Time-dependent toxic effects of N-ethyl-N-nitrosourea on the testes of male C57BL/6J mice: a histological and ultrastructural study.

N-ethyl-N-nitrosourea (ENU), a well known alkylating agent, is a powerful mutagen in mouse spermatogonia that is frequently used to generate mutant mice for the study of gene function. The present study was performed to investigate the toxic effects of a suggested ENU treatment protocol (100 mg/kg ENU once a week for three consecutive weeks) on the C57BL/6J mouse testis using light and transmission electron microscopy, with reference to testis weight and sperm count. Time-dependent changes in the weight of the testes, sperm counts and testicular morphology were observed, following an injury and recovery pattern; the most severe damage was observed in week four after the first injection of ENU, and then the testis gradually recovered. By the end of the experiment (week 12), the testis weights and sperm counts of the ENU-treated mice had restored to around 80% of the respective values in the control group. Histopathological alterations in the testis were identified by light and electron microscopy, which revealed that ENU led to a temporary depletion in the number of spermatogenic cells via direct and indirect toxic effects, including apoptosis and growth arrest in spermatogonia, Sertoli cell damage and peritubular cell injury. The results of this study complement the existing basic information on the toxicity of ENU in the testis, and provide scientific information for selecting the appropriate mating time for ENU-treated male mice.