Effect of Dietary Proteins on Isoniazid Induced

Subchronic Hepatotoxicity In SD rats.

Presented by
Rajesh . D . Hillal
Pharmacology & Toxicology
Reg.No.PT.2009.09

Guide Co-Guide
Dr. P. Uday Kumar. Dr. S. Ramakrishna .
Scientist E, Course Coordinator,
Department of Pathology Pharmacology and Toxicology,
NIN ,Hyderabad NIPER Hyderabad.
 Contents
Introduction

Literature Review

Hypothesis

Aim & Objectives

Work done

Plan of work

Expected Outcome

References

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 Introduction
Drug induced hepatotoxicity
• The liver is the very important body organ plays role in the
metabolism of drugs and other environmental chemicals.
• During this xenobiotic metabolism, liver is susceptible to the injury
due to the generation of toxic metabolites or hepatotoxic effects of
the drug.
• Drug induced hepatotoxicity accounts for more than 50% of acute
liver failure, including hepatotoxicity caused by overdose of
acetaminophen (APAP, 39%) and idiosyncratic liver injury triggered
by other drugs (13%).

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 Cont…

• The drug induced hepatotoxicity is the major problem in the western

countries and it is responsible for withdrawal of many drugs from
market like Troglitazone, Bromfenac, Ebrotidine.
• Majority of the liver toxicity caused by the therapeutic classes of the
drug is acute liver failure or idiosyncratic reactions .

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 Types of drug induced hepatotoxicity

Drug induced
Hepatotoxicity

Idiosyncratic Intrinsic

Allergic Nonallergic

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 Mechanisms of drug induced hepatotoxicity

12/09/2021 Stefan Russmann et al, Current Medicinal Chemistry, 2009, 16, 3041-3053 . 6
 Risk factors for hepatotoxicity

Toxic potential of drug

-Reactive
metabolites
-Acylglucuronide
-Mitochondrial
effects

Genetic factors Environmental factors

-Drug metabolism -Underlying disease
-Detoxification -Age
-Transport -Ethanol
-Others -Other

Neil kaplowitz, Drug induced liver diseases,2005 .

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 Isoniazid induced hepatotoxicity
• The cornerstone of tuberculosis management is a 6-month course of
isoniazid, rifampicin, pyrazinamide, streptomycin and ethambutol.
• The most frequent adverse effects of antituberculosis treatment are
hepatotoxicity,& neurological disorders.
• Asymptomatic transaminase elevations are common during
antituberculosis treatment, but hepatotoxicity can be fatal when not
recognized early and when therapy is not interrupted in time.
• Isoniazid (INH) is the most preferred drug in both chemoprophylaxis
and treatment of tuberculosis. Daily intake, however, causes moderate
elevation in liver enzymes and severe hepatic damage (especially
hepatic necrosis) in 3–20% of patients, respectively.
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 Metabolism of INH

12/09/2021 A Tostmann et al. Journal of Gastroenterology and Hepatology . 2008; 23:192–202 . 9

 Literature Review
Effect of Dietary components on drug metabolism & toxicity

Food and dietary components may affect the fate of a drug or

toxicant by the following mechanisms:

1) Altering the rate of its absorption and uptake,

2) Reacting or tightly binding with the drug,

3) Competing with the drug for binding to plasma proteins

4) Affecting phase I and phase II metabolism.

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 Influence of proteins on drug metabolism & toxicity

 Proteins, fat, carbohydrate, vitamins, and minerals can all have

pharmacological as well as physiological effects on a biological
system.
 Interfering with P450-dependent metabolism appears to be the
most selective mechanism by which dietary proteins exert their
effects on drug metabolism & toxicity.
 This enzyme system has been shown to increase as dietary level of
protein is increased up to around 25%, which shows a correlation
with the growth rates of the animals.

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 Effect of protein content on drug metabolizing enzymes

Seysuo Tomizawa et al. Jap. journal of pharmacology,1968,18,356-366 . 12

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 Effect of protein content on drug induced toxicity

Seysuo Tomizawa et al. Jap. journal of pharmacology,1968,18,356-366 .

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 Hypothesis

 It is known that dietary protein level influences the level of P450

enzymes
 It is well known that proteins have masking effect on toxicities of
various drugs.
 Keeping this in view, it has been hypothesized that, to study the
protective effect of dietary proteins , against isoniazid -induced
hepatotoxicity in rats.

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 Aim & Objectives

 To study the effect of NTP-2000 diet containing protein (14.5%), on

Hepatotoxic effect of Isoniazid in comparison to NIN stock diet
(20% protein), one commercially available diet (22-23% protein),
and a specially formulated diet containing (8.0% protein).
 It is also proposed to quantify levels of drug metabolizing enzymes

in response to the above situation.  

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 Work done
• Learning histopathology, hematology techniques as well as urine

parameters estimation which will help in project work.

• Histopathology techniques:
Sampling Sectioning
Processing Staining
Embedding Mounting
Trimming
• Hematology techniques:
RBC count Platelet count
WBC count Erythrocyte sedimentation rate.
Hemoglobin
Packed cell volume
• Urine parameter estimation:

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 Plan of work

• Materials and Methods

• Animals required
a. Species (Albino Rats) : Sprague Dawley rats
b. Age/Weight/Size : 3 months old
c. Gender : Males+ Females
 
• Materials: Isoniazid, NIN, NTP-2000, Commercial AND Low protein diets

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 Study Design

(Equal numbers of both sexes)

X = 50mg (INH)
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 Cont…

 Animals would receive the drug from 40th day to 54th day of
experiment and animals would be euthanized on day 90.
 Blood withdrawal at 30th , 40th ,54th , 90th day of the experiment.

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 Parameters

Feed intake Body weight Clinical

Hematology Histopathology
measurement Measurement Biochemistry

-Red Blood Cell

-Creatinine
-White Blood Cell
-Blood Urea Nitrogen
-Hemoglobin
-Total Protein
-Packed cell volume
-Albumin

-Aspartate amino transferase

-Alanine amino transferase

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 Expected Outcome
When the experiment will be completed, the following outcomes are
envisaged:

The higher protective effect of diet containing higher protein

content compared to diet containing lower protein content against
isoniazid induced hepatotoxicity in SD rats.

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 References
1 Alma Tostmann, Martin J Boeree, Rob E Aarnoutse, Wiel C M de
Lange, Andre J A M van der Ven, Richard Dekhuijzen.
Antituberculosis drug-induced hepatotoxicity: Concise up-to-date
review, Journal of Gastroenterology and Hepatology. 2008; 23:
192– 202.
2. Yakup Ergul,TulayErkan, HafizeUzun, HabibeGenc, Tuncay Altug,
Ethem Erginoz. Effect of vitamin C on oxidative liver injury due to
isoniazid in rats. Pediatrics International. , 2010; 52:69-76.
3. Troy C. Sarich, Mohammed YousseÞ, Ting Zhou Stephen P Adams,
Richard A Wall, James M. Wright. Role of hydrazine in the
mechanism of isoniazid hepatotoxicity in rabbits. Archieve of
Toxicology. 1996; 70: 835-840 .
4. William M. Lee, Drug-Induced Hepatotoxicity. The New England
journal of medicine. 2003; 349: 474-85.

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5. Sarich TC, Youssefi M, Zhou T, Adams SP, Wall RA, Wright JM.
Role of hydrazine in the mechanism of isoniazid hepatotoxicity in
rabbits. Archieve of Toxicology. 1996; 70: 835–40.
6. Huang YS, Chern HD, Su WJ. Cytochrome P450 2E1 genotype and
the susceptibility to antituberculosis drug-induced hepatitis.
Hepatology 2003; 37: 924–30.
7. Philip G. Reeves. Components of the AIN-93 Diets as Improvements
in the AIN-76A Diet. Journal of Nutrition.1997; 127: 838S-841S.
8. Newberne PM. Influence of pharmacological experiments of
chemicals and other factors in diets of laboratory animals.
Federation Proceedings.1986;34: 209-218.
9. Rao GN. Diet and Kidney diseases in rats. Toxicologic .pathology.
2002; 30: 651- 656.
10. Edward J Mosoro, Yu BP. Dietary modulation of the progression of
nephropathy in aging rats: An evaluation of importance of protein.
American journal of clinical nutrition.1989; 49:1217-27.
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11. Rao GN. New diet (NTP-2000) for rats in the national toxicology
program toxicity and carcinogenicity studies. Fundamental Applied
Toxicology, 1996; 32:102-108.
12. Guengerich FP. Influence of nutrients and other dietary materials on
cytochrome p-450 enzymes. American journal of clinical nutrition.
1995; 61:651S-656S.
13. Sude Eminzade. Fikriye Uraz, Fikret V Izzettin. Silymarin protects
liver against toxic effects of anti-tuberculosis drugs in experimental
animals, Nutrition & Metabolism, 200; 5: 1-7.
14. Stefan Russmann, Gerd A, Kullak-Ublick. Current Concepts of
Mechanisms in Drug-Induced Hepatotoxicity. Current Medicinal
Chemistry, 2009; 16: 3041-3053.

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 Backup slides

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G.N Rao.FUNDAMENTAL AND APPLIED TOXICOLOGY ,32, 102-108 (1996 )
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G.N Rao.FUNDAMENTAL AND APPLIED TOXICOLOGY ,32, 102-108 (1996 )