issn: 2454-1362, evaluation of histopathological studies ... · · 2016-10-27evaluation of...

Imperial Journal of Interdisciplinary Research (IJIR) Vol-2, Issue-11, 2016 ISSN: 2454-1362, http://www.onlinejournal.in

Imperial Journal of Interdisciplinary Research (IJIR) 940

Evaluation of Histopathological studies of Achyranthes aspera in mice, Swiss albino

Dr. K. Geetha

Department of Botany With Specialization In Plant Biotechnology The Standard Fireworks Rajaratnam College For Women, Sivakasi, Tmailnadu, India.

(An Autonomous Institution affiliated to Madurai Kamaraj University, accredited with ‘A’ Grade by NAAC)

Abstract: Medicinal plants are of great importance to the health of individuals and communities in general. The medicinal value of plants lies in some chemical substances that produce a definite physiological action on the human body. Achyranthes aspera L. (Amaranthaceae) is an indigenous medicinal plant of Asia, South America, and Africa. It is commonly used by traditional healers for the treatment of fever, especially malarial fever, dysentery, asthma, hypertension and diabetes.This paper explains evidence based-information regarding the histopathological activity of ethanol extract of Achyranthes aspera with alloxanised mice. A control group of mice with a number of six for each treatment were maintained. Mice were exposed to alloxan and then the alloxanised mice were administered with different extracts of A.aspera at a sublethal concentration of 12.5g/kg for a period of 30 days. After the exposure period the various tissues namely liver, spleen and muscle were taken from mice maintained as control, alloxanised mice and the lone effect of various extracts (hexane, butanol, ethanol, chloroform and water) of A.aspera on the alloxanised mice were processed for histological studies.Among the various extracts namely hexane, butanol, ethanol, chloroform and water in the extraction of A.aspera, the ethanol extract exhibited a protective effect on the histology of liver, spleen and muscle of alloxanised mice. The hepatoprotective nature of ethanol extract of A.aspera as evidenced through the histological studies can be substantiated with the findings of marked restoration in the level of liver enzymes AST, ALT and ALP in alloxanised mice after administering with plant extract in the present study.

1. INTRODUCTION Good health is a treasure that means it has

value. When a nation is healthy, it saves more than it spends and the standard of living is high. However, when a nation is not enjoying good health, this kind of nation experiences a low level of education and political and economic problems. Histopathological alteration can be used as indicator for the effect of chemicals on organisms. Study on Histology appears

to be a very sensitive parameter and is crucial in determining cellular changes that may occur in target organs (Priya Kannappan, 2009).

The liver is an important organ performing vital functions including bio transformation, migration of lipids, glycogen storage and release of glucose. Liver impairment is one of the leading causes of death in diabetes mellitus. The mortality rate due to hepatic dysfunction is greater than that of the cardiovascular complications. Liver, an insulin-dependent organ, plays a vital role in glucose and lipid homeostasis. It participates in the uptake, oxidation and metabolic conversion of free fatty acids and in the synthesis of cholesterol, phospholipids and triglycerides (Brixova, 1981; Moller, 2001). Spleen is the secondary lymphoid organ containing about one-fourth of the body’s lymphocytes and initiates immune responses to blood-borne antigens (Kuper et al., 2002; Nolte et al., 2002; Balogh et al., 2004). It brings about humoral and cell mediated immunity. Muscles are involved in contraction and relaxation and is the tissue receiving the major portion of blood supply. It is important that the therapeutic value of any isolated plant extract depends not only on its pharmacological potency but also on its lack of toxicity. This is important in the case of hypoglycemic drugs preparations. Considering this in mind, the influence of various extracts of A.aspera on the histology of mice maintained as control and the impact of extracts on the recovering or repairing the histological alterations caused by alloxan in mice were investigated. 2. MATERIALS AND METHODS A control group of mice with a number of six for each treatment were maintained. Mice were exposed to alloxan and then the alloxanised mice were administered with different extracts of A.aspera at a sublethal concentration of 12.5g/kg for a period of 30 days. After the exposure period the various tissues namely liver, spleen and muscle were taken from mice maintained as control, alloxanised mice and the

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Imperial Journal of Interdisciplinary Research (IJIR) Page 1941

lone effect of various extracts (hexane, butanol, ethanol, chloroform and water) of A.aspera on the alloxanised mice were processed for histological studies. The collected specimen of the various organs from mice treated with different extracts of A.apera were fixed in 10% buffered formalin solution then processed for histopathological studies in ascending grades of ethylalcohol, cleared in xylene, then embedded in paraffin wax. Sections of about 5-7 microns thickness were taken and then stained with hematoxylin and eosin (Drury and Wallington, 1967). The tissues were examined by a Nikon E600 light microscope and photograpthed by a Nikon DXM 1200 digital camera (Humason, 1972 and Bancrift et al., 1996). 3. RESULTS AND DISCUSSION In the present study, histology of liver, spleen and muscle tissues of the mice maintained as control and the changes caused in the said tissues of mice treated with alloxan were observed. The amelioration effect of plant extract on the normal well being of the histology of the liver , spleen and muscle were also noticed. LIVER The liver of mice maintained as control exhibited no pathological abnormalities (Plate - 1). Liver of alloxanised mice showed more hepatocytes with cytoplasmic vacuolar degeneration and pyknotic nuclei (Plate - 2) as observed in rat due to actominophen (Raj Kapoor et al., 2008). Mice treated with alloxan in combination with hexane extract of A.aspera showed a few hepatocytes, and fragmented nuclei (Plate - 3). Alloxanised mice administered with butanol extract of A.aspera showed disorganized hepatic cords and many hepatocytes with cytoplasmic vacuolization (Plate - 4) . In case of alloxanised mice treated with ethanol extract of A.aspera no inflammation and fibrosis were observed (Plate - 5). Further, regeneration of the histology of liver was evident due to the influence of ethanol extract of A.aspera in the alloxanised mice. Similar results were reported by Karthick et al., 2011 in alloxanised rat with ethanol extract of A.aspera and with different extract of Citrullus colocynthis by Mohammed et al., 2010. Liver of alloxanised mice after treating with chloroform extract of A.aspera showed cell infiltration and hepatocytes with cytoplasmic vacuolar degeneration and pyknotic nuclei. (Plate - 6). In alloxanised mice treated with water extract of A.aspera showed prominent nuclei (Plate - 7).Similar findings were reported by Badakhshan, 2011 in alloxanised mice due to Lantana camera. It is obvious from the present study that the ethanol extract of A.aspera has a telling effect on the

repairing of liver abnormalities that observed in mice exposed to alloxan. The amelioration effect of the A.aspera indicated its hepatoprotective nature. The hepatoprotectiveness of A.aspera observed in the study may be due to the presence of phytochemicals such as flavonoids (quercetin), saponins (oleanolic acid), hydroquinone and aminoacids etc. It is also to be considered that the phytochemicals present in the ethanol extract of A.aspera are not only hepatoprotective but also non-toxic to mice. Similar conclusion was arrived in respect to non-toxic response of flavonoid content of Artocarpus heterophyllus by Chandrika et al., 2006. SPLEEN

No histological alteration were observed in the spleen of mice maintained as control (Plate - 8). In alloxanised mice , cytoplasmic vacuolation, nuclear hypertrophy and degeneration of structural integrity of spleen were noticed (Plate - 9). Nuclear hypertrophy and infiltration of neutrophil were observed in alloxanised mice treated with various extracts of A.aspera in general (Plate - 10,11, 13 and 14). However, in alloxanised mice after exposing them to ethanol extract of A.aspera a tendency towards normal histological features of spleen was observed. (Plate - 12). The histology of spleen was affected to some extent by various treatments. Whereas the administration of ethanol extract of A.aspera to the alloxanised mice the integrity of spleen was found be normal as that of mice maintained as control. As mentioned in the introduction, the spleen is the sight of lymphoid organ and responsible for humoral and cell mediated immunity. In the present study the histology of spleen was protected in alloxanised mice particularly due to ethanol extract of A.aspera. It can be attributed to the immune potentiality of ethanol extract of A.aspera. This finding can be further substantiated by the result observed on the enhanced immunological response of cancer induced mice after treating with ethanol extract of A.aspera.

MUSCLE

No histological variation was observed in the muscle of mice maintained as control (Plate - 15). In muscle of alloxanised mice, severe infiltration of neutrophil was noticed (Plate - 16). No much variation was observed in muscle of alloxanised mice after individually treated with hexane and butanol extracts of A.aspera (Plate - 17 and 18). Muscle of alloxanised mice with ethanol extract of A.aspera a marked reduction in nuclear infiltration was seen (Plate - 19). Muscular atrophy was the result in alloxanised mice treated with chloroform extract of A.aspera (Plate - 20). In alloxanised mice after treating with water extract of A.aspera the muscle showed a reduction in neutrophil infiltration (Plate -





21). Similar observations were noticed by Manvi et al., 2010 in rat due to Annona squamosa. Plate 1 – 7 Histology of liver of Control mice and the

various histopathological abnormalities due to alloxan and different extracts of A.aspera

Plate 1 – Control

Plate 2 – Alloxan treated

Plate 3. Alloxan + Hexane extract of A.aspera

Plate 4. Alloxan + Butanol

extract of A.aspera

Plate 5. Alloxan + Ethanol extract of A.aspera

Plate 6. Alloxan + Chloroform

extract of A.aspera

Plate 7. Alloxan + Water

extract of A.aspera

Plate 8 – 14 Histology of spleen of Control mice and the various histopathological variations due to

alloxan and different extracts of A.aspera Plate 8 – Control







11. Alloxan + Butanol extract of A.aspera

Plate 12. Alloxan + Ethanol extract of A.aspera


extract of A.aspera

Plate 14. Alloxan + Water

extract of A.aspera

Plate 15 – 21 Histology of Muscle of Control mice

and the various histopathological changes due to alloxan and different plant extracts

Plate 15 – Control







Plate18. Alloxan + Butanol extract of A.aspera

Plate 19. Alloxan + Ethanol

extract of A.aspera


extract of A.aspera

Plate 21 Alloxan + Water extract of A.aspera

4. CONCLUSION Medicinal plants are of great importance to the health of individuals and communities in general. The medicinal value of plants lies in some chemical substances that produce a definite physiological action on the human body. From the result of the present study it is obvious that the alloxanised mice showed histological alterations at various extents in the liver in particular, and also in spleen and muscle. Regarding the amelioration efficiency of various extracts of A.aspera, the ethanol extract of A.aspera is found to have better effect. Further, the ethanol extract of A.aspera is non-toxic to mice at this concentration and also helps to recover the damage caused by alloxan in the histological features of said tissues. 5. REFERENCE: [1] Badakshan Mahdi Pour, Lachimanan Yoga Latha and

Sreenivasan Sasidharan. 2011. Cytotoxicity and Oral Acute Toxicity Studies of Lantana camara Leaf extract. Molecules. 16: 3663-3674.

[2] Bancrift, J., Steven, A. and D. Turner. 1996. Theory and practice of histological Techniques.4th (Ed). Churchill living stone, London, ISBN. 10:0443064350.

[3] Chandrika, U.G, Wedage, W.S., Wickramasinghe, S.M.D.N. and W.S. Fernando. 2006. Hypoglycaemic action of the flavonoid fraction of Artocarpus heterophyllus leaf. Afr.J.Trad CAM. 3(2): 42-50.

[4] Drury, R.A.B and E.A. Wallington. 1967. Carleton’s Histological Technique (Fourth Edition). Oxford University Press. 432 pp.

[5] Humason, G.L. 1972. Animal tissue techniques 3rd ed., W.H. Free man and co., San Francisco. ISBN: 9780716706922.

[6] Karthick, R., Rao, C.H., Trivedi, V., Pushpangadan, S. P. and G.D. Reddy. 2011. Amelioration effects against N-nitrosodiethylamine and CCl4 induced hepatocarcinogenesis in Swiss albino rats by whole plant extractbof Achyranthas aspera . Indian Journal of Pharmacology. 42(6): 370-375.

[7] Manvi, F.V., Nankawade, B.K. and Sanjiv Shing, 2011. Pharmological screening of combined extract of Annona Squamosa and Nigella sativa, International Journal of Pharma and bio Sciences. 2(2): 520-529.





[8] Mohammad Khalil, Gamal Mohamed, Mohammad Dallak, Fahaid Al-Hashem, Hussein Sakr, Refaat A.Eid, Mohamed A. Adly, Mahmoud Al-Khateeb, Saleh Banihani, Zuhair Hassan and Nabil Bashir. 2010. The effect of Citrullus colocynthis pulp extract on the liver of diabetic rats a light and scanning electron microscopic study. American Journal of Biochemistry and Biotechnology. 6(3): 155-163.

[9] Priya Kannappan and Krishnakumari Shanmuga Sundaram. 2009. Toxicity assessment of the medicinal plant Cyathuia prostrata. Journal of Applied Biosciences. 13: 681-687.

[10] Rajkapoor, B., Venugopal, Y., Anbu, J., Harikrishnan, N., Gobinath, M. and V. Ravichandran. 2008. Protective effect of Phyllanthus poluphyllus on Acetaminophen induced hepatotoxicity in rats. Pak.J.Pharm. 21(1): 57-62.



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