The document presents a study that examined the effects of aspirin administration on the liver, kidney, and reproductive system of female albino rats. Various parameters were measured including organ weights, hematological factors, enzyme levels, histopathology, and estrus cycle. The results showed that aspirin induced toxicity in the liver and kidney and caused morphological changes in the ovaries, oviduct, and uterus along with effects on the estrus cycle.

1. “Aspirin induced alterations in
the Liver, Kidney and Reproductive
System of female albino rat,
Rattus rattus norvegicus”.
2010-2011
Supervisor Co-supervisor Presented By
Prof. Arun Raghuwanshi Prof. Vinoy K. Shrivastava Neha Jain
Faculty of Life Science Head of the M.Phil.Bioscience
Department of Bioscience

2. CONTENTS
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 SUMMARY
 REFERANCES

3. INTRODUCTION
• What is Aspirin
• History of Aspirin
• Mechanism of action
• Common-side effects
• Indications

4. What is Aspirin
Aspirin is an analgesic the brand name Aspirin was
coined by the Bayer company of Germany
also called acetylsalicylic acid most frequently selled
acetylsalicylic acid compound Worldwide
Aspirin belongs to a class of drugs called
non-steroidal anti-inflammatory drugs
(NSAIDs).
They work by inhibiting the enzymes that
make prostaglandins, chemicals that promote
inflammation, pain and fever, and are
necessary for blood clotting.

5. History of aspirin
In the 5 th century people used the bitter
powder extracted from a willow bark to
relieve pain
salicylic acid was used since 1874 as a
drug
• in 1897 Felix Hofmann formed
acetylsalicylic acid by derivatizing one of
the hydroxyl functional groups in
salicylic acid with an acetyl group
• 'Aspirin' was patented in 1899
Felix Hofmann

6. Aspirin: Mechanism of Action
Aspirin inhibits an enzyme, called COX which is responsible for the production of
prostaglandines there are two forms : COX 1 and COX 2
COX 2
COX 1 • induced ( normally not present in cells)
• continuously stimulated by the body • built only in special cells (A549 lung
• its concentration in the body remain cells)
stable • used for signaling pain and
• creates prostaglandins used for basic inflammation
house keeping throughout body • produces prostaglandins for
• prostaglandins stimulate normal body inflammatory response
functions such as stomach mucous • stimulated only as part of immune
production, regulation of gastric acid and response
kidney water excretion

7. Aspirin: Mechanism of Action
Membrane Phospholipids
Aspirin
Arachidonic Acid
Cyclooxygenase
Lipooxygenase
PGH PGG2
2
Leukotrienes
Prostaglandins Thromboxanes

8. Aspirin: Mechanism of Action
Platelet Aggregation and Activation

9. Aspirin: Mechanism of Action
Available Antiplatelet Agents

10. Indications
• Aspirin used as,
• Local analgetic effect
• Antipyretic
• Anti-inflammatory
• Antiplatelet
• Analgesic &Antipyretic
• Rheumatic fever & heumatoid
Arthritis
• External applications Clinical aspirin resistance
• Corns & calluses
• Colon cancer • The inability of aspirin to protect a
• Cardiovascular applications person from cardiovascular events such as
heart attacks and strokes.
• Prophylactically to decrease the
incidence of ischemic attacks &
unstable angina

11. AIM
“Aspirin induced alterations in the Liver, Kidney and Reproductive
System of female albino rat, Rattus rattus norvegicus”

12. MATERIALS AND METHODS

13. Animals Selection
• The present experiment were performed on mature female albino
rat, Rattus rattus (n=20). The rats were obtained from the colony of the
animal house of Barkatullah University, Bhopal. The rats were 15-16
weeks old, the body weight range 125 ± 5 g and acclimatized up to
three weeks. The rats were maintained at suitable temperature of 23 ±
2ºC with an automatically controlled photo-period (12 hours light and 12
hours darkness) and the rats were fed with standard balanced pelleted
diet with tap water "ad libitum". The changes in vaginal smear were also
measured daily. The above experiment was performed during the month
of November-10 to March- 11.

14. Chemical and Dose Selection
• Aspirin tablets (Acetyl Salicylate, Ecosprin-
150) obtained from registered medical shop,
Bhopal has been used for present experiment.
The experimental animals were administered
Aspirin orally at a dose level of 100 mg/Kg
b.w./day through gavage.

15. Experimental design
For present investigation total 20 female rats were divided into 2 groups
(ten each), these different groups and dosing patterns as follows.
• Ist Group – The animals of this group were fed with normal diet,
were called SET A or Control Group.
• IInd Group – The animals of this group was fed with normal diet
and received Aspirin by oral administration for 15 & 30 days, were
called Treated Group. This group was divided into 2 sub groups
as following.
• Ist Sub Group – these groups of 5 animals was fed with normal
diet and received Aspirin by oral administration for 15 days, was
called SET B or Treated Group A.
• IInd Sub Group – these groups of 5 animals was fed with normal
diet and received Aspirin by oral administration for 30 days, was
called SET C or Treated Group B.

16. Method Design
• All animals were sacrificed after 24 hours of the last
dose according to their dosing patterns i.e. 15 days
treated animals killed 16th day and 30 days treated
animals killed 31st day along with control animals. After
sacrificing blood were collected through cardiac
puncture and serum were prepared for the different
parameters i.e. biochemical parameter etc. At the same
time body organs viz. liver, kidney, ovary, oviduct,
uterus were quickly removed, washed with physiological
saline, dried (by filter paper), weighed (by physical
precision balance). Then these organs either kept in
Bovines fixative for histopathological study or tissue
homogenate were prepared for enzymological study.

17. Parameter estimated
Body Weight study: From 1-30 days of control and treated animals on different time
interval i.e. initial, after 15 days and after 30 days.
Hematological Study: Hb %, RBCs and WBCs count
Enzymological Study:
1) Acid Phosphatase (ACP): King and Kings Method (1954).
2) Alkaline Phosphatase (ALP): King and Kings Method (1954).
3) Glutamate Oxalate Transaminase (GOT): Reitman and Frankel Method (1957).
4) Glutamate Pyruvate Transaminase (GPT): Reitman and Frankel Method (1957).
Biochemical Study:
1) Cholesterol Estimation: By Ferric Chloride and Sulphuric Acid Method
(Henly, 1957).
2) Protein Estimation: By Follin-Phenol Method (Lowry et al., 1951).
3) Glucose Estimation: By Dinitro Salicylic Acid Method (Miller, 1959).
4) Bilirubin Estimation: By Malloy and Evelyn Method (1937)
5) Calcium Estimation: By Eriochrome Dye Method (Preston Smith et al., 1966).
Histological study: Ovary, Oviduct, Uterus, Liver and Kidney.
Estrus cycle study: From 1-30 days (daily) of control and treated animals.

18. RESULTS

19. Body Weight study

20. Hematological Study

21. Enzymological Study

22. Biochemical Study

23. Histological study
Microphotograph of Ovary
AF
CL
AF CL
AF CL

24. Histological study : Microphotograph of Uterus (X 40/100/400)
L
EM
MM
PM
EM
L MM
PM
L EM
MM
PM

25. Histological study
Microphotograph of Liver
S
S
S

26. Histological study
Microphotograph of Kidney (Cortex Region)
T
T
T

27. Estrus cycle study
Microphotograph of Vaginal Smear
S
Proestrus Phase Estrus Phase
Metaestrus Phase Diaestrus Phase

28. Following phases are seen in rat vaginal smear
•Estrus lasts up to 12 h and is indicated by the Cell types in vaginal smears of Rattus rattus
presence of large cornified cells in the vaginal
smear. Epithelial cells
•Metestrus lasts 21 h and usually has many Х 80
Х 80
neutrophils in the smear and scattered squamous
epithelial cells.
•Diestrus lasts up to 57 hours and there are Cornified cells Leucocytes
abundant neutrophils and a few nucleated non-
cornified epithelial cells.
•Proestrus lasts 3-12 hours and has abundant
nucleated non-cornified epithelial cells.

29. The regular study of vaginal smears of control
and treated groups of rat (Rattus rattus).
DAYS 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30
C
O
N
T D E M P D E M P D E M P D E M P D E M P D E M P D E M P D E
R
O
L
G
R
O
U
P
T
S
R
E
A E M P D E M P D E M P D E M P D E M P D D D D D D D D D D D
T
E
D
Control Group = This Group contains 5 animals which receive D, E, M and P = Showing estrus cycle phases
only normal diet from 1 to 30 days. D = Diaestrus Phase
E = Estrus Phase
Treated Group = This Group contains 5 animals which receive M = Metaestrus Phase
normal diet and aspirin (100mg/Kg b.w.) by oral administration
P = Proestrus phase
from 1 to 30 days.

30. DISCUSSION

31. DISCUSSION
• Data of the present study indicate that, body weight of rats (n=5) were not significantly
affected by oral administration of Aspirin (100mg/Kg b.w.) for 30 days. One could suggest
that oral administration of Aspirin (0.05% w/v) do not affect body weight in rats (Ebuehi et
al., 2007; Kwon et al., 1997).
• Our present findings shows that, significant (<0.001) increase in the number of TC WBC in
30 days Aspirin treated rats. The auther suggested that, Aspirin increases the production of
blood eosinophils (and possibly blood basophils) (Hirai et al., 2001).
• After the administration of Aspirin (100mg/Kg b.w.) for 30 days do not significant altered
TC RBC and Hb %. Accoding to auther, Aspirin had showed no effect or alteration on
packed cell volume (Otimenyin, et al).
• In our present study, the serum protein content were not significantly (p<0.01) reduced by
oral administration of Aspirin. Ebuehi et al., (2007) suggested that, oral administration of
Aspirin may inhibit protein synthesis or many prevent the uptake of amino acids into the
tissues (such as in brain tissues).
• The study of Aspirin on the serum Ca level suggest that, the Ca++ concentration significantly
(p<0.01) decreased on the dose level 100mg/Kg b.w. of Aspirin. In addition to, Aspirin diet
enhances calcium absorption for the improvement in hen’s egg shell percentage and
thickness (Abdullah et al., 2010). Low levels of serum Calcium can be caused by low
albumin or total protein levels (www.questdiagnostics.com, 2004).
• The administration of Aspirin for 15 and 30 days on rats caused significant reduce the
glucose and cholesterol level. Aspirin is known to inhibit glycogenesis and
aminotransferase enzymes which may lead to decreased gluconeogenesis. The reduction of
glucose and cholesterol levels which accompanied the reduction of corticosterone and T3
levels that consequently may affect its metabolism (Mohammed et al., 2010).

32. DISCUSSION
• In our present study Aspirin significantly increases ACP and ALP content in Ovarian
Utricular tissues and not significantly altered ACP and ALP content in Serum. The
present investigation revealed that, Aspirin significantly increased serum GOT and
GPT content in 15 days and 30 days treated rats at the dose level 100 mg/Kg (b.w.). As
well as those patients, who are treated with NSAIDs, particularly with Aspirin, are at
more danger of developing hepatotoxicity than others as well as the liver enzymes
(ALT ,ALP, GOT and GPT) started to deteriorate gradually (Kawar et al, 2010).
• In the present data indicates, Aspirin significantly reduces serum bilirubin content in
rats compare to control rats. In the treatment of arthritis an anti-inflammatory drug
were given (Aspirin level was 200 mg/kg/24hr) for two days found that, the bilirubin
level were normal in patients (Kawar et al., 2010).
• After the 15 and 30 days treatment of Aspirin (100mg/Kg b.w.) causes, significantly
increase of serum Creatinine. Aspirin treated patients increase of serum Creatinine
(Muhammad et al., 2010).
• In the present investigation found that Aspirin induced toxicity on liver and Kidney
both along with patho-histological changes at the higher dose level (100mg/Kg
b.w./day). The results revealed that Aspirin made changes in antioxidant system even
at therapeutic doses, made no changes in hepatic and renal pathology (Balog et al.,
2000).

33. DISCUSSION
• Our present findings show that, the morphological changes found
in both ovaries and oviduct and uterus seen in the experimental
groups may be due to the vascular properties of Aspirin. long-term
administration of Aspirin for 30 days, developed an areas of hemorrhagic
spots, at the site of uterine wall vascularization and this may be due to
blood vessels weakness induced by the prolonged program of Aspirin-
induced treatment (Schafer. et al., 1995).
• Aspirin administration persistently arrest luteolysis (i.e. intact and
functional corpora lutea), which has been confirmed by daily vaginal
smear, that reflect a cellular picture of diestrous phase. Inhibitors of
PGF2α and synthesis (indomethacin and acetylsalicylic acid) as the
endogenous mediator of luteolysis, were shown to delay the regression
of the corpora lutea and to prolong the luteal activity in pseudopregnant
rats (Jezova et al., 1999).
• structural ovarian changes seen: significant increase in the number of
corpora lutea and a significant increase in the diameter of the granulose
lutein cells, with absence of regressed corpora luteal when compared to
the control animals.
• The structural uterine changes by Aspirin namely: the significant
decrease in the endometrial lining cell height, endometrial thickness and
diameter of endometrial glands in both periods of Aspirin treated rat (15
and 30 days) (Chaffin et al., 2000).

34. SUMMARY

35. SUMMARY
The data of the present study summarized as, the oral administration of Aspirin (100mg/Kg b.w.)
do not significantly affected to the body weight, Hb%, TC RBC and protein content. As well as
significantly increases TC WBC. In addition, oral administration of Aspirin significantly inhibited
the level of Glucose and cholesterol level. Therefore, it could be inferred the level of serum and
tissues enzymes such as ACP, ALP, GOT and GPT. And also Aspirin significantly caused
alteration in bilirubin and Creatinine concentration. Furthermore information about Aspirin is that
it also causes patho-histological changes in liver, kidney and reproductive organ (i.e. ovary,
oviduct and uterus).
It conclude from present study is that, the long term use of Aspirin at the dose
of 100mg/kg b. w. orally is responsible for damage to female reproductive
organs, if precautionary measures are not taken while using them. Structural
damage to mammalian oviduct may be lead to infertility cause. So in selecting a
safe drug, quantities and mode of usage must be strictly monitored to minimize
the possibility of injurious complications.
This can be achieved through public health education to make people aware of
the hazardous effect of these compounds. It is therefore recommended that
great precautions and supplements must be taken to minimize the harmful
side effects of such chemicals especially to human and animals.

36. REFERANCES

37. REFERENCES
• Abdullah A. Mohammed (2010): Effect of acetyl salicylic acid (Aspirin) in drinking water on productive performance and blood characteristic of
layer hens during heat stress. Int. J. Poult. Sci., vol. 9 (4), p382-385.
• Acta Veterinaria, Turja A. Panteli, Drenka et al., (2010): Effects of Aspirin on the number of peripheral white blood cells and spleen eosinophils in
guinea-pigs. Vol. 60, No. 2-3, p355-362.
• Bonses, R.W. & Tausskay, H.H. (1945); J.Biol, Chem., 158, 581. Slot, C. (1965).
• Scand J. Clin, Toro, G. & Ackermann P, (1975): Practical Clinical Chemistry, p154.
• Buck ML, Grebe TA, Bond GR (1993): Toxic reaction to salicylate in a newborn infant: similarities to neonatal sepsis. J. Pediatr., 122:955-8.
• Cheng Y, Austin SC, Rocca B, Koller BH, Coffman TM, Grosser T, Lawson JA, Fitz Gerald GA (2002): Role of prostacyclin in the cardiovascular
response to thromboxane A2. Science, 19, 296(5567), p539-41.
• Chignard M, Concalves de, Moraes VL, Lefort J, Meager A, Vargafting B (1996): Effect of cyclooxygenase inhibitors and modulators of cyclic
AMP formation on lipopolysaccharide-inducedneutrophil infiltration in mouse lung. Br. J. Pharmacol., 117, 8.
• Daud AN, De Silva KI, Deng J, Gamelli RL, Jones SB, Shanker R.(2003): Prostaglandin E2 mediates growth arrest in NFS-60 cells by down-
regulating interleukin-6 receptor expression. Biochem. J, 370, Pt 1.
• Decaterina R, Giannessi D, Bernini W (1985): Low-dose Aspirin in patient recovering from myocardial infarction: evidence for a selective
inhibition of thromboxane-related platelet function. Eur. Heart. J., 6, p409-417.
• Ebuehi, O.A.T., Q.C. Opara and A.I. Akinwande, (2007). Effects of oral administration of Aspirin and paracetamol on plasma and brain
protection, tryptophane levels and monoamine oxidase activity in rats. Journal of Biological Sciences, 7 (8), p1468-1472.
• Endo T, Yamamoto H, Tanaka S (1988): Effect of prostaglandins on progesterone production by human luteal cells & their ability of
prostaglandin production. Nippon. Naibunpi Cakkai Zaschi, 20, 64(8), p687-97.
• Ganong WF ((2003): Review of medical Physiology. A Lange medical book, 21 ed., McGraw-Hill.
• Garcia-Rodriguez LA (2001): The effect of non steroidal anti-inflammatory drugs on the risk of coronary heart disease: fusion of clinical
pharmacology and pharmacoepidemiologic data. Clin. Exp. Rheumatol, 19, (6 suppl. 25), p41-4.
• Hall JE, Schoenfeld DH, Martin KA, et al (1992): Hypothalamic gonadotropin-releasing hormone secretion & folliclestimulating hormone
dynamics during the luteal-follicular transition. J. Clin. Endocrinol. Metab., 74, p600-7.
• Hall, D., et al. (1992): Counteraction of the Vasodilator Effects of Enalapril by Aspirin in Severe Heart Failure. J. Am. Coll. Cardiol., 20:1549-
1555).
• J. M. Balog, 2 G. R. Huff, N. C. Rath, and W. E. Huff (2000): Effect of Dietary Aspirin on Ascites in Broilers Raised in a Hypobaric Chamber.
Poultry Science 79:1101–1105.
• Jezova M, Scsukova S, Vranova J, Kolena J (1999): Modulation of LH/hCG receptors & physical state of ovarian membranes in rat
psuedopregnancy. Gen. Physiol. Biophys., 18(4), p347-56.
• Lowry O.H., Rosebrough N.J., Farr A.L., Randal R.J. (1951): Protein measurement with the Folin phenol reagent. J. Bio. Chemistry, 193, 265.
• Luigi Di Luigi, Laura Guidetti, Francesco Romanelli, Carlo Baldari, and Domenico Conte (2001): Aspirin, Exercise, And Pituitary Hormones.
Official Journal of the American College of Sports Medicine, p2029-2035.
• Marcia L. Buck, (2007): Use of Aspirin in Children with Cardiac Disease. Pediatric Pharmacotherapy,Vol. 13, Number 2.
• McCannce KL, Huether SE (1994): Pathophysiology, the biologic bases for disease in adults and children. 2nd ed. Mosby year book, Inc., The
menstrual cycle, p723.

38. Thanks !!!