Possible protective effects of crocin on destructive side effects of cyclo-phosphamide in mice ovarian tissue: Evaluation of histomorphometrical and biochemical changes

Document Type : Original Article


Department of Basic Sciences, Faculty of Veterinary Medicine, Urmia University, Urmia, Iran


One of the side effects of cyclophosphamide (CP) is low fertility. In this study, we investigated the protective role of crocin (Cr) against CP chemotherapy-induced changes in ovarian tissue. In the current study, we treated 15 female mice aged 6-8 weeks old for 21 days. The mice were distributed into three groups including control received normal saline (0.10 mL; IP), CP or sham-control group (CP once a week, 15.00 mg kg-1; IP) and experimental (CP + Cr) group received CP along with Cr (200 mg kg-1 daily; IP). After completing the procedure, levels of total anti-oxidant capacity (TAC), superoxide dismutase (SOD) and sex hormones in serum as well as malondialdehyde (MDA) in the left ovarian tissue were measured. The right ovaries were used for histological and morphological tests. The obtained data were statistically analyzed by SPSS software using ANOVA and Tukey follow-up studies. Results showed that in the CP group a significant decrease was observed in ovarian follicles, the number of corpus luteum, levels of TAC, SOD and sex hormones; while, there was a significant increase in the number of atretic follicles and mast cells and level of MDA compared to control group. Administration of Cr along with CP caused a significant ameliorative effect on the studied parameters. In conclusion, the Cr could significantly decrease the side effects caused by CP chemotherapy in mice ovarian tissue.


  1. Himelstein-Braw R, Peters H, Faber M. Morphological study of the ovaries of leukaemic children. Br J Cancer 1978; 38 (1): 82-87.
  2. Khan Mohammadi Ghane F, Ahmadi A, Shahrooz R, et al. Evaluation of protective effects of ethyl pyruvate on embryo developing process in in vitro fertilization (IVF) in cyclophosphamide treated mice. Urmia Med J 2014;25(8):760-768.
  3. Vaskivuo T. Regulation of apoptosis in the female reproductive system. Thesis, University of Oulu. Oulu, Finland: 2002.
  4. Ben-Yehuda D, Krichevsky S, Caspi O, et al. Microsatellite instability and p53 mutations in therapy- related leukemia suggest mutator phenotype. Blood 1996;88(11):4296-4303.
  5. Apperley JF, Reddy N. Mechanism and management of treatment-related gonadal failure in recipients of high dose chemoradiotherapy. Blood Rev 1995; 9 (2):93-116.
  6. Abdoon AS, Kandil OM, Otoi T, et al. Influence of oocyte quality, culture media and gonadotropins on cleavage rate and development of in vitro fertilized buffalo embryos. Ani Reprod Sci 2001;65(3-4):215-223.
  7. Kianbakht S, Mozaffari K. Effects of saffron and its active constituents, crocin and safranal, on prevention of indomethacin induced gastric ulcers in diabetic and nondiabetic rats. J Med Plants 2009;8(Suppl 5):30-38.
  8. Abe K, Saito H. Effects of saffron extract and its constituent crocin on learning behavior and longā€term potentiation. Phytother Res 2000; 14(3):149-152.
  9. Zarei Jaliani H, Riazi GH, Ghaffari SM, et al. The effect of the crocus sativus L. carotenoid, crocin, on the polymerization of microtubules, in vitro. IJBMS 2013;16(1):101-107.
  10. Alizadeh F, Bolhassani A. In vitro cytotoxicity of Iranian saffron and two main components as a potential anti-cancer drug. SM J Pharmac Ther 2015;1(1):1001.
  11. Hosseinzadeh H, Younesi H. Petal and stigma extracts of Crocus sativus L. have antinociceptive and anti-inflammatory effects in mice. BMC Pharmacol 2002; 2:1-8.
  12. Hosseinzadeh H, Khosravan V. Anticonvulsant effects of aqueous and ethanolic extracts of Crocus sativus L. stigmas in mice. Arch Irn Med 2002; 5(1):44-47.
  13. Hosseinzadeh H, Karimi G, Niapoor M. Antidepressant effect of Crocus sativus L. stigma extracts and their constituents, crocin and safranal, in mice. Acta Hortic. 2004;650:435-445;
  14. Esalatmanesh S, Biuseh M, Noorbala AA, et al. Comparison of saffron and fluvoxamine in the treatment of mild to moderate obsessive-compulsive disorder: a double blind randomized clinical trial. Iran J Psychiatry 2017; 12(3):154-162.
  15. Zargari A. Medicinal plants [Persian]. Tehran, Iran: Tehran University Press 1990; 574.
  16. Selvakumar E, Prahalathan C, Sudharsan PT, et al. Protective effect of lipoic acid on cyclophosphamide-induced testicular toxicity. Clinia Chimica Acta2006; 367(1-2): 114-119.
  17. Hesari AK, Shahrooz R, Ahmadi A, et al. Crocin prevention of anemia-induced changes in structural and functional parameters of mice testes. J Appl Biomed 2015;13(3):213-223
  18. Humson GL. Animal tissue techniques. 4th ed. San Francisco, USA: W. H. Freeman, Macmillan Publishers 1979; 137-138.
  19. Benzie IF, Strain JJ. Ferric reducing/antioxidant power assay: direct measure of total antioxidant activity of biological fluids and modified version for simultaneous measurement of total antioxidant power and ascorbic acid concentration. Methods Enzymol 1999; 299: 15-27.
  20. Garcia YJ, Rodríguez-Malaver AJ, Peñaloza N. Lipid peroxidation measurement by thiobarbituric acid assay in rat cerebellar slices. J Neurosci Methods 2005; 144(1):127-135.
  21. Niehaus Jr WG, Samuelsson B. Formation of malondialdehyde from phospholipid arachidonate during microsomal lipid peroxidation. Eur J Biochem 1968; 6(1):126-130.
  22. Lowry OH, Rosebrough NJ, Farr AL, et al. Protein measurement with the Folin phenol reagent. J Biol Chem 1951; 193(1):265-275.
  23. Hassanpour A, Yousefian S, Askaripour M, et al. Ovarian protection in cyclophosphamide-treated mice by fennel. Toxicol Rep 2017;4: 160-164.
  24. Johari H. Mahmoudinejad F, Amjad G. Evaluate the effect of ginger extract on the axis hypothalamus - pituitary - gonadal axis in adult female rats treated with cyclophosphamide [Persian]. J Pzhvda 2011; 6 (20): 62-70.
  25. Agarwal A, Saleh RA, Bedaiwy MA. Role of reactive oxygen species in the pathophysiology of human reproduction. Fertil Steril 2003; 79(4):829-843.
  26. Eurell JA, Frappier BL. Dellmann's textbook of veterinary histology. 6th ed. New Jersey, USA: John Wiley & Sons 2006; 256-278.
  27. Ahmadi A, Chafjiri SB, Sadrkhanlou RA. Effect of satureja khuzestanica essential oil against fertility disorders induced by busulfan in female mice. Vet Res Forum 2017; 8(4): 281-286.
  28. Das UB, Mallick M, Debnath JM, et al. Protective effect of ascorbic acid on cyclophosphamide-induced testicular gametogenic and androgenic disorders in male rats. Asian J Androl 2002; 4(3):201-207.
  29. Li D-J, Xu Z-S, Zhang Z-H, et al. Antagonistic effects of vitamin E on the testicular injury by cyclophosphamide in mice [Chinese]. Zhonghua Nan Ke Xue 2006; 12(4):318-322.
  30. Douglas DC, Nakhuda GS, Sauer MV, et al. Angiogenesis and ovarian function. J Fertil Reprod 2005; 13(4): 7-15.
  31. Mescher AL. Junqueira's basic histology: text and atlas. 13th ed. California, USA: McGraw-Hill, 2013; 449-477.
  32. Tsai-Turton M, Luong BT, Tan Y, et al. Cyclo-phosphamide-induced apoptosis in COV434 human granulosa cells involves oxidative stress and glutathione depletion. Toxicol Sci 2007; 98(1):216-230.
  33. Chaudière J, Ferrari-Iliou R. Intracellular antioxidants: from chemical to biochemical mechanisms. Food Chem Toxicol 1999; 37(9-10):949-962.
  34. Krishna A, Terranova PF. Alterations in mast cell degranulation and ovarian histamine in the proestrous hamster. Biol Reprod 1985; 32(5):1211-1217.
  35. Karaca T, Yörük M, Uslu S. Distribution and quantitative patterns of mast cells in ovary and uterus of rat. Arch Med Vet 2007; 39(2):135-139.
  36. Prussin C, Metcalfe DD. 4. IgE, mast cells, basophils, and eosinophils. J Allergy Clin Immunol 2003; 111(S2): S486-S494.
  37. Roaiah MMF, Khatab H, Mostafa T. Mast cells in testicular biopsies of azoospermic men. Andrologia 2007; 39(5):185-189.