Glol Journl of Phrmology 8 (3): 284-293, 2014 ISSN 1992-0075 IDOSI Pulitions, 2014 DOI: 10.5829/iosi.gjp.2014.8.3.83176 Moultion of Gentmiin-inue Testiulr n Brin Dmge in Rts Zeyn Kh. El-Mwy Deprtment of Phrmology, Fulty of Veterinry Meiine, Alexnri University, Egypt Astrt: The effets of e minoglyosie ntiioti gentmiin, on loo, reproutive system n rin tissue of rts n e onurrent ministrtion of L-rnitine wi e rug were stuie. Rts were intrmusulrly injete wi gentmiin t ose of 20 or 80 mg/kg.wt. i.m., ivie into ree equl oses ily for 10 ys n kille t e 11 or 57 y of experiment. Gentmiin inue signifint reution in inex weight of testes, epiiymis, essory sex orgns, sperm ell onentrtion, motility, live sperms perentges n testosterone hormone level. While testiulr totl holesterol n sperm normlities were inrese. The rug lso use signifint erese in e tivity of testiulr n rin tlse n reue glutione (GSH) ontent. Also e level of lipi peroxition prout mlonilehye (MDA) showe signifint inrese. Some histopologil hnges in testiulr n rin tissues in ition to verse effets on hemtologil prmeters were lso reore. The intensity of ese verse effets inrese signifintly wi e high ose of e rug. The suutneous injetion of L-rnitine (200 mg/kg.wt.) one/y for 56 suessive ys improve signifintly e previously mentione prmeters ompre to gentmiin trete groups in o perios of experiment. Moreover, ll stuie prmeters were restore to norml vlues in L-rnitine wi gentmiin (20 mg/kg.wt.) trete group t e 57 y of experiment. In onlusion, injetion of erpeuti or high ose of gentmiin inue verse effets on reproutive system, loo n rin tissue of mle rts. Interestingly ministrtion of L-rnitine wi gentmiin meliortes ese verse effets y improving ntioxint sttus n reuing lipi peroxition. Key wors: Gentmiin Testes Brin Oxitive Stress L-rnitine Antioxint sttus INTRODUCTION tissues [1].Gentmiin oes not pper to e metolize n exrete lmost entirely y renl glomerulr filtrtion Gentmiin is n minoglyosie ntiioti, it ts y n exrete unhnge in e urine. A mjor inhiiting protein synesis in suseptile teri omplition of gentmiin tretment is nephrotoxiity resulting in ell e (teriil). It hs rpi onset of even wi e erpeuti oses [3]. Gentmiin inue tion, low rte of true resistne [1], high ntiteril nephrotoxiity oul e ue to e rug genertion of effiy n is ommonly use for e tretment of severl retive oxygen speies (ROS) [4-7]. pogeni Grm-negtive teril infetions. It is lso ROS estroy e ell omponents (lipis, proteins tive ginst some Grm-positive orgnisms, e.g. n DNA). Peroxition of memrne lipis uring Stphyloous (inluing meiillin n peniillin oxitive stress inues e frgmenttion of resistnt strins). In vitro gentmiin is tive ginst polyunsturte ftty is n relese of vrious Slmonell n Shigell [2]. lehyes n lkenes [7]. Gentmiin is rpily sore fter i.m. injetion L-rnitine (4Ntrimeylmmonium3 hyroxyutyri n pek serum levels re usully hieve wiin 30 to i), L-lysine erivtive, is nturlly ourring 90 minutes n re mesurle for 6-8 hours. It is wiely mino i-like ompoun n it is n enogenous istriute into oy flui n lso n e etete in mitohonril memrne ompoun [8]. L-rnitine plys Corresponing Auor: Zeyn Khmis EL-Mwy, Deprtment of Phrmology. Fulty of Veterinry Meiine Alexnri University, Efin-Rshi-Beher, P.O. Box: 22758, Egypt. Tel: +20-1007284201, Fx: +20-452960450. 284
Glol J. Phrmol., 8 (3): 284-293, 2014 n importnt role for filitting or trnsport of long hin ftty is from ytosol to mitohonri in orer to enter e -oxition yle [9-11]. Crnitine is lso ssoite wi uffering of exess etyl-co A whih is potentilly toxi to e ells [9]. It is minly use s n effetive nutritionl supplement n lso for ntioxint tivity. There were few stuies investigting e role of gentmiin in reproutive system n rin tissue so is work ws esigne to evlute some phrmoynmi effets of gentmiin on mle lino rts, rough stuying e effets of rug on semen hrters, some hemtologil, iohemil, hormonl n histopologil hnges. Also is stuy extene to investigte e protetive effets n possile mehnism of tion of L-rnitine ginst gentmiin inue verse effets. MATERIALS AND METHODS Gentmiin ws otine from Memphis Compny, Egypt uner uority of Shering-plough orportion/usa. L-rnitine provie y Ar Compny for Phrmeutils n Meiinl Plnts (Mepo- Meifoo), Egypt. The ignosti kits etermining vlues of H, totl holesterol, tlse level, GSH n MDA ontents were otine from Bio-ignosti Compny, Egypt. Oer hemils purhse from EL-Gomhori Compny, Egypt. Animls n Experimentl Design: Seventy two Alino mle rts of 180-200 ys ol n weighing 190-220 g were otine from e Meil Reserh Institute of Alexnri University, Egypt. Rts reeive lne rtion, wter n humne re in ompline wi e guielines of e Ntionl Institutes of Hel (NIH) of Animl Cre n e lol ommittee pprove is stuy. Rts were limtize 2 weeks prior to e experiments. Rts were rnomly ssigne into 6 equl groups (12 rts / group) Group 1: Rts reeive sline (6 ml/ kg.wt. i.m.) ivie into ree equl oses ily for 10 ys togeer wi single s/ injetion of sline (2 ml/ kg.wt.) ily for 56 suessive ys (ontrol group). Group 2: Rts reeive L-rnitine t ose of 200 mg/kg.wt. s.. in 2 ml/ kg.wt. sline [12] one/ y for suessive 56 ys. Group 3: Rts reeive gentmiin t ose of 20mg/kg.wt. i.m. [13] ivie into ree equl ily oses for 10 ys. Group 4: Rts reeive gentmiin t ose of 80 mg/kg.wt. i.m. oring to e moifie meo of Kopple et l. [12] ivie into ree equl ily oses for 10 ys. Group 5: Rts reeive gentmiin t ose of 20 mg/kg.wt. i.m. ivie into ree equl oses, ily for 10 ys togeer wi L-rnitine t ose of 200 mg/kg.wt. s.. one/ y for 56 suessive ys. Group 6: Rts reeive gentmiin t ose of 80 mg/kg.wt. i.m. ivie into ree equl oses ily for 10 ys togeer wi L-rnitine t ose of 200 mg/kg.wt. s.. one/ y for 56 suessive ys The urtion of e present stuy ws 56 ys to over e spermtogeni yle in rts whih rnges from 48-56 ys [14]. Six rts from eh trete n ontrol groups were kille t e 11 y from rugs n sline ministrtion to follow up some phrmoynmi effets of e rugs on mle rts. At e 57 y of experiment ll rts were kille. All experimentl rts were eprive of foo overnight n iniviully weighe t e speifi killing time. Bloo, oy orgns n epiiyml ontents were otine from trete n ontrol rts. Bloo Smples: Fsting loo smples were ollete vi retro oritl plexus leeing t speifi time for killing. From eh rt 2 loo smples were ollete one on EDTA for hemtologil stuies n e oer ws left to lot en entrifuge t 3000 rpm for 15 min to otin serum whih store t -20 C for hormonl ssy. Weight of Internl Boy Orgns: After olletion of e loo smples, e nimls were kille, immeitely srifie y ervil epittion. The testes, epiiymis, prostte n seminl vesile glns were issete out, grossly exmine n weighe. The inex weight (I.W.) of eh orgn ws lulte oring to Mtousek [15] where, I.W. = orgn weight (g) / oy weight (g) x 100. Exmintion of Epiiyml Sperm: Epiiyml Sperm Count: Epiiyml spermtozo were ounte y moifie meo of Yokoi et l. [16]. The epiiymis ws mine in 5 ml sline, ple in roker for 10 min n inute t room temperture for 2min. The superntnt ws ilute 1: 100 in solution 285
Glol J. Phrmol., 8 (3): 284-293, 2014 ontining 5 g NHCo 3 (for reking up e muous prout oring to Ohkw et l. [23]. Ctlse tivity roplets), 1 ml formlin 35% n 25 mg eosin per100 ml ws estimte se on e retion of e enzyme wi istille wter. Approximtely, 10 µl of e ilute semen known quntity of H2O2 oring to Aei [24]. ws trnsferre to eh ounting hmer of n improve Neuur hemoytometer (Dep 0.1 mm; LABART, Histologil Exmintion: One testis from eh rt were Munih, Germny) n ws llowe to stn for 5 min remove, wshe wi physiologil sline, lotte on efore ounting uner light mirosope t high power filter pper, rpily fixe in moifie Dvison s flui for lens. 48 h n store in 70%lohol until furer proessing [25]. The remining prts of rins speimens were Epiiyml Sperm Motility, Anormlities n Live- ollete n fixe in 10% neutrl uffere formlin sperm Perentges: A rop of freshly unilute semen solution. Five miron ik prffin setions were from e u epiiymis ws mixe wi one rop of prepre from o orgns en stine wi hemtoxylin physiologil sline on e slie. The progressively motile n eosin (HE) n lter exmine histopologilly sperm perentge ws evlute mirosopilly uner [26]. high power lens [17]. The perentges of live sperm n morphologilly norml spermtozo were etermine Sttistil Anlysis: Results were sttistilly nlyze fter stining on e sme slie wi eosin nigrosin stin. y ANOVA followe y Dunn s multiple rnge test. A totl of 300 sperm were ounte on eh slie uner Dt re presente s mens plus or minus e stnr high power lens of e light mirosope n perentges error. The minimum level of signifine ws set t p 0.05 of norml sperm n live ones were reore oring [27]. to Beren n Fuquy [17]. RESULTS Hemtologil Stuies: Hemogloin onentrtion ws etermine y e meo esrie y Drkin n Orgns Weights: e otine t revele signifint Austin [18] using ommerilly ville ignosti kits. erese (P 0.05) in inex weight vlues of testes, Pke ell volume perentge ws etermine y epiiymis n essory sex orgns in gentmiin t mirohemtorite tehnique oring to Die n Lewis ifferent oses n perios of experiment n L-rnitine [19]. Eryroyti n totl leukoyti ounts were + gentmiin (20 mg/kg t 11 y or 80mg/kg.wt. t estimte using Doule improve Neuuer o perios) trete groups ompre to ontrol. hemoytometer [19]. The reution ws more evient in gentmiin (80mg/kg.wt.) n less evient in L-rnitine wi gentmiin Hormonl Assy: Serum testosterone ws etermine (20mg/kg.wt. t 11 y) trete groups (Tle 2). using n enzyme immunossy kit (Immunometris Lt., However, L-rnitine ompletely restore e inex Lonon, UK) oring to Demetrious [20]. weight vlues of reproutive orgns in gentmiin (20 mg/kg.wt.) trete group t 57 y ompre to Totl Cholesterol Assy: Testiulr totl holesterol ws ontrol (Tle 1). extrte wi etone-lohol (1:1). Totl holesterol is etermine fter enzymti hyrolysis n oxition. Seminl, Hormonl n Testiulr Totl Cholesterol The initor quinoneimine is forme from hyrogen Assys: There ws signifint (P 0.05) erese in peroxie n 4-minophenzone in e presene of phenol sperm ell onentrtion, motility n live perentges, n peroxise oring to Allin et l. [21]. testosterone hormone level n signifint elevtion in sperm normlities n testiulr totl holesterol vlues Antioxint Sttus n Oxitive Stress Assys: One in rts trete wi gentmiin t ifferent oses n testis n prt of rin of eh rt were kept frozen t 70 perios of experiment n L-rnitine + gentmiin (20 o C for ssessment of GSH n LPO ontents n tlse mg/kg t 11 y or 80mg/kg.wt. t o perios) tivity. GSH level ws mesure se on e reution ompre to ontrol (Tles 2&3). The ministrtion of of 5,5 iiois (2-nitroenzoi i) (DTNB) wi L-rnitine wi gentmiin improve e previously glutione oring to e meo of Gltzle et l. [22]. mentione vlues exept sperm ell onentrtion vlue LPO ws represente s MDA whih ws mesure fter wi gentmiin ose (80mg/kg t 11 y) ompre to e retion wi iorituri i in ii meium t gentmiin lone trete groups. Moreover, Tle 2&3 95 C for 30 minutes to form iorituri i retive showe t, ll e ove mentione prmeters were 286
Glol J. Phrmol., 8 (3): 284-293, 2014 Tle 1: Effet of gentmiin n / or L-rnitine on inex weight of testes, epiiymis n essory sex orgns of rts Inex weight of Inex weight of Inex weight of essory testes (g/100g.wt.) epiiymis (g/100g.wt.) sex orgns (g/100g.wt.) Prmeter -------------------------------------- --------------------------------------- ----------------------------------------- Group At 11 y At 57 y At 11 y At 57 y At 11 y At 57 y Control 1.74±0.02 1.74±0.01 0.81±0.01 0.79±0.02 1.15±0.01 1.19±0.01 L-rnitine 1.75±0.01 1.73±0.01 0.81±0.01 0.80±0.01 1.18±0.01 1.17±0.01 Gentmiin (20 mg /kg.wt.) 1.51±0.02 1.50±0.02 0.56±.01 0.57±0.01 0.94±0.01 0.89±0.01 e e Gentmiin (80mg/kg.wt.) 1.12±0.01 1.08±0.01 0.45±0.02 0.46±0.02 0.46±0.01 0.51±0.02 Gentmiin (20 mg/kg.wt.) + L-rnitine 1.42±0.01 1.72±0.01 0.64±0.01 0.76±0.02 1.11±0.02 1.17±0.01 Gentmiin (80mg/kg.wt.) + L-rnitine 1.21±0.02 1.30±0.01 0.52±0.01 0.59±0.01 0.64±0.02 0.80±0.02 Vlues re expresse s men ± S.E. N= 6. Vlues wi ifferent letters t e sme olumn re signifintly ifferent t P 0.05. Tle 2: Effet of gentmiin n / or L-rnitine on sperm ell onentrtion, motility n normlities of rts. 6 Sperm ell onentrtion (x10 /ml) Sperm motility (%) Sperm normlities(%) Prmeter ----------------------------------------- ---------------------------------------- -------------------------------------- Group At 11 y At 57 y At 11 y At57 y At 11 y At 57 y Control 430.00±7.75 442.3±6.09 91.67±1.05 90.0±1.83 8.89±0.83 9.89±0.83 L-rnitine 437.50±8.54 440.0±7.30 90.83±1.54 90.8±1.54 9.8±0.54 9.33±0.54 Gentmiin (20 mg /kg.wt.) 365.17±4.48 356.3±6.10 55.00±1.83 59.2±2.39 18.2±0.79 17.2±0.89 Gentmiin (80mg/kg.wt.) 325.33±5.00 315.0±5.32 35.83±2.39 35.8±2.39 35.2±1.39 32.2±1.39 Gentmiin (20 mg/kg.wt.) + L-rnitine 391.67±2.79 436.3±7.78 63.33±1.67 85.0±1.83 15.0±0.83 8.5±0.83 Gentmiin (80mg/kg.wt.) + L-rnitine 340.83±2.39 371.3±5.53 50.83±1.54 65.8±1.54 25.8±1.23 23.8±1.46 Vlues re expresse s men ± S.E. N= 6. Vlues wi ifferent letters t e sme olumn re signifintly ifferent t P 0.05. Tle 3: Effet of gentmiin n / or L-rnitine on sperm live, testosterone n totl holesterol vlues of rts. Testiulr totl holesterol Sperm live (%) Testosterone (ng/ml) (mg/g wet tissue) Prmeter ---------------------------------------- ---------------------------------------- -------------------------------------- Group At 11 y At 57 y At 11 y At 57 y At 11 y At 57 y e Control 92.17±0.98 93.2±1.19 2.29±0.01 2.30±0.01 5.55±0.51 6.38±0.45 e L-rnitine 93.00±0.37 93.3±1.26 2.28±0.01 2.30±0.01 5.00±0.70 6.10±0.42 Gentmiin (20 mg /kg.wt.) 70.33±1.15 69.5±2.49 2.15±0.01 2.17±0.011 15.20±0.71 12.5±0.88 e Gentmiin (80mg/kg.wt.) 55.17±2.98 58.7±3.99 1.04±0.01 1.10±0.01 33.55±1.88 30.6±1.12 Gentmiin (20 mg/kg.wt.) + L-rnitine 81.50±0.99 91.2±1.08 2.21±0.01 2.29±0.03 10.08±0.68 6.18±0.4 Gentmiin (80mg/kg.wt.) + L-rnitine 65.33±2.39 74.2±1.70 1.13±0.01 1.18±0.02 23.17±1.66 18.6±1.09 Vlues re expresse s men ± S.E. N= 6. Vlues wi ifferent letters t e sme olumn re signifintly ifferent t P 0.05. Tle 4: Effet of gentmiin n / or L-rnitine on testiulr lipi peroxition (MDA) level, reue glutione (GSH) ontent n tlse tivity of rts. Testiulr LPO Testiulr GSH Testiulr tlse (nmol MDA/g wet tissue) (umol/g wet tissue) (unit/g wet tissue) Prmeter ---------------------------------------- ---------------------------------------- -------------------------------------- Group At 11 y At 57 y At 11 y At 57 y At 11 y At 57 y e Control 4.00±0.08 4.05±0.08 10.12±0.03 10.3±0.06 6.20±0.18 5.95±0.17 e L-rnitine 3.95±0.05 3.97±0.09 11.92±0.09 12.2±0.13 6.38±0.14 6.13±0.10 Gentmiin (20 mg /kg.wt.) 6.10±0.09 6.13±0.16 8.23±0.12 8.32±0.14 4.22±0.10 4.19±0.06 f e Gentmiin (80mg/kg.wt.) 10.27±0.18 10.3±0.15 5.23±0.10 5.38±0.1 2.38±0.12 2.33±0.11 Gentmiin (20 mg/kg.wt.) + L-rnitine 4.73±0.12 3.83±0.12 9.40±0.16 10.6±0.09 5.10±0.09 6.03±0.04 e Gentmiin (80mg/kg.wt.) + L-rnitine 8.15±0.11 6.84±0.13 7.60±0.07 8.52±0.07 3.40±0.14 4.35±0.10 Vlues re expresse s men ± S.E. N= 6. Vlues wi ifferent letters t e sme olumn re signifintly ifferent t P 0.05. 287
Glol J. Phrmol., 8 (3): 284-293, 2014 Tle 5: Effet of gentmiin n / or L-rnitine on rin lipi peroxition (MDA) level, reue glutione (GSH) ontent n tlse tivity of rts. Brin LPO Brin GSH Brin tlse (nmol MDA/g wet tissue) (umol/g wet tissue) (unit/g wet tissue) Prmeter --------------------------------------- ---------------------------------------- ------------------------------------- Group At 11 y At 57 y At 11 y At 57 y At 11 y At 57 y Control 13.50±0.28 13.1±0.31 4.76±0.11 4.96±0.19 3.03±0.04 3.31±0.1 L-rnitine 13.20±0.23 13.1±0.27 5.21±0.14 5.92±0.07 3.08±0.08 3.37±0.15 Gentmiin (20 mg /kg.wt.) 17.88±0.81 18.9±0.51 e 3.16±0.07 3.15±0.06 1.93±0.05 1.96±0.03 Gentmiin (80mg/kg.wt.) 40.67±1.83 34.8±1.15 f 2.12±0.06 2.09±0.13 1.29±0.08 1.36±0.1 Gentmiin (20 mg/kg.wt.) + L-rnitine 15.40±0.37 12.6±0.22 4.07±0.06 5.83±0.12 2.43±0.06 3.42±0.19 Gentmiin (80mg/kg.wt.) + L-rnitine 26.33±0.76 18.8±0.52 3.56±0.10 3.25±0.33 1.93±0.06 2.11±0.05 Vlues re expresse s men ± S.E. N= 6. Vlues wi ifferent letters t e sme olumn re signifintly ifferent t P 0.05. Tle 6: Effet of gentmiin n / or L-rnitine on hemogloin (H g%), pke ell volume (PCV %), re loo orpusles (RBCs) n white loo ells (WBCs ) ounts of rts. H (g%) PCV(%) 6 RBCs (x10 mm) 3 WBCs (x10 mm) Prmeter ---------------------------------- ------------------------------- ------------------------------- ------------------------------- Group At 11 y At 57 y At 11 y At 57 y At 11 y At 57 y At 11 y At 57 y Control 13.00±0.12 13.1±0.13 43.73±0.34 43.5±0.33 7.02±0.11 7.00±0.09 9.93±0.08 10.5±0.33 L-rnitine 12.80±0.12 13.0±0.12 44.03±0.31 44.0±0.31 7.03±0.10 7.02±0.07 9.98±0.09 10.6±0.30 Gentmiin (20 mg /kg.wt.) 10.53±0.27 10.6±0.08 40.27±0.53 39.9±0.63 6.30±0.13 6.42±0.09 9.02±0.07 9.35±0.17 Gentmiin (80mg/kg.wt.) 9.10±0.28 9.57±0.23 35.20±0.29 36.5±0.43 5.87±0.11 5.82±0.2 8.83±0.08 9.09±0.39 Gentmiin (20 mg/kg.wt.) + L-rnitine 10.87±0.34 12.8±0.09 41.80±0.36 43.8±0.21 6.72±0.14 7.02±0.13 9.20±0.09 10.30±0.9 Gentmiin (80mg/kg.wt.) + L-rnitine 9.83±0.27 10.3±0.11 39.70±0.67 39.4±0.39 6.50±0.15 6.40±0.08 9.10±0.05 9.35±0.10 Vlues re expresse s men ± S.E. N= 6. Vlues wi ifferent letters t e sme olumn re signifintly ifferent t P 0.05. ompletely restore to norml vlues in L-rnitine wi (20mg/kg t 57 y) trete groups ompre to ontrol gentmiin(20 mg/kg.wt.) trete group t e 57 y of one. While ere ws signifint reution (P 0.05) in experiment. eir vlues in oer trete groups ompre to ontrol. The reution ws more evient in gentmiin (80mg/kg Antioxint Sttus n Oxitive Stress Assys:.wt.) n less evient in L-rnitine wi gentmiin Gentmiin t ifferent oses n perios of experiment (20mg/kg.wt. t 11 y) trete groups (Tle 6). n L-rnitine + Gentmiin (20 mg/kg t 11 y or 80mg/kg.wt.. t o perios) trete groups use Histopologil Finings signifint (P 0.05) inrese in testiulr & rin LPO Testes: Norml testiulr rhiteture, well-orgnize represente y MDA level n erese in GSH seminiferous tuules wi omplete spermtogenesis n ontent n tlse tivity in e sme tissues norml interstitil onnetive tissue oserve in o ompre to ontrol group (Tles 4&5).However, ontrol nimls n rts trete wi L-rnitine eier t ministrtion of L-rnitine lone inue signifint 11 or 57 y of experiment. (P 0.05) inrese in testiulr & rin GSH ontent Testiulr tissue of rts trete wi gentmiin ompre to ontrol. (20mg/kg)lone t 11 or 57 y of experiment showe Moreover, ministrtion of L-rnitine + gentmiin ongestion of e interstitil loo vessels.(fig.1a), reue e intensity of inrese LPO use y e esies egenertive hnges of some of e rug n prtilly restore ntioxint sttus, even e seminiferous tuules s inomplete spermtogenesis MDA level, GSH ontent n tlse tivity were whih represente y lk of e epielil multilyers on ompletely restore to norml vlues in gentmiin e sement memrne of e seminiferous tuules. (20 mg/kg.wt.) trete group t e 57 y of Testiulr tissue of rts trete wi gentmiin experiment (Tles 4&5). (80mg/kg) t o perios of experiment showe eem of e interstitil loo vessels. (Fig.1B), signifint Hemtologil Prmeters: The otine results showe egenertion of e seminiferous tuules represente y t, ere ws non signifint effet (P 0.05) on H sever esqumtion of germinl epielium wi nerosis g%, PCV%, RBCs n WBCs ounts in L-rnitine t o of spermtoytes, spermtis, moreover ere ws perios of experiment n L-rnitine + gentmiin hyliniztion of e luminl ontents (Fig.1C). 288
Glol J. Phrmol., 8 (3): 284-293, 2014 Fig. 1: Photomirogrph of rt testes n rin stine wi HE(x 200): (A) Testiulr tissue of rt trete wi gentmiin (20mg/kg) lone t 11 y of experiment showe ongestion of e interstitil loo vessels(rrow). (B)Testiulr tissue of rt trete wi gentmiin (80mg/kg) lone t11 y showe eem of e interstitil loo vessels (strs).(c)testis of rt trete wi gentmiin (80mg/kg) lone t 57 y showe hyliniztion of e luminl ontents (rrows). ( D) Testis of rt trete wi L-rnitine +gentmiin (80mg/kg) t 57 y showe mil esqumtion of germinl epielium (rrow) wi mrke improvement of spermtogenesis.(e) Brin of rt trete wi gentmiin (80mg/kg) t 11 y showe hemorrhge in Purkinje ell lyer of e ereellum(rrows).(f).brin of rt trete wi L-rnitine +gentmiin (80mg/kg)t 57 y showe mil eem in Purkinje ell lyer of e ereellum. In nimls trete wi L-rnitine +gentmiin eem in Purkinje ell lyer of e ereellum (Fig.1F) in (20mg/kg) ere ws protetive effet on e testiulr some ses. tissue where ongestion of e interstitil loo vessels In nimls trete wi L-rnitine +gentmiin ws sent wi mil ellulr egenertion of e tuulr (20mg/kg) ere ws omplete protetive effet in e germinl epielium t11 of experiment. Moreover, rin tissue (similr to ose of ontrol) t o perios of reltively norml testes wi lw ensity of spermtozo experiment. in whih struturlly n funtionlly tive seminiferous There ws protetive effet of L-rnitine in e tuules wi presene of elongte spemtis n group trete wi gentmiin (80mg/kg) t o perios spermtozo (wi lw ensity) in e mjority of e in e rin tissue s it h e norml histologil tuules were etete t 57 of experiment. pperne wi ongestion of some meningel loo In nimls trete wi L-rnitine +gentmiin vessels in few ses. (80mg/kg) ere ws protetive effet in testiulr tissue strt t 11 n eome more pronoune t 57 y DISCUSSION eviene y mil esqumtion of germinl epielium wi mrke improvement of spermtogenesis(fig.1d). Oxitive stress is onsiere one of e importnt uses of vrious iseses n orgn toxiities. Brin: Mirosopilly, e rin showe slight Aministrtion of gentmiin t o oses (20 or 80 ongestion in loo vessels of ererum gry mtter s mg/kg.wt.) for o perios (11&57 ys of experiment) well s in meninges in rts trete wi gentmiin inue some fertility normlities ppere s (20mg/kg) t 11 or 57 y of experiment. Moreover in signifint reution in testes, epiiymis n essory rts trete wi gentmiin (80mg/kg)t o perios of sex glns weights, testosterone hormone level, sperm experiment e rin showe hemorrhge (Fig.1E) n mil ell onentrtion, motility n livility perentges. 289
Glol J. Phrmol., 8 (3): 284-293, 2014 Moreover, ere ws signifint inrese in totl provie signifint enefiil effet on gentmiin sperm normlities % n testiulr totl holesterol. inue testiulr oxitive stress. L-rnitine, onsiere These finings go sie y sie wi e histologil s n importnt exmple for ntioxints t hve een finings in testes. All ese fertility troules were more lrey proven s effetive gents in vrious moels o pronoune wi high ose of e rug. in vitro n in vivo. The reution of testosterone hormone n elevtion The results showe t ministrtion of L-rnitine of testiulr totl holesterol my e resulte from e wi gentmiin tretment eier 20 or 80mg/kg.wt. t e effet of gentmiin on e steroiogeni enzymes, 11 or 57 y of experiment improve e reproutive using n ltertion in e formtion of testosterone n ltertion inue y e rug. It reue oxitive stress umultion of holesterol in e testes [28]. Also, e n improve ntioxint sttus n histologil struture hnges in testiulr lipi profile were strongly orrelte of testiulr tissue. Also L-rnitine inue signifint wi testiulr egenertion, histologil n iohemil erese in testiulr totl holesterol n testosterone ltertion [29]. The etiology my e srie to e ft hormone levels. Sperm ount n perentges of sperm t lipis, in prtiulr holesterol is e preursor of motility n viility were improve, Moreover, ll mle sex hormones n us inrese testiulr mesure prmeters were ompletely restore to norml holesterol my result from impire utiliztion in vlues in group trete wi gentmiin (20mg/kg.wt.) t steroiogenesis [30], ssoite wi impire testiulr 57 y of experiment. Histopologil finings in e tivity. groups trete wi L-rnitine lso onfirme e Khki et l. [31] suggeste t e nerosis of e previous improvement in testiulr tissues. interstitil ells my e resulte in erese synesis of It hs een foun t L-rnitine improve testosterone hormone. This stuy showe n inrese in ntioxint sttus in rts n inrese free ril testiulr MDA levels, reution in ntioxint reserves svenging from e ellulr sites [36,37].L-rnitine oul GTH n tlse in gentmiin trete groups in ose protet HK-2 ells from H2O2-inue injury vi inhiition epennt t ifferent perios s ompre to e ontrol of oxitive mge, ell poptosis n mitohonri group. It inites oxitive stress n its implitions on ysfuntion [38]. testiulr tivity. Antioxints protet DNA n oer importnt Gentmiin inue reproutive normlities s moleules from oxition n mge, us improve sperm result of its iret toxi effets on testes inue oxitive qulity n onsequently inrese fertility rte [39,40]. stress [32]. Spermtozo hve high ontent of This stuy showe t n inrese in rin MDA polyunsturte ftty is in eir plsm memrne level, reution in ntioxint reserves GTH n tlse us ey re highly suseptile to mge y exessive togeer wi some histologil hnges in rin tissue in onentrtions of ROS. gentmiin (20 or 80 mg/kg.wt.) trete groups t The lipi peroxition mges e struture of lipi ifferent perios s ompre to e ontrol group. e mtrix in spermtozo memrnes le to loss of motility ltertions in rin tissue inrese wi high ose. n impirment of spermtogenesis [33]. Gentmiin Gentmiin ws foun to penetrte into e inhiits germ ells ivision n protein synesis in e ererospinl flui in hely ogs. The inrese of CSF testis. It lso, inue ell e in e seminl vesile onentrtion oul e expete fter repetitive [32].These results re omptile wi ose of Nryn gentmiin ministrtion. Also, meningitis inreses e [32] n Akoni et l. [34]. penetrtion of prenterlly ministere gentmiin into Agrwl n Sleh [35] reporte t exessive e CSF [41, 42]. genertion of free rils my ttk integrity of DNA in Derese of ntioxint enzyme my e ue to rpi e sperm nuleus, us elerte e proess of germ onsumption n exhustion of storge of is enzyme in ell poptosis, leing to eline in sperm ounts n fighting free rils generte uring e evelopment of normlities ssoite wi mle infertility. Gentmiin rin mge. inue inrese in intrellulr ROS ontent whih n The results of is investigtion showe t reh toxi level, us using ell e n ministrtion of L-rnitine wi gentmiin tretment mlfuntioning of e orgn. eier 20 or 80mg/kg.wt. t 11 or 57 y of experiment There were no reports s fr s known esriing e reue oxitive stress n improve e ntioxint protetive role of L-rnitine ginst gentmiin inue sttus n histologil hnges in rin tissue. ese testiulr oxitive mge. Inhiitors of oxitive stress results re in line wi previous stuies of Pllor et l. [43] 290
Glol J. Phrmol., 8 (3): 284-293, 2014 who emonstrte t L-rnitine inue protetive ACKNOWLEDGEMENT effets on e toxiity of rin injury in neontes relte to hypoxi n ishemi n uring ishemi-reperfusion The uor woul like to nk Dr. Hossm G. ses. Tohmy (leturer of Pology, Fulty of Gopl et l. [44] foun t -Meionine (-Met), Veterinry Meiine, Alexnri University), for his sulfur-ontining nuleophili ntioxint, hs een vlule help to e histopologil exmintion of is shown to prevent ispltin-inue entrl nervous work. system tissue mge from ute ispltin toxiity. L-rnitine reue e oxitive stress n REFERENCES rottion ehvior stimulte y quinolini i n 3-nitropropioni i eh lone or in omintion in rin 1. Chmers, H.F., 2008. Aminoglyosies. In: Goomn of rts, e protetive properties of L-rnitine in e n Gilmn s. The Phrmologil Bsis of neurotoxi moels teste re mostly meite y its Therputis (Brunton, L.L., J.S. Lz n K.L. Prker). hrteristis s n ntioxint gent [45]. 11 E. Meil Pulition Division, New The present investigtion showe t ere ws York, Chiho, Sn Frneso, Lison, Lonon, signifint erese in H g%, PCV%, RBCs n WBCs pp: 1165-1166. ounts in gentmiin (20or 80mg /kg.wt.) trete groups 2. Reiter, R.J., D. Tn, R.M. Sinz, J.C. Myo n t e 11 or 57 y of experiment, e reution ws more B.S. Lopez, 2002. Meltonin reuing e toxiity n pronoune wi high ose. These results oul e inresing e effiy of rugs. Journl of Phrmy ttriute to leukopeni, nemi, erese retiuloyte n Phrmology, 5: 1299-1321. ounts n romoytopeni use y e rug. 3. Sweileh, W.M., 2009. A prospetive omprtive Gentmiin ws shown to helte mitohonril iron stuy of gentmiin n mikin inue forming iron-gentmiin omplex whih is potent nephrotoxiity in ptients wi norml seline renl tlyst of free ril formtion ple of inhiiting funtion. Funmentl n Clinil Phrmology, mitohonril respirtion using ell e [46]. 23(4): 515-520. These results re omptile wi ose reore y 4. Amini, F.G., M. Rfiein-Kopei, M. Nemtkhsh, El Bwi et l. [47], Hsn et l. [48] in rts n mie, A. Brrn n H. Nsri, 2012. Ameliortive effets respetively. of metformin on renl histologi n iohemil Tretment wi L-rnitine improve ese ltertions of gentmiin-inue renl toxiity in hemtologil prmeters ue to its ntioxint Wistr rts. Journl of Reserh in Meil Sienes, prmeters t enhne hemtopoiesis. 17(7): 621-625. The present results etete t ministrtion of 5. Rfiein-Kopei M., H. Nsri, M. Nemtkhsh, L-rnitine ily inue signifint inrese in testiulr A. Brrn, A. Gheissri n H. Rouhi, 2012. n rin GTH t e 11 or 57 y of experiment s Eryropoietin meliortes genetmyin-inue ompre to ontrol group. So, L-rnitine oul offer renl toxiity: A iohemil n histopologil protetion y iretly inresing tissue ntioxint stuy. Journl of Nephropology, 1: 109-116. pities. 6. Stojiljkovi, N., M. Stoiljkovi, P. Rnjelovi, It oul e onlue t gentmiin t erpeuti S. Veljkovi n D. Mihilovi, 2012. ose or high ose inue testiulr n rin oxitive Cytoprotetive effet of vitmin C ginst stress in ition to verse effets on loo piture, e gentmiin-inue ute kiney injury in rts. severity of ese verse effets inrese wi high Experimentl n Toxiologi Pology, 64( ose. However, L-rnitine eliite signifint protetive 1-2): 69-74. effiy role ginst gentmiin-ssoite oxitive 7. Hlliwell, B. n S. Chirio, 1993. Lipi peroxition: stress owing to its ntioxint property. So, e present its mehnism, mesurement n signifine. stuy reommens t L-rnitine oul e inlue in Amerin Journl of Clinil Nutrition, 57: 715-725. presriptions long wi gentmiin to meliorte its 8. Kelly, G.S., 1998. L-rnitine: Therpeuti verse effets on hemtologil prmeters, rin tissue pplitions of onitionlly-essentil mino i. n mle fertility. Alterntive Meiine Review, 3: 345-360. 291
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