THE POTENTIAL PROTECTIVE EFFECT OF NATURAL HONEY AGAINST.pdf

8/11/2019 THE POTENTIAL PROTECTIVE EFFECT OF NATURAL HONEY AGAINST.pdf

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Vol. XV, No. 2, July 2007 Mansoura J. Forensic Med. Clin. Toxicol.

Abdel-Moneim & Ghafeer

ment that occurs naturally in ores togetherwith zinc, lead and copper or is emittedinto the air through the process of volcanic

INTRODUCTlON

Cadmium (Cd) is a relatively rare ele-

THE POTENTIAL PROTECTIVE EFFECT OF NATURAL HONEY AGAINST

CADMIUM-INDUCED HEPATOTOXICITY AND NEPHROTOXICITY BY

Wafaa M. Abdel-Moneim* and Hemmat H. Ghafeer

Departments of Forensic Medicine and Clinical Toxicology* and

Histology, Faculty of Medicine, Assiut University.

ABSTRACT

The therapeutic properties of natural honey once considered a form of folk or preventive medicine. It

is important for the treatment of acute and chronic free radical mediated diseases and toxicity. Oxidative

stress can play a key role in cadmium-induced dysfunction. The aim of this work was to study the effect of

natural honey on cadmium-induced liver and kidney damage. A total of 30 adult male rats were divided

into three groups. Group I animals served as control were injected daily I.P. by 1 ml saline. Group II an-

imals were injected daily I.P. with 0.5mg/kg cadmium chloride dissolved in 1 ml saline for 4 weeks. Ani-

mals of group III were treated with 0.5 mg /kg cadmium chloride I.P. and 0.05ml of natural honey mixed

with water orally concurrently for 4 weeks. Liver function (SGOT),(SGPT), (ALP) and kidney function

(creatinine and urea nitrogen) tests were measured. In addition lipid peroxidation,reduced glutathione

(GSH) and glutathione peroxidase (GPx) were estimated in liver and kidney tissues samples. Light and

transmission electron microscopic examination were used for histological changes. The results revealed

that treatment with Cd caused marked elevation in the level of free radicals (lipid peroxidation) and kid-

ney and liver enzymes, and a decline in GPx activity and GSH level. Administration of honey with Cd in-

duced improvement in all examined parameters. On the other hand, light microscopic examination of kid-

ney cortex of Cd treated group revealed swelling of the cells lining the convoluted tubules and vaculation

of their cytoplasm. Variable degrees of glomerular degeneration were present. The liver showed different

degrees of cell degeneration, necrosis, dilatation and congestion of blood vessels. Results obtained by

EM examination revealed that there were affection of mitochondria and partial loss of microvilli of some

kidney tubules. Furthermore, electron dense mitochondrea, depletion of glycogen granules in a rarified

vaculated cytoplasm were seen in the hepatocytes. It is noticed that concurrent administration of honey

with cadmium improved histological changes in both kidney and liver by light and electron microscope. It

could be concluded that honey via its antioxidant activity has the ability to protect against cadmium-

induced hepatotoxicity and nephrotoxicity.


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Abdel-Moneim & Ghafeer 76


terial, bacteriostatic, anti-inflammatory,

wound and sunburn healing activities. Re-cent views propose honey not only as ahealth promoting dietary supplement, butshed light on antioxidant, non-peroxidedependent properties. This makes honeymore than just a nourishment of high val-ue but a valuable dietary source of antioxi-dants (Beretta et al., 2005).

In recent years there has been an in-creased interest in the application of anti-oxidants to medical treatment as informa-tion is available linking the developmentof human diseases to oxidative stress (Al- Jadi and Kamaruddin, 2004).

Honey is a natural antioxidant which

may contain flavinoids, ascorbic acid, to-copherols, catalase, and phenolic com-pounds all of which work together to pro-vide a synergistic antioxidant effect,scavenging and eliminating free radicals(Johnston et al., 2005).

Little information is available on protec-tive effect of honey against cadmium in-

duced hepatotoxicity and nephrotoxicity.In the present study an experimental mod-el of rats treated with Cd during fourweeks as a model of Cd induced hepato-toxicity and nephrotoxicity were used. Inthis model the protective effect of concur-rent administration of honey on Cd-induced hepatic and renal damage wereassessed and if the protective effect of

emission. It became commercial in the

20th century due to agricultural and in-dustrial applications (WHO, 2000 and Jar-up, 2003).

Occupational exposure to cadmium,such as working with cadmium con-taining pigments, plastic, glass, metal al-loys and electrode material in nickel -cadmium batteries, and non occupation-al exposure, such as food, water andcigarette smoke induces uptake of Cdfrom the environment into the bodythrough pulmonary and enteral pathways(Waisberg et al., 2003). Cadmium ab-sorbed and accumulates mainly in the kid-ney and liver, then it is bound to the apo-protein metallothionein (Morales et al.,

2006).

The intracellular release of cadmium isresponsible for the generation of reactiveoxygen species, glutathione depletion, lip-id peroxidation, protein cross-liking, DNAdamage, culminating ultimately in oxi-dant-induced cell death (Brennan, 1996;Shaikh et al., 1999; Jurezuk et al., 2004 and

Babu et al., 2006).

Honey has been an ingredient of tradi-tional medicine on account of its dietaryand curative properties since ancienttimes. Starting in the early 1970 research-ers from different scientific fields have in-vestigated the chemical and biologicalproperties of honey, including antibac-


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- Group III (Cd + honey) rats were con-

current by administred of 0.5 ml/kgCdCl2 I.P. and 0.05 ml natural honey(Paget and Barnes, 1964) dissolved inwater orally by gastric tube daily forfour weeks.

Blood collection :1- Animals were anesthetized and

blood samples were collected from occularvascular bed using capillary tubes.

2- Blood samples were collected intodry clean tubes on EDTA then centrifugedfor 10 min to isolate plasma and stored at -70oC.

Tissue samples : animals were killed by

cervical decapitation. The liver and kid-neys were taken and each organ was di-vided into two parts.

The 1st specimen was homogenized inice cold 100 mM phosphate buffer (pH7.4). Homogenates were centrifuged andthe resulting supernatant was storted at -70oC for biochemical analysis.

The 2nd specimen was preserved in10% formaline for histological examina-tion.

Biochemical measurements:A) The plasma samples were taken for

determination of :1- Liver function tests: glutamyl - oxal-

honey is based on its antioxidant proper-

ties.

MATERIAL AND METHODS

Chemicals: All chemicals and reagentsused were of analytical grades. Cadmiumchloride (CdCl2) was purchased from ICNpharmaceutical company (USA). Reducedglutathione and Ellman's reagent 5,5 Di-thiobis-2 nitrotenzoic acid (DTNB) wereobtained from ICN Biomedica. Inc. (USA).

Animals: Thirty male albino wister ratsweighing 150-200 gm each obtained fromthe Animal House, Faculty of Medicine,Assiut University were used. Rats werehoused in stainless steel cages at a con-

stant temperature 25 + 2oC with alternat-ing 12 hours light and dark cycles and al-

lowed water and food (laboratory chow)ad libitum. The research was conducted inaccordance with the internationally ac-cepted guidelines for laboratory animaluse and care. The experiment was ap-proved by the Institutional Ethics Com-mittee.

Treatment: Rats were divided intothree groups 10 animals each.

- Group I (control) animals were inject-ed intraperitoneally (I.P.) with 1 mlsaline daily for four weeks.

- Group II animals were injected IP dai-ly with 0.5 mg/kg CdCl2 for fourweeks (Satarug and Moore, 2004).


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and glutathione reductase. The en-

zyme activity is expressed as U/mgprotein.

3- Reduced glutathione (GSH): An equalvolume of 10% metaphosphoric acidwas added to a part of the homogen-ates and mixed by vortexing. The mix-ture was allowed to stand for 5 min atroom temperature. After centrifuga-tion for 5min, the supernatant was col-lected carefully without disturbing theprecipitate. The GSH contents of theneutralized supernatant were assayedusing Ellman's reagent (5, 5'-dithiobis-2-nitrobenzoic acid (DTND solution)according to the method of Griffith(1980). A standard reference curve was

prepared for each assay.

4- Protein contents were determined us-ing the method of Lowry et al., (1951).

Preparation of histological sections:Specimens of the liver and kidney were

taken and fixed in formalin and processedfor light microscope. The specimens were

embedded in paraffin and sectioned at athickness of 7 microns. The sections werestained with haematoxyline and eosinestain according to Drury and Wallington(1980). Other small pieces were fixed inglutraldehyde and processed for electronmicroscope examination. Semithin sec-tions were cut at 1/2-1 micron and stainedwith toluidine blue. Selected sections were

oacetic acid transaminase (SGOT), glu-

tamyl pyruvic transaminase (SGPT)and alkaline phosphatase (ALP).

2- Kidney function tests: creatinine andurea nitrogen.

The two tests were measured spectro-photometrically using standardizedcommercially available Diamond Kits(Modern Laboratory, Egypt).

3- Oxidative stress indices: lipid peroxi-dation level was estimated by themeasurement of malondialdehyde(MDA), an end product of lipid perox-idation level were determined usingspectrophotometre employing thiobar- bituric acid reactive substances de-

scribed by Thayer, (1984). Levels wereexpressed as nmol/ml plasma.

B) Liver and kidney homogenates: wereutilized for determination of

1- Lipid peroxidation was measured asdescribed by Thayer (1984), levelswere expressed as nmol/mg protein.

2- Glutathion peroxidase (GPx): Activitywas measured by the method of Tap-ple (1978). The method is based on oxi-dation of glutathione by cumen hydro-peroxide via glutathione peroxidase.The reaction in the form of decrease inabsorbance at 340nm was followedwhen oxidized glutathione convertedto reduced glutathione via NADPH


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Table (3) shows the levels of lipid per-

oxidation in plasma, and tissues of liverand kidney. In Cd-group the level of LPOin plasma, tissues of liver and kidney wassignificantly higher than controls.In groupreceived Cd + honey the level of LPO wassignificantly reduced to near control levelscompared with the Cd - group.

The effects of Cd and Cd + honey onglutathione peroxidase activity (GPx) intissues of liver and kidney are shown intable (4). The GPx activity level was signif-icantly lower in tissues of liver and kidneyin Cd-group compared with the controlgroup.In comparison with Cd group therewas significant increase in the level of GPxactivity in Cd + honey group in tissues of

liver and kidney. Table (5) shows that re-duced glutathione level in tissues of liverand kidney of cadmium treated groupwas significantly decreased in comparisonto control group.In group received Cd +honey the level of reduced glutathioneshowed significant increase in tissues of liver and kidney in comparison to Cdgroup.

Histological Results :I- Liver :

A) Light microscope results (Hx&E stain) Group I: control group :The control livers show normal lobular

architecture with central vein and radiat-ing cords of hepatocytes, separated by blood sinusoids. Hepatocytes are large

contrasted and electron micrographs were

taken with Jeol transmission electron mi-croscope (TEM).

Statistical analysis:* The data were presented as means +

standard errors.* For the comparison of statistical sig-

nificance between two groups studentNewman-keuls t-test was used.

* Values were accepted as being statisti-cally significant if a P value was lessthan 0.01.

RESULTS

Biochemical Results :Table (1) displays the effects of Cd

alone and Cd with honey on plasma liverenzymes. Levels of SGOT, SGPT and ALPwere significantly increased in the cadmi-um treated group in comparison to thecontrol group. Animals of group III re-ceived Cd + honey showed a significantreduction in SGOT, SGPT, ALP level com-pared with Cd group.

The effects of Cd alone and Cd + honeyon plasma level of kidney function test areshown in table (2). The plasma levels of urea and creatinine were significantlyhigher in Cd-group than controls. Levelsof urea and creatinine in plasma exhib-ited significant reduction in group re-ceived Cd + honey in comparison with Cdgroup.


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formed of rough endoplasmic reticulum

and mitochondria. These cytoplasmic or-ganelles were separated by large areas of rarified cytoplasm. The cisternae of therough endoplasmic reticulum showedfragmentation in the form of short seg-ments and loss of their attached ribosomes(Fig. 5). Whorly appeared structures weredemonstrated in the rarified cytoplasm.Mitochondriae were polymorphic con-tained electron-dense granular matrix.Some of them were swollen, lost their cris-tae and contained vacuoles (Fig. 6).

Group III : combined treatment withCdCl2 and honey led to improvement inthe ultrastructure of hepatocytes whichappeared nearly similar to the control.

Most of the mitochondriae were intact.The cisternae of rough endoplasmic reticu-lum acquired their normal appearance.The nuclei of some hepatocytes appearedlarge, rounded and euchromatic withprominent nucleoli (Fig. 7).

II- Kidney : A) Light microscope results (Hx & E

stain).Group I : the cortex of control kidney is

mostly occupied by renal corpuscles andsurrounding proximal and distal convo-luted tubules. The renal corpuscle isformed of glomerular tuft of blood capil-laries surrounded by capsular space andBowman's capsule.

and polyhedral in shape with slightly

acidophilic granular cytoplasm. They havelarge, rounded, vesicular nuclei withprominent nucleoli (Fig. 1) .

Group II : animals that received CdCl2only :

The liver cells of group II animalsshowed obvious histological changes, inthe form of distortion in the hepatic organ-ization, dilatation and congestion of the blood sinusoids and central vein. Somehepatocytes showed signs of degenerationin the form of hypertrophy with highlyvacuolated cytoplasm and deeply stainednuclei. Other hepatocytes exhibited hyali-nized cytoplasm with pale nuclei andprominent nucleoli (Fig.2).

Group III: CdCl2 + honey :The liver cells appeared more or less

similar to those of the control apart fromfew hepatocytes appeared with vacuolat-ed cytoplasm and pyknotic nuclei (Fig.3).

B) Electron microscope results:Group I : the hepatocytes of control ani-

mal exhibit a large rounded euchromaticnucleus. The cytoplasm contains numer-ous cisternae of rough endoplasmic reticu-lum, numerous well-preserved mitochon-dria and glycogen rosettes (Fig. 4).

Group II : after treatment with CdCl2the hepatocytes showed clumping of thecellular organelles which were mainly


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tes are situated between glomerular capil-

laries, and their secondary foot processeswere extended and rest on the basementmembrane. Numerous filtration slit-membranes were observed between theprocesses of glomerular capillaries(Fig.11).

Regarding the cells of the proximal con-voluted tubule: The apical cell membraneis occupied by numerous microvilli. Thecytoplasm contains numerous mitochon-dria of various sizes. Few cisternal profilesof rough endoplasmic reticulum and ribo-somes were detected in the cytoplasm.The nuclei is rounded euchromatic withprominent nucleolus The basement membrane exhibited basal infoldings with elon-

gated mitochondria between them(Fig.13).

Group II : in cadmium treated animals,most of the renal corpuscles were highlyaffected. There was dilatation and swell-ing of the glomerular capillaries accompa-nied with narrowing of the urinary spaces.The glomerular basement membranes

showed obvious thickening in most of re-nal glomeruli. Most of the secondary footprocesses of the podocytes appeared flat-tened and amulgumated with each otherwith complete disappearance of their slitmembranes. Some of these processescontained dense granules and dense irreg-ular bodies. The lumina of the capillariescontained few exudates and degenerated

The proximal convoluted tubules are

lined with large cuboidal cells with deeplystained acidophilic cytoplasm with apical brush border and rounded vesicular nu-clei. The boundaries between the adjacentcells are indistinct. The distal convolutedtubules are lined by low cuboidal cellswith distinct cell boundaries and less acid-ophilic cytoplasm (Fig.8).

Group II : the glomerular capillaries of some renal corpuscles of animals showedcongestion and dilatation. There wasswelling of some cells of the proximal con-voluted tubules leading to diminution oreven obliteration of the tubular lumina.Cytoplasmic vacuolation and deeplystained nuclei were observed compared to

the control group. Destruction of the brush borders of the proximal convolutedtubules was also detected. The distal con-voluted tubules showed degenerativechanges in the form of pyknotic nuclei andvacuolated cytoplasm (Fig.9).

Group III : combined treatment withhoney and CdCl2 led to obvious improve-

ment in the histological structures of thekidney compared to group II. The renalcorpuscles appeared nearly similar to thecontrol. Most of kidney tubules exhibitedacidophilic cytoplasm and rounded vesic-ular nuclei (Fig.10).

B) Electron microscope results:Group I : ultrastructurally the podocy-


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and genetic information (Patrick 2003 and

DeBurbure et al., 2006). Therefore, someauthors have postulated that antioxidantsshould be one of the important compo-nents of an effective treatment of cadmi-um poisoning (El-Demerdash et al., 2004).

The present study concentrates on thepossible protective effect of honey on oxi-dative damage generated by cadmium in-duced hepatotoxicity and nephrotoxicity.Liver and kidney function tests were donefor different studied groups to assess theirstatus. Biochemical analysis was done foroxidative stress indices such as lipid per-oxidase level. The activity of antioxidantswas measured e.g. glutathione peroxidase(GPx) and reduced glutathione (GSH), be-

cause these antioxidants are the common-est to be affected by cadmium toxicity(Jurezuk et al., 2004). Histological changesof kidney and liver were examined bylight and electron microscopes.

In this work the liver enzymes SGOT,SGPT and ALP in the Cd treated groupwere significantly elevated compared

with the control group, denoting thepresence of liver dysfunction (Shimadaet al., 2004). In concurrent administrationof honey and Cd, the levels of liver en-zymes activity were significantly reducedas compared with the Cd-group. Thisfinding indicates the protective effects of honey in ameliorating the hepatotoxic ef-fect of Cd.

endothelial cells. There was apparent in-

crease in the amorphous secretion of me-sangial matrix (Fig. 12).

Regarding the cells of the proximalconvoluted tubules the apical brush bor-der exhibited partial loss or erosion of their microvilli. The cytoplasm showedmany vacuoles. The mitochondriae weremarkedly affected. They appeared smallwith dense matrix and distructed cristaein some of them. The tubular basementmembrane displayed obvious demolish-ing of their basal infoldings (Fig.14).

Group III : combined treatment withCd and honey. The glomerular capillariesand the podocytes appeared more or less

similar to the control group. The cell of theproximal convoluted tubule exhibited nor-mal cellular organelles and intact microvil-lous border. The apical cytoplasm of somerenal tubules was rarified with somedense bodies (Fig.15).

DISCUSSION

Available literature indicates that noprevious studies have been done to evalu-ate the antioxidant capacity of honey andits protective effect against cadmium in-toxication.

The mechanisms of cadmium-induceddamage include the production of freeradicals that alter mitochondrial activity


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cadmium is thought to induce lipid perox-

idation and this has often been consideredto be the main cause of its deleterious in-fluence on membrane-dependent function.

In the present study the elevation in thefree radicals (LPO) induced by cadmiumalone was very high significantly de-creased in the presence of honey. Thismeans that honey minimized the toxic ef-fect of cadmium via its antioxidant activi-ty. These results are in line with the viewheld by Beretta et al., (2005) who con-firmed the role of honey as an antioxidantagent in blood. Also Manuela et al., (2007)found that there was a direct link betweenthe honey consumption and the level of polyphenolic antioxidants in the plasma.

In agreement with a previous study, thelevel of GSH was significantly decreasedin the liver and kidney extracts of cadmi-um treated group compared to the controlgroup. This decrease in GSH levels may bedue to its consumption in the preventionof free radical-mediated lipid peroxidation(Demopoulos, 1973; Koyuturk et al., 2006).

Also, GSH may be consumed in the detox-ification of heavy metals (Kim et al., 1998and Thevenod, 2003). Furthermore, it has been suggested that the decrease in GSHlevels upon cadmium exposure might im-pair the degradation of lipid peroxides,thereby leading to its accumulation in thetarget organs (Sarkars et al., 1997). In con-troversy to the current results, Kamiyama

The plasma level of creatinine and urea

was significantly increased after cadmiumtreatment compared to the control groupindicating the impairment in the kidneyfunction. Similar observation was ob-tained by Novelli et al., (1998). In fact,urea is the first acute renal marker whichincreases when the kidney suffers anykind of injury. Otherwise, creatinine is themost trustable renal marker and increaseonly when the majority of renal function islost (Borge et al., 2005). The changes inurea and creatinine level in the presentstudy concluded the severe injured effectof CdCl2 on kidney. Moreover, in thepresent study honey co-administrationwith cadmium significantly amelioratedthe increased plasma levels of creatinine

and urea.

In the current study, lipid peroxidationlevel was significantly elevated in plasma,liver and kidney tissues of rats treatedwith cadmium compared to control groupthus suggesting increased oxidative stress.These results were supported by Manca etal., (1991) who reported that LPO is an

early and sensitive consequence of Cd ex-posure. Also, Hassoun and Stohs (1996)demonstrated that oxidative stress was in-duced following oral administration of cadmium chloride to rats. A similar datahad been reported by Jurezuk et al.,(2004). In addition, Elizabeth et al., (2003); Jahangir et al., (2005); Eybl et al., (2006)and Kawamoto et al., (2007) reported that


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ultrafiltered cadmium is taken up, largely

by the proximal tubules of the kidney andis accumulated mainly in kidney cortexleading to proximal tubule lesions (Lars,2002). These findings are in agreementwith the current study.

The nephrotoxicity of Cd has been ex-tensively studied in various experimentalmodels (Mitsumori et al., 1998; Ohta et al.,2000). Recent papers show that tubulardamage may appear at lower levels of Cdexposure than previously anticipated (Jar-up et al., 2000; Noonan et al., 2002).

The Cd concentrations applied in thisstudy was in the range of previous studies(Brzoska et al., 2003; Choi and Rhee, 2003)

according to average human intake data,soil Cd concentrations from contaminatedsites (Blanusa et al., 2002; Satarug andMoore, 2004) and Cd levels from animalscaught in polluted areas (Damek-Proprawa and Sawicka-Kapusta, 2003).

In the present study administration of Cd 0.5 mg/kg I.P. daily for four weeks in-

duces significant damage in function andstructure of kidney assessed by increasedcreatinine and urea concentrations in plas-ma and existence of atrophy of and swell-ing of some glomerular capillaries as wellas proximal tubular necrosis and apopto-sis (Damek-Proprawa and Sawicka - Ka-pusta, 2004). The distal convoluted tubules showed degenerative changes with

et al., (1995) reported an increase in GSH

level in liver and kidney tissues after Cdinjection which could be explained as aprotective mechanism.

The co-administration of honey withcadmium increased the level of the antiox-idant (GSH), and approximated to the nor-mal values of the control group. Al-Waili,(2003) confirmed that honey increased thelevels of antioxidants and this effect might be attributed to the composition of honey,which contains many nutrient elementsand antioxidants as vitamin C, which is apotent antioxidant agent.

The activities of GPx was significantlydecreased in the liver and kidney extracts

of cadmium treated rats. The decline inthe level of GPx activity resulted from Cdtoxicity was previously demonstrated inliver tissues (Sidhu et al., 1993; Ossola andTomara, 1995; Sarkars et al., 1997) and inkidney tissues (Bragadin et al., 2004). Inthe present study reduced glutathione per-oxidase activity was recovered after co-administration of honey with cadmium in

liver and kidney tissues, which in turn de-structs the lipid peroxidase (Eaton et al.,1980). This confirm that honey increase an-tioxidant activity and able to protect fromoxidative stress.

Epidemiological studies have revealedthat cadmium is one of the most toxic of the heavy metals to humans; 70% of the


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(1996) and Shaikh et al., (1999) have re-

ported the mechanism of Cd-induced re-nal damage include increased oxidativestress. Whereas increased oxygen free rad-icals production seems to be induced bythe interaction of Cd and mitochondrialstructure (Tang and Shaikh, 2001).

In the current study treatment withCdCl2 caused liver damage demonstratedfunctionally by increasing the activity of SGOT, SGPT and ALP, and histological al-terations. These alterations were in theform of dilatartion in the hepatic organiza-tion, dilatation and congestion of centralveins and blood sinusoids. Hypertrophyor degeneration of some hepatocytes witheither hyalinized and vacuolated cyto-

plasm with pale nuclei and prominent nu-cleoli. These results coincide with resultsof Borges et al., (2005). These alterationswere observed with electron microscopy.Most of the hepatocytes showed clumpingof the cellular organelles. The cisternae of the rough endoplasmic reticulum showedfragmentation in the form of short seg-ments and loss of their attached ribo-

somes. Mitochondriae were polymorphiccontained electron-dense granular matrix.Some of them were swollen, lost their cris-tae and contained vacuoles. These resultsare in agreement with study of Mousa,(2004). Co-administration of honey withcadmium markedly ameliorated thesehistopathological changes induced by Cd.Most of mitochondrie were intact. The

pyknotic nuclei, and cytoplasmic vacuola-

tion. Ultrastructrually the glomerular basement membranes showed obviousthickening in most of the renal gomeruli.The filteration slits were markedly de-creased in number. Tubular lesions con-sisted in partial lost of the brush bordermicrovilli, altered mitochondrial struc-ture some of them appeared small withdense matrix and districted cristae. Thisis in contrast with the study of Sandy etal., (2007) who revealed that the toxic ef-fects of Cd in the kidney are confinedto proximal tubular cells. With no signs of glomerular and mitochondrial damagewere detected. One possible explanationis that Cd was administrated via thedrinking water not by injection as in the

current study.

These results reveal that honey has amarked protective effect on renal tubulartoxicity and showed decreased lipid per-oxidation and increased tissue levels of GSH and GPx activity a potent endoge-nous antioxidant (Meister and Anderson,1983). This protective effect of honey was

confirmed when renal tissues were ob-served by electron microscopy. Most of kidney tubules exhibited acidophilic cyto-plasm and rounded vesicular nuclei.Moreover, the observation that Cd+honeytreated animals did not have mitochondri-al damage might be seen as supportingthe anti-oxidant properties of honey. Inagreement with this suggestion, Brennan,


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ing free radicals (Johnston et al., 2005).

In conclusion: The present study dem-onstrated that honey administered incombination with cadmium minimized itshazards. Honey can protect against oxida-tive stress induced by cadmium, by lower-ing the free radicals and increased the lev-els of antioxidants. Further studies arerequired to recommend the use of honeyand its therapeutic potential in human.Several antioxidant assays were utilized inorder to evaluate the biological and chemi-cal properties of honey. In addition, theexposure to cadmium should be reducedand attention paid to sources of cadmiumin food, water and personal-care products.Furthermore using diets rich in honey

could be beneficial in alleviating cadmiumtoxicity.

cisternae of rough endoplasmic reticulum

acquired their normal appearance.

In the present study CdCl2 exposure in-creased LPO levels in plasma and tissueswith alterations in antioxidant defenses(GPx and GSH). Thus it may be possiblethat oxidative stress and disturbance inantioxidant defenses were the causes forliver and kidney damage induced byCdCl2 in this experimental model.

On hypothesis to explain the benefi-cial effects of honey in ameliorating bi-ochemical parameters and histologicalchanges is that honey may contains flav-onoids, ascorbic acid, tocopherols, catalaseand phenolic compounds. All of which

work together to provide a synergistic an-tioxidant effect, scavenging and eliminat-


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Table (1): The effect of cadmium alone and with honey on the liver enzymes.

Items

Groups

SGOT

U/L

SGPT

U/L

ALP

U/L

Group I (Control) 12.08 0.74 10.12 0.56 142.21 9.63

Group II (Cd) 58.20 4.98** 65.21 3.42** 1278.404.92**

Group III (Cd+Honey) 15.11 1.1*** 12.34 1.20*** 150.28 6.63***

** P< 0.01 group II vs control. *** P< 0.01 group III vs group II.

Table (2): The effect of cadmium alone and with honey on kidneyfunction tests (creatinine and urea).

Items

Groups

Urea (mg/dl) Creatinine (mg/dl)

Group I 32.24 2.13 0.92 0.04

Group II 86.5 6.42** 3.01 0.3**

Group III 35.47 2.26*** 0.95 0.06***


Table (3): The effect of cadmium alone and with honey on lipid peroxidation level.

Groups

Organs

GroupI

Control

GroupII

Cd

GroupIII

Cd + honey

Plasma nmol/ml 3.56 0.12 9.10 0.53** 4.82 0.11***

Liver nmol/mg 0.79 0.08 2.76 0.38** 0.8 0.04***

Kidney nmol/mg 1.28 0.09 2.67 0.10** 1.58 0.03***



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Table (4): The effect of cadmium alone and with honey on glutathioneperoxidase activity (GPx).

Groups

Organs

GroupI

Control

GroupII

Cd

GroupIII

Cd + honey

Liver U/mg protein 328.50 5.18 231.28 15.38** 331.92 10.36***

Kidney U/mg protein 333.06 4.80 224.20 16.63** 331.20 10.18***


Table (5): The effect of cadmium alone and with honey on reduced glutathione(GSH) level.

Groups

Organs

Group I

Control

Group II

Cd

Group III

Cd + honey

Liver nmol/mg 6.64 0.35 2.65 0.16** 6.22 0.43***

Kidney nmol/mg 2.96 0.21 1.28 0.09** 2.66 0.29***



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Fig. (1) : A photomicrograph of a section in theliver of control adult male albino rat(group I) showing; central vein (C) andhepatocytes with vesicular nuclei ( )separated by blood sinusoids(S). (H&E

X200).

Fig.(2): A photomicrogaph of a section in theliver of rat of (group II) showing; dis-organization of the hepatocytes withvacuolated cytoplasm(V) and deeplystained nuclei. Some of them are hy-pertrophied with deep acidophilic cyto-plasm ( ). Note: dilated and congestedcentral vein (C). (H&E X400).

Fig. (3): A photomicrograph of a section in theliver of rat of (group III) showing; hep-atocytes appeared more or less similarto control apart from few cells withvacuolated cytoplasm ( )

(H&E X400).

Fig.(4): An electron micrograph of control livercells showing; large nucleus with euch-romatin (N). The cytoplasm containnumerous mitochondriae(M), strandsof rER and fine glycogen granules (g)

(X5000).


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Fig.(5): An electron micrograph of liver cell of (group II) animals showing : clumpingof cellular organelles, whorly appearedstructures ( ) in a rarified cytoplasm

(X4000).

Fig.(6): A magnified part of a hepatocyte ob-tained from the previous group. Show-ing : polymorphic mitochondria withelectron dense matrix. Note: vacuole(v) and dense bodies (D) inside the de-generated mitochondriae (X8000).

Fig.(7): An electron-micrograph of the hepato-cyte of (group III) showing : more orless normal ultrastructure with largevesicular nucleus (N), strands of rERand numerous mitochondria (M).

Fig.(8): A photomicrograph of a section of con-trol renal cortex (group I) of adult malealbino rat showing : renal corpuscle( ), Proximal (P) and distal (D) con-voluted tubules. (H&E X200).


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Fig.(9): A photomicrograph of a section of renalcortex of (group II) showing : conges-tion of the glomerular capillaries (G),cytoplasmic vacuolation (v) and pyk-notic nuclei (p) in most of the tubules.Note: some tubular cells with highlyacidophilic cytoplasm ( )(H&E X200).

Fig.(10): A photomicrograph of a section of re-nal cortex of (group III) showing :more or less normal structure

(H&E X200).

Fig.(11): An electron micrograph of a renal cor-puscle of (group I) showing : podocy-tes (P) with their minor or secondaryfoot processes (P2) that rest on normalbasement membrane ( ) of capillaryendothelium. (X 6700).

Fig.(12): An electron micrograph of a renal cor-puscle of (group II) showing : parts of podocytes with fused minor processes( ) rest on a thick basement mem-brane (B). (X6700).


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Fig.(13): An electron-micrograph of proximalconvoluted tubule of (group I) show-ing : long microvillous border (MV),many elongated mitochondria (M), ba-sal spherical nucleus with euchromatin(N). Note: normal basal infoldings.

Fig.(14): An electron-micrograph of proximalconvoluted tubule of (group II) show-ing : loss of the microvilli ( ) smalldense mitochondria (M) and cytoplas-mic vacuoles (V). Note: loss of basalinfolding (X5000).

Fig.(15): An electron-micrograph of proximalconvoluted tubules of (group III) show-ing : intact microvillous border (MV),intact mitochondria. Note: rarifactionof apical cytoplasm with some denseparticles (X5000).


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