Mouse Spermatogonia Research Articles

Dose-effect relationship for X-ray-induced reciprocal translocations in mouse spermatogonia following pretreatment with 3-aminobenzamide

The influence of 3-aminobenzamide (3-AB) pretreatment on the dose-response relationship for radiation-induced reciprocal translocations in mouse spermatogonial stem cells was studied. The results show that at doses of 3–10 Gy of X-rays the frequencies of translocations were higher in 3-AB-pretreated animals as compared to animals that received X-rays only. The 3-AB pretreatment was not effective at dose levels of 1 and 2 Gy. The shape of the dose-effect curve was similar to that obtained without 3-AB pretreatment, i.e., a humped curve, but the initial slope was clearly steeper and the position of the peak was shifted from 7 to 9 Gy. The effects observed can be explained by a 3-AB-mediated sensitization of normally radioresistant stem cells that are at the stage of stimulation to enter the mitotic cycle, thus increasing the population of radiosensitive spermatogonial stem cells.

Inhibition by radioprotectors of radiation-induced chromosome damage in mouse spermatogonia

Inhibition by radioprotectors of radiation-induced chromosome damage in mouse spermatogonia

Towards an understanding of the nature and fitness of induced mutations in germ cells of mice: homozygous viability and heterozygous fitness effects of induced specific-locus, dominant cataract and enzyme-activity mutations

A total of 219 specific-locus, 35 dominant cataract and 44 enzyme-activity mutations induced in spermatogonia of mice by radiation or ethylnitrosourea (ENU) treatment were characterized for homozygous viability as well as fitness effects on heterozygous carriers. For all 3 genetic endpoints, the frequency of homozygous lethal mutations was higher in the group of radiation-induced mutations than in the ENU-treatment group. The observations are consistent with the hypothesithat radiation-induced mutations recovered in the mouse are mainly due to small deletions while ENU induces mainly intragenic mutations. The overall fitness of mutant heterozygotes was reduced for the group of radiation-induced specific-locus, dominant cataract and enzyme-activity mutations while the ENU-induced mutations exhibited no reduction in fitness. The fitness reduction of heterozygous carriers for a newly occurring mutation in a population is important in determining the persistence of the mutation in a population, and thus the total number of individuals affected before a mutation is eventually eliminated from the population. For the present results a maximal persistence of 12 generations and a minimal persistence of 3 generations is estimated. These results are consistent with the 6–7-generation persistence time assumed by UNSCEAR (1982) in an estimate of the overall effects of radiation-induced mutations in man.

N-Propyl- N-nitrosourea-induced recessive mutations in mouse spermatogonia

N-Propyl- N-nitrosourea-induced recessive mutations in mouse spermatogonia

Induction of specific-locus and dominant-lethal mutations by cyclophosphamide and combined cyclophosphamide-radiation treatment in male mice

Cyclophosphamide is the most widely used antineoplastic agent. It is also used to condition patients for bone-marrow transplantations. Because of the general interest of this compound we initiated a systematic study of the induction of dominant-lethal and specific-locus mutations in male mice. In addition, we investigated the induction of specific-locus mutations by the combined treatment of cyclophosphamide and ionizing radiation. A dose of 40 mg/kg bw of cyclophosphamide caused dominant-lethal mutations in male mice only in the 1st and 2nd week after treatment. A dose of 120 mg/kg induced dominant-lethal mutations in the mating intervals 1–21 days posttreatment. No dominant lethal mutations were observed after the 3rd week. The same differential spermatogenic response was observed for the induction of specific-locus mutations. Cyclophosphamide induced recessive mutations exclusively in spermatozoa and spermatids. No mutations were recovered from treated spermatocytes and spermatogonia. In contrast to cyclophosphamide, radiation induces specific-locus mutations in all germ-cell stages. The pretreatment with cyclophosphamide 24 h before radiation enhanced the frequency of specific-locus mutations in spermatogonia. The distribution of the observed mutations among the 7 loci and their viability supports the hypothesis that these mutations were induced by radiation rather than by cyclophosphamide. The compound causes an immediate inhibition of DNA and RNA synthesis in spermatogonia. The inhibition very likely interferes with the repair process. The disturbance of the repair process is probably the cause of the synergistic effect for the induction of specific-locus mutations in spermatogonia of mice after pretreatment with cyclophosphamide 24 h before irradiation.

Induction of specific-locus and dominant-lethal mutations by cyclophosphamide and combined cyclophosphamide-radiation treatment in male mice

Cyclophosphamide is the most widely used antineoplastic agent. It is also used to condition patients for bone-marrow transplantations. Becaused of the general interest of this compound we initiated a systematic study of the induction of dominant-lethal and specific-locus mutations in male mice. In addition, we investigated the induction of specific-locus mutations by the combined treatment of cyclophosphamide and ionizing radiation. A dose of 40 mg/kg bw of cyclophosphamide caused dominant-lethal mutations in male mice only in the 1st and 2nd week after treatment. A dose of 120 mg/kg induced dominant-lethal mutations in the mating intervals 1–21 days posttreatment. No dominant lethal mutations were observed after the 3rd week. The same differential spermatogenic response was observed for the induction of specific-locus mutations. Cyclophosphamide induced recessive mutations exclusively in spermatozoa and spermatids. No mutations were recovered from treated spermatocytes and spematogonia. In contrast to cyclophosphamide, radiation induces specific-locus mutations in all germ-cell stages. The pretreatment with cyclophosphamide 24 h before radiation enhanced the frequency of specific-locus mutations in spermatogonia. The distribution of the observed mutations among the 7 loci and their viability supports the hypothesis that these mutations were induced by radiation rather than by cyclophosphamide. The compound causes an immediate inhibition of DNA and RNA synthesis in spermatogonia. The inhibition very likely interferes with the repair process. The disturbance of the repair process is probably the cause of the synergistic effect for the induction of specific-locus mutations in spermatogonia of mice after pretreatment with cyclophosphamide 24 h before irradiation.

Induction of translocations in mouse spermatogonia after fractionated exposure to 60Co γ-rays

Male mice of the Q strain were exposed to 60 Co γ-rays at 2 Gy and 2 × 2 Gy separated by increasing time intervals (from 0 min to 4 min). The chromosome translocations induced in spermatogonia were scored at diakinesis-metaphase I. A significant decrease of the translocation frequency at time intervals higher than 2 min was observed, confirming results obtained with plant materials.

Induction of translocations in mouse spermatogonia after fractionated exposure to 60Co γ-rays

Male mice of the Q strain were exposed to 60Co γ-rays at 2 Gy and 2 × 2 Gy separated by increasing time intervals (from 0 min to 4 min). The chromosome translocations induced in spermatogonia were scored at diakinesis-metaphase I. A significant decrease of the translocation frequency at time intervals higher than 2 min was observed, confirming results obtained with plant materials.

Identification of the specific proteins associated with differentiation of spermatogonia in mice by two-dimensional gel electrophoresis.

Protein synthesis in testicular germ cells was studied during the differentiation of Type A spermatogonia in testes from adult cryptorchid mice to Intermediate and Type B spermatogonia in organ culture by two-dimensional gel electrophoresis. By comparing the patterns of Type A spermatogonia with those of differentiated (Intermediate and Type B) spermatogonia, we found two newly synthesized protein spots that correlated with differentiation. The isoelectric points (pIs) and molecular weights (MWs) of these proteins were estimated to be approximately pI 5.4, MW 67,000, and pI 5.8, MW 28,000, respectively.

Dominant lethal mutations induced in mouse spermatogonia by mechlorethamine, procarbazine and vincristine administered in 2-drug and 3-drug combinations

Male mice were treated with mechlorethamine (2.0 mg/kg), procarbazine (100 mg/kg) and vincristine (0.67 mg/kg) alone, or in 2-drug and 3-drug combinations. 5 weeks later and continuing for 5–8 weeks, embryos fertilized by spermatozoa that were derived from drug-treated spermatogonia were evaluated for drug-induced dominant lethal mutations. Significant mutagenesis was detected for mechlorethamine alone, for 2-drug combinations including mechlorethamine and for 3-drug combinations. Combinations where mechlorethamine was given first were mutagenic whereas combinations where mechlorethamine was not given first were not. Some combinations were more cytotoxic to the germinal epithelium than others. The data suggest that mutagenesis by this combination of drugs which is used extensively in treating Hodgkin's disease is due primarily to the mechlorethamine and that the frequency of mutation-induction may be a function of the order of administration.

Antimutagenic properties of WR 2721 and of a radioprotective mixture, ATP-AET-serotonin, with regard to X ray induced reciprocal translocations in mouse spermatogonia

Pretreatment by intraperitoneal administration of WR 2721 at 400 mg/kg body weight in mice receiving 4.0 Gy X rays was found to have an appreciable antimutagenic effect with regard to reciprocal translocation induction in spermatogonia. The effectiveness of the product tested proved superior to that of a radioprotective mixture of ATP-AET-serotonin given at optimal dose ratio—360, 24, and 8 mg/kg body weight, respectively. The RF (Reduction Factor) was 2.4 for WR 2721 and 1.8 for the mixture. The effect observed indicated WR 2721 to have potential capabilities for reducing the genetic risk of radiation in male individuals.

Antimutagenic properties of selected radioprotective drug mixtures with regard to X-ray-induced reciprocal translocations in mouse spermatogonia

The radioprotective drugs AET, serotonin, and ATP were tested for antimutagenic activity against induction by 4.0 Gy X-rays of reciprocal translocations in mouse spermatogonia. Single drugs administered in doses of 8, 24 and 360 mg/kg b.wt., respectively, had no effect on translocation yields recorded in diakinesis-metaphase I spermatocytes. Two-drug mixtures afforded insignificant protection. Three-drug mixtures, however, were found to reduce radiation damage considerably, and the extent of protection was dependent in part on the amount of ATP. The best effect was obtained with formulations of serotonin-AET-ATP at the following doses, respectively: 8 + 24 + 360 mg/kg, 16 + 24 + 336 mg/kg, and 16 + 32 + 264 mg/kg. Less effective were the serotonin-AET-ATP formulations: 16 + 32 + 120 mg/kg, and 8 + 24 + 480 mg/kg. Treatment with drugs omitting radiation exposure was observed to raise, though insignificantly, the level of spontaneous translocation frequency.

Occurrence of premature chromosome condensation in mouse spermatogonia treated with busulfan.

Busulfan induction of premature chromosome condensation (PCC) was examined in mouse spermatogonia. Adult (C3H X SWV) F1 male mice were injected intraperitoneally with busulfan at 10 or 30 mg/kg of body weight and killed 18-72 h later. Polyploid-like mitoses were frequent (ca. 1/4 of all spermatogonial mitoses examined) in both the untreated and treated groups. Most of these were considered to have been derived from normal spermatogonia. In busulfan-treated mice, polyploid-like mitoses with PCC were seen. The frequency of PCC-containing mitoses increased with increasing dose and exposure time. A possible interpretation for PCC induction in mouse spermatogonia is discussed.

Modification of radioresponse of intermediate and B-type spermatogonia by a phosphorothioate.

Spermatogonia of mouse present a wide range of radiosensitivity. Among the three major types of spermatogonia, intermediate and B-type spermatogonia are the more radiosensitive groups having comparable radiosensitivity (Oakberg, 1955; Partington et al, 1962). In the present study, radioresponse of intermediate (I) and B-type spermatogonia was observed in vivo with and without pretreatment of S-2 (3- aminopropylamino) ethylphosphorothioic acid, popularly known as WR-2721. It is an effective radioprotector known to alter radiation effects in a number of tissues in vivo and in vitro (Yuhas & Storer, 1969) including mouse testes (Milas et al, 1982). It also protected A-type spermatogonia of the mouse in similar conditions, as in the present experiment (Jaimala et al, 1984). The present investigation seeks to ascertain the extent to which this drug could alter the radioresponse of intermediate and B-type spermatogonia after exposure to different doses of gamma radiation.

Dose-repetition increases the mutagenic effectiveness of N-ethyl-N-nitrosourea in mouse spermatogonia.

In order to maximize the mutagenic effectiveness of N-ethyl-N-nitrosourea in mouse stem-cell spermatogonia, advantage was taken of the fact that these cells can accumulate mutations from repeated doses given over relatively long time periods. Repeated doses (100 mg/kg) of ethylnitrosourea injected intraperitoneally into male mice at weekly intervals were found to allow adequate survival and fertility with total dosages of 300 and 400 mg/kg. The specific-locus mutation frequencies at these dosages were, respectively, 1.8 and 2.2 times that obtained with the maximal practicable single dose of 250 mg/kg. The mutation frequency induced by a 400 mg/kg dosage of ethylnitrosourea is 12 times the maximal mutation frequency achievable with a single exposure to x-rays and 36 times that reported for procarbazine, the most effective chemical mutagen previously known for mouse stem-cell spermatogonia. Ethylnitrosourea is already the mutagen of choice in deliberate attempts to create mouse models for human disease and in any experiments in which a maximal mutation rate is desired. Repeated-dose regimens similar to the ones reported here should increase the efficiency of such studies.

Dominant lethal mutations induced in mouse spermatogonia by antineoplastic drugs

Male mice were treated with nitrogen mustard (2.25 mg/kg), procarbazine (100 mg/kg) or vincristine (0.67 mg/kg). 5 weeks later they were mated to untreated females and the dominant lethal mutant rate was determined in the offspring by an in vitro analysis. Nitrogen mustard and procarbazine induced dominant lethal mutations to significant levels. Mutations were expressed, throughout early development, especially as a failure to form a trophectoderm outgrowth and to differentiate and proliferate an inner cell mass. Variations in the dominant lethal mutant rate were found from one week to the next, suggesting that differentiated and stem cell spermatogonia have differential sensitivity to mutation induction and cell killing by each drug.

No synergism between caffeine and arsenic in the induction of chromosomal aberrations in bone-marrow cells and spermatogonia of mice

No synergism between caffeine and arsenic in the induction of chromosomal aberrations in bone-marrow cells and spermatogonia of mice

Effects of unequally fractionated X-ray exposures on the induction of chromosomal rearrangements in mouse spermatogonia

The induction of reciprocal translocations in mouse stem-cell spermatogonia was determined following different unequally fractionated X-ray exposures with 24 h between the fractions. The results indicate that quite variable levels of chromosomal damage can be induced using the same exposure schemes. This variation does not seem to be correlated with radiation set-up or mouse strain. In general, with higher conditioning exposures from 12.5 to 300 rad (or R), higher frequencies of translocations were produced using a larger second challenging exposure of 700–900 rad (or R). The increase in yields of aberrations was more or less paralleled by a shift from a strong deviation from a Poisson fit of the number of translocations per spermatocyte, to a good fit, suggesting a transition from the original heterogeneity of the stem cell population to a radiation induced, more homogeneous stage. Earlier observations concerning a threshold dose for sensitization (van Buul and Léonard, Mutation Res., 70 (1980) 95–101) could not be confirmed.

Sensitivity of somatic tissues and spermatogonia of the mouse for induction of point mutations by alkylating agents

Sensitivity of somatic tissues and spermatogonia of the mouse for induction of point mutations by alkylating agents

Genotoxicity of garlic, turmeric and asafoetida in mice

The genotoxic effects of orally administered garlic and turmeric were evaluated in bone-marrow cells of mice by performing the micronucleus test. Another spice, asafoetida, was tested for the induction of sister-chromatid exchanges (SCEs) in spermatogonia of mice. Results of the micronucleus test with garlic and turmeric were not significantly different from control values. Orally administered asafoetida, however, showed a weak SCE-inducing effect in spermatogonia.