Pediatric Surgery International

, Volume 29, Issue 2, pp 179–184 | Cite as

Methotrexate induces germ cell apoptosis and impairs spermatogenesis in a rat

  • Igor SukhotnikEmail author
  • Omri Nativ
  • Alex Roitburt
  • Daniel Bejar
  • Arnold G. Coran
  • Jorge G. Mogilner
  • Ofer Nativ
Original Article



The primary toxic effects of methotrexate (MTX) are myelosuppression and/or intestinal mucositis. The objective of the present study is to investigate the effect of MTX on germ cell apoptosis and spermatogenesis in a rat.


Male Sprague–Dawley rats were divided into three experimental groups: control rats treated with vehicle; MTX-2 rats treated with one dose (20 μg/kg) of MTX given IP and killed on the second day; and MTX rats treated with IP MTX (20 μg/kg) and killed on day 4. Johnsen’s criteria and the number of germinal cell layers in the testes were used to categorize the spermatogenesis. TUNEL assay was used to determine germ cell apoptosis. Western blotting was used to determine Bax and Bcl-2 protein levels. Statistical analysis was performed using the non-parametric Kruskal–Wallis ANOVA test, with p less than 0.05 considered statistically significant.


On day 2, MTX-treated animals demonstrated minimal changes in the histological parameters of spermatogenesis, but germ cell apoptosis increased significantly (threefold increase, p = 0.002) compared to control rats. On day 4, MTX-treated rats demonstrated a trend toward a decrease in germ cell apoptosis, compared to day 2, and showed histological signs of impaired spermatogenesis (decreased number of germ cell layers and Johnsen’s criteria). A significant increase in cell apoptosis in MTX-treated rats was correlated with higher Bax/Bcl-2 protein levels.


MTX induced germ cell apoptosis and impaired spermatogenesis in rat testes.


Methotrexate Germ cell Apoptosis Spermatogenesis Rat 



This work was supported by a research grant from Israeli Ministry of Health No. 3/6164 from 01/06/2010.


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Copyright information

© Springer-Verlag Berlin Heidelberg 2012

Authors and Affiliations

  • Igor Sukhotnik
    • 1
    • 2
    Email author
  • Omri Nativ
    • 1
  • Alex Roitburt
    • 1
  • Daniel Bejar
    • 1
  • Arnold G. Coran
    • 4
  • Jorge G. Mogilner
    • 2
  • Ofer Nativ
    • 3
  1. 1.The Bruce Rappaport Faculty of Medicine, Laboratory of Intestinal Adaptation and RecoveryTechnion-Israel Institute of TechnologyHaifaIsrael
  2. 2.Department of Pediatric SurgeryBnai Zion Medical CenterHaifaIsrael
  3. 3.Department of UrologyBnai Zion Medical CenterHaifaIsrael
  4. 4.Section of Pediatric Surgery, C.S. Mott Children’s HospitalUniversity of Michigan Medical SchoolAnn ArborUSA

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