• Dina R Gad El-Karim
  • Gamal El-Amrawi



Cyclophosphamide (CPA) is a widely used anti-neoplastic drug, but its usage is associated with several side effects including hepatotoxicity. This study was designed to evaluate the ameliorative effect of Ganoderma lucidium (GL) extract as a natural antioxidant against CPA hepatotoxicity in male Wistar albino rats. Four groups of rats (n = 7) were used in this study, Group I: served as a control group, group II: was administrated CPA (5 mg/kg), group III: received GL extract (50 mg/kg), and group IV: received CPA and GL extract at the same doses of groups II and III. All the treatments were orally administrated via gastric intubation for 21 consecutive days. Serum activities of the hepatic enzymes (AST, ALT and GGT) in addition to serum proteinogram were evaluated. Moreover, hepatic oxidant-antioxidant markers (4- hydroxynonenal (4-HNE), GSH, CAT and SOD) besides activity of cytochrome C oxidase enzyme were detected. Also, histopathological examination of the hepatic sections (H&E stained) was performed. Serum activity of liver enzymes was elevated, proteinogram was disturbed in CPA-treated animals which accompanied by an increase in hepatic content of 4-HNE, depletion of hepatic antioxidant (GSH, CAT, SOD) and inhibition of hepatic cytochrome C oxidase activity, also hepatic histopathological disruptions were noticed in this group. All these hepatic disturbances were partially ameliorated upon co-administration of GL extract with CPA. In conclusion, production of 4- HNE and inhibition of cytochrome C oxidase enzyme may play a role in occurrence of CPA hepatotoxicity and GL extract may have an ameliorative role against this toxicity.

Key words: cyclophosphamide; hepatotoxicity; Ganoderma lucidium; 4-hydroxynonenalon; cytochrome C oxidase


Dollery C. Cyclophosphamide. In: Thera-peutic Drugs. Edinburgh: Churchill Living-stone1999; 349–53.

Paolo AD, Danesi R, Tacca MD. Phar-macogenetics of neoplastic disease: New Trends. Pharmacol Res 2004; 49: 331–42.

Stankiewicz A, Skrzydlewska E , Makiela M. Effects of amifostine on liver oxidative stress caused by cyclophosphamide administra-tion to rats. Drug Metabol Drug Interact 2002; 19: 67–82.

Haque R, Bin-Hafeez B, Parvez S, Pan-dey S. Aqueous extract of walnut (Juglans regia L.) protects mice against cyclophosphmide in-duced biochemical toxicity. Human Exp Toxi-col 2003; 22:473–80.

Bhatia K, Ahmad F, Rashid H, Raisuddin S. Protective effect of S-allylcysteine against cyclophosphamide-induced bladder hemor-rhagic cystitis in mice. Food Chem Toxicol 2008; 46(11):3368–74.

Chakraborty P, Hossain U, Murmu N, Das JK, Pal S, Bhattacharya S. Modulation of cy-clophosphamide-induced cellular toxicity by diphenylmethyl selenocyanate in vivo, an en-zymatic study. J Cancer Mol 2009; 4(6): 183–9.

Mousa NK, Muhsin EA, Sabbar SS, Ish-rak A. Ahmed IA. Liver histopathological of purification cinnamic acid activity against en-doxan in mice. J Gen Enviro Reso Cons 2014; 2(1): 22–9.

Weijl NI, Cleton FJ, Osanto S. Free radi-cals and antioxidants in chemotherapy-induced toxicity. Cancer Treat Rev 1997; 23 (4): 209–40.

Wasser SP. Reishi or Lingzhi (Ganoderma lucidium). Encyclopedia of dietary supple-ments 2005; 1: 603–22.

Watchel-Galore S, Yuen J, Buswell JA. Ganoderma Lucidium (Lingzhi or Reishi): a Medicinal Mushroom. In: Benzie IFF, Watchel-Galore S, editors. Herbal Medicine: Bimolecu-lar and Clinical Aspects. 2nd edition. Boca Ra-ton (FL): CRC Press/Taylor &Francis; 2011. Chapter 9.

Lin ZB, Zhang H. Anti-tumor and im-munoregulatory activities of Ganoderma lucid-um and its possible mechanisms. Acta Pharma-col Sin; 2004, 25 (11): 1387–95.

Lee JM, Kwon H, Jeong H, Lee JW, Lee SY, Baek SJ, Surh YJ. Inhibition of Lipid Pe-roxidation and Oxidative DNA Damage by Ganoderma lucidum. Phytother Res 2001; 15:245–9.

Zhu MQ, Chang LK, Wong FS, Li RC. Triterpene antioxidants from Ganoderma lucid-um. Phytotherapy Res1999; 13(6): 529–31.

Kao CH, Jesuthasan AC, Bishop KS, Glucina MP, Ferguson LR. Anti-cancer activi-ties of Ganoderma lucidum: active ingredients and pathways. Fun Foods Health Dis 2013; 3(2):48–65.

Lin Z. Cellular and Molecular Mecha-nisms of Immuno-modulation by Ganoderma lucidum. J Pharmacol Sci 2005; 99: 144–53.

Kim KC, Kim IG. Ganoderma lucidum extract protects DNA from strand breakage caused by hydroxyl radical and UV irradiation. Int J Mol Med 1999; 4(3): 273–7.

Zhang GL, Wang YH, Ni W, Teng HL, Lin ZB, Lin ZB. Hepatoprotective role of Ganoderma Lucidium polysaccharide against BCG induced immune liver injury in mice. World J Gastroenterol 2002; 8 (4): 728–33.

Lin WC, Lin WL. Ameliorative effect of Ganoderma lucidum on carbon tetrachloride-induced liver fibrosis in rats. World J Gastroen-terol 2006; 12(2):265–70.

Wang X, Zhao X, Li D, Lou YQ, Lin ZB, Zhang GL. Effects of Ganoderma lucidum Polysaccharide on CYP2E1, CYP1A2 and CYP3A Activities in BCG-Immune Hepatic Injury in Rats. Biol. Pharm. Bull 2007; 30 (9): 1702–6.

Shieh YH, Liu CF, Huang YK, Yang JY, Wu L, Lin CH , Lin SC. Evaluation of the He-patic and Renal-protective Effects of Ganoderma lucidum in Mice. Am Chin Med 2001; 29(3): 501–7.

Ghosh S, Ghosh D, Chattopadhyay S, Depnath J. Effect of Ascorbic Acid Supple-mentation on Liver and Kidney Toxicity in Cyclophosphamide Treated Female Albino Rats. The J toxicol Sci 1999; 24(3): 141–4.

Reitman S, Frankel S. A coloremetric method for determination of oxaloacetic trans-aminase and serum glutamic pyruvic transami-nase. Am. J. Clin Pathol 1957; 28: 56–63.

Szasz ZG. A kinetic photometric method for serum gamma glutamyltranspeptidase. Clin. Chem 1969; 15:124–36.

Doumas BT, Bayso DD, Caster RJ, Let-ers T, Schaffer R. Determination of serum total protein. Clin. Chem 1981; 27: 1642.

Drupt F. Dosage del albumin serque part vert de bromocresol. Pharma. Biol 1974; 111: 77.

Watson D. Albumin and “total globulin†fraction of blood. In: Advances in Clinical Chemistry. Academic Press Inc. 1965; 8: 238–49.

Aebi H. Catalase in vitro. Methods En-zymol 1984; 105: 121–6.

Ukeda H, Maeda S, Ishii T, Sawamura M. Spectrophotometric assay for superoxide dismutase based on tetrazolium salt 3'{1-[(phenylamino)-carbonyl]-3,4- tetrazolium}-bis(4-methoxy-6-nitro) benzenesulfonic acid hydrate reduction by xanthine-xanthine oxi-dase. Anal Biochem1997; 251: 206–9.

Beutler E, Duron O, Kelly MB. Reduced glutathione determination. J Lab Clin Med 1963; 61:882.

Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding, Anal Biochem1976; 72 (1-2): 248–54.

Bancroft JD, Stevens A. Theory and Practice of Histological Technique. 4th ed. Churchil livingestone; 1996.

SAS. Statistical analysis system. SAS User’s Guide, SAS incorp Inst 2004.

Patel JM. Stimulation of cyclophospha-mide-induced pulmonary microsomal lipid pe-roxidation by oxygen. Toxicology 1987; 45: 79–91.

McDonald GB, Slattery GH, Bouvier ME, Ren S, Batchelder AL et al. Cyclophos-phamide metabolism, liver toxicity, and mortal-ity following hematopoietic stem cell transplan-tation. Blood J 2003;101: 2043–8.

Sheeja K, Kuttan. Ameliorating Effects of Andrographis Paniculata Extract Against Cyclophosphamide-Induced Toxicity in Mice. Asia Pac Can Prev 2006; 7: 609–14.

Alenzi FQ, El-Bolkiny YE, Salem ML. Protective effects of Nigella sativa oil and thy-moquinone against toxicity induced by the an-ticancer drug cyclophosphamide. BRIT J BIO-MED SCI 2010; 67 (1):20–8.

Jain R, Jain SK. Effect of Buchanania lanzan Spreng bark extract on cyclophospha-mide induced genotoxicity and oxidative stress in mice. Asia Paci J Trop Med 2012; 187–91.

Shokrzadeh M, Ahmadi A, Naghshvar F, Chabra A, Jafarinejhad M. Prophylactic Effica-cy of Melatonin on Cyclophosphamide-Induced Liver Toxicity in Mice. BioMed Res Int 2014; Article ID 470425.

Bhattacharya A , Lawence RA, Krishnan A, Zaman K, Sun D, Fernandes G. Effect of dietary n-3 and n-6 oils with and without food restriction on activity of antioxidant enzymes and lipid peroxidation in livers of cyclophos-phamide treated autoimmune-prone NZB/W female mice . J Am Coll Nutr 2003; 22: 388–99.

Kehrer JP, Biswal SS. The molecular ef-fects of acrolein. Toxicol Sci 2000; 57:6–15.

Werts ED, Gould MN. Relationships be-tween cellular superoxide dismutase and sus-cepti-

bility to chemically induced cancer in the rat mammary gland. Carcinogen1986; 7:1197–201.

Shokrzadeh M, Chabra A, Ahmadi A , Naghshvar F, Habibi E , Salehi F , Assadpour S. Hepatoprotective Eff ects of Zataria Multifl ora Ethanolic Extract on Liver Toxicity Induced by Cyclophosphamide in Mice. Drug Res 2014; 64: 1–7.

Kim DH, Shim SB, Kim NJ, Jang LS. Beta Glucoronidase Inhibitory Activity and Hepatoprotective Effect of Ganoderma Lucidi-um. Biol Pharm Bull 1999; 22(2): 162–4.

Zhong D, Wang H, Liu M, Li X, Huang M, Zhou H et al. Ganoderma lucidum polysac-charide peptide prevents renal ischemia reper-fusion injury via counteracting oxidative stress. Sci Rep 2015; 5:16910.

Latimer KS, Mahaffey EA, Prasse KW. Plasma Protein. In: Veterinary Laboratory Med-icine, Clinical Pathology. 4th Ed., Iwoa State Press; 2003. p. 162–71.

Vandenberghe J. Hepatotoxicology: Mechanisms of liver toxicity and methodologi-cal aspects. In: Niesink RJM, De Vries J, Hol-linger MA. Toxicology: Principle and Applica-tions. New York: CRC Press, 1996; 703–23.

Chen JJ, Schenker S, Henderson GI. 4-Hydroxynonenal Levels Are Enhanced in Fetal Liver Mitochondria by In Utero Ethanol Expo-sure. HEPATOL 1997; 25(1):142–7.

Chen J, Robinson NC, Schenker S, Frosto T, George I. Henderson GI. Formation of 4-Hydroxynonenal Adducts With Cyto-chrome c Oxidase in Rats Following Short-Term Ethanol Intake. HEPATOL 1999; 29 (6): 1792–8.

Srinivasan S, Avadhani NG. Cyto-chrome c Oxidase dysfunction in oxidative stress. Free Rad Biol Med 2012; 53(6): 1252–63.

Wikstrom M. Cytochrome C Oxidase. eLS. 2001. 015902.a00


Sohal RS, Toroser D, Bre´ge`re C, Mockett RJ and Orr WC. Age-related decrease in expression of mitochondrial DNA encoded subunits of cytochrome c oxidase in Drosophi-la melanogaster. Mech Ageing Dev 2008; 129: 558–61.

Senthilkumar S ,Devaki T, Manohar BM, Babu MS. Effect of squalene on cyclophos-phamide-induced toxicity . Clin Chim Acta 2006; 364: 335–42.




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Veterinary Medicine and The One Health Concept