Environmental Science and Pollution Research

, Volume 26, Issue 15, pp 15559–15572 | Cite as

Protective effects of garlic extract against hematological alterations, immunosuppression, hepatic oxidative stress, and renal damage induced by cyclophosphamide in rats

  • Ahmed M. El-Sebaey
  • Fatma M. AbdelhamidEmail author
  • Osama A. Abdalla
Research Article


Cyclophosphamide is an alkylating agent widely used as anticancer drug, reported to exert cytotoxic effects attributed to oxidative stress. Therefore, this study aimed to explore the protective effect of ethanolic extract of garlic (EEG) against cyclophosphamide (Cyp)-induced hematological disturbance and immunosuppressive and hepatotoxic effects. Forty male Wistar albino rats were randomized into four equal groups: the normal control one, the Cyp-treated group (50 mg/kg BW/IM, once weekly), the EEG-treated group (300 mg/kg BW, orally, daily), and the Cyp & EEG group. All rats received their relevant treatments for four consecutive weeks. This study revealed that Cyp significantly decreased erythrocyte count, hemoglobin (Hb), packed cell volume (PCV), and total leukocyte and lymphocyte counts. However, the counts of neutrophils, eosinophils, and toxic neutrophils were elevated. Additionally, hepatic malondialdehyde (MDA) and levels of liver and renal biomarkers were significantly elevated in the Cyp-treated group. Otherwise, hepatic catalase (CAT), reduced glutathione (GSH), superoxide dismutase (SOD), and serum total antioxidant capacity (TAC) were significantly lower than the control rats. Furthermore, Cyp significantly reduced whole blood respiratory burst activity (NBT), serum lysozyme and bactericidal activities, interlukin-12 (IL-12), and interferon-γ. In contrast, the levels of nitric oxide (NO), tumor necrosis factor-α (TNF-α), and interlukin-1β (IL-1β) were elevated. Additionally, Cyp induced hepatic and renal histopathological alterations. Data in the present study demonstrated that EEG has immunomodulatory and antioxidant effects and has the ability to diminish the alterations induced by Cyp.


Ethanolic extract of garlic Cyclophosphamide Hematological alterations Immunosuppression Hepatic oxidative stress Renal damage 



The authors are deeply grateful to Professor Dr. Mohamed El-Boshy, Laboratory Medicine Department, Faculty of Applied Medical Science, Umm Al-Qura University, PB 7296, Makkah 21955, Saudi Arabia, for his kind help in statistical analysis.

Compliance with ethical standards

Animals were handled in accordance with animal welfare and under ethical protocols by the Faculty of Veterinary Medicine, Mansoura University, Egypt.

Conflict of interest

The authors declare that they have no conflict of interest.


  1. Abdul Qadir M, Shahzadi SK, Bashir A, Munir A, Shahzad S (2017) Evaluation of phenolic compounds and antioxidant and antimicrobial activities of some common herbs. Int J Anal Chem 6:1–6. CrossRefGoogle Scholar
  2. Akinbi HT, Epaud R, Bhatt H, Weaver TE (2000) Bacterial killing is enhanced by expression of lysozyme in the lungs of transgenic mice. J Immunol 165:5760–5766. CrossRefGoogle Scholar
  3. Alabbassi MG (2010) Melatonin ameliorates hepatic damage induced by cyclophosphamide in rats. J Pharm Sci 17:47–54. Google Scholar
  4. Al-Malki AL (2014) Synergestic effect of lycopene and melatonin against the genesis of oxidative stress induced by cyclophosphamide in rats. Toxicol Ind Health 30:570–575. CrossRefGoogle Scholar
  5. Anusuya N, Durgadevi P, Dhinek A, Mythily S (2013) Nephroprotective effect of ethanolic extract of garlic (Allium sativum) on cisplatin induced nephrotoxicity in male Wistar rats. Asian J Pharm Clin Res 6:97–100Google Scholar
  6. Asdaq SMB (2015) Antioxidant and hypolipidemic potential of aged garlic extract and its constituent, S-allyl cysteine, in rats. Evid Based Complement Alternat Med 7:1–7. CrossRefGoogle Scholar
  7. Bancroff JP, Stevenes A, Turner DR (1990) Theory and practice of histological techniques, 3rd edn. Clurechill Livingston, Edinburgh, LondonGoogle Scholar
  8. Berger J, Slapničková M (2003) Circadian structure of rat neutrophil phagocytosis. Comp Clin Path 12:84–89. CrossRefGoogle Scholar
  9. Boonpeng S, Siripongvutikorn S, Sae-wong C, Sutthirak P (2014) The antioxidant and anti-cadmium toxicity properties of garlic extracts. F Sci Nut 2:792–801. CrossRefGoogle Scholar
  10. Casanova NA, Simoniello MF, López Nigro MM, Carballo MA (2017) Modulator effect of watercress against cyclophosphamide-induced oxidative stress in mice. Medicina (BA) 77:201–206Google Scholar
  11. Chang CC, Yang MH, Wen HM, Chern JC (2002) Estimation of total flavonoid content in propolis by two complementary colorimetric methods. J Food Drug Anal 10:178–182Google Scholar
  12. Dame C, Kirschner KM, Bartz KV, Wallach T, Hussels CS, Scholz H (2006) Wilms tumor suppressor, Wt1, is a transcriptional activator of the erythropoietin gene. Blood 107:4282–4290CrossRefGoogle Scholar
  13. Dong Q, Sugiura T, Toyohira Y, Yoshida Y, Yanagihara N, Karasaki Y (2011) Stimulation of IFN-γ production by garlic lectin in mouse spleen cells: involvement of IL-12 via activation of p38 MAPK and ERK in macrophages. Phytomedicine 18:309–316CrossRefGoogle Scholar
  14. Duggina P, Kalla CM, Varikasuvu SR, Bukke S, Tartte V (2015) Protective effect of centella triterpene saponins against cyclophosphamide-induced immune and hepatic system dysfunction in rats: its possible mechanisms of action. J Physiol Biochem 71:435–454. CrossRefGoogle Scholar
  15. El-Banna SG, Attia AM, Hafez AM, El-Kazaz SM (2009) Effect of garlic consumption on blood lipid and oxidant-antioxidant parameters in rat males exposed to chlorpyrifos. Slolvak J of Animal Science (Slov Rep) 42:111–117Google Scholar
  16. El-Olemy MM, Al-Muhtadi FJ, Afifi AFA (1994) Experimental phytochemistry: a laboratory manual. Department of Pharmacognosy, College of Pharmacy, King Saud University Press, Saudi Arabia, pp 21–27Google Scholar
  17. Fahmy SR, Amien AI, Abd-Elgleel FM, Elaskalany SM (2016) Antihepatotoxic efficacy of Mangifera indica L. polysaccharides against cyclophosphamide in rats. Chem Biol Interact 244:113–120. CrossRefGoogle Scholar
  18. Fandrey J, Frede S, Jelkmann W (1994) Role of hydrogen peroxide in hypoxia-induced erythropoietin production. Biochem J 303:507–510CrossRefGoogle Scholar
  19. Feldman BF, Zinkl JG, Jain VC (2000) Schalm’s veterinary hematology, 5th edn. Lippincott Williams and Wilkins, Toronto, Canada, pp 1145–1146Google Scholar
  20. Förstermann U, Schmidt HH, Pollock JS, Sheng H, Mitchell JA, Warner TD, Nakane M, Murad F (1991) Isoforms of nitric oxide synthase characterization and purification from different cell types. Biochem Pharmacol 42:1849–1857. CrossRefGoogle Scholar
  21. Fotakis G, Timbrell JA (2006) In vitro cytotoxicity assays: comparison of LDH, neutral red, MTT and protein assay in hepatoma cell lines following exposure to cadmium chloride. Toxicol Lett 160:171–177. CrossRefGoogle Scholar
  22. Germoush MO, Mahmoud AM (2014) Berberine mitigates cyclophosphamide-induced hepatotoxicity by modulating antioxidant status and inflammatory cytokines. J Cancer Res Clin Oncol 140:1103–1109. CrossRefGoogle Scholar
  23. Goyal S, Gupta N, Chatterjee S (2016) Investigating therapeutic potential of Trigonella foenum-graecum L. as our defense mechanism against several human diseases. J Toxicol 2:1–10. CrossRefGoogle Scholar
  24. Hamlaoui-Gasmi S, Mokni M, Limam N, Limam F, Amri M, Aouani E, Marzouki L (2014) Effect of garlic’s mode of administration on erythrocytes and plasma parameters in Wistar rat. Afr J Biotechnol 11:8259–8263Google Scholar
  25. Hickman-Davis JM, Lindsey JR, Matalon S (2001) Cyclophosphamide decreases nitrotyrosine formation and inhibits nitric oxide production by alveolar macrophages in mycoplasmosis. Infect Immun 69:6401–6410. CrossRefGoogle Scholar
  26. Higuchi T, Bodin JI (1961) In: Higuchi T, Hansen EB (eds) Alkaloid and other basic nitrogenous compounds; in pharmaceutical analysis (edns.). Interscience, New York, pp 315–345Google Scholar
  27. Huang JQ, Pang MR, Li GY, Wang N, Jin L, Zhang Y (2017) Alleviation of cyclophosphamide-induced immunosuppression in mice by naturally acetylated hemicellulose from bamboo shavings. Food Agric Immunol 28:328–342. CrossRefGoogle Scholar
  28. Inoue LAKA, Oliveira Maciel P, Gusmão Affonso E, Lima Boijink C, Tavares-Dias M (2016) Growth, parasitic infection and hematology in Colossoma macropomum Cuvier, 1818 fed diets containing Allium sativum. J Appl Ichthyol 32:901–905. CrossRefGoogle Scholar
  29. Khalil R, El-Hofy H, Yehya MM, Selim KM (2014) Some biochemical and immunological changes associated with use of garlic extract (allicin) in combating some fish pathogens. Alex J Vet Sci 41:1–18. Google Scholar
  30. Khan MA (2017) Immune potentiating and antitoxic effects of camel milk against cyclophosphamide-induced toxicity in BALB/C mice. Int J Health Sci 11:1–18Google Scholar
  31. Kilikdar D, Mukherjee D, Mitra E, Ghosh AK, Basu A, Chandra AM, Bandyoapdhyay D (2011) Protective effect of aqueous garlic extract against lead-induced hepatic injury in rats. Indian J Exp Biol 49:498–510Google Scholar
  32. Kim SH, Lee IC, Ko JW, Moon C, Kim SH, Shin IS, Kim JC (2015) Diallyl disulfide prevents cyclophosphamide-induced hemorrhagic cystitis in rats through the inhibition of oxidative damage, MAPKs, and NF-κB pathways. Biomol Ther (Seoul) 23:180–188. CrossRefGoogle Scholar
  33. Klaassen CD, Casarett LJ, Doull J (2008) Casarett and Doull’s toxicology: the basic science of poisons, 7th edn. McGraw-Hill, New YorkGoogle Scholar
  34. Kocahan S, Dogan Z, Erdemli E, Taskin E (2017) Protective effect of quercetin against oxidative stress induced toxicity associated with doxorubicin and cyclophosphamide in rat kidney and liver tissue. Iran J Kidney Dis 11:124–131Google Scholar
  35. Kokate CK, Purohit AP, Gokhale SB (1995) Pharmacognosy, 3rd edn. Niralin Prakashan, Pune, p 125Google Scholar
  36. Kumar VP, Venkatesh YP (2016) Alleviation of cyclophosphamide-induced immunosuppression in Wistar rats by onion lectin (Allium cepa agglutinin). J Ethnopharmacol 186:280–288. CrossRefGoogle Scholar
  37. Lindsay H (1973) A colorometric estimation of reducing sugar in potatoes with 3-5 dinitrosalicylic acid. Potatoes Res 16:176–179. CrossRefGoogle Scholar
  38. Madondo MT, Quinn M, Plebanski M (2016) Low dose cyclophosphamide: mechanisms of T cell modulation. Cancer Treat Rev 42:3–9. CrossRefGoogle Scholar
  39. Mahmoud AM (2014) Hesperidin protects against cyclophosphamide-induced hepatotoxicity by upregulation of PPARγ and abrogation of oxidative stress and inflammation. Can J Physiol Pharmaco 92:717–724. CrossRefGoogle Scholar
  40. Mahmoud AM, Wilkinson FL, Jones AM, Wilkinson JA, Romero M, Duarte J, Alexander MY (2017) A novel role for small molecule glycomimetics in the protection against lipid-induced endothelial dysfunction: involvement of Akt/eNOS and Nrf2/ARE signaling. Biochim Biophys Acta Gen Subj 1861:3311–3322. CrossRefGoogle Scholar
  41. Merwid-Ląd A, Ksiądzyna D, Hałoń A, Chlebda-Sieragowska E, Trocha M, Szandruk M, Sozański T, Magdalan J, Kopacz M, Kuźniar A, Nowak D, Pieśniewska M, Szeląg A (2015) Impact of morin-5′-sulfonic acid sodium salt on cyclophosphamide-induced gastrointestinal toxicity in rats. Pharmacol Rep 67:1259–1263. CrossRefGoogle Scholar
  42. Mikaili P, Maadirad S, Moloudizargari M, Aghajanshakeri S, Sarahroodi S (2013) Therapeutic uses and pharmacological properties of garlic, shallot, and their biologically active compounds. Iran J Basic Med Sci 16:1031–1048Google Scholar
  43. Mirfardi M, Johari H (2015) The effect of hydro-alcoholic Allium sativum extract on sexual hormones in mature male rats under chemotherapy with cyclophosphamide. Zahedan J Res Med Sci 17:29–33CrossRefGoogle Scholar
  44. Mohammed NA, Abd El-Aleem SA, El-Hafiz HA, McMahon RF (2004) Distribution of constitutive (COX-1) and inducible (COX-2) cyclooxygenase in postviral human liver cirrhosis: a possible role for COX-2 in the pathogenesis of liver cirrhosis. J Clin Pathol 57:350–354CrossRefGoogle Scholar
  45. Montgomery HAC, Dymock JF (1961) Determination of nitric oxide in water. Analyst (London) 86:414–416Google Scholar
  46. Motawi TM, Sadik NA, Refaat A (2010) Cytoprotective effects of DL-alpha-lipoic acid or squalene on cyclophosphamide-induced oxidative injury: an experimental study on rat myocardium, testicles and urinary bladder. Food Chem Toxicol 48:2326–2336. CrossRefGoogle Scholar
  47. Nasr AY (2014) Protective effect of aged garlic extract against the oxidative stress induced by cisplatin on blood cells parameters and hepatic antioxidant enzymes in rats. Toxi Rep 1:682–691. Google Scholar
  48. Ndong D, Fall J (2011) The effect of garlic (Allium sativum) on growth and immune responses of hybrid tilapia (Oreochromis niloticus x Oreochromis aureus). J Clin Immunol Immunopathol Res 3:1–9Google Scholar
  49. Neboh EE, Ufelle SA (2015) Myeloprotective activity of crude methanolic leaf extract of Cassia occidentalis in cyclophosphamide-induced bone marrow suppression in Wistar rats. Adv Biomed Res 4:5. CrossRefGoogle Scholar
  50. Niki E, Noguchi N, Tsuchihashi H, Gotoh N (1995) Interaction among vitamin C, vitamin E, and beta-carotene. Am J Clin Nutr 62:1322S–1326SCrossRefGoogle Scholar
  51. Olaniyan OT, Meraiyebu AB, Anjorin YD, Shekins O, Dare BJ, Shafe MO (2013) Effects of aqueous extract of onion (Allium sativum) on blood parameters in adult wistar rats (Rattus novergicus). IJPSI 2:42–45Google Scholar
  52. Ozougwu JC (2011) An investigation of the effects of Allium Sativum (garlic) extracts on haematological profile of white albino rats. Pharmacologonline 2:299–306CrossRefGoogle Scholar
  53. Padiya R, Khatua TN, Bagul PK, Kuncha M, Banerjee SK (2011) Garlic improves insulin sensitivity and associated metabolic syndromes in fructose fed rats. Nut Metab (Lond) 8:53. CrossRefGoogle Scholar
  54. Park KJ, Lee BC, Lee JS, Cho MH (2014) Angelica gigas Nakai extract ameliorates the effects of cyclophosphamide on immunological and hematopoietic dysfunction in mice. J Med Plant Res 8:657–663. CrossRefGoogle Scholar
  55. Peng JP, Yao X, Kobayashi H, Ma C (1995) Novel furastonol glycosides from Allium macrostemon. Planta Med 6:58–61. CrossRefGoogle Scholar
  56. Price ML, Butter LG (1977) Rapid visual estimation and spectrophotometric determination of tanning content of sorghum grain. J Agric Food Chem 25:1268–1273CrossRefGoogle Scholar
  57. Rubinstein MP, Su EW, Suriano S, Cloud CA, Andrijauskaite K, Kesarwani P, Schwartz KM, Williams KM, Johnson CB, Li M, Scurti GM, Salem ML, Paulos CM, Garrett-Mayer E, Mehrotra S, Cole DJ (2015) Interleukin-12 enhances the function and anti-tumor activity in murine and human CD8+ T cells. Cancer Immunol Immunother 64:539–549. CrossRefGoogle Scholar
  58. Sahoo PK, Mukherjee SC (2001) Immunosuppressive effects of aflatoxin B1 in Indian major carp (Labeo rohita). Comp Immunol Microbiol Infect Dis 2:143–149. CrossRefGoogle Scholar
  59. Sasaki G, Satoh T, Yokozeki H, Katayama I, Nishioka K (2000) Regulation of cyclophosphamide-induced eosinophilia in contact sensitivity: functional roles of interleukin-5-producing CD4+ lymphocytes. Cell Immunol 203:124–133. CrossRefGoogle Scholar
  60. Seckiner I, Bayrak O, Can M, Mungan AG, Mungan NA (2014) Garlic supplemented diet attenuates gentamicin nephrotoxicity ın rats. Int Braz J Urol 40:562–567. CrossRefGoogle Scholar
  61. Selvakumar E, Prahalathan C, Mythili Y, Varalakshmi P (2005) Mitigation of oxidative stress in cyclophosphamide-challenged hepatic tissue by DL-α-lipoic acid. Mol Cell Biochem 272:179–185CrossRefGoogle Scholar
  62. Senthilkumar S, Devaki T, Manohar BM, Babu MS (2006) Effect of squalene on cyclophosphamide-induced toxicity. Clin Chim Acta 364:335–342. CrossRefGoogle Scholar
  63. Singleton VL, Rossi JA (1965) Colorimetry of total phenolics with phosphomolybdic-phosphotungstic acid reagents. Am J Enol Vitic 16:144–158Google Scholar
  64. Song Y, Zhang C, Wang C, Zhao L, Wang Z, Dai Z, Ma X (2016) Ferulic acid against cyclophosphamide-induced heart toxicity in mice by inhibiting NF-κB pathway. Evid Based Complement Alternat Med 8:1–8. Google Scholar
  65. Sowjanya BL, Devi KR, Madhavi D (2009) Modulatory effects of garlic extract against the cyclophosphamide induced genotoxicity in human lymphocytes in vitro. J Environ Biol 30:663–666Google Scholar
  66. Swamy AV, Patel UM, Koti BC, Gadad PC, Patel NL, Thippeswamy AHM (2013) Cardioprotective effect of Saraca indica against cyclophosphamide induced cardiotoxicity in rats: a biochemical, electrocardiographic and histopathological study. Indian J Pharmacol 45:44–48. CrossRefGoogle Scholar
  67. Wani AA, Sikdar-Bar M (2014) Ameliorative efficacy of taurine and garlic extract on copper induced immunotoxic effect on total and differential leucocyte counts in African catfish, Clarias gariepinus. Asian J Med Pharm Res 4:122–129Google Scholar
  68. Wójcik R (2014) Reactivity of the immunological system of rats stimulated with Biolex-Beta HP after cyclophosphamide immunosuppression. Cent Eur J Immunol 39:51–60. CrossRefGoogle Scholar
  69. Wojcik R, Dabkowska A (2010) The effect of cyclophosphamide on the selected parameters of immunity in rats. Cent Eur J Immunol 35:1–9Google Scholar
  70. Zarei M, Shivanandappa T (2013) Amelioration of cyclophosphamide-induced hepatotoxicity by the root extract of Decalepis hamiltonii in mice. Food Chem Toxicol 57:179–184. CrossRefGoogle Scholar
  71. Zhao R, Ma C, Tan L, Zhao X, Zhuang D (1994) The effect of acupuncture on the function of macrophages in rats of immunodepression. Zhen Ci Yan Jiu 19:66–68Google Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Clinical Pathology Department, Faculty of Veterinary MedicineMansoura UniversityMansouraEgypt
  2. 2.Clinical Pathology Department, Faculty of Veterinary MedicineSuez Canal UniversityIsmailiaEgypt

Personalised recommendations