuclei (L) and peripherally located heterochromatin. The cytoplasm contains multiple lysosomes (S) and many mitochondria (arrow). Fig. (4D). Improvement of the appearance of germ cells. The Spermatids (Sd) with rounded or oval euchromatic nuclei and peripherally ar- ranged mitochondria with preserved cristae (ar- row) and the characteristic acrosomal cap (Ac) can be observed. Minimal empty spaces (star) in the affected intercellular bridge. Conclusions Fenugreek and α-tocopherol could represent a promising medicinal combination to amelio- rate the toxic effects of cadmium exposure. References 1. Ognjanovic, B., Markovic, S., Pavlovic, S., Zikic, R., Stajn, A., & Saicic, Z. (2008). Effect of chronic cadmi- um exposure on antioxidant defense system in some tis- sues of rats: protective effect of selenium. Physiologi- cal Research, 57(3), 403. 2. Farina, M., Soares, F. A., Feoli, A., Roehring, C., Brusque, A. M., Rotta, L., Rocha, J. B. T. (2003). In vit- ro effects of selenite and mercuric chloride on liver thiobarbituric acid–reactive substances and non-protein thiols from rats: Influences of dietary cholesterol and polyunsaturated and saturated fatty acids. Nutrition, 19(6), 531-535. 3. Flammang, A., Cifone, M., Erexson, G., & Stankows- ki Jr, L. (2004). Genotoxicity testing of a fenu- greek extract. Food and chemical toxicology, 42(11), 1769-1775. 4. Renugadevi, J., & Prabu, S. M. (2009). Naringenin pro- tects against cadmium-induced oxidative renal dysfunc- tion in rats. Toxicology, 256(1-2), 128-134. 5. Aslanturk, A., Uzunhisarcikli, M., Kalender, S., & Demir, F. (2014). Sodium selenite and vitamin E in preventing mercuric chloride induced renal toxicity in rats. Food Chem Toxicol, 70, 185-190.
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Ameliorative Potentials of a Combination of Fenugreek and Α-Tocopherol on Cadmium Induced Testicular Toxicity an Ultrastructural Study
Transcript of Ameliorative Potentials of a Combination of Fenugreek and Α-Tocopherol on Cadmium Induced...
Ameliorative Potentials of a Combination of Fenugreek andα-tocopherol on Cadmium induced Testicular toxicity: An
Ultrastructural StudyAdel M. Hussein, Hesham N. Mustafa, Mohamed H. Badawoud
Anatomy Department, Faculty of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia
IntroductionCadmium is an ecological pollutant which clas-sified amongst the 10 most poisonous elementsfor human beings. Due to its long biologicalhalf-life, exposure to cadmium causes toxiceffects after long periods of a cessation ofexposure due to its residual effects. Perox-idation induced by Cd is the cause of therelease of free oxygen radicals. The oxygenradicals lead to the destruction of sensitivemolecules and tissues [1]. α-tocopherol causesthe elimination of the free oxygen radicals andthe inhibition of the peroxidation [2]. Fenu-greek seed demonstrate an antioxidant impactdue to presence of different ingredients likeflavonoids, alkaloids and amino acids [3].
ObjectivesThe purpose of this study is to evaluate thecytoprotective effcts of fenugreek and α-toco-pherol on cadmium chloride induced testiculardamage.
Materials & MethodsForty male adult Wistar rats weighed 175-200gm, randomly placed into four groups (N=10).Group I served as the control group. Group IIreceived 5 mg/kg/day cadmium chloride (1/15LD50) [4]. Group III received 5 mg/kg/day cad-mium chloride and 150 mg/kg/day fenugreekand 100 mg/kg/day of α-tocopherol [5]. Theprophylactic combination started one weekbefore CdCl2 administration to acquire thebenefis of both curative and preventive regi-mens. All groups were treated by oral gavagefor 60 consecutive days. Testis where subject-ed to histology using H&E. Immunohistochem-istry (IHC) using Anti-CD68+ for macrophages,Anti CD3+ for T-lymphocytes, and Anti CD20+for B-lymphocytes. Morphometry and Ultra-structural examination.
Results
Graph (1) Mean testes weight and body weight bygrams
Fig. (1A): Control. Fig. (1B): CdCl2 showed disin-tegration of the testicular architecture, vacuola-tion (V), a separation of the spermatogonia fromthe basal lamina (↑), and a widening of the inter-stitial spaces (W). Fig. (1C): Higher magnificationshowed multinucleated giant cells (G) and con-gestion of the capillaries (C). Fig. (1D): CdCl2 andcombined fenugreek and α-tocopherol showedpreservation of the architecture of the testiculartissues.
Fig. (2A). Control. Fig. (2B) CdCl2 showed a no-ticeable reduction in epithelial cells. Vacuolations(V) and intercellular areas (sp) among the germcells can be observed. Fig. (2C) CdCl2 showeddepletion, and separation of the spermatogonialayer, vacuolization (V) of the germinal epitheli-um. (2D) Combined fenugreek and α-tocopherolwith CdCl2 shows an apparent improvement ofthe testicular architecture with a partial restora-tion of a low germinal epithelium.
Graph (2): Mean Diameter of SeminiferousTubules and Germinal Epithelial Height.
Graph (3): Mean area percentage of CD3, CD20,and CD68 immunostaining.
Fig. (3A). CdCl2 group showed a Sertoli cell withan irregular euchromatic nucleus (St) with thecharacteristic indentation of the nucleus. Prima-ry spermatocyte (Sc) is separated by intercellu-lar spaces (white star) and the vacuolated distort-ed mitochondria (black arrows). Vacuoles (V) allover the cytoplasm. Note the apparently thick-ened basal lamina (BL) containing collage (blackstar) and a myoid cell (curved arrow). Fig. (3B).A mid piece of the sperm (MP) with distorted andswollen mitochondrial sheaths (black arrow) sur-round the central axoneme (white arrow). End
(EP) pieces can be observed. Fig. (3C). A Leydigcell (L) with an oval, indented nucleus. An elec-tron dense bodies, vacuoles (V), an empty space(black star), lymph vessels (Ly), and the rupturedvacuolated mitochondria can be observed (whitearrow). Spermatogonia (Sg) and Sertoli cell (St)are resting on an apparently thickened basal lam-ina (BL) with a myoid cell (star). Fig. (3D). Sper-matids (Sd) with rounded or oval nuclei that havedisturbed chromatin and peripherally arrangedthe mitochondria with lost cristae (black arrow)and the characteristic acrosomal cap (Ac) can beobserved.
Fig. (4A). Combined fenugreek and α-tocopheroland CdCl2 showed a Sertoli cell (St) with a eu-chromatic-indented nucleus and apparent nu-cleolus (N) and the spermatogonia (Sg) restingon the thickened basal lamina (BL) with an ap-parent myoid cell (black star). Some mitochon-dria are affected, and others are healthy (arrow).Fig. (4B). Reconstruction of the structure of thecross-sections of the mid pieces (MP) with ax-oneme (white arrow) surrounded by a mitochon-drial sheath (black arrow). Fig.(4C). Leydig cellswith oval euchromatic nuclei (L) and peripherallylocated heterochromatin. The cytoplasm containsmultiple lysosomes (S) and many mitochondria(arrow). Fig. (4D). Improvement of the appearanceof germ cells. The Spermatids (Sd) with roundedor oval euchromatic nuclei and peripherally ar-ranged mitochondria with preserved cristae (ar-row) and the characteristic acrosomal cap (Ac)can be observed. Minimal empty spaces (star) inthe affected intercellular bridge.
ConclusionsFenugreek and α-tocopherol could representa promising medicinal combination to amelio-rate the toxic effects of cadmium exposure.
References1. Ognjanovic, B., Markovic, S., Pavlovic, S., Zikic, R.,
Stajn, A., & Saicic, Z. (2008). Effect of chronic cadmi-um exposure on antioxidant defense system in some tis-sues of rats: protective effect of selenium. Physiologi-cal Research, 57(3), 403.
2. Farina, M., Soares, F. A., Feoli, A., Roehring, C.,Brusque, A. M., Rotta, L., Rocha, J. B. T. (2003). In vit-ro effects of selenite and mercuric chloride on liverthiobarbituric acid–reactive substances and non-proteinthiols from rats: Influences of dietary cholesterol andpolyunsaturated and saturated fatty acids. Nutrition,19(6), 531-535.
3. Flammang, A., Cifone, M., Erexson, G., & Stankows-ki Jr, L. (2004). Genotoxicity testing of a fenu-greek extract. Food and chemical toxicology, 42(11),1769-1775.
4. Renugadevi, J., & Prabu, S. M. (2009). Naringenin pro-tects against cadmium-induced oxidative renal dysfunc-tion in rats. Toxicology, 256(1-2), 128-134.
5. Aslanturk, A., Uzunhisarcikli, M., Kalender, S., & Demir,F. (2014). Sodium selenite and vitamin E in preventingmercuric chloride induced renal toxicity in rats. FoodChem Toxicol, 70, 185-190.
