Document Type : Original Article


1 Pharmacy Student, Department of Basic Sciences, Pharmacy and Pharmaceutical Sciences, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran.

2 Department of Basic Sciences, Faculty of Pharmacy and Pharmaceutical Sciences, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran

3 Department of Biology, Faculty of Basic Sciences, Hamedan Branch, Islamic Azad University, Hamedan, Iran



Introduction and Aim:  One of the most well-known side effects of anticancer drugs is disruption of spermatogenesis, which in many cases causes infertility. The present study aimed to evaluate the expression change of Nrf-2 and Keap-1 genes due to peritoneal injection of busulfan in male Wistar rats.
Methods: In this study, 20 two-month-old adult male Wistar albino rats weighing approximately 200-150 g were studied in two groups: the control group (healthy) and the busulfan group. After this period, the testicular tissue of the mice was isolated and evaluated for the expression of Nrf-2 and Keap-1 genes, as well as changes in testosterone, FSH, and LH. Data were analyzed using SPSS software and a one-way ANOVA statistical test.
Results: The statistical analysis of the results showed that the expression of Nrf-2 and Keap-1 genes in the treatment group (busulfan) decreased compared to the control group, but this decrease was not statistically significant (0.2951 and P = 0.3528). LH and testosterone levels were not significantly different between the control and treated groups (0.1327 and P = 0.0809) but FSH levels were significant between the two groups (P = 0.0189).
Conclusion: The results of this study showed that busulfan can affect the level of FSH, which is effective in spermatogenesis, by causing changes in the expression of keap1 and Nrf-2 genes and causing infertility. Therefore, these genetic and hormonal factors can be used to predict infertility.


Main Subjects

Abidemi, A.A. 2011. Protective effect of tahitian noni juice on the reproductive functions of male wistar rats traeted with cyclophosphamide. Iranian Journal of Pharmacology and Therapeutics, 10(1): 39-30.
Adamian, M., Hekmat, A. and Hajebrahimi, Z. 2021. The impacts of simulated microgravity on the cell viability and claudin-1 and claudin-3 expression of mcf-7 breast cancer cells. Journal of Sciences, Islamic Republic of Iran, 32(2): 105-114.
Alahmar, A.T. 2019. Role of oxidative stress in male infertility: An updated review. Journal of human reproductive sciences, 12(1): 4.
Anjam, R.S., Movahedin, M., POURBEYRANVAND, S. and MOULA, S.J. 2007. Assessment of morphological and functional changes in the mouse testis and epididymal sperms following busulfan treatment.
Babu, S.R., Sadhnani, M., Swarna, M., Padmavathi, P. and Reddy, P. 2004. Evaluation of fsh, lh and testosterone levels in different subgroups of infertile males. Indian Journal of Clinical Biochemistry, 19(1): 45-49.
Chen, J., Bhandar, B. and Kavdia, M. 2015. Interaction of ros and rns with gsh and gsh/gpx systems. The FASEB Journal, 29: 636.637.
Choi, Y.-J., Ok, D.-W., Kwon, D.-N., Chung, J.-i., Kim, H.-C., Yeo, S.-M., Kim, T., Seo, H.-G. and Kim, J.-H. 2004. Murine male germ cell apoptosis induced by busulfan treatment correlates with loss of c-kit-expression in a fas/fasl-and p53-independent manner. FEBS letters, 575(1-3): 41-51.
Craft, I., Bennett, V. and Nicholson, N. 1993. Fertilising ability of testicular spermatozoa. The Lancet, 342(8875): 864.
Ferdosi, K.A., Bakhtiari, M., SOLEIMANI, R.J., Koroji, M., Roshangar, L., Janzadeh, A., Kerdari, M. and Jameie, B. 2013. Study of the effect of exogenous melatonin on sperm fertility in busulfan induced oligospermic of pinealectomeized rat.
Foresta, C., Bettella, A., Spolaore, D., Merico, M., Rossato, M. and Ferlin, A. 2004. Suppression of the high endogenous levels of plasma fsh in infertile men are associated with improved sertoli cell function as reflected by elevated levels of plasma inhibin b. Human Reproduction, 19(6): 1431-1437.
Forouzesh, F., Hajimirza Shafiesoltani, P., Ghiaghi, M. and Shabani, M. 2021. Anti-proliferative effect of sodium butyrate against caco-2 cell line of human colorectal cancer by reducing the expression ratio of bcl-2 to bax mrna. Research in Karyotic Cell & Tissue, 1(3): 16-24.
Gharagozloo, P., Gutiérrez-Adán, A., Champroux, A., Noblanc, A., Kocer, A., Calle, A., Pérez-Cerezales, S., Pericuesta, E., Polhemus, A. and Moazamian, A. 2016. A novel antioxidant formulation designed to treat male infertility associated with oxidative stress: Promising preclinical evidence from animal models. Human Reproduction, 31(2): 252-262.
Ghasemi, M., Bahadori, M., Faghani, M., Nasiri, E. and Soleimani Rad, J. 2009. Buserelin inhibits apoptosis in male germ cells induced by busulfan in mouse testis. J Iran Anat Sci, 7: 45-54.
Gutierrez, K., Glanzner, W.G., Chemeris, R.O., Rigo, M.L., Comim, F.V., Bordignon, V. and Gonçalves, P.B. 2016. Gonadotoxic effects of busulfan in two strains of mice. Reproductive Toxicology, 59: 31-39.
Hayes, J., Chanas, S., Henderson, C., McMahon, M., Sun, C., Moffat, G., Wolf, C. and Yamamoto, M. 2000. The nrf2 transcription factor contributes both to the basal expression of glutathione s-transferases in mouse liver and to their induction by the chemopreventive synthetic antioxidants, butylated hydroxyanisole and ethoxyquin. Portland Press Ltd.
Homma, S., Ishii, Y., Morishima, Y., Yamadori, T., Matsuno, Y., Haraguchi, N., Kikuchi, N., Satoh, H., Sakamoto, T. and Hizawa, N. 2009. Nrf2 enhances cell proliferation and resistance to anticancer drugs in human lung cancer. Clinical Cancer Research, 15(10): 3423-3432.
Itoh, K., Chiba, T., Takahashi, S., Ishii, T., Igarashi, K., Katoh, Y., Oyake, T., Hayashi, N., Satoh, K. and Hatayama, I. 1997. An nrf2/small maf heterodimer mediates the induction of phase ii detoxifying enzyme genes through antioxidant response elements. Biochemical and biophysical research communications, 236(2): 313-322.
Kaspar, J.W., Niture, S.K. and Jaiswal, A.K. 2009. Nrf2: Inrf2 (keap1) signaling in oxidative stress. Free Radical Biology and Medicine, 47(9): 1304-1309.
Matin-du-Pan, R. and Bischof, P. 1995. Increased follicle stimulating hormone in infetile men: Is increased plasma fsh always due to damaged germinal epithelium? Human Reproduction, 10(8): 1940-1945.
Nakamura, B.N., Lawson, G., Chan, J.Y., Banuelos, J., Cortés, M.M., Hoang, Y.D., Ortiz, L., Rau, B.A. and Luderer, U. 2010. Knockout of the transcription factor nrf2 disrupts spermatogenesis in an age-dependent manner. Free Radical Biology and Medicine, 49(9): 1368-1379.
Nieto, Y., Thall, P., Valdez, B., Andersson, B., Popat, U., Anderlini, P., Shpall, E.J., Bassett, R., Alousi, A. and Hosing, C. 2012. High-dose infusional gemcitabine combined with busulfan and melphalan with autologous stem-cell transplantation in patients with refractory lymphoid malignancies. Biology of Blood and Marrow Transplantation, 18(11): 1677-1686.
Olooto, W.E., Amballi, A.A. and Banjo, T.A. 2012. A review of female infertility; important etiological factors and management. J Microbiol Biotech Res, 2(3): 379-385.
Probin, V., Wang, Y. and Zhou, D. 2007. Busulfan-induced senescence is dependent on ros production upstream of the mapk pathway. Free Radical Biology and Medicine, 42(12): 1858-1865.
Qin, Y., Liu, L., He, Y., Wang, C., Liang, M., Chen, X., Hao, H., Qin, T., Zhao, X. and Wang, D. 2016. Testicular busulfan injection in mice to prepare recipients for spermatogonial stem cell transplantation is safe and non-toxic. PLoS One, 11(2): e0148388.
Schafer, M., Dütsch, S., Auf dem Keller, U. and Werner, S. 2010. Nrf2: A central regulator of uv protection in the epidermis. Taylor & Francis: pp: 2917-2918.
Simon, L., Proutski, I., Stevenson, M., Jennings, D., McManus, J., Lutton, D. and Lewis, S. 2013. Sperm DNA damage has a negative association with live-birth rates after ivf. Reproductive biomedicine online, 26(1): 6: 78-88.
Wu, C.-H., Yang, J.-G., Chang, Y.-J., Hsu, C.-C. and Kuo, P.-L. 2013. Screening of a panel of steroid-related genes showed polymorphisms of aromatase genes confer susceptibility to advanced stage endometriosis in the taiwanese han population. Taiwanese Journal of Obstetrics and Gynecology, 52(4): 485-492.
Wu, C., Zhang, Y., Shen, Q., Zhou, Z., Liu, W. and Hua, J. 2016. Resveratrol changes spermatogonial stem cells (sscs) activity and ameliorates their loss in busulfan-induced infertile mouse. Oncotarget, 7(50): 82085.