Pathomorphological changes of aceclofenac toxicity in layer chicks

Aim: The present research work was undertaken to study the patholomorphogical changes induced by repeated doses of aceclofenac. Materials and Methods: 100 BV-300 day old layer chicks were randomly divided into four equal groups and treated with aceclofenac at the dose rate of 0, 10, 20 and 30 ppm orally through feed for 21 consecutive days. Results: The abnormal behavioral clinical signs like dullness, lethargy, lameness, anorexia, feather plucking and uneven growth were noticed only in chicks of treatment groups III and IV with varied severity. Maximum mortality was observed in group IV (16%) followed by group III (4%). A dose dependent reduction in body weight was found in all the treatment groups. Grossly, on surface of visceral organs white chalky urate deposits of varying degree were observed in chicks which died during experiment from treatment groups III and IV. Histopathologically, the lesions were characterized by congestion, degeneration, haemorrhage and deposition of uric acid crystals. Kidney was the main target organ affected. Conclusion: The present study indicates that aceclofenac is hepatotoxic as well as nephrotoxic which causes visceral gout in layer chicks.


Introduction
meal, bone based DCP, mutton tallow etc. which are likely to have aceclofenac residues and are probably A recent study showed that aceclofenac is a responsible for visceral gout in birds.Birds' being potential threat to critically endangered vultures in uricotelic and lacking the enzyme uricase, the end India [1].Despite the ban on diclofenac in veterinary product of protein metabolism is uric acid and practice, other non-steroidal anti-inflammatory drugs therefore birds have the potential to develop gout.Uric such as aceclofenac, ketoprofen, carprofen, flunixin acid is produced by the liver and excreted through the meglumine, meloxicam, phenylbutazone etc. are still kidneys.The condition occurs as a consequence of widely used.Vultures feeding on the carcasses of severe renal dysfunction, where a reduction in renal recently treated livestock with aceclofenac suffer acute filtration leads to an increase in the blood uric acid kidney failure within few days of exposure and show levels [3]. the same toxic effect when fed on the carcasses of Thus, to further understand the toxic effects of animals treated with diclofenac.As a result, acecloaceclofenac, we conducted the present study to fenac might also be contributing to further decline of elucidate by administering aceclofenac at variable the remaining vulture population.Researchers doses to commercial layer chicks.formerly thought that aceclofenac would not be as harmful because it is metabolized faster by cows, and

converted within hours into a product that is not
Ethical approval: The prior approval from the dangerous to avian species, but with recent advance-Institutional Animal Ethical Committee was obtained ments scientists found that doses given to cattle in India for use of the animals in this study.were sufficient to kill the birds.Hinz et al [2] showed that aceclofenac could slowly but sustainably Chemicals and experimental animals: Aceclofenac in biotransform into diclofenac.Aceclofenac is lethal to tablet form containing 100 mg of the active drug per birds in the doses that would be administered to livestock tablet was used for the present investigation.The study to reduce pain and swelling due to rheumatism or was conducted on day old BV-300 layer chicks.A total arthritis.Poultry feed also contains many animal of 100 BV-300 layer chicks were procured from a local source byproducts like meat and bone meal, blood commercial hatchery (Shakti Hatcheries Pvt. Ltd., Sarsa, Anand, Gujarat, India) and were maintained under standard managemental conditions.Layer chicks were kept in cages and housed in the Experimental Unit, Department of Veterinary Pathology, College of Veterinary Science and Animal Husbandry, Anand.

Statistical analysis:
The data obtained from feed Environmental temperature and lighting regimens consumption, body weight and mortality were were applied according to the BV-300 guidelines.
subjected to one way ANOVA test where a p < 0.05 was considered as statistically significant and p < 0.01 as Determination of the dose: Aceclofenac is less highly significant.potential than diclofenac and most of the studies were

Results and Discussion
conducted in animals, especially in lab animals (mice and rat) and has been not yet conducted in layer chicks.
Chicks under group I and II did not reveal any Aceclofenac is structurally and pharmacologically observable clinical signs whereas chicks of groups III closely related to diclofenac.Hence the dose used and IV showed similar clinical signs which gradually during this study was double that of the previous became more pronounced in group IV.Birds exhibited diclofenac study on broiler chicks [4].
a tendency to remain standing at one place with apathy, unthriftiness with ruffled feathers, dullness and Experimental design: The day old 100 layer chicks drooping of the wings.Birds appeared emaciated, were randomly divided into four different groups, dehydrated, depressed and lethargic with shrunken comprising of 25 chicks in each group.Aceclofenac eyes.They also exhibited signs like anorexia, feather tablet was crushed to fine powder and mixed in the plucking and uneven growth.Such birds showed mash feed in graded levels for inducing toxicity in BVshifting leg lameness and inability to stand.Most of the 300 layer chicks.The groups II, III and IV were given clinical signs observed in aceclofenac treated chicks graded doses of aceclofenac at the rate of 10 ppm (low were in conformity with earlier reports in layer chicks dose), 20 ppm (mid dose), 30 ppm (high dose), [5,6].In albino rats, aceclofenac caused gastro respectively through feed every day for 21 consecutive intestinal ulcers and gastro intestinal bleeding leading days where as group I served as control and fed only to anaemia [7] whereas Darbar et al [8] observed layer mash feed without aceclofenac.Weighing of depression, anorexia, jaundice, fever and nausea due to chicks was done at weekly intervals.Mortality and acute hepatitis following exposure to aceclofenac in morbidity were recorded throughout the experimental rats.Maximum mortality was observed in group IV period.After completion of the experiment, the (16%) followed by group III (4%).In group IV surviving birds from all the four groups were subjected mortality started from day 16 onwards and was to a terminal sacrifice by means of the cervical continued up to day 19 whereas in group III it started on dislocation.The postmortem examination of each bird th 19 day after aceclofenac treatment.Similar results from different groups was performed and gross lesions were observed by Sehgal et al [9] and Reddy et al [10].were recorded.Tissue samples from organs like The dose dependent reduction in body weight was kidney, liver, heart, lung, spleen and intestine were st collected in 10 % formalin for histopathological observed in all the treatment groups.At the end of 1 examination.The tissue pieces from kidney and liver week only group IV showed significant (p < 0.05) were fixed in absolute alcohol and were stained using decrease in body weight.The groups III and IV showed nd De-Galantha's special staining method for demonssignificant reduction in body weight at the end of 2 rd tration of urate crystals.The different parameters like week.At the end of 3 week group III showed significant clinical signs, mortality, body weight, kidney: body whereas group IV showed highly significant (p < 0.01) weight ratio, feed consumption, feed conversion ratio reduction in body weight as compared to the control.(FCR) were studied.
The mean values of kidney: body weight ratio were  significantly increased in groups III and IV as ketoprofen sodium induced toxicity changes in layer compared to the control.There was reduction in the and broiler chicks [12,13].Microscopically, kidney feed consumption observed in group IV followed by lesions showed mild to moderate congestion, intergroup III and II when compared to control during the tubular haemorrages, parenchymatous degeneration, entire experimental period.Shinde [11] observed necrosis of tubules and deposition of uric acid crystals similar results in broiler chicks.The average FCR (Fig- 3 and 4) which appeared black with degalantha's calculated at the end of experiment was higher in staining (Fig- 5). .Liver from the dead dehydration with deposition of white urates in the form birds showed presence of chalky white deposits on its of dry platery patches over various visceral organs surface.Deposition of chalky white urate crystals on upon opening the abdomen (Fig- 1).The affected the liver, a diagnostic feature of visceral gout was also kidneys were more enlarged and frosted with chalky reported by Jana et al [21].Microscopically, liver white urate deposits and pin point haemorrages on the lesions were characterized by necrosis of hepatic cells surface (Fig- 2).Gross pathological lesions of and infiltration of heterophils and mononuclear cells intoxicated layer chicks of the present study were in with focal deposition of uric acid crystals (Fig- 6).agreement with the earlier reports of diclofenac and Focal uric acid crystals were also visible as black    The authors declare that they have no competing interests.gizzard, spleen, lung and intestine and were specific of

Figure- 1 .
Figure-1.Layer chick died from group IV showing white urate deposition on the surface of pericardium (a), abdominal air sacs (b), proventriculus (c) and gizzard (d).

Figure- 2 .
Figure-2.Layer chick died from group IV showing white chalky urates deposition on pericardium and enlarged frosted kidneys with dilated ureters filled with urates.
Madhuri et al [14], Meteyer et al [15], treatment groups as compared to control.Highest FCR Mulcahy et al [16] and Naidoo et al [17] have reported of 1.84 was observed in group IV followed by 1.82 in similar results related to kidney during their studies.group III, 1.80 in group II and lowest 1.79 was recorded The kidney lesions of the nature of gouty nephritis with in group I. uric acid and urates deposition were also reported by Post mortem examination of birds which died Irtaza et al [18], Sharma and Vegad [19] and Senthil during the experiment showed emaciation and kumar and Thirumurugan [20]

Figure- 3 .
Figure-3.Section of kidney from chick died of group III showing marked deposition of needle shaped uric acid crystals in renal parenchyma.

Figure- 4 .
Figure-4.Section of kidney from chick died of group IV showing marked inter tubular haemorrhages, deposition of uric acid crystals degeneration and necrosis of renal tubules.H & E X 300.

Figure- 5 .
Figure-5.Section of kidney from chick died of group IV showing deposition of black uric acid crystals in renal parenchyma.De galantha's X 300.

Figure- 6 .
Figure-6.Section of liver from chick died of group IV showing deposition of uric acid crystals in hepatic parenchyma.H & E X 300.

Figure- 7 .
Figure-7.Section of liver from chick died of group IV showing deposition of black uric acid crystals in hepatic parenchyma.De Galantha's X 300.

Figure- 8 .
Figure-8.Section of heart from chick died of group IV showing severe engorgement of blood vessel in the cardiac parenchyma along with uratic deposition on the pericardium (arrow).H & E X 300.

Figure- 9 .
Figure-9.Section of spleen from chick died of group IV showing depletion of lymphoid cells.H & E X 150.

Figure- 10 .
Figure-10.Section of lung from chick died of group IV showing moderate congestion and focal to diffuse haemorrhages in alveolar parenchyma.H & E X 300.