BIOLOGIA (PAKISTAN) 2014, 60 (1), 135-138
PKISSN 0006 – 3096 (Print)
ISSN 2313 – 206X (On-Line)
Histological comparison of natural lung injury in Rattus norvegicus induced by a
natural herb (Nerium oleander) and a known carcinogen (thioacetamide)
MUDDASIR HASSAN ABBASI1, SANA FATIMA2 AND *NADEEM SHEIKH3
1
Department of Zoology, Govt. College of Science, Wahdat Road, Lahore
2
University of Health Sciences, Lahore.
3
Cell and Molecular Biology Lab, Department of Zoology, University of the Punjab, Lahore.
ABSTRACT
Plant extracts have extensively been studied to look for the treatment of various diseases but their
poisoning effects have also been reported. The current study was undertaken with the objective of comparing
acute toxic effect of Thioacetamide (TAA), a proven toxicant and Nerium oleander (N. oleander), a herb reported
to have multiple therapeutic properties including anti-inflammatory, antifungal, antibacterial and antioxidative
effects using histological parameters. Animals in group A were treated with 10 ml/kg of aqueous decoction of
leaves of the plant while 300mg/kg of TAA was administered in group B and control (Con.) did not receive any
treatment (n=3). Histological alterations in the lungs were then studied after 6 hours of respective treatment.
There was an onset of more or less similar destruction of the tissue in both the experimental groups. Disruption
of bronchus mucosal folds and alveolar cells along with significant nodule like accumulation of macrophages and
mononuclear cells around arteriole were noticed. These results indicated that use of ethno-medicines, like N.
oleander caused similar histological alterations in lungs as that of known carcinogen TAA. Thus, application of
N. oleander may pose a serious risk for health.
Keywords. Lung, Nerium oleander, Thioacetamide.
____________________________________________________________________________________
INTRODUCTION
Plants are the largest source of herbal
medicines worldwide and these have the potential to
be both therapeutic and harmful but still masses of
population rely on such remedies.
Nerium oleander, (N. oleander), (Apocynaceae) an
ornamental shrub commonly known as “Kaner”, has
been used as a traditional medicine and is reputed
to have wide spectrum of bioactivities including
pharmaceutical products (Anvirzel), molluscicide,
rodenticide and insecticides (Wang et al., 2000;
Adome et al., 2004; Turan et al., 2006; Derwich et
al., 2010). But poisoning action of this plant has also
been reported with significant time-dependent
cytotoxic effects in cattle, goat, man and other
experimental animal models (Langford & Boor,
1996; Aslani & Rezakhani, 2000; Hughes et al.,
2002; Smith et al., 2003; Barbosa et al., 2008;
Abbasi et al., 2013b). Animals exposed to the plant
are often found suddenly dead owing to cardiac
dysfunction. Oral doses of N. oleander proved fatal
to animals within 1 to 24 hours with clinical toxicosis
(Aslani et al., 2007).
Thioacetamide (TAA) a proven carcinogen
has been extensively reported to produce toxic
effects in animals and is suitable for studying and
comparing the histological alterations (Abbasi et al.,
2013a).
*Corresponding author: [email protected]
The aim of this study was to compare acute
toxic effect of TAA and N. oleander produced
histological variations in lungs of albino rats and to
find out whether the injury caused by TAA
resembles with N. oleander induced injury.
MATERIALS AND METHODS
Animals
Male wistar rats of about 200g body
weights, were kept in a well ventilated hygienic
animal house under standard conditions with 12-h
light/dark cycles and access to fresh water and
food pellets ad libitum. All the animals were
acclimatized for a period of 2 weeks before the
experiment.
Plant Material
Fresh leaves of N. oleander were collected
from Govt. College of Science (GCS), Lahore,
Pakistan. The plant material was identified by
comparing with the reference collection available in
the Herbarium of GCS, Lahore, Pakistan and had
been preserved in Department of Botany, GCS for
further reference.
Extraction protocol
Air-dried leaves were processed according
to the method of Rashan et al., 2011, with
modifications. Briefly, the leaves decoction was
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M. HUSSAN ET AL
BIOLOGIA (PAKISTAN)
prepared by boiling in 0.9 % NaCl solution (1:1, w/v)
for 3h by steam distillation. The decoction was
filtered and used for the experimental animals.
Experimental design
The rats were grouped into three viz. A, B
and control (Con.) (n=3); A: was injected
intramuscularly (i.m) with 10 ml/kg of N. oleander
leaves decoction, B:
300mg/kg of TAA was
administered intraperitoneally (i.p) and Con. did not
receive any treatment. After 6h of administration, all
the animals were anesthetized with i.p injection of
ketamine–distilled water mixture (1:1).
Tissue Sampling and Histological Examination
Lungs of all the animals were excised,
immediately after sacrifice, rinsed with physiological
sodium saline and portion was fixed in 10% formalin
for histological studies. The fixed tissue specimens
were sectioned by standard methods and stained
with Hematoxylin and Eosin (H & E) for detecting
the degree of damage.
Fig., 1: Hematoxylin–Eosin (H&E) staining of lung
section of control (Con.; a= 100x &b=400x). The
normal architecture of section showing numerous
clear alveoli (A) and alveolar sacs (AS). Bronchus
(Br) with mucosal folds in a close proximity to blood
vessel can also be seen in the section.
Microscopic Analyses
To assess organ damage, microscopic
images were captured using a trinocular IRMECOGmbH model 1M-910, 21493 Schwarzenbek/
Germany displayed on a computer via a Scope
®
Tek (scope photo 3.0).
RESULTS AND DISCUSSION
The present study investigated the
histopathological changes in the lung tissues of
Wistar rats that were treated with N. oleander
leaves extract and TAA separately. Since lung is a
target organ of many toxicants, an understanding of
pulmonary architecture in health and disease is of
paramount importance.
H&E stained lung sections of control group
showed normal histological architecture; numerous
clear alveoli (A) with thin inter-alveolar septa and
alveolar sacs (AS). A bronchus (Br) with mucosal
folds in a close proximity to blood vessel can also
be seen in the section (Fig., 1).
Alterations in the pulmonary architecture
were observed in group A with evident disruption of
bronchus mucosal folds. Alveolar cells with extreme
widening of lumen of the bronchiole and significant
signs of vascular lesions were also noticed (Fig., 2).
Fig., 2: Section of lung tissue stained with H&E
compared after 6h of N. oleander (A) and TAA (B)
administration in wistar rats. Widening of lumen of
the bronchiole with significant signs of vascular
lesions is evident in group A while massive cell
infiltration in sections of the TAA treated animals is
shown by black arrow (x100).
The damage observed might be due to an
oxidative stress produced by toxicants, oleanderin
and neriine, cardiac glycosides (cardenolides) which
can be isolated from all parts of the N.oleander and
are similar to the toxin in foxglove (Digitalis) (AlFarwachi et al., 2008).These toxins may lead to an
imbalance in the production/consumption level of
reactive oxygen species (ROS).
There was an onset of more or less similar
destruction of the tissue in group B with the damage
of alveolar cells along with significant nodule like
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HISTOLOGICAL COMPARISON OF NATURAL LUNG INJURY
accumulation of macrophages and mononuclear
cells around arteriole (Fig., 2B). It may be explained
that during inflammatory response, accumulation of
macrophages occurs which ultimately release some
chemotactic factors that attract neutrophils which is
evident in the sections. Influx of inflammatory cells
plays a major role in increasing the permeability of
alveolar capillary barrier providing the cellular
toxicity.
Neutrophils
together
with
other
inflammatory cells must frequently found in the
broncho-alveolar region in the current study. Sheikh
& Javed, 2009 observed similar changes along with
pulmonary edema in lung sections of swiss albino
mice, exposed to 0.5% dilution of cypermethrin in an
inhalation chamber. Similar histopathological
changes in the sections of lungs were observed
during Fenvalerate (Fen), a synthetic pyrethroid
exposed rats with influx of mononuclear cells
admixed with a few giant cells in alveolar lumen
(Mani et al., 2001). More pronounced histological
changes were noted in pesticide mixture inhaled
rats for consecutive 28 days (Noaishi et al., 2013).
CONCLUSION
137
oleander aqueous leaf extract in rabbits.
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