Effect of Antox on Paraquat Changes in Kidney of Albino Rats

Journal of Applied Sciences Research, 3(10): 988-993, 2007
© 2007, INSInet Publication
Effect of Antox on Paraquat - Induced Histological and Biochemical
Changes in Kidney of Albino Rats
1
1
Hawazen A. Lamfon and 2Maisaa M.Al-Rawi
Biology Department, Faculty of Applied Sciences, Umm Al-Qura University.
2
Sciences Department, Girls Collage, Makkah, Saudi Arabia.
Abstract: Paraquat is a widely used and effective herbicide. Treating rats with paraquat at a dose level
of 1/36 LD 50 3 d/week for 3 weeks induced many histological changes in the kidney.The renal tubules
lost their characteristic appearance and their linning epithelial cells appeared with cytoplasmic
vacuolation. The glomeruli were degenerated and the renal blood vessels were congested. The intertubular
spaces were infiltrated by inflammatory leucocytic cells. Paraquat also caused marked elevation in serum
creatinine and blood urea nitrogen. These alterations were time-dependent. Treating rats with paraquat
and the antioxidant,antox led to an improvement in both the histological and biochemical alterations
induced by paraquat. This inhibitory effect of antox is attributed to its antioxidant and free radicals
scavenging properties of its components (selenium, vitamin A acetate,ascorbic acid and vitamin E).
Keywords:
INTRODUCTION
M ATERIALS AND M ETHODS
Paraquat (1,1'-dimethyl-4,4'-bipyridylium dichloride)
is a widely used and effective herbicide with a broad
spectrum of activity. However, paraquat is quite toxic;
the toxicity on animals and humans has been well
documented [5,17]. A mouthful of the herbicidal
compound usually results in death from caustic burns,
renal tubular necrosis, and circulatory failure due to
pulmonary fibrosis Campbell[4]. Experimental studies
have shown that it is accumulated in the lung and
kidney epithelial cells, leading eventually to pulmonary
fibrosis and acute renal failure [8,10]. M any studies
showed that paraquat poisoning was accompaninig
by histological changes in different organs of
laboratory animals [13,20].
Antioxidents nutrients, as ascorbic
acid,
tocopherol, B-carotene, etc., are considered to give
protection against oxidative damage induced by
different toxicants and reduce the activity of free
radical-induced reactions [16]. Antox is an antioxidant
drug composed of selenium, vitamin A acetate,ascorbic
acid and vitamin E. Antox was used in therapy
of different liver diseases [24,11,25,19]. The aim
of
the present work
is
to
explore
the
effect
of antox on paraquat –induced kidney
injury
in albino rats.
Adult male albino rats weighing 120 ± 5 g were
used. Animals were kept in the laboratory under
constant temperature (24±2 oC) for at least one week
before and throughout the experimental work. They
were maintained on a standard diet and water was
available ad libitum. Animals were divided into 4
groups. Group1: animals of this group (20 rats) were
given orally the herbicide paraquat dissolved in
mammalian saline solution at a dose level of 1/36
LD 50 (3.46 mg/kg body weight) 3 times per week for
3 weeks. Group 2: animals in this group (20 rats)
were given the same dose of paraquat given to
animals of group 1 followed by antox dissolved in
water at a dose level of 3.4 mg/kg body weight 3
times weekly for 3 weeks.Antox tablets composed of
selenium, medicinal yeast,ascorbic acid,vitamin A
acetate and vitam in E (A rab C ompany for
Pharmaceuticals and Medicinal Plants). Rats in the
third group (20 animals) were given antox only and
those in the fourth group(10 animals) were given
saline. The treated animals and their controls were
killed by cervical disslocation, quickly dissected and
their kidneys were fixed in Bouin's fluid, dehydrated,
embedded in wax and 5 micrometers thick sections
were stained with haematoxylin and counterstained
with eosin. for creatinine and urea nitrogen
Corresponding Authr: Hawazen A. Lamfon, Biology Department, Faculty of Applied Sciences, Umm Al-Qura University
988
J. Appl. Sci. Res., 3(10): 988-993, 2007
determination, sera were obtained by centrifugation of
the blood samples and stored at -20 0C until essayed
for the biochemical parameters. Serum creatinine and
urea nitrogen were measured using a fully automated
Hitachi 911 analyzer (Tokyo, Japan). Commercial
Randox kits (Randox Laboratories Ltd, Ardmore,
Crumlin, United Kingdom) were used in these
analyses. The results were analysed statistically using
Student’s “t” test.
RESULTS AND DISCUSSIONS
Data in table (1) shows that treating animals with
paraquat induced an elevation in creatinine in the
serum compared with that of control. This increase
was significant (p < 0.05) after 2 and 3 weeks of
treatment. Animals treated with paraquat and antox
showed a significant decrease in creatinine values in
comparison with those treated with paraquat alone.
Results in table (2) shows that blood urea nitrogen
exhibited a significant increase after 1,2 and 3 weeks
of treatment with paraquat.On the other hand, rats
treated with paraquat and antox revealed a significant
decrease in urea nitrogen after 2 and 3 weeks in
comparison with rats treated with antox.
Fig. 2: Section in the kidney of a rat treated with
paraquat for 2 weeks showing dilated and
congested renal vein (CR), x 400.
Fig. 3: Section in the kidney of a treated rat
showing renal tubules desquameted from the
underlying basement membrane (arrow) x
400.
capsule called Bowman’s capsule. Between the two
layers of the capsule is the urinary space. The
proximal convoluted tubule is lined by simple cuboidal
or columnar epithelium. Its cells have an acidophilic
cytoplasm and the apex possesses abundant microvilli
which form a brush border. The distal convoluted
tubule is lined by simple cuboidal epithelium. The
cells of the collecting tubules and ducts stain weakly
with the used stains.
Histological examination of kidneys of animals
after one week of treatment showed insignificant
changes. After 2 weeks, examination of the kidney
tissue showed that most of the renal blood vessels
were dilated, congested and engorged with blood
(Fig. 2).
The linning cells of the renal tubules exhibited
signs of cloudy swelling and most of them showed
Fig. 1: Section. in the kidney of a control rat
showing a glomerulus (G) and renal tubules
(R), x 400.
Figure (1) shows the histological structure of the
kidney of control rat. The kidney is divided into an
outer, granular appearing cortex and an inner, striatedappearing medulla. It is composed of a huge number
of functional filtering units, nephrone. Each nephrone
consists of a dilated portion, the renal corpuscle; the
proximal convoluted tubule; the thin and thick limbs
of the loop of Henle; and the distal convoluted tubule.
The renal corpuscle consists of a tuft of capillaries,
the glomerulus, surrounded by a double walled
epithelial
989
J. Appl. Sci. Res., 3(10): 988-993, 2007
Table 1: Effect of different treatments on serum creatinine.
Period of treatments (weeks)
Control
Antox
Paraquat
Paraquat +Antox
1
0.45 ± 0.02
0.55 ±0.01
0.67 ±0.03
0.70 ±0.2
-----------------------------------------------------------------------------------------------------------------------------------------------------------------2
0.43 ±0.01
0.51 ±0.02
1.80* ±0.3
1.21±0.1
-----------------------------------------------------------------------------------------------------------------------------------------------------------------3
0.51 ±0.01
0.53 ±0.2
2.3* ±0.7
1.45 ±0.4
-Values are expressed as mean ± SD (mg/dl).
(*) Significant at P<0.05
Table 2: Effect of different treatments on blood urea nitrogen
Period of treatments (weeks)
Control
Antox
Paraquat
Paraquat +Antox
1
31.6 ± 1.2
32.4 ±2.2
54.6* ±1.4
43.5 ±1.4
-----------------------------------------------------------------------------------------------------------------------------------------------------------------2
33.2 ± 2.3
31.8 ±3.1
68.7* ±2.4
36.7 ± 3.4
-----------------------------------------------------------------------------------------------------------------------------------------------------------------3
33.6 ±1.2
30.9 ±2.2
77.3* ±1.5
41.4 ±5.2
-Values are expressed as mean ± SD (mg/dl).
(*) Significant at P<0.05
Fig. 4: Section in the kidney of a treated rat showing
leucocytic infiltration (F) and degenerated
glomeruli (G), x 400
Fig. 5: Section in the kidney of a rat 3 weeks posttreatment with paraquat showing degenerated
tubules x 400.
separation of the tubular cells from the underlying
basement membranes (Fig. 3). Large number of
glomeruli were degenerated and inflammatory
leucocytic cells were abundant in the intertubular
spaces (Fig. 4). These histopathological changes
were increased after three weeks of treatment. M ost
of the renal tubules were damaged and lost their
characteristic appearance. Their lining epithelial cells
became undistinguished and their contents were
intermixed with each other (Fig. 5). The walls of
Bowman’s capsule were eroded and the glomeruli
were atrophied and in some sections appeared as
empty spaces containing amorphous cellular derbis
(Fig. 6). Kidney sections of animals treated with
paraquat and antox for 2 weeks revealed
certain degree of improvement in the tubular
appearance, but few glomeruli were atrophied (Fig.7).
An obvious degree of improvement were observed
after 3 weeks and the kidney tissue appeared normal
(Fig.8). Animals treated with antox for 3 weeks
showed normal appearance of the kidney tissue.
Discussions: The present results showed that paraquat
induced many histological alterations in the kidney of
albino rats. These results are similar to those shown
by many investigators. Abdel-M agid [1] reported that
kidney of p araquat intoxicated rats revealed
degeneration of renal tubules, shrunken glomeruli and
congested dilated blood vessels. Damin et al.,[7]
emphasized that as a result of paraquat poisoning,
990
J. Appl. Sci. Res., 3(10): 988-993, 2007
Fig. 6: Section in the kidney of a rat 3 weeks posttreatment with paraquat showing atrophied
glomerulus (G) x 400.
Fig. 8: Section in the kidney of a rat treated with
paraquat and antox for3 weeks showing
normal appearance of the tubules and the
glomerulus (G) x 400.
increased blood urea nitrogen and creatinine
concentration are caused by paraquat poisoning. Hyo
et al.,[14] reported a case of Korean woman who
p r e se n te d w ith ge n era lized p ro xim a l tu b u la r
dysfunction including aminoacidiuria, phosphaturia and
glycosuria after paraquat intoxication.Paraquat is
known to be filtered at the glomerulus and actively
secreted by tubular cells [2]. In addition, it has been
reported that the kidney has the highest concentration
of paraquat in rats [21]. Therefore, the histological and
biochemical results observed in the present work
proved that paraquat affected both function and
structure of rat kidney.
Animals treated with both paraquat and antox
revealed an improvement in histopatho logical
alterations induced by paraquat alone.Moreover, the
elevation of blood urea nitrogen and creatinine was
significantly reduced in rats treated with paraquat and
antox. This proved the effectiveness of antox in
prevention of paraquat-induced renal toxicity. The
protective effect of antox was studied by many
investigators. Antox succeeded in minimize cadmiuminduced toxicity in albino rats and increase the activity
of endogenous antioxidants
including
glutathione,superoxide dismutase and catalase [11].
W atson et al., [24] reported that bio-antox have
protective effect against primary biliary cirrhosis. A
significant reduction in oxidative stress parameters as
well as in blood and hepatic lead level and in hepatic
8-oxodeoxyguanisone phosphate were recorded after
giving antox to Schistosoma-infected and chronic lead
Fig. 7: Section in the kidney of a rat treated with
paraquat and antox for 2 weeks showing
somewhat normal appearance of the tubules
and a degenerated glomerulus (G) x 400.
renal damage occurs. Laurence and Bennett[15]
reported that toxicity of paraquat caused renal tubular
necrosis followed by kidney failure. Inflammatory cells
were observed in the interstitial tissue of kidney of
paraquat-treated
rats. Similar observation was
described in human cases of intoxication with
paraquat in the lung and considered as sign
of
toxicity
and
consequent
activation
of
defensive mechanism [3].
Regarding the biochemical results, administration
of paraquat caused marked elevation in creatinine and
blood urea nitrogen. Similarly, Cobe [6] reported that
991
J. Appl. Sci. Res., 3(10): 988-993, 2007
exposed hamsters [9]. Antox was found to maintain
blood glutathione,plasma vitamin C and serum
selenium levels towards the normal range [12].
Some studies showed that antioxidents are usefull
in prevention of renal damage induced by different
toxicants including paraquat. Nagano et al.,[18] reported
that dimethylthiourea,deferoxamine and alphatocopherol have a protective action against paraquatinduced acute renal failure in mice. Shukla and
Chandra [22] reported that antioxidants (vitamin E and
ascorbic acid) produced a significant reduction in lipid
peroxidation induced by cadmium and prevent kidney
damage in rats. Antox is a multivitamin compound
(ascorbic acid,vitamin A acetate and vitamin E). The
protective of these vitamines rests with strong
antioxidants, free radical scavenging activity and
inhibition of lipid peroxidation [23].Thus the preventive
effect of antox against paraquat-induced renal damage
recorded in the present work is attributed to its
antioxidant properties.
10. Haley, T.J., 1979. Review of the toxicology of
paraquat. Clin. Pharmacol., 14: 1-19.
11. Hamooda H., A. baalash, K. Saad and A. Emara,
2 0 0 3 . o x id a tive stre ss; aha llm a rk o f
cadmiuminduced toxicity in different tissues, Drug
Metabolism Rev., 35(1): 231-237.
12. Heaney A.P., N. Sharer, B. Rameh, J.M. Bregnza
and P.N. Durrington, 1999. Prevention of recurrent
pancreatitis in familial lipoprotein lipase deficiency
with high-dose antioxidant therapy. J. Clinic.
Endocrinol. Metabol., 84(4): 1203-1205.
13. Hirai, K.I., H. W itschi and M.G. cote, 1985.
Mitochondrial injury of pulmonary alveolar
epithelial cells in acute paraquat intoxication. Exp.
Mol. Pathol., 43: 242-245.
14. Hyo, W .G., O.Y. Jong, Y.L. Eun and Y.H. Sae,
2005. Paraquat-ind uced F anco ni syndrome.
Nephrology, 10 (5): 430-432.
15. Laurence, D.R. and D.N. Bennett, 1992. Clinical
phanmacology, 7 th ed., Churchill livingstone, oxord
univ. press.
16. McCall MR, F. Balz, 1999. Can antioxidant
vitamins materially reduce oxidative damage in
humans? Free Rad. Biol. Med., 26: 1034-1053.
17. Murray, R.E. and J.E. Gibson, 1972. A
comparative study of paraquat intoxication in rats,
guinea pigs and monkeys. Exp. Mol. Pathol., 17:
317-325
18. Nagano, N., M. Yagi and N . K oji, 1992.
Protective effects of antioxidants on paraquat induced acute renal failure in mice. Japan. J.
pharmacol., 59: 481-483.
19. Oz, H.S., C.J. McClain, H.T. Nagasawa, M.B.
Ray, W .J. deVilliers and T. Chen, 2004. Diverse
antioxidants pro tect against acetam inophen
hepatotoxicity. J. Biochem. Mol. Toxicol.,
18(6): 361-368.
20. Sakr, S.A., M.M. El-Mofty, E.I. Agamy and Y.A.
Okdah, 1999. Electron microscopic study on the
effect of the herbicide “paraquat” on the ileum of
albino rat. J Egypt. Germ. Soc. Zool., 30: 173180.
21. Sharp, C.W ., A. Ottolenghi and H.S. Posner,
1972. Correlation of paraquat toxicity with tissue
concentration and weight loss of rat. Toxical.
Appl. Pharmacal., 22: 241-251.
22. Shukla, G.S. and S.V. Chandra, 1989. Cadmium
toxicity and bioantioxidants:status of vitamin E
and ascorbic acid of selected organs in rat. J.
Appl. Toxico. 9: 119-122.
23. Upasani, C.D., A. Khera, R. Balaraman, 2001.
Effect of lead with vitamins E,C, or spirulina on
m a lo n d ia ld e h y d e : c o n j u g a te d d i e n e s a n d
hydroperoxides in rats. Indian J. Exp. Biol.,
39(1): 70-74.
REFERENCES
1.
2.
3.
4.
5.
6.
7.
8.
9.
Abdel-Mageid, S.A., 1994. Structural changes in
the kidney of albino rat in response to the
administration of paraquat herbicide. J. Egypt.
Germ. Soc. Zool., 15: 153-175.
Bismuth, C., R. Garnier, F.J. Baud, J. Muszynski
and C. Keyes, 1990. Paraquat poisoning: An
overview of the current status. Drug Saf.,
5: 243-251.
B orchard, F., 1974.
U ltrastructural
and
microscopical findings in 3 cases of paraquat
p o is n in g w ith p r o lo n g e d l e t h a l c o u r s e .
Pneumonologie, 150-185.
Campbell, S., 1968. Paraquat poisoning. Clin.
Toxical., 1: 245-249.
Clark, D.G., T.F. Elligott and E. W eston-l lurst,
1966. the toxicity of paraquat. Br. J. Ind.
Med., 23: 126-132.
Cobe, R.B., 2004. Helping animals exposed to the
herbicide paraquat. Vetrinary Med, 110: 755-762.
Damain, F., B. Frank, H. W in fried, M. Hartmut
and W . Laus, 1992. Failure of radio therapy to
resolve fatal lung damage due to paraquat
poisoning. Chest., 100: 1146-1165.
Davis, D.S., Hawks G.M. worth and P.N. Bennet,
1977. Paraquat poisoning. Proc. Eur. Soc.
Toxicol., 18: 21-26.
El-Gohary A.E. M.,Yassin and M.A. Shalaby,
2003. The effect of chronic lead exposure on the
effect of schistosomasis in hamsters and the
protective effect of the antioxidant preparation
"Antox". Hum.Exp.Toxicol., 22(9): 481-490.
992
J. Appl. Sci. Res., 3(10): 988-993, 2007
24. W aston J.P., D.E. jones, O.F james, P.A. cann
and M.G. Bramble, 1999. Oral antioxidant therepy
for the treatment of primary biliary cirrhosis :a
pilot study. J. Gastroenterol Hepatol., 14(10):
1034-1040.
25. Zhong Z. and J.J. Lemasters, 2004. Role of free
radicals in failure of fatty liver grafts caused by
ethanol. Alcohol, 34(1): 49-58.
993