Comparison of methods for detecting mitomycin C‐ and ethyl nitrosourea‐induced germ cell damage in mice: Sperm enzyme activities, sperm motility, testis weight

Abstract

Testes weights, sperm motility and enzyme activities in single sperm were compared with respect to their ability to detect either developmental or mutational damage to germ cells. Male mice were injected i.p. with 2.5 mg/kg mitomycin C (MC) or 50 or 100 mg/kg ethylnitrosourea (ENU) or saline and were then killed at times such that sperm derived from treated vas sperm (SZ), spermatids (ST), preleptotene-late-spermatogonial cells (PLSG), spermatogonial cells (SG), or spermatogonial stem cells (SGS) could be evaluated. Testis weights decreased significantly as early as 1 wk after treatment, with the greatest decrease reached 3-4 wk after treatment, followed by recovery to normal levels 10-15 wk after treatment. We conclude that testis weight, which is easily obtained, is a sensitive indicator of germ cell damage by these agents. Sperm from each animal were evaluated for sperm motility, acrosin activity, succinic dehydrogenase (SDH) activity with or without the competitive inhibitor malonate or after exposure to 60 degrees C for 10 min. The latter two assays were to detect sperm enzymes resistant to the inhibitor or heat. The presence of the acrosin protein was also detected immunologically. Sperm motility decreased most from treatment of PLSG and SG. After MC or ENU treatment, the greatest loss of acrosin activity and of the acrosin protein was also noted in sperm derived from treated PLSG and SG. MC and ENU failed to induce SDH activity in single sperm resistant to 60 degrees C heat inhibition or to inhibition by malonate. Of the sperm assays, acrosin activity proved to be the most sensitive indicator of germ cell damage and was the simplest to measure.

Male-mediated teratogenesis: Spectrum of congenital malformations in the offspring of A/J male mice treated with ethylnitrosourea

“…In contrast, preimplantation loss was significantly induced following mating with males treated at 5 x 50 mgkg. Preimplantation losses induced after treatment of spermatogonial stem cells are probably due to oligospermy or non-functioning sperm, reflecting a strong cytotoxic effect by ENU at the stem cell stage [12,13]. In this context, it should be noted that the values for the frequency of preimplantation losses given in Table I are underestimates because the number of corpora lutea was not counted in females with no implants.…”

Use of Sperm Enzymes to Detect Genotoxic Agents.

“…These multifunctional chemicals undoubtedly caused their effects in PLSG cells by a different mechanism than the one by which they affect already developed SZ (Ficsor et al, 1984a). Therefore, it is important to sample at least two populations of germ cells representing pre-and post-meiotic cells, as was done in the present study.…”

Initial Insights Regarding the Role of p53 in Maintaining Sperm DNA Integrity Following Treatment of Mice with Ethylnitrosourea or Cyclophosphamide

“…Numerous studies have shown that ENU alters spermatogenesis. Ficsor et al (1984) reported altered mouse sperm morphology forty-nine days after a single ip treatment with 100 mg/kg ENU. Skare and Schrotel (1984) have shown DNA single-strand breaks in rat testicular DNA following ip injection of 85.2 mg/kg ENU.…”