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BRIEF RESUME OF INTENDED WORK:
6.1 Need for the study:
Cardiac
toxicity is
a
major
adverse
effect
caused
by
Doxorubicin/Adriamycin (Dox) therapy. Doxorubicin(Dox), an anthracycline
commonly used against a broad spectrum of human cancers, has well known dose
dependent cytotoxic effects like myocardial injury leading to potentially lethal
congestive heart failure.1
Although the mechanism underlying the severe toxicity of doxorubicin
and other anthracyclines are not fully understood, there is evidence that toxicity
may ensue through drug free radical formation2 and subsequent redox cycle with
O2 resulting in the generation of reactive oxygen species(ROS) such as superoxide
anion, hydroxyl radicals and hydrogen peroxide. Tissues with less developed
activities of antioxidant enzymes [superoxide dismutase (SOD) and catalase
(CAT)] and glutathione dependent antioxidant enzymes [glutathione peroxidase
(GPX) and glutathione-s-transferase (GST)] have been reported3 in Adriamycin
induced cardio toxicity in rats. The antioxidant defenses of heart are particularly
susceptible to injury by Dox induced oxygen radicals.
Available reports suggest that some plants like Phyllanthus Urinaria L.4
and Centella Asiatica5 show cardioprotective effect against Dox induced cardio
toxicity as they may have free radical scavenging property and also promote
antioxidant activity. Therefore any plant extract or drug showing free radical
scavenging property and promote antioxidant activity may show cardioprotective
effect.
Phyllanthus Niruri L. belongs to Phyllanthus species but no scientific
reports are available on cardioprotective and antioxidant effect.
Literature survey reveals that Phyllanthus Niruri has a paucity of
information regarding its cardioprotective effect.
Therefore, the present work is planned to investigate the antioxidant and
cardioprotective effects of Phyllanthus Niruri L. on Dox induced cardio toxicity in
albino rats.
6.2 Review of literature:
Phyllanthus Niruri Linn. is a traditional herbaceous plant of
Euphorbiaceae family and is widely distributed all over India.
The herb is bitter in taste and is reported to possess astringent,
deobstruent, stomachic, diuretic, febrifugal, antiseptic, antibacterial8 and antiviral
properties.9 In the indigenous system of medicine, it has been used in stomach
troubles such as dyspepsia, colic diarrhoea and dysentery, in dropsy and diseases
of urogenital system.10
Phytochemical
evidence
reveals
the
presence
of
phyllanthin,
hypophyllanthin,6 gallic acid, kaempferol,9 alkaloids, flavonoids, diterpenes,
triterpenes, sterols, isoquercetin and potassium in the whole plant.10
Niruriflavone, a new antioxidant flavone sulfonic acid exhibited potent
free radical scavenging property.6 Therefore one may speculate that it can be
useful against Dox-induced cardiotoxicity.
Therefore, the present attempt has been made to investigate the role of
aqueous extract Phyllanthus Niruri L. on Dox- induced cardiotoxicity in rats
6.3 Objective of the present study:
1) To find out the preventive role of Phyllanthus Niruri on Dox induced
myocardial toxicity.
2) To evaluate the possible mode of action of Phyllanthus Niruri by
measuring various markers in Dox treated rats.
3) To study histopathological parameters against Dox induced myocardial
toxicity.
4) Evaluation of oxidative status in described models.
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METHODS AND MATERIALS:
7.1 Source of data :
Data will be collected using laboratory experimental techniques on
animals as described in the present study. The results obtained from the present
study will be analysed using the one way analysis of variance (ANOVA) test
followed by Dunnet’s multiple ‘t’ test.
7.2 Method of collection of data:
Following data’s are collected from the experimental protocol:Animals:Adult Wistar albino rats of either sex weighing between 150-200 gms
will be used and divided into four groups of six animals each.
Doxorubicin induced myocardial toxicity11 :
Group I: CONT (Control group): Animals will be treated with vehicle alone; on
the same regimen as doxorubicin.
Group II: DOX (Doxorubicin treated): Doxorubicin will be administered
intraperitonially in 6 equal injections (each dose containing 2.5 mg/kg body
weight) for a cumulative dose of 15 mg/kg body weight.
Group III: PN extract administered (1/10 of LD50 i.e. effective dose): Extract will
be pretreated orally for 2 weeks and then alternatively with vehicle for next 2
weeks.
Group IV: PN extract + DOX (Phyllanthus Niruri extract + Doxorubicin treated):
Extract will be pretreated for 2 weeks and then alternatively with Doxorubicin
injection (for a cumulative dose of 15mg/kg body weight) for next 2weeks.
Control as well as treated animals shall be observed for a period of 3 weeks
after the last injection for the general appearance, behaviour and mortality. At the
end of 3 weeks post treatment period, animals will be assessed for described
parameters. The animal will be killed under ether anesthesia and a midline
abdominal incision will be performed and heart tissue will be quickly dissected
out, washed in ice cold saline, dried on filter paper and weighed. A 30% w/v heart
tissue homogenate will be prepared in 0.9% buffered potassium chloride (pH-7.4)
for the estimation of glutathione and malondialdehyde. The remaining portion of
the heart tissue will be used for the assay of cardiac damage enzyme markers
(AST, ALT, ALP)12 and antioxidant enzymes (SOD, CAT)13.
The result obtained from the present study shall be analysed by one way
ANOVA followed by Dunnet’s ‘t’ test.
7.3 Does the study require any investigation or interventions to be conducted
on patients or other humans /animals? If so please described briefly.
Yes, the above study requires investigation to be done on the albino rats of
Wistar strain for the determination of myocardial toxicity.
7.4 Has animal ethical committee clearance been obtained from your
institution in case?
The study has been referred to the ethical committee of the institution for
clearance.
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References:
1. Singal PK, Li T, Kumar D, Danelisen I, Iiskovic N. Adriamycin-induced
heart failure: Mechanisms and modulations. Mol. Cell Biochem 207:77-85
2000
2. Karim S, Bhandari U, Kumar H, Salam A, Siddiqui MAA, Pillai KK. Dox
induced cardiotoxicity and its modulation by drugs. Ind J of Pharmacol
2001: 3: 203-207
3. Rajaprabhu D,Rajesh R, Jeyakumar R, Buddhan S, Ganesan B, Anandan R.
Protective effect of Piccrorhiza kurroa on antioxidant defense status in
Adriamycin- induced cardiomyopathy in rats J of Medicinal Plant Research
Vol. 1(4) 2007: 080-085.
4.
Chularojmontri Linda, Suvara Kimnite, Wattanapitayakul, Angkana
Herunsalee, Suphan Charuchongkolgwongse, Somchit Niumsakul, Supatra
Srichairat. Antioxidant and cardioprotective effects of phyllanthus Niruri l.
on Dox- induced cardiotoxicity. Biol. Pharm. Bull 2005; 28(7) :1165-1171
5.
Gnanapragasam A. Yogeeta S, Subhashini R, Ebenezar KK, Satish V,
Devaki T. Adriamycin induced myocardial failure in rats: Protective role of
Centella Asiatica. Mol. And cell. Biochem. 294,2007: 55-63
6.
Than NN, Fotso S, Poeggeler B, Hardeland R, Laatsch H. Niruriflavone, a
new antioxidant Flavone Sulfonic acid from phyllanthus Niruri.
Z. Naturforsch. 61b, 2006: 1-4.
7.
Nwanjo HU, Oze G, Okafor MC, Nwosu D, Nwankpa P.Protective role of
phyllanthus Niruri extract on serum lipid profiles and oxidative stress in
hepatocytes of diabetic rats. African J Biotech.2007; 6(15): 1744-1749
8.
The Wealth of India Vol. III. 2003: 34-35.
9.
Unander DW, Venkateswaran PS, Millman I, Byran HH, Blumberg BS.
1990. Phyllanthus species: Sources of new antiviral compounds. 518-521.
in: J. Janick and JE Simon (eds), Adv. in new crops. Timber press,
Portland.
10. Kirtikar KR, Basu BD. Indian medicinal plants. Vol III.1988. India. Lalit
Mohan Basu Allahabad:2225-6
11. Outomuro D, Grana DR, Azato F, Milei J, Adriamycin- induced
myocardial toxicity: new solutions for an old problem. Int J Cardio 2006;
08470:1-10
12. Deepa PR, Veralakshmi P. Protective effect of low molecular weight
heparin on oxidative injury and cellular abnormalities in Adriamycininduced cardiac and hepatic toxicity. Chemico Biol Int 2003; 146:201-210.
13. Balaram R, Pallavi A, Kolhapure SA. Antioxidant activity of DHC-1, a
herbal formulation. J Ethno Pharmacol 2004; 94: 135-141.