Method for testing mutagenic effects of chemicals on spermatogonia of the Chinese hamster: results obtained with cyclophosphamide, saccharin, and cyclamate.

The action of different cyclophosphamide doses on spermatogonia of the Chinese hamster was examined. Two oral treatments at an interval of 24 h were carried out and spermatogonia were prepared for examination 24 or 48 h after the second dose. Accordingly the effects of 5 oral cyclophosphamide doses given on five consecutive days were tested on spermatogonia and preparations were made 24 or 72 h after the last treatment. The results so obtained form the basis of reference for findings following oral administration of saccharin sodium, sodium cyclamate, or trimethylphosphate. Male Chinese hamsters, 6-8 per group, were used, from each of which about 100 metaphases were evaluated. Preparation was carried out essentially according to Hoo and Bowles [Mutation Res. 13, 85-88 (1971)]. gaps, breaks, fragments, deletions and translocations were rated as structural aberrations. For every dose and every time of preparation the incidence of metaphases with aberrations, with or without gaps, and with translocations were assessed. The different experiments led to the following conclusions: 1. By analysis of spermatogonial metaphases of treated Chinese hamsters chemically induced chromosome aberrations can be proved with certainty. 2. Incidence of metaphases with translocations is a sensitive measure which is distinctly superior to the summary determination of all aberrations. In this way it was possible to show a mutagenic influence of 2 times 8 mg/kg cyclophosphamide p.o. 3. Following two cyclophosphamide doses administered at an interval of 24 h it was found that preparations of spermatogonia 48 h after the second dose was better suited for the evaluation than that at 24 h, for aberrations were more frequent with the same treatment. After five cyclophosphamide treatments at 24-h intervals, aberrations were somewhat more frequent 24 h after the last dose than at 72 h; in any case the values exceeded significantly the results of untreated controls. 4. A conclusive numeric chromosome analysis is not possible with the spermatogonia test, since a relatively high percentage of non-diploid cells is apparently of methodological origin. 5. Tests with 2 times 500 or 5 times 1000 mg/kg trimethylphosphate orally showed an increase in chromosome aberrations compared with controls indicating mutagenic effects in both cases; with 5 times 1000 mg/kg p.o., however, the figures were low as a result of marked mitotic inhibition. 6. The results of the spermatogonia test on Chinese hamsters revealed no mutagenic effects of saccharin sodium 2 times 5000 mg/kg orally, and of sodium cyclamate 5 times 2000 mg/kg orally. This is based on comparisons of the results both with untreated controls and positive controls treated with trimethylphosphate or cyclophosphamide.