Original Article

Effect of cimetidine on hepatotoxicity induced by

isoniazid-rifampicin combination in rabbits
Bhupinder Singh Kalra, Sarita Aggarwal,* Nita Khurana,** Usha Gupta
Departments of Pharmacology, *Biochemistry and ** Pathology,
Maulana Azad Medical College, New Delhi 110 002

Objective: To evaluate the hepatoprotective potential of cimetidine in hepatotoxicity induced by isoniazid-rifampicin combination in albino rabbits. Methods:
Six groups of six rabbits each were studied. Three
groups received saline (control), isoniazid (50 mg/
Kg/d) alone or isoniazid with rifampicin (100 mg/Kg/
d) daily orally for 7 days. Other groups received
intraperitoneal cimetidine (50 mg/Kg/d) alone, or
cimetidine (50 or 120 mg/Kg/d) along with isoniazidrifampicin combination. Serum levels of liver enzymes
were measured at baseline and on day 8, and liver
histology was studied on day 8. Results: Rabbits
receiving isoniazid alone for 7 days showed no increase in serum ALT and AST levels, whereas those
receiving isoniazid-rifampicin combination had a 3-4fold increase in these levels (p=0.02). Animals receiving cimetidine pre-treatment did not show a significant increase in ALT and AST levels. Histological
changes in the liver were more common with isoniazid-rifampicin combination than with isoniazid only.
These changes were reduced in animals receiving
low-dose cimetidine and prevented in those receiving
high-dose cimetidine. Conclusion: Cimetidine in high
dose can prevent hepatotoxicity induced by isoniazidrifampicin combination. [Indian J Gastroenterol
2007;26:18-21]

he use of isoniazid and rifampicin in the treatment

of tuberculosis is limited by their potential for
hepatotoxicity. The incidence of hepatotoxicity is
higher with isoniazid and rifampicin combination
than with isoniazid alone. 1
The exact mechanism responsible for liver injury caused by these drugs is not clear. Isoniazid
is acetylated and then hydrolyzed, resulting in
isonicotinic acid and monoacetylhydrazine; the latter
compound can be activated to a toxic species by
cytochrome P-450. 2 In vitro studies indicate that
metabolic oxidation of acetylhydrazine leads to a
reactive acylating species that binds covalently to
microsomal protein. It is postulated that
acetylhydrazine and hydrazine act as acetylating
agents by binding covalently with liver cell macromolecules, causing hepatocyte injury. 3

Copyright 2007 by Indian Society of Gastroenterology

Rifampicin, a powerful inducer of mixed-function oxidase, increases the hepatotoxicity of isoniazid by enhancing the production of toxic metabolites from acetylhydrazine. 4 Cimetidine, a microsomal P-450 enzyme-system inhibitor, has been
shown to reduce the hepatotoxicity of several drugs
in rats by inhibiting oxidative drug metabolism. 5,6,7
Rabbits show a similar genetically determined
acetyltransferase activity as in humans and are
more sensitive to isoniazid-induced hepatotoxicity
due to a high amidase activity, which results in
release of large amount of acetylhydrazine, which
induces hepatotoxicity. 8 We therefore used a rabbit model to study whether cimetidine could ameliorate the hepatotoxicity of isoniazid and rifampicin.
Methods
This study was approved by the institutional animal ethical committee and Committee for Prevention of Cruelty and Supervision of Experiment on
Animals. Albino rabbits of either sex, weighing 23 Kg, were procured from the central animal house
of the institute and housed in individual cages in
air-conditioned environment. They were allowed
to acclimatize for 7-10 days before start of study,
and were provided normal diet and free access to
water throughout the study.
Rabbits were divided into 6 groups of six animals
each. Animals in Group 1 (control) received normal saline orally. Animals in Group 2 received
isoniazid (50 mg/Kg/d), and those in Group 3 received a combination of isoniazid (50 mg/Kg/d)
and rifampicin (100 mg/Kg/d) orally for 7 days.
Group 4 received only cimetidine (120 mg/Kg/d)
intraperitoneally for 7 days. Animals in Group 5
and Group 6 received isoniazid-rifampicin combination with intraperitoneal cimetidine in doses of
50 or 120 mg/Kg/d, respectively, for 7 days.
Cimetidine was given 30 minutes before the isoniazid-rifampicin combination.
A baseline blood specimen (2 mL) was drawn
from the lateral ear vein of each rabbit, after mop-

Cimetidine salvage of isoniazid-rifampicin hepatotoxicity

Kalra, Aggarwal, Khurana, Gupta

Table 1: Body weight, liver weight, liver morphology and food intake in different treatment groups
(n = 6 each)

Group

signed rank test;

p values <0.05
Food intake Liver weight Animals with changes in liver (n)
were taken as
(g)
(g)
Inflammation Necrosis
significant.
90 (5.5)
24.67 (2.66)
0
0
70 (2.0)a
24.50 (3.39)
2
0
Results

Body weight (Kg)

Baseline
Day 8
Group 1
2.4 (0.4)
2.3 (0.4)
Group 2
2.4 (0.5)
1.7 (0.3)*
26.67 (2.07)
4
2
Group 3
2.6 (1.7)
1.8 (0.3)*
65 (6.0)b
Group 4
2.5 (0.4)
2.2 (0.3)
80 (2.5)
24.83 (2.48)
1
0
Group 5
2.8 (0.3)
2.1 (0.4)*
75 (6.5)c
26.00 (2.45)
2
0
Group 6
2.4 (0.3)
2.0 (0.3)
75 (5.0)d
25.00 (2.45)
1
0
Data as mean (SD). Group 1: control; Group 2: isoniazid; Group 3: isoniazid + rifampicin; Group 4:
cimetidine (120 mg/Kg/d); Group 5: isoniazid + rifampicin + cimetidine (50 mg/Kg/d); Group 6:
isoniazid + rifampicin + cimetidine (120 mg/Kg/d)
*p=0.02 as compared to baseline values. a p=0.009, bp=0.004, cp=0.01, d p=0.02 as compared to
Group 1

ping the ear with xylene, on day 1 (before administration of any drug). On day 8 (the day after
last day of drug administration), the animals were
sacrificed by cervical dislocation followed by
exsanguination; from each animal, a blood specimen was collected through cardiac puncture and
liver tissue was collected. Blood was centrifuged
at 2500 rpm for 10 min and extracted serum was
stored at -20 0 C till estimation of liver enzymes. 9
The liver tissue was examined with naked eye
and preserved in 10% buffered formalin for histological examination. Two wedge sections were
made from each liver, one each from the center
and the periphery. Tissue was processed by cycling it through different chambers of a histokinette
(Leica Corp, Germany) over 15 hours followed
by preparation of paraffin blocks. Sections from
the blocks were stained with hematoxylin-eosin
and examined under a light microscope for portal
inflammation, ballooning degeneration, necrosis and
fatty changes.
Statistical analysis was done using Wilcoxons

Food intake
and weight of
animals
Animals receiving
isoniazid alone
and in combination with rifampicin
had
decreased food intake (Table 1). Animals in the isoniazid-rifampicin combination group were also
quieter, had less motor activity, and roughening
of hair coat. Rabbits in Groups 2, 3 and 5 showed
reduction in mean body weight (Table 1).
Liver enzymes (Table 2)
Animals who received isoniazid alone had no significant increase in ALT and AST levels. However, rabbits receiving isoniazid-rifampicin combination showed a 3-4-fold increase in ALT and
AST levels (p=0.02). Concomitant administration
of cimetidine at the higher dose prevented this
increase, but that of low-dose cimetidine did not.
Liver morphology
Mean liver weights were similar in all the groups.
Livers from the control group did not show portal inflammation, necrosis, fatty change or ballooning degeneration (Table 3). Animals receiving
isoniazid or isoniazid-rifampicin combination showed
portal inflammation, fatty changes (Fig 1) and
liver cell necrosis (Fig 2); the changes were more

Table 2: Effect of different treatments on AST and ALT

serum levels (n = 6 each)

Groups

AST (IU/L)
ALT (IU/L)
Baseline
Day 8
Baseline
Day 8
1
27 (13-58) 28 (13-43)
28 (12-70) 30 (12-38)
2
34 (13-109) 83 (13-200)
27 (12-38) 38 (27-153)
3
24 (13-47) 124 (27-200)* 23 (6-52) 60 (12-200)*
4
20 (13-47) 20 (13-47)
12 (12-70) 20 (12-38)
5
27 (13-57) 74.5 (20-181) 23 (12-38) 63 (27-153)#
6
20 (13-57) 42 (20-200)
23 (6-52) 33 (12-200)
Data as median (range); *p<0.02, #<0.04 as compared to
baseline
Group 1: control; Group 2: isoniazid; Group 3: isoniazid +
rifampicin; Group 4: cimetidine (120 mg/Kg/d); Group 5:
isoniazid + rifampicin + cimetidine (50 mg/Kg/d); Group 6:
isoniazid + rifampicin + cimetidine (120 mg/Kg/d)

Fig 1: Portal inflammation (grade 1). Mild portal inflammation seen in livers of animals treated with isoniazid-rifampicin
combination (Group 3) (H&E, 100X)

Indian Journal of Gastroenterology 2007 Vol 26 January - February 19

Kalra, Aggarwal, Khurana, Gupta

Cimetidine salvage of isoniazid-rifampicin hepatotoxicity

mild portal triaditis. 14
Similar changes were
s e e n i n o u r s t u d y,
confirming the validity of our animal
model.
Cimetidine has
been reported to have
a hepatoprotective
effect in animals receiving toxic doses of
p a r a c e t a m o l . 5,15
These studies showed a marked reduction in serum liver enzyme levels; however, histology was
not studied. To the best of our knowledge, the
effect of the microsomal-enzyme inhibitor
cimetidine on isoniazid- and rifampicin-induced
hepatotoxicity has not been reported in the literature. In the current study, we found that cimetidine
at the higher dose level prevented an increase in
AST and ALT levels as well as the histological
changes associated with isoniazid-rifampicin combination. In addition, it also prevented the reduction in body weight caused by isoniazid-rifampicin combination. This may be due to inhibition of
enhanced production of toxic reactive metabolites.
In conclusion, our data show that, in a rabbit
model, rifampicin potentiates the hepatotoxicity
caused by isoniazid, and that cimetidine administration in a high dose ameliorates biochemical and
histological changes associated with administration of isoniazid-rifampicin combination.

Table 3. Histological changes in livers in different treatment groups (n = 6 each)

Group

Portal inflammation
Necrosis
Fatty change
Ballooning
Mild
Moderate
Mild
Moderate
Mild
Moderate
degeneration
Group 1
0
0
0
0
0
0
0
Group 2
3
2
3
0
2
1
5
Group 3
2
4
2
4
3
3
2
Group 4
2
0
0
0
0
0
2
Group 5
4
0
1
0
3
1
2
Group 6
3
0
0
0
1
0
1
Data shown as number of animals affected
Group 1: control; Group 2: isoniazid; Group 3: isoniazid + rifampicin; Group 4: cimetidine (120
mg/Kg/d); Group 5: isoniazid + rifampicin + cimetidine (50 mg/Kg/d); Group 6: isoniazid +
rifampicin + cimetidine (120 mg/Kg/d)

marked in the latter group. Animals receiving only

cimetidine did not show any necrotic or fatty
changes in their livers. Cimetidine pre-treatment
was associated with reduction in changes induced
by isoniazid-rifampicin combination; this effect
was more marked in animals receiving a higher
dose of cimetidine.
Discussion
In a study of 427 hospital employees receiving
isoniazid chemoprophylaxis for tuberculosis, abnormal liver function test results correlated well
with histological evidence of hepatocellular damage. 10 Further, several studies suggest that hepatitis is more frequent and more severe in patients
receiving isoniazid-rifampicin combination treatment than in those receiving isoniazid alone. 11,12
Our study confirms these findings using an animal model.
Isoniazid-induced hepatitis is associated with
ballooning degeneration, focal hepatocyte necrosis, with minimal cholestasis. 13 Another study reported diffuse microvesicular fatty infiltration with

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Fig 2: Normal liver parenchyma with normal portal tract.

Normal parenchymal cells seen in animals administered
high dose of cimetidine along with isoniazid and rifampicin
(Group 6) (H&E, 200X)

6.

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rifampicin and INH on liver function. Br Med J 1972;1:14850.
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20 Indian Journal of Gastroenterology 2007 Vol 26 January - February

Kalra, Aggarwal, Khurana, Gupta

Cimetidine salvage of isoniazid-rifampicin hepatotoxicity

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Correspondence to: Dr Kalra, A-45, Surya Nagar, Ghaziabad

201 011, U P. E-mail: [email protected]
Acknowledgements: The authors thank Lupin Research
Laboratories for providing pure isoniazid and rifampicin
powders and Cadila Healthcare Ltd for providing pure
cimetidine powder
Received June 15, 2006. Received in final revised form
October 31, 2006. Accepted November 15, 2006

Image
Jejunal duplication cyst:
diagnosis aided by scintigraphic detection of ectopic gastric mucosa
A 2.5-year-old girl was referred with history of abdominal
pain, vomiting and recurrent melena. Ultrasonography
revealed a tangle of tubular structures in the descending colon. Despite de-worming twice, the pain persisted.
Meckels scan was performed with Tc99m-pertechnetate
(200 mCi/Kg IV). The stomach was visualized in the
first 15 minutes and later another area of tracer localization was seen below the stomach in the left hypogastric region. Subsequent static images until 3 hours
(Fig) showed an oblong area of activity, suggesting
ectopic gastric mucosa in a duplication cyst from the
gut, most probably in the jejunum.
Exploratory laparotomy revealed a diverticulumlike 4.5-cm structure arising from the antimesenteric
border approximately 30 cm from the duodeno-jejunal
junction, with a narrow communication with the lumen
of the jejunum. Histology showed an endogenous cyst
lined by gastric mucosa arising from the jejunum.

Of all alimentary tract duplications, jejunal

duplications are more likely to contain gastric
mucosa. Unlike in Meckels diverticulum, intestinal duplications commonly arise from the mesenteric border, 1 unlike in this case. Abdominal scintigraphy (Meckels scan) using Tc99m-pertechnetate
is a well-established diagnostic technique to evaluate
children with lower gastrointestinal bleeding to
look for ectopic gastric mucosa. 2
A Zakir Ali, K Kumaresan
Department of Nuclear Medicine, CARE
Hospital, Banjara Hills, Hyderabad

Fig: Scan at 3 hours shows persisting activity in oblique

linear structure outside stomach in left para-umbilical
region

References
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2.

Emamian SA, Shalaby-Rana E, Majd M. The spectrum of

heterotopic gastric mucosa in children detected by Tc99m
pertechnetate scintigraphy. Clin Nucl Med 2001;26:52935.
Sfakianakis GN, Anderson GF, King DR. The effect of
intestinal hormones on the Tc99m pertechnetate imaging
of ectopic gastric mucosa in experimental Meckels diverticulum. J Nucl Med 1981;22:678-83.

Correspondence to: Dr. Zakir Ali, Jasmine, Plot No. 81,

Happy Homes Colony, Upparapally, Hyderabad 48. E-mail:
[email protected]

Indian Journal of Gastroenterology 2007 Vol 26 January - February 21