doi:10.5455/vetworld.2013.534-537 Serum

Background: Zinc is an essential co-factor for many enzymatic activities and its deficiency may produce clinical signs of parakeratosis, loss and failure of growth of wool and hair in sheep. 
 
Aim: The present study was designed to evaluate the effect of zinc oxide nano-particles (instead conventional zinc oxide) on serum biochemical factors and histopathological changes in liver and kidney of lambs. 
 
Materials and Methods: One group of lamb (4 lambs each group) received zinc oxide nano-particles suspension daily for 25 day. Whole blood sample and serum were collected at the start and the end of study. Activities of lactate dehydrogenase (LDH), alanine aminotransferas (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), blood urea nitrogen (BUN), and creatinine (CR) were measured in serum of lambs. Sections of liver and kidney were stained with hematoxylin-eosin and examined by light microscopy. 
 
Results: Activity of ALP (p=0.011) was significantly decreased and creatinine level (p=0.002) was significantly increased by zinc oxide nano-particles. Cell swelling, eosinophilic necrosis of hepatocytes, and multifocal interstitial nephritis was observed during histopatological examination. 
 
Conclusion: Results of study suggest that zinc oxide nano-particles may be toxic for use in sheep in zinc deficiency status.


Introduction
of the dermis have also been reported [5].However, excessive Zn may reduce the absorption of calcium or In the recent years the application of nanophosphorus.Zinc deficiency occurs in cattle, sheep, technology in human and veterinary medicine has and goats.The disease in sheep is characterized by loss shown a great progress.Also, nanotechnology has of wool and development of thick and wrinkled skin.revolutionized the commercial application of products Wool eating also occurs in sheep and may be one of the in the fields of medicine, engineering, manufacturing, earliest signs noticed in lambs [6].However, Zinc and information and environmental technology [1,2].
causes a high incidence of abortions and stillbirths in Zinc oxide (ZnO) nanoparticles are used for industrial the ewes [7].Zinc toxicity in sheep induces and cosmetic purposes.Zinc is an integral component pathological changes in the pancreas, kidney, liver, of a wide range of metalloenzymes and acts as a cofactor rumen, abomasum, small intestine, and adrenal gland for RNA and DNA polymerases.It is of particular [8].The potential hazard of high concentration of importance in rapidly-dividing cells, including those of manufactured nanoscale zinc powder is still unknown the epidermis [3,4].Zinc is also essential for the and their toxicological data are rather sparse, although biosynthesis of fatty acids and participates in both the the toxicity of normal zinc and zinc compounds has inflammatory and immune systems.It is also involved been much reported [9][10][11][12]. in the metabolism of vitamin A [3,4].Zinc deficiency The aim of this study was to evaluate the oral results in failure of keratinization, which leads to toxicity of nanoscale Zn oxide in lambs.Additionally, parakeratosis, loss and failure of growth of wool and the effects of these particles on the serum biochemical hair, lesions of coronary bands, retarded testicular level and tissues were also investigated.development and cessation of spermato-genesis in zinc deficient animals [5].The lesions of the arteriolar walls used for statistical analysis.Groups' variance was analyzed by one-way Analysis of Variation (ANOVA) The administered particles were suspended in normal and Fisher least significant difference test (LSD) was saline and dispersed by vibration for 5 min.The tested for significant differences between groups.A concentration of Zn in the suspension was 200 mg/ml.significant difference was considered to be p< 0.05.In order to avoid the aggregation of the particles, a few glass beads were added in the suspension and then

Results and Discussion
stirred on vortex agitator before every use.
There was no mortality in nanoparticle group.Animals: The experiment was began in early July 2011 The amount of serum zinc and the effect of oral and finished in September 2011; using eight male administration of zinc nanoparticles on the serum lambs (aged 5 -6 months and weighed 18-20 kg).The biochemical levels of lambs are in Table -1

. animals were purchased from Bervayeh village
When zinc level results were evaluated it was observed (Ahvaz, Iran).The animals were housed in clean room that, before and after oral exposure to zinc and maintained at 32±35°C, 30-45% relative humidity nanoparticles showed an increase zinc level but this and 12-h light/dark cycle.The lambs were allowed free sharp elevation was not significant (p<0.05).Table-1 access to diet and water.After one week acclimation, shows The mean concentration of Zn in serum at the the suspension of zinc nanoparticle was oral end of the experiment was 2.02 ìmol/l in nano zinc administration to lambs by dose 20mg/kg body weight.
group whereas before nano zinc supplement the mean This group received zinc nanoparticles suspension values were around 0.79 ìmol/l .daily for 25 day.Blood was collected from vein before The results indicated that activity of creatinine and in the end of study and serum was separated by was significantly increased compared with the before centrifuge.These samples were stored at -20 C until treatment (P= 0.002).The level of the serum ALP was analysis.The biochemical levels including lactate significantly decreased after administration of zinc dehydrogenase (LDH), alanine aminotransferas nanoparticles (p=0.011).The level of other factors did (ALT), aspartate aminotransferase (AST), alkaline not change significantly (Table -

1). phosphatase (ALP), blood urea nitrogen (BUN),
The histopathological pictures of liver are illustrated creatinine (CR), were assayed by an automatic in Figure -1.At the microscopic level, the liver of lambs biochemical analyzer (Cobas Mira Plus ,Roch nano zinc group presented the reversible histopatho-Diagnsotics, Germany).Level of zinc in serum of logical changes as cell swelling (about 50% of lambs lambs was determined at first day and 25 days by was seen) and irreversible histopathological changes as Atomic Absorbtion Spectrometry (Varian AA-240; eosinophilic necrosis of hepatocytes (about 50% of Varian Corp, California, USA).lambs was seen).Figure -2 shows histopathological The animals were sacrificed on 25th day.The findings of kidney control and nano zinc group.Significant liver and kidneys were collected, while all tissues were histopathological alteration of kidney tissues nano zinc fixed in 10% formalin-buffer for histopatho-logical group was multifocal interstitial nephritis (about 75% examination.Then, tissues embedded into paraffin, of lambs was seen), but remainder of kidney nano zinc sectioned for 5 ìm thick, and mounted on the glass group was normal.microscope slides using standard histopatho-logical Serum level of zinc was increased by oral adminitechniques.The sections were stained with hematoxylinstration of nano zinc particles in lambs in our study but eosin and examined by light microscopy.Our study suggests that the oral administration of widely used in monitoring the response to the nano zinc may cause toxic effects on the liver and exogenous toxic exposure.In ruminants AST is often kidney in lambs.tested along with LDH and ALP to evaluate whether the liver is damaged or diseased.When the liver is in Authors' contribution dysfunction, the levels of the above enzymes will rise.
All authors contributed equally.All authors read ALT is considered to be liver specific in cat and dog but and approved the final manuscript.there is little hepatic ALT activity in sheep.Clearly shown the value of serum ALT as an indicator of

Figure- 1 .
Figure-1.Liver tissue from lambs control group, lambs exposed to nano zinc at dose of 20mg/kg body weight on 25 days oral administration.(A) Liver in the control group, showing normal structure (10x).(B) Liver in the nano zinc group arrow shows eosinophilic necrosis of hepatocytes (10x).(C) Liver in the nano zinc group arrows show eosinophilic cells (40x).(D) Liver in the nano zinc group arrow shows cell swelling hepatocytes (40x).(H&E).

Figure- 2 .
Figure-2.Histopathological findings of kidney of lambs exposed to nano zinc at dose of 20mg/kg body weight on 25days oral administration.(A) Kidney of the control group.There are no significant lesions (10x).(B) Kidney of the nano zinc group arrows show multifocal interstitial nephritis (infiltration of inflammatory cells) (10x).(C) Kidney of the nano zinc group arrow shows glomerular fibrosis (40x).(H&E).