Parveen et al                                   Tropical Journal of Pharmaceutical Research April 2010; 9 (2): 127-133    ...
Parveen et alINTRODUCTION                                         accumulation   of   polycyclic  aromatic                ...
Parveen et altransferred    to     normal     saline     for     morphological and pathological changes inhistopathalogica...
Parveen et al Table 1: Gross features of untreated and treated liver and kidney tissues of albino rats.Tissue             ...
Parveen et alKidney changes in rats treated with a high                The findings are based on the morphologicaldose of ...
Parveen et alkidney tissues were probably due to the               ACKNOWLEDGEMENTmetabolic changes arising from the inhib...
Parveen et al        Molt-4 and HL-60 cells. Nitric Oxide. 2006;                   hydrocarbons. J. Chem. Soc. Pak. 1990; ...
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The Effect of Jatropha Curcas Root Oil on Liver and Kidney Cancer

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The Effect of Jatropha Curcas Root Oil on Liver and Kidney Cancer

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The Effect of Jatropha Curcas Root Oil on Liver and Kidney Cancer

  1. 1. Parveen et al Tropical Journal of Pharmaceutical Research April 2010; 9 (2): 127-133 © Pharmacotherapy Group, Faculty of Pharmacy, University of Benin, Benin City, 300001 Nigeria. . All rights reserved Available online at http://www.tjpr.orgResearch ArticleEffect of Cedrus deodara (Pinaceae) Root Oil on theHistopathology of Rat Liver and Kidney 1 2*Rehana Parveen , Muhammad A Azmi , Syed Naeemul H 1 3 4Naqvi , Syed M Mahmood and Ijaz H Zaidi1 2Department of Pharmacology, Institute of Pharmaceutical Sciences, Department of Physiology, Baqai Medical 3University, 51 Deh Tor, Gadap Road, Near Toll Plaza, Super Highway, P.O. Box 2407, Karachi-74600, Department 4of Oncology, Agha Khan University Hospital, National Stadium Road, Department of Casualty, P.N.S. Shifa Hospital,DHA, Phase 2, Karachi, Pakistan.AbstractPurpose: To evaluate the effect of Cedrus deodara (Pinaceae) root oil, used traditionally for thetreatment of various ailments, on the histopathology of the liver and kidney of Wistar albino rats.Methods: The oil was extracted from the plant root by dry destructive distillation method andadministered orally at doses of 0.5 ml and 2.5 ml per rat. Each animal group was comprised of 30 rats.Treated and untreated group of rats were sacrificed and organs (liver and kidney) were taken out fortissue processing and staining with haematoxylin and eosin according to the method of Bancroft andStevens. Microscopic examination was then carried out to observe any pathological changes in theanimals.Results: The root oil produced atrophic changes and haemolysis in the glomerular region as well asshrinkage and shedding of glomerular epithelium in the kidney tissues. Marked dilation of the centralvein and congested blood vessels in the rat liver at both doses tested was also seen.Conclusion: The root oil of C. deodara (Pinaceae), at the doses administered, produced some adverseeffects on the tissues tested but these effects were not lethal. Some toxic changes in the tissues of liverand kidney were also found.Keywords: Histopathology, Cedrus deodara root oil, Liver, Kidney, Albino rats.Received: 15 October 2009 Revised accepted: 25 February 2010*Corresponding author: E-mail: azmiahmed@hotmail.com; Tel: +9221-4410434; Fax: +9221-4410439 Trop J Pharm Res, April 2010; 9 (2): 127
  2. 2. Parveen et alINTRODUCTION accumulation of polycyclic aromatic hydrocarbons (PAHs) in two kinds of pineCedrus deodara (Deodar - Pinaceae) is found (Cedrus deodara and Pinus thunbergii)in the hilly areas of North Western Frontier needles [10].Province (N.W.F.P.) of Pakistan and also inthe Western Himalaya and the Mediterranean Considerable work has been done on theregion. It is a tall evergreen tree with chemistry of stem bark in Pakistan and Indiaspreading horizontal branches having circular to analyse stem bark extract or oil [11-14] butleaves. In the Indus-Unic system (previously to the best of our knowledge, no scientificUnani or Greek system), the plant’s extracts, studies have been reported in the literatureoils, etc, have been used in Eastern medicine on the analysis of Cedrus deodara root oilfor a long time in the treatment of patients for and its pathological effects on animal tissues.various ailments. For example, the wood Therefore, the objective of this study was toextract is carminative, diaphoretic and investigate the effect of Cedrus deodara rootantipyretic, and has also been used to treat oil on the liver and kidney tissues of albinoflatulence, pulmonary and urinary disorders, rats (Wistar Strain) at low and high doserheumatism, piles and kidney stones. In levels.addition, the bark extract is astringent andalso useful for treating fever, diarrheoa and EXPERIMENTALdysentery [1]. The root extract or oil,alsocontains terpenoids such as himachalol, Materials and animalsatlantone and trans-atlantone; these aresimilar to those found in the trunk oil [2]. In The root oil of Cedrus deodara was collectedthe frontier province (N.W.F.P) of Pakistan, from Brooke’s Pharma (Pvt) Ltd, Karachi,the root oil is used as an anti-ulcer drug by Pakistan. The animals used for thisHakims (old Unic-medical personals). experimental study were adult albino ratsThere is a report that one of the major (Wistar strain), weighing 200 ± 5 g. Theycomponent in the wood of Cedrus deodara, were housed in cages (2 rats per cage) inhimachalol, has antispasmodic and animal house and a well-balanced laboratoryspasmolytic activities similar to those of diet in the form of biscuits prepared inpaparverine that also induces spasm in Hussain Ebrahim Jamal (HEJ) Researchdifferent tissues such as ileum, jejunum and Institute of Chemistry, University of Karachi.bronchial muscles when given in different The animals were kept in a 12 h light / 12 hdoses [3]. Another study reported the use of dark cycle and water was given freely.wood essential oil of Cedrus deodara, as a15% dispersion in castor oil, as a dermal Treatment protocolapplication in rabbit for the treatment of skindiseases [4]. Similarly, mast cell-stabilizing A total of 90 rats, divided into three groupsand lipoxygenase-inhibitory activities [5] as (30 per group), were used. Group A ratswell as the immunological properties of (control) did not receive any treatment whileCedurs deodara oil in allergy caused by Groups B and C were administered low (0.5pollen antigens, have also been evaluated [6]. ml/rat) and high (2.5 ml/rat) doses of Cedrus deodara root oil orally by gauze syringe. AllIn vitro cytotoxic activity against human the animals were fasted overnight prior tocancer cell lines, in vivo induction of being sacrificed. The animals wereintracellular caspases, DNA fragmentation anaesthetized with chloroform, placed on aand DNA cell cycle analysis have been dissection board and their abdomen wasreported [7-9]. Recently, a study investigated opened via a long midline incision, the organsthe lipid contents, specific surface areas and (liver and kidney) were taken out, blotted drystomata density as well as the levels of with tissue paper and then immediately Trop J Pharm Res, April 2010; 9 (2): 128
  3. 3. Parveen et altransferred to normal saline for morphological and pathological changes inhistopathalogical studies. the tissues of the animals.The approval for conducting the experimental RESULTSprocedures on the animals was given byBoard of Advance Studies & Research The general characteristic features of the(BASR) and Ethics Committee of Baqai liver and kidney tissue sections of theMedical University, Karachi – Pakistan with animals examined microscopically areapproval ref. no. BMU-VC/145. presented in Table 1.Routine tissue processing Liver changes in rats with root oil high doseThe tissues were fixed in normal saline for 24– 48 h and then processed through a series Normal liver tissues were observed as normalof ethyl alcohol of ascending strength (70, 80 central vein, sinusoids and hepatic cells inand 95 %) for a period of 1 h; twice in the control group (see Fig 1a). However,absolute alcohol (for 1 h each); and twice in histopathological changes were seen in thexylene (for 1 h) in order to render the tissue liver tissues of albino rats treated with a highelements transparent. The tissues were then dose of Cedrus deodara root oil (2.5 ml/rat). оinfiltrated with molten paraplast at 58 C. This The general architecture of liver tissue waswas done twice (for one hour on each found intact but fatty changes (++) were seenoccasion). The transparent tissues, after (Figure 1b); almost intact portal vein andclearing all elements from it, were embedded markedly dilated central vein were also seenin a solid mass of paraplast. The blocks were (Figure 1c). Congested (+) blood vessels andlabelled, allowed to cool and the metal blocks necrotic and fibrotic changes were not seen.were removed. The solid mass was then In addition, few inflammatory cells (+) weretrimmed free of excess paraplast, leaving also found on examination.some free margins around the embeddedtissues. [a]Three microns thick longitudinal sectionswere cut with a rotary microtome. Thesections were mounted on thoroughlycleaned gelatinized slides and the slides оwere then placed on hot plates at 37 C for (a) Control liver24 h for proper fixation. The slides werestained by H & E stain according to theprescribed staining method [15]. The stain [b] [c] [cwas prepared by dissolving hematoxylin inabsolute alcohol. The mixture was boiledrapidly and mercuric oxide was then added.The stain was cooled rapidly in a cold waterbath; glacial acetic acid was then added andthe stain was ready for immediate use. (b) Treated liver (c) Treated liverSeveral slides were prepared accordingly.The stained slides, after drying and labelling, Figure 1: Sections (a) of rat liver (Control)were preserved and stored for showing normal central vein (C), sinusoid (S) andhistopathological studies which entailed hepatic cells (H); (b) fatty changes (F); and (c)microscopical examination for comparative dilated central vein (C), following treatment of rats with Cedrus deodara root oil (2.5 ml) Trop J Pharm Res, April 2010; 9 (2): 129
  4. 4. Parveen et al Table 1: Gross features of untreated and treated liver and kidney tissues of albino rats.Tissue Control Treatment with 0.5 ml root oil Treatment with 2.5 ml root oil Type • Appeared as dark brown bilobed organ. • General Architecture: found intact • General Architecture: found intact • Showed normal hexagonal lobular pattern. • Central Vein: markedly dilated. • Central Vein: markedly dilated .. Fig 1c. • Also showed normal central vein [C], sinusoids • Portal vein: almost found intact. • Portal vein: almost found intact. [S] and hepatic cells [H] … Fig 1a. • Blood vessels: slightly Congested (+). • Blood vessels: Congested (+).Liver • Sinusoids lined by fenestrated endothelium. • Fatty changes: prominent (++) • Fatty changes: prominent (++) .. Fig 1b. • Hepatocytes appeared as large polyhedral cells • Necrosis: no changes were seen. • Necrosis: not seen. with round nuclei dispersed chromatin and prominent nucleoli. • Fibrosis: no changes were seen • Fibrosis: not seen • Cytoplasmic hepatocytes found granular and • Inflammatory cells: few cells (+) seen • Inflammatory cells: few cells (+) seen. deep eosinophilic. • Appeared normally consisting of cortex and • Glomeruli: atrophic changes, hemolysis • Glomeruli: atrophic changes, hemolysis seen in medulla. and shrinkage. Fig 2b. • Cortex showed normal glomeruli appeared as • Basement membrane: no significant • Basement membrane: normal. dense rounded structure .. Fig 2a. changes • Vascular basement and glomerular basement • Cellularity: normal. membranes have uniform thickness and • Cellularity: normal. • Inflammatory cells: few cells (+) found. uninterrupted. • Inflammatory cells: not found • Afferent and efferent arterioles were seen at • Tubules: atrophic changes (+), shedding ofKidney vascular pole of renal corpuscles. • Tubules: atrophic changes (+), hemolysis epithelium, edema .. Fig 2b. desquamated cells • Proximal convoluted tubule (PCT) showed seen. and hemorrhage also seen .. Fig 2c. simple cuboidal epithelium with brush border and • Necrosis: negligible • Necrosis: negligible narrow lumen. • Cytoplasm of PCT stained dark pink. • Blood vessels: normal appearance. • Blood vessels: dilated, thickened and congested • Distal convoluted tubule (DCT) contains few (++) • Interstitium: edematous (+), fibrotic cuboidal cells with wide lumen. • Interstitium: slightly edematous and few changes not seen, inflammatory cells • Cytoplasm of DCT stained light pink. inflammatory cells (+) found. (++) were found • Medulla consists of loop of Henle, collecting tubules vasa recta and collecting duets. Trop J Pharm Res, April 2010; 9 (2): 130
  5. 5. Parveen et alKidney changes in rats treated with a high The findings are based on the morphologicaldose of root oil and pathological effects of the root oil on the animal tissues as indicated in Table 1. TheKidney tissues in the control group showed features include prominent fatty changes,normal renal corpuscles (see Figure 2a) but marked dilation of central vein, andsome morphological and pathological congested blood vessels in liver tissues atchanges occurred in the kidney tissues of both doses. The kidney tissues also showedalbino rats (Group C) treated with a high dose marked changes at both doses such asof the root oil (2.5 ml/rat). In the glomerular hemolysis and shrinkage of glomeruli as wellregion of the kidney, some atrophic changes as the shedding of tubular epithelium andand haemolysis were seen (Figure 2b) but hemorrhage.cellularity and basement membrane werenormal. No inflammatory cells were found. Thus Cedrus deodara root oil, at both lowThe tubular portion also showed atrophic (0.5 ml) and high (2.5 ml) doses, producedchanges, shedding of epithelium and oedema some toxic changes in the different regions of(Figure 2b). Furthermore, desquamated cells liver and kidney tissues when compared withand haemorrhage (Figure 2c) were seen in the untreated animals. However, at low dose,the tubular region. The blood vessels became these changes are mild.dilated, thickened and congested (++), and afew inflammatory cells (+) and slightly DISCUSSIONoedematous interstitial regions were seen. To the best of our knowledge, the [a] pathologically effects of Cedrus deodara root oil on the liver and kidney tissues of albino (Wistar) rats are being reported for the first time in this study. Marked dilation of the central vein and prominent fatty changes were observed in (a) Control kidney liver tissues at both low and high (Figure 1 and Table 1). Prominent morphological and pathological changes were also found in [b] [c] [c] kidney tissues (Figure 2). One of the important compounds, himachalol, in the wood of C. deodara has been reported to exert antispasmodic activity on tissues such as the ileum, jejunum and bronchial muscles of guinea pigs and rabbits [3]. Significant changes have also been noticed (b) Treated kidney (c) Treated kidney [16] in the reproductive organs and tissues of rats, such as the luminal epithelium of uterusFigure 2: Sections (a) of rat kidney (Control) and uterine glands, following theshowing normal renal corpuscles (R); (b) showing administration of neem oil subcutaneously. Inatrophy of glomeruli and haemorrhage in glomeruli the present study, the liver and kidney of the(A), oedema in tubules (B); and (c) shrunken rats experienced shedding of tubularglomeruli (A), hemorrhage in tubules (B) and epithelium of the glomeruli, congestion ofdesquamated cells in tubules (C) following blood vessels as well as atrophic changes intreatment of rats with Cedrus deodara root oil (2.5 renal tubular epithelium, as shown earlier.ml). The changes that took place in both liver and Trop J Pharm Res, April 2010; 9 (2): 131
  6. 6. Parveen et alkidney tissues were probably due to the ACKNOWLEDGEMENTmetabolic changes arising from the inhibitionof enzymes when treated with root oil of The authors gratefully acknowledge thedifferent doses. On the other hand, no assistance of Mr. Abdul Haseeb Khan,significant damage to organs were noticed Chairman Brooke’s Pharma (Pvt) Ltd.,following the oral administration of bitterless, Karachi, Pakistan for providing the Cedrusodourless and colourless neem oil and deodara oil used in this study. We are alsogroundnut oil in male and female rats up to grateful to Prof. Dr. Syed Azhar Ahmed, Vicethree generations. These oils are generally Chancellor, Baqai Medical University andaccepted as safe for human consumption Prof. Dr. Syed Fazal Hussain of Baqai[17]. However, the present study indicates Institute of Pharmaceutical Sciences for theirthat cedar wood oil may not be safe in technical support and cooperation. Mr. Arifhumans since it damaged the liver and Jawaid, Baqai Institute of Hematology, Baqaikidney tissues of Wistar rats following oral Medical University is thankfullyadministration. acknowledged for assisting in putting this manuscript together.The pathological and cytotoxic activities ofAP9-Cd, a standardized lignan compositionderived from Cedrus deodara, in human REFERENCESleukaemic Molt-4 and HL-60 cells, has also 1. Baquar S.R. Medicinal and poisonous plants ofbeen noticed [8]. Similarly, the plant’s lignan Pakistan. Printas, Karachi, 1989; pp. 94-95.mixture has cytotoxic potential, in vitro, 2. Rehana P. Analysis of Cedrus deodara root oil andagainst human cancer lines from different its pharmacokinetic and pharmacodynamic studes with reference to anti-ulcer andtissues such as breast, cervix, colon, liver antifungal effects. Ph.D. Thesis Baqai. Med.and prostate; it was also found to induce Univ. 2006; pp. 76-79.tumor regression in vivo, induction of 3. Kar K, Puri VN, Patnanik G.K, Sur RN, Dhawan BN,intracellular DNA fragmentation and DNA cell Kulshrestha DK, Rastogi RP. Spasmolytic constituents of Cedrus deodara:cycle analysis [9]. No evidence can, however, Pharmacological evaluation of Himachalol. J.be adduced from the present study that links Pharm. Sci. 1975; 64: 258-262.the plant’s lignan mixture to the root oil 4. Tandan CK., Singh R., Gupta S, Chandra S, Lal J.effects on the liver and kidney tissues of the Subacute dermal toxicity study of Cedrus deodara wood essential oil. Indian. Vet. J.animals. 1989; 66: 1088-1091. 5. Shinde UA, Kulkarni KR, Phadke AS, Nair AM,CONCLUSION Mungantiwar AA, Dikshit VJ, Saraf MN. Mast cell stabilizing and lipoxygenase inhibitory activity of Cedrus deodara (Roxb) Loud woodScientific analysis of these materials in oil. Indian. J. Exp. Biol. 1999; 37: 258 – 261.relation to their pharmacological studies was 6. Rawat A, Singh A, Singh AB, Gaur SN, Kumar L, Roy I., Ravidrun P. Clinical and immunologicnot carried out in this study. In India, Pakistan evaluation of Cedrus deodara Pollen: a newand other countries, Colleges of Eastern allergen from India. Allergy. 2000; 55: 620-Medicine commenced such studies before 626.using these materials as drugs. The root oil 7. Dimri M, Sharma MC. Effects of sarcoptic manage and its control with oil of Cedrus deodara,used in the present work may be categorised Pangamia glabra, Jatropha curcas and benzylunder ‘least toxic’ group, according to the benzoate both with and without ascorbic acidstandard used by toxicologists. Thus, Cedrus on growing sheep, assessment of weight gain,deodara root oil may not be safe for human liver function, nutrient digestibility, wood production and meat quality. J. Vet. Med. A.consumption, but this assertion requires Physiol. Pathol. Clin Med. 2004; 51: 79−84.further investigation. 8. Shashi B, Jaswant S, Madhusudana RJ, Kumar SA, Nabi QG. A novel lignan composition from Cedrus deodara induces apoptosis and early nitric oxide generation in human leukemia Trop J Pharm Res, April 2010; 9 (2): 132
  7. 7. Parveen et al Molt-4 and HL-60 cells. Nitric Oxide. 2006; hydrocarbons. J. Chem. Soc. Pak. 1990; 12: 14: 72 - 78. 282-284.9. Singh SK, Shanmugavel M, Kampasi H., Singh R, 13. Adinarayana D, Seshadri TR. Stem bark of Cedrus Mondhe DM., Rao JM, Adwankar MK, Saxena, deodara. Isolation of new dihydroflavonol, AK, Qazi GN. Chemically standardized deodarin. Tetrahedron. 1965; 21: 3727-3730. isolates from Cedrus deodara stem wood 14. Agarwal PK, Rastogi RP. Terpenoids from Cedrus having anticancer activity. Planta. Med. 2007; deodara. Phytochemistry. 1981; 20: 1319- 73: 519 - 526. 1321.10. Yang P, Wang Z, Chen JW, Tian FL, Ge Lk. 15. Bancroft JD, Stevens A. Theory and practice of Influence of pine needles physiological histological techniques, 3rd Ed, Newyork, properties on the PAH accumulation. Huan Churchill Livingstone 1990; pp. 281 - 282. Jing Ke Xue. 2008;29: 2018. 16. Tewari RK, Pathak S, Parkash AO. Biochemical11. Khan N, Naheed S. Chemical investigation of and histological studies of reproductive organs Cedrus deodara stem-bark. I. Isolation and in cyclic and ovariectomized rat supporting a structure of branched chain fatty acid ethyl non- hormonal action for neem oil. J. ester. J. Chem. Soc. Pak. 1988; 10: 63-65. Ethnopharmacol. 1989; 25: 281 - 293.12. Khan N, Naheed S. Chemical investigation of 17. Chinnasamy N, Harishankar N, Kumar PU, Rukmini Cedrus deodara stem bark. II. Isolation and C. Toxicological studies on debitterized neem identification of some sesquiterpene oil (Azadirachta indica). Food Chem. Toxicol. 1993; 31: 297 - 301. Trop J Pharm Res, April 2010; 9 (2): 133

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