Biodiversity and its Conservation and Environmental Impact on Global Waming


EFFECTS OF Euphorbia antiquorum latex ON HISTOLOGICAL AND
BIOCHEMICAL CHANGES IN FEMALE ALBINO RATS
R.DHANAPAL & P.MURUGAIAN
Biomedicine and Computational Biology Lab, PG and Research Department of
Zoology,Rajah Serfoji Govt. College (Autonomous), Thanjavur-613 005.

C.KANDEEPAN
PG and Research Department of Zoology, A.P.A.College of Arts and Culture,
Palani-624601.

Abstract
The present study the influence of the Euphorbia antiquorum latex of
histological and biochemical changes on female albino rat. The test rats were
maintained 0.2ml of latex/body weight organs treated for 48 days. Creatinine, total
protein, albumin, globulin, total lipids, total cholesterol, calcium and glucose were
induced significant elevations of serum. However, the latex produced significant
alterations in albumin, globulin, cholesterol, calcium and glucose then the treated
animals of the latex treated animals showed a decrease in the organ weight and body
weight ratio compared to the control animals. The result concluded that latex of
Euphorbia antiquorum toxicological effect of the liver, kidney, spleen were changed
in architecture of cell arrangement of compared to control.
Keywords: Euphorbia antiquorum latex, toxic effect.

Introduction
The pharmaceutical imbalance between remedies that protect the
liver and have antioxidant capacity and drugs that induced hepatotoxicity has
promoted and accelerates search into plants used in folk medicine to treat
liver diseases and increases liver functions (Chatterjee, 2000). The growing
interest in herbal medicine therefore demands toxicity risk assessment of the
various indigenous preparations used in the treatment of diseases (Yakubu et
al., 2005). Stress reduces blood parameters such as haematocrit,
haemoglobin, erythrocyte count but increased leucocytes count (Gbore et al.,
2006). Circulating levels of alanine transaminase (ALT) and aspartate
transaminase (AST) increase under psychological and toxic stress; reflecting

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liver injury. This suggests that chronic stress relates to hepatic damage
(Roland et al., 2009). The toxicants and pathological conditions may produce
oxidative stress and induce damage to the liver, kidney and brain tissue (Yu
et al., 2008; El-Gendy et al., 2010).
The plant Euphorbia antiquorum Linn. belongs to the family of
Euphorbiaceae. It is a small fleshy shrub or small tree distributed throughout
the hotter parts of India and Sri Lanka. Previous studies reported that the
latex of plant has hepatoprotective, antioxidative and insecticidal property.
Studies also reported that the plant contains terpenoids which posses anti-
inflammatory and anti-arthritic property (Silva et al., 2008; Jyothi et al.,
2007; Anand Nirmal Harpalani et al., 2011).
Material and Methods
Plant Euphorbia antiquorum fresh (Latex) was collected from
Thirumangalakkottai Keelaiyur, Thanjavur District, in Tamil Nadu.
Wistar strain albino female rats 90 days old 160-190gm weight was
used in this study. An animal was kept in standard conditions. The diet pellet
is standard (Sai Enterprisei, Chennai) and provided ad libitum normal water.
Experimental Design
These female albino rats were divided into two groups (each group 4
rats) the first group- I is control group normal water and feed received daily
another group- II daily dose of 0.1 ml/kg/body weight of latex and group-III
dose of 0.2 ml/kg/body weight of latex is all the groups were orally
administrated for 48 days.
Biochemical Tests
The blood samples of different categories were subjected to different
biochemical tests in order to determine any biochemical change in treated
female rat blood serum. The following biochemical tests were performed for
this purpose: Serum creatinine, total protein, albumin, globulin, total lipid,
total cholesterol, calcium, glucose, andalkaline phosphatase, HDL and LDL
were estimated by standard kit method (Randox, United Kingtom).

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Biodiversity and its Conservation and Environmental Impact on Global Waming

Histopathological Examinations
Spleen, liver and kidney of the treated rats were taken and fixed in 10
% neutral formalin solution. The fixed specimens were then trimmed, washed
and dehydrated in ascending grades of alcohol. These specimens were cleared
in xylene, embedded in paraffin, sectioned at 4-6 microns thickness and
stained with Hematoxylen and Eosin (H &E) then examined microscopically
according to the method described by (Luna 1968).
Results
Liver
The liver histology of the group II and III rats confirmed the
occurrence of liver cirrhosis with disorganization of the normal lobular
pattern accompanied by the formation of well-defined pseudolobules (Fig.5,
6). Thick fibrous connective tissue septa were seen bridging the portal areas
and frequently extending into the lobules (Fig.7, 8) when compare to control
(Fig.1, 2,3,4).
Kidney
Histological study of the normal kidney of the nondiabetic rats
revealed normal glomerulus surrounded (Fig. 9,10,11,12) by the Bowman’s
capsule, proximal and distal convoluted tubules without any inflammatory
changes. The kidneys of untreated diabetic rats showed degenerated
glomeruli infiltrated by the inflammatory cells and thickening of the
basement membrane. The proximal convoluted tubule exhibited edematous
changes with deposition of mucopolysaccharide and hyaline substances. All
the necrotic changes observed in the proximal and distal convoluted tubules
along with the deposits were found to be absent in the diabetic rats treated
with the latex extract. The group that was treated with latex extract showed
features of healing i.e. normal glomerulus, absence of inflammatory cells,
normal basement membrane and capillaries, decrease in the
mucopolysaccharide and hyaline deposit, respectively. The tissue necrosis
was also observed to decrease in the group treated with latex extract
(Fig.13,14,15).

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Spleen
We can take notice that at latex of E. antiquorum treated rats both
alkaline phosphatase and BCE amount are a bit decreased in comparison with
the control (Table- 1) and (Table-2). The results of our study reveal a normal
structure of white and red pulps within the spleen of rats belonging to the
control group (Fig.16,17). The spleen follicles are big, centered on arterioles
and their majority represents a well defined germinative center. The splenic
sinuses are well loaded with red blood cells and lymphoid cells. The splenic
belts (Billroth) are well evidenced and cells density that makes them up is
found under normal limits. We find out very rare isolated megakaryocytic on
the section surface. In the second test group we observed that the spleen
structure is serious changed. Comparing the second group with control we
remark the following: most spleen follicles are almost equal in size with those
of the control, even if they have a severe shifted structures; the density of the
cells is much reduced in spleen follicles and none of these display a
germinative center; a high number of lymphocytes have in their structure
necrobiosis various stages ; the loading of the sinuses with erythrocytes and
lymphoid cells is very weak; the spleen belts are very thin and there are much
less cells in their structure; at the electron microscopy examination there is a
severe cells number reduction in those rats spleen where Cyclophosphamide
was applied. Also we can observe the wide intercellular occurrence (Fig. 18).
Concerning the third test group we have the following remarks: the action of
the cancer drug seems to stop; the spleen is in a full process of structural
recovery, at the level of both the red and the white pulp; at the level of the
white pulp, the spleen follicles are still smaller; they have a relatively
homogenous aspect, that is they are drawn on by entire young cells even if
they do not display a germinative center; in the red pulp, the spleen sinuses
are relatively well loaded with red blood cells and lymphoid cells; the spleen
belts are well represented, they are thicker and contain a great deal of cells
featured with emphasized polymorphism; there are numeorus
megakaryocytes all over the section surface (Fig. 19) ; many granulocytes,
neutrophils and eosinophils have polymorphous structure, and they can be
seen in layers (Fig. 20) ; numerous limphoblasts in various stages of
evolution.
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Biodiversity and its Conservation and Environmental Impact on Global Waming

Biochemical Analysis
The Latex of E.antiquorum had effects on total protein, albumin,
ALT activity, sodium and calcium levels in the serum. But all the
concentrations investigated caused significant reduction in total bilirubin and
globulin. The latex caused significant increase in serum level of conjugated
bilirubin at 0.1ml/kg body weight and 0.2ml/kg body weight of the animals
compared with that in the control group. The administration of the latex also
led to reduction in 0.1ml/kg body weight and 0.2ml/kg body weight, the
serum level of ALP increased significantly. The latex exhibit any significant
effect on the serum level of GGT but led to a significant reduction in this
parameter. The extract caused a significant increase in AST activity
parameter reduced significantly (Table- 3). The extract at all the doses
investigated led to a significant decrease in the serum level of cholesterol,
triacylglycerol, HDL-C and LDL-C. Administration of the extract also caused
significant reduction in artherogenic index of all animals but the parameter
compared with that in the control group.
Table.1 Euphorbia antiquorum latex effect of animal weight (SD± SE, n=10)
Experiment Initial weight Final Gain of weight(gm)
(gm) weight(gm)
Control 4.32±2.16 6.54±3.27 2.07±0.30
0.1ml Treatment 7.59±2.40 8.16±2.58 2.68±0.84
0.2ml Treatment 9.66±2.62 8.25±2.59 2.72±0.89

Table.2 Euphorbia antiquorum latex effect of organs weight (SD± SE, n=10)

Experiment Spleen(mg) Kidney(mg) Liver(mg)

Control 0.57±0.21 1.23±0.19 1.87±0.21
0.1ml Treatment 0.43±0.13 0.96±0.30 1.01±0.31
0.2ml Treatment 0.39±0.11 0.91±0.27 0.96±0.29

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Table .3 Biochemical changes of Euphorbia antiquroum latex treated animal

Experiment Total Albumin Globulin Creatinine Total HDL LDL Triglycerides
protein (mg/dl) (mg/dl) (mg/dl) cholesterol Cholesterol Cholesterol (mg/dl)
(mg/dl) (mg/dl) (mg/dl) (mg/dl)

Control 8.82±1.61 4.42±0.54 4.37±0.33 1.06±0.21 72.58±5.78 138.24±10.95 1.66±0.33 4.23±0.23

0.1ml
Treatment 2.68±0.68 1.10±0.38 1.52±0.48 1.07±0.33 25.31±8.01 10.01±3.16 0.75±0.23 5.43±1.71
0.2ml 2.63±0.62 1.06±0.33 1.49±0.43 1.02±0.29 25.26±8.09 9.84±3.09 0.71±0.19 5.39±1.67
Treatment

ϲ
ϭ Ϯ ϱ ϲ

ϯ ϰ ϳ ϴ

ϭ
ϵ
ϵ ϭϬ

ϭϮ
ϭϭ

ϭϯ ϭϲ ϭϳ

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Biodiversity and its Conservation and Environmental Impact on Global Waming


ϭϴ ϭϵ
ϭϰ

ϭϱ ϮϬ

1. Shows liver control

2. Shows enlarged portion of hepatocytes
3.&4. Shows enlarged portion with hepatocytes
5. Shows the treated liver
6. Shows degranulated blood vessles and hepatocytes
7.&8. Shows shrinkage nucleus of hepatocytes
9. Shows control kidney
10. Shows the treated kidney
11.&12. Shows granulated disorganization of glomerulus
13. Shows Bowmam’s capsule
14.&15. Shows the granulocytes disorganized
16. Shows control spleen
17. Shows the treated spleen disorganized
18.&19.Shows the inner nucleus disorganized
20.Shows the enlarged of gland cell wall dislocated nucleus

Discussion
After the last given dose, the histopathological examination of the
spleen indicates the the cancer drug has a visible effect on the lymphocyte
populations, both to red and white pulp, and most of them were found out in

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different stages of apoptosis or necrobiosis. According to the previous

studies, all cytostatic, including CFA, can block cell division that is grounded
on tissue development and substitution, especially in germinal ones (Ardelean
et al., 2000). Similar results were obtained in vivo experiments when
cyclophosphamide decreased almost all of peripheral blood cell counts and
lymphocyte subset counts in the thymus and spleen at all ages (Blankenberg
et al., 2001; Michuyuki, 2007). The presence of many dividing cells, at white
and red pulps shows the same thing. The unusual polymorphisms of the cells
in red pup belts are more numerous as compared to the control. That suggests
the body direct involvement to contribute to qualitative and quantitative
recovery of populations treated with Cyclophosphamide. Many
megakaryocytes with a marked polymorphism are present in the whole
section surface. It means the beginning at the organ level, of a compensatory
hematopoesis process for a short time. Hematopoiesis reactivation is also
present in the spleen and liver of the adult mice after giving them anti-
inflammatory compounds. This phenomenon was noticed in case report also,
in the case of human patients after the granulocyte colony-stimulating factor
was given (Litam et al, 1999).
These enzymes can be used to assess damage to the liver (cytolysis)
and heart. The non significant effect of the extract on the serum ALT and no
specific pattern of effect on AST could suggest partial disruption of the
plasma membrane of the organs of the animals and this may not have
consequential effect on the metabolism and regulation of certain enzymes in
the liver. Our findings agree with earlier reports of (Adeneye 2008; Oduola et
al., 2007) where M. lucida leaf extracts at 2000 and 3200 mg/kg body
weights were neither lethal, hepatotoxic nor nephrotoxic in experimental
animals. The concentration of total proteins, bilirubin and albumin in the
serum may indicate the state of the liver and the type of damage. The fact that
the extract did not exhibit any significant effect on the total protein, albumin
and globulin further confirms that the extract is likely not hepatotoxic, while
the reduction in the serum levels of total bilirubin and fluctuation in the
conjugated bilirubin could be attributed to adaptation of the animals to the
physiologic changes exerted by the extract.

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Biodiversity and its Conservation and Environmental Impact on Global Waming

Disruption or alteration in the concentration of major lipids such as
cholesterol, HDL - C, LDL-C and triacyglycerol can give useful information
on lipid metabolism as well as predisposition of the heart to atherosclerosis
and its associated coronary heart diseases. Triacyglycerol, LDL - C and HDL
- C are associated with lipolysis, carrier of plasma cholesterol and
atherosclerotic tendency, respectively. The reduction in the serum levels of
cholesterol, triacyglycerol, LDL - C and HDL - C at all the doses investigated
could suggest that the extract at 50, 100, 200 and 300 mg/kg body weight
may not predispose the animals to cardiovascular risk. This is supported by
the reduction in the computed atherogenic index, a useful indicator of
cardiovascular diseases. The significant reduction in the lipid profile could
also mean that the extract possesses anti - lipidemic potential. The plant could
be employed in the management of ailments associated with
hypercholesterolemia.

Conclusion
The histopathological examination of latex of E. antiquorum
administration indicates the latex lead to spleen, kidney, liver tissues damages
and biochemical changes of albino rats. A further study of E. antiquorum is
necessary to fully establish its safety profile.
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