PROTECTIVE INFLUENCE OF OLIVE OIL ON REPRODUCTIVE PARAMETERS IN MALE RAT TREATED WITH CADMIUM

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1 G.J.B.B., VOL.2 (4) 2013: ISSN PROTECTIVE INFLUENCE OF OLIVE OIL ON REPRODUCTIVE PARAMETERS IN MALE RAT TREATED WITH CADMIUM Nwl Khiry Hussin AL-Ani High Institute of Infertility Dignosis nd Assisted Reproductive Technology, Al-Nhrin University, Bghdd/Irq ABSTRACT Cdmium is no-essentil toxic trce element, it ffects reproductive orgns. It's ction my e either direct, ffecting the gonds nd ccessory orgns, or indirect vi interference with the hypothlmus-pituitry-gondl xis. Olive oil is ft otined from the olive, its helth enefits re due to its ftty cid composition nd minor compounds, tocopherols, polyphenols, ortho-diphenols crotenoids, nd Vitmin E which possesses strong rdicl scvenging ctivity. The purpose of this study ws to investigte the protective effect of olive oil ginst cdmium-induced testiculr dysfunction in dult mle rts. Forty five mture lino rt (Rttus rttus norvegicus linos) ws used s mmmlin model. These nimls were divided into three groups, 15 rt/group.group1, served s control ws orlly dministrted with distilled wter. Testiculr dysfunction ws induced y n orl dministrtion of cdmium chloride ( 2mg/kg ody weight, for four weeks dily) group2. Group3 ws orlly dministrted with cdmium in dose of (2mg/kg ody weight)plus one ml of extr virgin olive oil, for four weeks To ll nimls, hormones (LH, FSH nd Testosterone) were mesured, histologicl study ws done nd semen prmeters ws studded. The effects of dministrtion of cdmium showed significnt decrese in semen prmeters, highly significnt increse in FSH level, significnt decrese in LH level, significnt decrese in testosterone level nd testiculr histologicl normlities. Administrtion of olive oil with cdmium shows cused n elevtion in sperm counts, their motility, their normlity, improvement in histologicl findings nd hormonl results when compred with treted group with cdmium only. Olive oil plys significnt role in the improvement of testiculr dmge long with reduces oxidtive stress nd protect from reproductive orgns injury during cdmium toxicity. KEY WORDS: Cdmium, Olive oil, LH, FSH, Testosterone. INTRODUCTION It is well estlished tht mny trce elements ply essentil role in numer of iologicl processes through their ction s ctivtors or inhiitors of enzymtic rection y competing with other elements nd proteins for inding sites, nd influencing the permeility of cell memrne or through other mechnisms (1).A lrge numer of trce elements could exert stimultory or inhiitory effects on the sperm depending on the concentrtion of ech divlent ctions (2). Cdmium (Cd) is non-essentil toxic trce element tht hs no known iologicl function. Its most undnt nturlly-occurring isotope is nonrdioctive. It is found in nture in minerl forms nd is otined for commercil uses principlly from cdmium ore, clled greenockite, which is commonly found in ssocition with zinc ore. Commercil production of cdmium ore depends on the mining of zinc.cd is commercilly ville s n oxide, chloride, or sulfide. Cd metl (Cd2+) refined from the ore is silver -white, lue-tinged lustrous hevy metl solid t room temperture (3). Cd ffects reproductive orgns (4). It s ction my e either direct, ffecting the gonds nd ccessory orgns, or indirect vi interference with the hypothlmus-pituitrygondl xis (5). Cd hs een shown to concentrte in testiculr tissue fter occuptionl exposure.acute exposure to lrge doses of Cd induces gondl necrosis wheres chronic tretment with low doses hs een shown to produce testiculr trophy (6). Olive oil is ft otined from the olive, it is used throughout the world. Olive oil is composed minly of the mixed triglyceride esters of oleic nd plmitic cid long with trces of squlene (up to 0.7%) nd sterol (out 0.2% phytosterol nd tocosterols) (7).Virgin olive oil is chrcterized y its sensoril nd nutritionl properties, which re different from those of other edile oils. Its helth enefits re due to oth its ftty cid composition nd minor compounds, tocopherols, polyphenols, sterols, nd crotenoids (8). A numer of studies hve shown tht polyphenolic compounds possess strong rdicl scvenging ctivity nd pper to e t lest, s if not more, effective thn other importnt dietry ntioxidnt (9). The ftty cid composition of virgin olive oil hs gret importnce from helth point of view. The purpose of this study ws to investigte the protective effect of olive oil ginst cdmium-induced testiculr dysfunction in dult mle rts. MATERIALS & METHODS All experiments were performed on 45 mture lino rt (Rttus rttus norvegicus linos) s mmmlin model with ody weight (B. wt) rnging etween g. Rt were otined from the colony of the niml house of the High Institute for Infertility Dignosis & ART. AL- Nhrin University/Irq. These nimls were divided into three groups, 15 rt/group fter leling them with er or til mrking nd weighing them using n electricl lnce, s follows: group (G1): This group ws 500

2 Olive oil on reproductive prmeters in mle rt treted with cdmium orlly dministrted with distilled wter during the four weeks period of the experiment. Experimentl group (G2): This group ws orlly dministrted with cdmium in dose of(2mg/kg ody weight) (10) dily during the four weeks period of the experiment. Experimentl group (G3): This group ws orlly dministrted with cdmium in dose of (2mg/kg ody weight) plus one ml of extr virgin olive oil) (11). All tretments were given y intrgstric tue dily. Blood smples were spirted from the hrt nd put in plstic centrifuge tues, left to clot nd serum ws otined y centrifugtion t 1500 rpm (revolution per minute) for15 min, the smple were kept frozen t-20 0 C until LH, FSH nd testosterone were mesured. Then rts were scrificed the weight of reproductivee orgns were recorded, reproductive orgns (testis nd epididymes) were quickly excised nd immersed in few drops of norml sline which ws plced in Petri dish to e clered from surrounding dipose tissue under dissecting microscope using fine surgicl scissors. Both right nd left testes were fixed with 10% forml sline for susequent histologicl study. The right nd left epididymes were put in smll Petri dish contined 1ml of gloll culture medi then the epididymes were minced y microsurgicl scissor out 200 times until we got homogenized solution which contined the spermtozo. Sttisticl nlyze The dt were sttisticlly nlyzed using SPSS/PC version 10 softwre (SPSS, Chicgo) ). Level of FSH, LH nd testosterone were nlyzed using complete rndomized design (CRD). The Sttisticl model ws Yij = µ + Ti + eij. Where Yij= dependent vriles (Level of FSH, LH nd testosterone), µ= overll men, Ti= effect of tretments (cdmium), eij= error term. Differences mong mens were computed using the Duncn multiple rnges test) (12). RESULTS The effects of dministrtion of cdmium showed significnt decrese in sperm concentrtion, highly significnt decrese in progressive sperm motility, significnt decrese in non progressive motility, significnt increse in immotile sperms nd highly significnt increse in sperm normlity. While the dministrtion of cdmium plus olive oil showed non significnt differences in sperm concentrtion nd sperm normlity, with significnt decrese in progressive nd non progressive motility nd significnt increse in immotile sperm when compred with control s shown in Tle 1. TABLE 1. Effect of cdmium tretment nd cdmium plus olive oil on Sperm Prmeters in mle rt Sperm prmeters sperm concentrtion millions/ml Progressive sperm motility (%) Non progressive sperm motility (%) Immotile sperm (%) ± ± ± ±7.246 Cdmium ± ± ± ±4.254 c Cdmium & Olive oil ± ± ± ±3.141 Sperm normlity (%) c ±3.329 ±1.667 ±2.386 cdmium cdmium +olive oil FSH level ng/ml c cdmium 6.04 cdmium +olive oil Tretments FIGURE 1. Effect of cdmium tretment nd cdmium plus olive oil on FSH level in mle rt serum. * Mens with different superscripts within ech columns re significntly different (P<0.05) ** Mens with semi superscripts within ech columns re non significntly different (P>0.05) Figure 1showed highly significnt increse e in FSH level fter cdmium dministrtion with significnt increse in FSH level fter cdmium plus olive oil dministrtion when compred with control. Figure 2showed significnt

3 G.J.B.B., VOL.2 (4) 2013: decrese in LH level fter cdmium dministrtion with non significnt deference in LH level fter cdmium plus olive oil dministrtion when compred with control. Figure 3showed significnt decrese in testosterone level ISSN fter cdmium dministrtion with non significnt deference in testosterone level fter cdmium plus olive oil dministrtion when compred with control. cdmium cdmium +olive oil LH level ng/ml cdmium cdmium +olive oil Tretments FIGURE 2. Effect of cdmium tretment nd cdmium plus olive oil on LH level in mle rt serum. * Mens with different superscripts within ech columns re significntly different (P<0.05) ** Mens with semi superscripts within ech columns re non significntly different (P>0.05) cdmium cdmium +olive oil Testesterone level ng/ml cdmium cdmium +olive oil Tretments FIGURE 3. Effect of cdmium tretment nd cdmium plus olive oil on testosterone level in mle rt serum. * Mens with different superscripts within ech columns re significntly different (P< <0.05). ** Mens with semi superscripts within ech columns re non significntly different (P> P>0.05). Histologicl oservtion Testiculr sections of the control group showed seminiferous tuules with norml germ cell popultion lyer thickness with norml orderly rrnged pttern up to mture spermtid. No mlignnt or norml cell ws seen within the germinl epithelium, lso no vcuoles is present in the tuules. There were dequte Sertoli cells popultions. The interstitil spces contined dequte norml Leydig interstitil cell, lood vessels were lso norml nd no edem ws present. The pertuulr surrounding tissues were norml s shown in figure 4 nd figure5.in cdmium treted group (G2), sections showed decrese in thickness of germ cell lyer, widening of the centrl seminiferous tuules lumen nd prominent germ cell popultion necrosis. Multiple rounded vcuoles were seen within the tuules, with the decreses inspermtogenesis nd stroml interstitil Leydig cell hypertrophy. Also Widening of the interstitil spces ws detected nd edem within the interstitil spces ws mrkedly present. A perituulr firotic chnge hd een seen lso in the testiculr sections s shown in figure 6.In the group treted with cdmium plus olive oil (G3), sections of tuules showed recovery of germ cell popultion, norml germinl epithelium thickness with order rrngement of the germ cell up to spermtid, dequte Sertoli nd Leydig interstitil cells popultion. Mild edem ws seen t the periphery of the testiculr section in this group s shown in figure 7 nd figure

4 Olive oil on reproductive prmeters in mle rt treted with cdmium FIGURE 4. Cross sections of testes were stined with hemtoxylin nd eosin (H nd E) of control Group 1 x10 FIGURE 5. Cross sections of testes were stined with hemtoxylin nd eosin (H nd E) of control Group 1 x 40 FIGURE 6. Cross sections of testes were stined with hemtoxylin nd eosin (H nd E) of Group2 treted with cdmium x40 FIGURE 7. Cross sections of testes were stined with hemtoxylin nd eosin (H nd E) of Group3 treted with cdmium plus Olive oil x4 FIGURE 8. Cross sections of testes were stined with hemtoxylin nd eosin (H nd E) of Group3 treted with cdmium plus Olive oil x10 DISCUSSION The present study showed the efficcy of olive oil in preventing the toxic effects of Cdmium in the rt testes. To our knowledge, this is the first report showing these effects. Our dt showed tht the dministrtion of cdmium to mle rts produced generlized degenertion of the germ, sertoli nd Leydig cells which grees with the dt reported y Lskey Jw et l nd Foote RH (13,14) nd lso showed decrese in sperm concentrtion, progressive motility, sperm normlity nd testiculr structure, this my e ttriuted to testiculr germ cell destruction cused either y memrne dmge or mcromoleculr degrdtion incurred y rective oxygen species (ROS) leding to significnt d ecline in spermtogenesis or perhps due to the low production of sperm in testes nd tht could e relted to the low levels of gondotrophins nd/or serum testosterone level nd my e due to testiculr necrosis (15).It hs een 503

5 G.J.B.B., VOL.2 (4) 2013: ISSN demonstrted tht Cd stimultes free rdicl production, resulting in oxidtive deteriortion of lipids, proteins nd DNA, nd inititing vrious pthologicl conditions in humns nd nimls (16). Cdmium ct on the testes, cusing prolems in spermtogenesis nd spermiogenesis (17).The dmge is due to cpillry stsis followed y mssive thromosis, nd is highly specific to the testes (18).The primry insult seems to e n increse in cpillry permeility nd rekdown of the lood testes rrier (19). The net effect of decresed spermtogenesis is the comintion of diminished lood flow nd the ility of cdmium to compete with zinc for inding to enzymes essentil for cell repliction (20).When Cd is dministered orlly nd chroniclly, the testis ccumulted out 4 times more Cd thn fter single olus, high dose Cd injection. Acute exposure to lrge doses of Cd induces gondl necrosis wheres chronic tretment with low doses hs een shown to produce testiculr trophy (21). The effect of cdmium on the testes ppers to e mnifested minly in the Sertoli cells, which present more morphologicl chnges under scnning electron microscopy. Cdmium cn lso interfere with the norml functions of mitochondril enzymes (22).Cd decreses ndrogen iosynthesis, possily y ltering progesterone synthesis nd metolism through direct interction of Cd with DNA nd competitive inhiition of essentil enzymes (23). Testiculr lesion from cdmium is primrily vsculr, nd the vsculr dmge determines the degree of lesion in the germ cells nd Leydig cells. This lesion cn generte Leydig cell tumors, tuulr degenertion (in high-dose exposure), nd trophy (24), in ddition to inducing tissue necrosis nd deficient ndrogen production (25). An increse in serum levels of FSH fter cdmium dministrtion ws shown in this study. This finding could e due to the ccumultion of cdmium in the testis.in this sis, it ws shown tht cdmium ffects Sertoli cell ctivity y decresing inhiin synthesis nd relese. Thus, the incresed plsm levels of FSH could e explined, s this peptide is the min inhiitory signl for FSH secretion (26),nother explntion my e ecuse spermtogenesis is more sensitive to toxic chemicls which disrupt spermtogenesis often cuse rised FSH (27). Metl exposure lso decresed serum levels of LH nd testosterone. This decrese my reflect direct effects of the metl t the testis s this metl ccumultes in this tissue. This effect my explin tht the metl disrupted the interreltionship mechnisms etween LH nd testosterone (28). Mny investigtions showed tht chronic tretment with cdmium induced oxidtive dmge cusing destruction of cell memrnes nd incresed lipid peroxidtion (29). The current study lso shows tht dministrtion of Olive oil with cdmium cused n elevtion in sperm counts,their motility, their normlity,improvement in histologicl findings nd hormonl results, The virgin olive oil,contins precious sustnces of high iologicl vlue such s phenols, ortho-diphenols nd vrious nturl ntioxidnts, tocopherol (Vitmin E), flvones s well s high content of oleic cid, lcohols,sterols, chlorophyll, voltile romtic sustnces. (30,31). The phenolic compound, hydroxytyrosol hs een identified s one of the most potent ntioxidnts found in olive oil (32). The ntioxidnt effects of hydrophilic phenols nd tocopherols on the oxidtive stility in virgin olive oils nd in purified olive oil hve een evluted y Bldioli M et l,(30) they found tht purified olive oil hd good oxidtive stility. Vitmin-E present in significnt mounts in olive oil. (33), Vitmin E is the primry liposolule ntioxidnt, which my hve n importnt role in scvenging free oxygen rdicls nd in stilizing the cell memrnes, thus mintining its permeility. (34). Vitmin E my lso ffect oxidtive chnges which occur in other cell orgnelles. (35),nd prevent lipid peroxidtion nd cell destruction. (33) The scvenging properties of olive oil is sid to e the min importnt effects on the sperm prmeters. Our results showed the ility of olive oil to reverse the testiculr dysfunction cused y cdmium, s represented y normliztion of epididyml sperm counts nd testosterone level nd improvement of sperm motility long with restortion of spermtogenesis in seminiferous tuules s shown vi the histologicl testiculr exmintion. CONCLUSION On sis of ove results nd findings it is concluded tht olive oil plys significnt role in the improvement of testiculr dmge long with reduces oxidtive stress nd protect from reproductive orgns injury during cdmium toxicity. REFERENCES [1]. Rizk, S.L. & sky-peck, H. H. (1984) Comprison etween concentrtion of trce elements in norml nd neoplstic humn rest tissue. C. Res., 44: [2]. Stegmyer, B. nd Ronquist, G. (1982) Stimultion of Sperm progressive, motility y orgnelles in humn seminl plsm. Scnd. J. Urol. Nephrol., 16: [3]. Anonymous (1999 ) World Helth Orgniztion WHO: Reference vlues of semen vriles. In: Lortory Mnul for the Exmintion of Humn Semen nd Sperm-Cervicl Mucus Interction. 4 th ed. Cmridge University Press, Cmridge, UK. Pp [4]. Tomn, R., Mssnyi, P. nd Uhrin, V. ( 2002) Chnges in the testis nd epididymis of rits fter n intrperitonel nd perorl dministrtion of cdmium, Trce Elements nd Electrolytes, 19: [5]. Pksy, K., Vrg, B. nd Lzr, P. (1992) Cdmium interferes with steroid iosynthesis in rt grnulose nd lutel cells in vitro. Bio. Metls, 1992; 5: [6]. Lee, I.P., nd Dixon, R.L. (1973) Effects of cdmium on spermtogenesis studied y velocity sedimenttion cell seprtion nd seril mting. J. Phrmcol. Exptl. Therp., 1973; 187: 641. [7]. Griel, Crmin ARMEN Del R. Sestin Snd Leopoldo, M. (2004) Influence of Hrvest Dte nd Crop Yield on the Ftty Acid Composition of Virgin Olive Oils from Cv. Picul J. Agric. Food Chem.52, [8]. Mrtı nez de Victori, E.; Mn s, M. El ceite de oliv en l diet y slud humns. In ElCultiVo del 504

6 Olive oil on reproductive prmeters in mle rt treted with cdmium OliVo; Brrnco, D.,Fern ndez-escor, R., Rllo, L., Eds.; Mundi Prens: Mdrid,Spin, 2001; pp [9]. Beecher, G., Wrdern, B. & Merken H. (1999) Anlysis of te polyphenols. Proc. Soc. Exp. Biol. Med. 220: [10]. Anuncicion, L., Nuri, M., Mri, P-L, Dvid, P., nd An, I.E. (2001) Cdmium Effect on Hypothlmic- Pituitry- Testiculr Axis in Mle Rts. Experimentl Biology nd Medicin, 226: [11]. Gison, K., Oriji Hrry, R. Keiser (1998) Role of Nitric Oxide in Cyclosporine A Induced Hypertension. Hypertension; 32; [12]. Duncn, D B. (1995) Multiple rnge nd multiple F tests. Biometrics, 11:1 42, [13]. Lskey, J.W., Rehnerg, G.L., Lws, S.C., Hein, J.F. (1984) Reproductive effects of low cute doses of cdmium chloride in dult mle rts. Toxicol Appl Phrmcol. 1984; 73: [14]. Foote, R.H. (1999) Cdmium ffects testes nd semen of rits exposed efore nd fter puerty. Reprod Toxicol., 13: [15]. Mnn, P., Sinh, M. nd Sil, P.C. ( 2008) Cdmium induced testiculr pthophysiology: Prophylctic role of turine. Reprod. Toxicol. 26: [16]. Wiserg, M., Joseph, P., Hle, B., Beyersmnn, D. (2003)Moleculr nd cellulr mechnisms of cdmium crcinogenesis. Toxicology, 192: [17]. Boscolo, P., Scchettoni-Logroscino G., Rnelletti, F.O., Gioi, A., Crmignni, M. (1985) Effects of long-term cdmium exposure on the testis of rits: ultrstructurl study. Toxicol Lett, 24: [18]. Prizek, J., Zhor, Z. (1956) Effect of cdmium slts on testiculr tissue, Nture, 177:1036. [19]. Setchell, B. P., Wites, G. M. H. (1970) Chnges in the permeility of the testiculr cpillries nd of the 'lood testis rrier' fter injection of CdCl2 in the rt, J. Endocrinol., 47: [20]. Elcote, P.V., Fischer, M. I., Mwson, C.A., Millr, M. J. (1955) The effect of zinc deficiency on the mle genitl system, J. Physiol., 129: [21]. Lee, I. P., nd Dixon, R. L. (1973) Effects of cdmium on spermtogenesis studied y velocity sedimenttion cell seprtion nd seril mting. J. Phrmcol. Exptl. Therp.1973, 187: 641. [22]. Jequier, A.M. (2002) Mle infertility guide for the clinicin. Oxford: Blckwell Science; [23]. Ghosh, P. K., Ghosh, A. K. nd Bisws, N. M. (1987) Effect of Cdmium on 17-hydroxysteroid Dehydrogense in totl testis. Andrologi, 19(2): [24]. Wissmnn, W. Endocrinoptologi ssocid o trlho. In: Mendes R, orgnizdor. Ptologi do trlho. São Pulo: Editor Atheneu; [25]. Wlkesm M.P., Anver, M., Diwn, B.A. (1999) Crcinogenic effects of cdmium in the nole (NBL/Cr) rt: induction of pituitry, testiculr, nd injection site tumors nd intrepithelil prolifertive lesions of the dorsolterl prostte. Toxicol Sci., 52: [26]. Esquifino, A., Lfuente, Nuri M., Mrí, P-L, Dvid, P. nd An, I. (2001) Cdmium Effects on Hypothlmic-Pituitry-Testiculr Axis in Mle Rts Experimentl Biology nd Medicine, 226: [27]. Rees (1993) The Toxicology of Mle Reproduction. Thesis Literture Review in Applied Toxicology Portsmouth University Sumitted in prtil fulfillment of the requirements for the Degree of Mster of Science. [28]. Lfuente, A., González-Crrcedo, A., Romero, A. Cno, P., Esquifino, A.I. (2004) Cdmium exposure differentilly modifies the circdin ptterns of norepinephrine t the medin eminence nd plsm LH, FSH nd testosterone levels Toxicology Letters [29]. Bldioli, M., Servili, M., Perretti, G., Montedoro, G.F. (1996) Antioxidnt ctivity of tocopherols nd phenolic compounds of virgin olive oil. Journl of the Americn Oil Chemists Society,73: ) [30]. De Fveri, D., Alikrin, B., Avogrdo, M., Perego, P., Converti, A. (2008) Improvement of olive oil phenolics content y mens of enzyme formultion: Effect of different enzyme ctivities nd levels. Biochemicl Engineering Journl , [31]. Piv-M F, Rodrigues, V., Clheiros, R., Mrques M.P.J. (2011) Chrcteriztion of ntioxidnt olive oil iophenols y spectroscopic methods Sci Food Agric. (2): [32]. Rfehi, H., Ververis, K., Krginnis, T. C. (2012) Mechnisms of ction of phenolic compounds in olive. J. Diet Suppl June 9(2): [33]. Nvrro, F., Arroyo, A., Mrtin, S.F., Bello, RI, De Co, R., Burgess, S.J.R., Nvs, P., Villl, J.M. (1999) Protective role of uiquinone in vitmin E nd selenium-deficient plsm memrnes. Bio Fctors; 9: [34]. Pvlovic, S.Z., Ognjnovic, B.I., Tjn, A., Ikic, R.V., Sicic, Z.S., Petrovic, V.M. (2001) The effect of coenzyme Q10 on lood scoric cid, vitmin E, nd lipid peroxide in chronic cdmium intoxiction. J Environ Pthol Toxicol Oncol., 20: ,. [35]. Irhim, W. H., Blgvn, H.N., Chopr, R.K., Chow, C. K. (2000) Dietry coenzyme Q10 nd vitmin E lter the sttus of these compounds in rt tissues nd mitochondri. J Nutr, :