Mouse minisatellite mutations induced by ionizing radiation.

The detection of changes in germline mutation rate in human populations remains extremely difficult. Estimating the genetic hazards of radiation and other mutagens in humans therefore depends on extrapolation from experimental systems. Because of the very low frequency of spontaneous mutation at most loci, enormous samples are required to detect increases of mutation rate. A very high rate of spontaneous germline mutation altering the length of minisatellite loci has been found in human populations and therefore this system might be useful for detecting induced mutations in relatively small samples. Here we present evidence that minisatellite mutation rate in mice is increased by low doses of ionizing radiation.

[The 1xC3 panel of recombinant inbred mouse strains. Presence or absence of pathologic neurologic traits of one of parental strains].

Mice from the earlier developed recombinant inbred strains (RIS), which were derived by crossing 101/HY mice (carrying the mut-1 allele determining increased susceptibility to the mutagenic action of alkylating compounds) with C3H/Sn mice (lacking this trait), were tested for the presence of two neurological pathologies, audiogenic epilepsy and splitting of pyramidal cell layer of the CA3 hippocampal field (specific only to 101/HY mice). It was demonstrated that segregation of RIS relative to these traits was independent from the presence or absence of the mut-1 allele. These findings suggested the appearance of mut-1-independent mutations in the 101/HY mice, which resulted in the development of neurological pathologies. The appearance of such mutations can be the consequence of the genetic repair defects, earlier observed in the mice with the same genotype, or they can be caused by other reasons.

[The cellular effects of functional unloading and passive stretch on m. soleus of dystrophin-deficient mdx mice].

Dystrophin, subsarcolemmal protein communicating muscle fiber cytoskeleton to extracellular matrix, is believed to participate in mechanical signal transduction. Recent works testify possible signaling role of this protein to prevent development ofproteolytic processes accompanying muscle fiber atrophy and to stimulate the passive stretch anabolic effects. The experiment was carried out to assess the role of dystrophin in these processes. The study was performed on two months old C57 black and mdx (dystrophin-deficient) mice. Passive stretch resulted in attenuating atrophy development in two fiber types of both C57 black and mdx mice, at the same time fiber type slow-to-fast transformation did not occur in mdx soleus. We established ablatitious effect of chronic hindlimb unloading on SC proliferative activity in soleus muscle and drastic increase of proliferation under effect of passive stretch. We observed no relationship between altered dystrophin synthesis and satellite cell proliferation activity in soleus muscle under conditions of simulated microgravity and concurrent passive stretch. It is concluded that altered dystrophin synthesis partly retarded slow myofibers atrophy and had virtually no effect on passive stretch preventive action. Thus, the hypothesis about dystrophin key role in downregulation of atrophy signaling mechanisms has not found its confirmation concerning gravitational unloading atrophy.

[Ribosomal genes in inbred mouse strains: interstrain and intrastrain variations of copy number and extent of methylation].

Quantitative dot hybridization was used to estimate the rDNA copy number in brain tissues of five inbred mouse strains (AKR/JY, NZB/B1OrlY, CBA/CaLacY, 101/HY, and 129/JY), which were obtained from the collection of the Research Center of Biomedical Technologies (Y). In each strain, 9-12 mice aged 1-2 months were examined. The rDNA copy number per diploid genome in strains AKR (range 105-181, mean +/- SD 136 +/- 27) and NZB (129-169, 148 +/- 12) was significantly lower than in strains CBA (172-267, 209 +/- 31), 101 (179-270, 217 +/- 30), and 129 (215-310, 264 +/- 33). Mice of strain NZB were relatively homogeneous in this trait (CV = 8.1%). Strains AKR, CBA, 101, and 129 displayed significant between-group differences, CV varying from 12.5 to 19.9%. The same DNA specimens were digested with MspI or HpaII and used to estimate the extent of methylation of the 28S rDNA region. Regardless of the strain, all mice could be classed into two groups. One group (20 mice) had a methylated fraction accounting for less than 8% of rDNA and included all nine mice of strain NZB, seven out of nine mice of strain 101, and three out of ten mice of strain 129. In the other group (29 mice), the methylated fraction varied from 18 to 38%. A possible role of methylation and the genome dosage of ribosomal genes in phenotypic variation (quantitative trait variation) of inbred mouse strains is discussed.

[The effect of mutation dominant spotting-Yurlovo (KitW-Y) on spermatogenesis, early embryogenesis, and fertility of C57BL/6JY mice].

The effect of mutation KitW-Y found in C57BL/6 mice on fertility, spermatogenesis, and early embryogenesis of mice have been studied. If heterozygotes KitW-Y/+ are crossed with wild-type mice, fertility decreases by 20%. Homozygotes Kitw-Y/KitW-Y and compounds KitW-Y/KitSsm are nonviable. The study of spermatogenesis in KitW-Y/+ mice has demonstrated a negative effect of this mutation on spermatocytes. Histological examination of the testes of mutant males has shown local empty spaces in seminal ducts. Electron microscopic examination of synaptonemal complexes have demonstrated desynapsis disturbance in some nuclei at the diplotene stage of meiotic prophase I. However, these disturbances do not cause a decrease in the number of fertilized oocytes/ova. The decrease in fertility is accounted for disturbances of early embryogenesis. In vivo and in vitro analyses of early embryogenesis have demonstrated that cleavage divisions are asynchronous in KitW-Y/+ heterozygous embryos. Some of these embryos die before implantation, and others cleave more rapidly than wildtype embryos, which give them selective advantage during the postimplantation period of embryogenesis. The pattern of KitW-Y expression during spermatogenesis and embryogenesis mimics potential human pathology, which makes these mutants an interesting and valuable object for genetics and developmental biology.

Cytogenetic analysis of translocations induced by the chemical mutagen thio TEPA in spermatids of male mice.

Cytogenetic analysis of meiotic cells at diakinesis/metaphase I was carried out in 36 reciprocal translocation heterozygotes. Translocations were induced in late spermatids of C57BL/6 male mice with 1:25 mg/kg body weight of thioTEPA. The prevalent types of multivalent configuration were Ring IV in 15F1 males, chain IV in 20 males, and one male showed the same proportion of cells with chains and rings. The conclusion can be made that an intensive mutation process with may induce as much as 30% heterozygous translocation males in characterized by a random distribution of breaks along the chromosome.

Cyclophosphamide-induced chromosomal aberrations in meiotic cells of male mice.

Single doses of 40, 80 and 160 mg of cyclophosphamide per kg body weight were administered intraperitoneally to C57BL/6 male mice. Cytogenetic analysis of spermatocytes at diakinesis/metaphase I, influenced as spermatogonia A and B and pachytene spermatocytes, showed the chromosomal damage. Cyclophosphamide induced structural chromosomal aberrations including translocations and interfered with the normal development of bivalents. The dose of 80 mg/kg yielded 0.6% spermatocytes with translocations and 3% translocation carriers in F1 males. Robertsonian translocations were detected in 3 males.

[Effects of para-aminobenzoic acid on radiosensitivity of mice of different strains]. / Vliianie paraaminobenzoinoi kisloty na radiochuvstvitel'nost' myshei raznykh linii.

The aim of the work was to study the possible protective effect of para-aminobenzoic acid (PABA) on the radiation lethality in mice of three inbred lines (BALB/cLacY, C3H/HeY, 101/Hy), stock YT1 and hybrids (C3H/He x 101/H)F1. The PABA solution was given to the mice intraperitoneally in single doses of 10, 50 and 100 mg/kg 40-50 min prior to irradiation with doses of 6 to 8 Gy depending on the line and sex of mice. The used doses of gamma-radiation were roughly LD75/30. The radioprotective effect of PABA was observed in all variants of the experiment but it was relatively low. The protection coefficient varied from 0 to 0.45. The protective effect depended on the line and sex of mice and on the dose of the injected substance.

[Specificity of the sensitivity of inbred mouse strains TPS, WR and CBA/LacY to the action of chemical mutagens]. / Spetsifika chuvstvitel'nosti myshei inbrednykh linii TPS, WR i CBA/LacY k deistviiu khimicheskikh mutagenov.

The cytogenetic effect of thioTEPA, ethyleneimine, mitomycin C, cyclophosphamide and phtorafurum in bone marrow cells of mouse strains TPS, WR, CBA/LacY was studied. Mice of the TPS strains were most sensitive to the action of all mutagens. Mice of the WR strain were sensitive to thioTEPA and phtorafurum but relatively resistant to mutagens which require metabolic preactivation, i. e. mitomycin C and cyclophosphamide. It was suggested to carry out the study of the cytogenetic activity of chemical compounds using the panel of TPS, WR, 101/H, C57BL/6Y, CBA/LacY strains.

[Interstrain differences in the sensitivity of male sex cells to induction of dominant lethals by thiophosphamide]. / Mezhlineinye razlichiia v chuvstvitel'nosti muzhskikh polovykh kletok u myshei k induktsii dominantnykh letalei tiofosfamidom.

Male mice of strains 101/HY, C57BL/6JY, CBA/LacY were injected intraperitoneally with 2.5 mg/kg dose of thioTEPA. The males were mated with tetrahybrid CBWA females during 5 weeks after the treatment. The same frequencies of dominant lethals were revealed in postmeiotic germ cells of the males of chosen strains. The strong influence of genotype on mutagenic effect in spermatocytes was found. The frequencies of dominant lethals in spermatocytes of 101, B6 and CBA males came to 42.7, 22.8 and 11.9% respectively. The differences in sensitivity of spermatocytes to dominant lethal induction correlated with variations in activity of genetic reparation in oocytes of these mouse strains females. The differences of translocations induced in spermatocytes were not found, that was due to low rate of these mutations (< 1%).

[Mutagenic effect of small doses of diethylsulfate in laboratory mice studied by specific locus method]. / Issledovanie mutagennogo effekta malykh doz diétilsul'fata u laboratornykh myshei metodom opredelennykh lokusov

Diethylsulphate solution was injected intraperitoneally twice a week to male mice B10.C3H having the wild-type colour at a 5 mg/kg dose. The treatment lasted 10 weeks, so that the total dose administered was 5-20 = 100 mg/kg. In fifth week after the end of the treatment these males were crossed to YT females homozygous for seven recessive mutant genes. In the control group all the F1 descendants were of wild-type colour, while in the experimental group among 5042 F1 mice derived from DES-treated spermatogonia there was one mutant d and 4 mosaics (3 with respect to the locus c and 1 with respect to the locusa). These results are interpreted as the evidence of the mutagenic effect of DES.

[Clastogenic effect of thiophosphamide in spermatogonia of inbred strains of mice]. / Klastogennyi éffekt tiofosfamida v spermatogoniiakh myshei inbrednykh linii.

Sensitivity of spermatogonia of 11 mouse inbred strains to induction of chromosome damages by thiophosphamide (thioTEPA) was studied. Metaphase chromosome preparations were made 24 h after treatment with thioTEPA (at 2.25 mg/kg, i/p). With respect to frequency of cells with chromosome damages, strains were ranked as follows: A/Sn (17.5 + 4.4%) greater than 101/H greater than TPS greater than WR = C57BL/6 = AKR = NZB greater than CBA/Lac greater than C3H/Sn greater than MRL greater than BALB/c (5.0 + 2.2%). This distribution does not coincide with that for sensitivity of bone marrow cells, though the data support, in general, the estimations obtained earlier for strains' mutability. Comparison of the data presented with those from literature demonstrates that the sensitivity to clastogenic effect of thioTEPA (and other alkylating agents) correlates neither with spontaneous level of SCE, nor with unscheduled DNA synthesis, nor with radiosensitivity of inbred mice. The frequency of induced chromosome aberrations in spermatogonia is relatively low and spermatogonia cannot substitute bone marrow cells as a test system when assaying chemical mutagens.

[Mutations in mouse minisatellite DNA, induced by radiation]. / Mutatsii v minisatellitnoi DNK myshei industirovannye radiatsiei.

Male (101/HY x C3H/SnY) F1-hybrid mice were given an acute exposure to 0.5 or 1 Gy gamma-irradiation using 60Co, and DNA fingerprints were produced from parents and progeny using probes 33.6 and 33.15. The frequency of mutations in the offspring of irradiated males was 1.72 times higher than in offspring of nonirradiated males. Possible implications of the DNA fingerprint technique in monitoring of mutations in human populations are discussed.

[Interlineage differences in murine sensitivity to thiotepa: an experiment with recombinant strains]. / Izuchenie mezhlineinykh razlichii u myshei po chuvstvitel'nosti k tioTEF: opyt s rekombinantnymi liniiami.

The study is devoted to the clastogenic effect of thio-TEPA in raising recombinant strains 1 x C3 in mice (obtained from 101/H x C3H/Sn crossing). By the summer of 1993, 15 strains had reached 10 generations of inbreeding. From five to six siblings of each strain (males and females) aged 6-8 weeks were treated with the mutagen (3 mg per kg, i/p). Strain sensitivity was estimated by the proportion of marrow cells exhibiting chromosome lesions (including gaps). According to this characteristic, the strains were divided into two unequal groups. The first one included 12 less-sensitive strains (chromosome aberrations were present, on average, in 33.5% of the cells, and lay in the range from 27.0 +/- 4.5 to 39.8 +/- 4.9). The second group included three more-sensitive strains with 51.2, 51.7, and 59.5% of the cells being affected (54.1% on average). A similar difference between the groups was found with respect to the number of breaks per cell: 1.15 in resistant strains and 2.88 in sensitive ones. This difference was significant and similar to that found earlier for C3H and 101 strains. Thus, the results obtained support the hypothesis that mice of the 101/H strain carry a gene enhancing sensitivity to chemical mutagens (mut-1). Some strains of the first group were eliminated; the remaining ones will be bred until the 20th generation, when they will be investigated again in more detail.

[Thiophosphamide induction of chromosome disorders in the spermatogonia of 101/HY, TPS/Y and CBA/LacY inbred mouse strains]. / Induktsiia khromosomnykh narushenii tiofosfamidom v spermatogoniiakh myshei inbrednykh linii 101/HY, TPS/Y i CBA/LacY.

The sensitivity of spermatogonia of 101, TPS and CBA inbred mice to induction of chromosome damages by thiophosphamide (thioTEPA) was studied. With respect to sensitivity of bone marrow cells, these strains were ranked as follows: TPS greater than 101 greater than CBA. When used at the dose 2.25 mg/kg i/p thioTEPA induced chromosome damages in 11.4, 11.6 and 6.0% of spermatogonia of TPS, 101 and CBA mice, respectively. Thus, selection which increased sensitivity of bone marrow cells of TPS mice poorly affected germ cells. To obtain more correct estimations of genetically determined mouse strain sensitivity to chemical mutagens, the estimations should be based on frequencies of induced mutations in germ cells. The spectrum of mutations in spermatogonia differs from that in bone marrow cells: the frequencies of isochromatid breaks and symmetrical chromatid exchanges are higher in spermatogonia. This can be due to close situation of homologous chromosomes in interphase nuclei of spermatogonia.

[Demonstration of dominant lethal mutations in early mouse embryogenesis]. / Proiavlenie dominantnykh letal'nykh mutatsii v rannem émbriogeneze myshi.

Male mice of C57Bl/6Y strain were injected intraperitoneally with 2.5 and 5 mg/kg doses of thioTEPA. Males were mated to tetrahybrid CBWA females during the second week after the treatment. Embryonic mortality was studied by two methods: by standard dominant lethal method on the 15-17th day of pregnancy and cytologically on the 4th day. The rate of fertilization was not affected by thioTEPA. After treatment with 2.5 mg/kg of thioTEPA the frequency of induced dominant lethals was 89.8%; preimplantation losses were 78,5% in treated and 13,8% in control group. The cytological analysis revealed that preimplantation embryonic death is equal to 63,9%. The death of embryos before implantation occurred at 2-20 blastomere stages. After treatment with 5 mg/kg of thioTEPA all embryos died before implantation at 2-16 blastomere stages. It was demonstrated that dominant lethal method gave more complete estimation of dominant lethal frequency, and that cytological analysis is the correct estimation of preimplantation death. Thus the methods used supplement each other.

[Role of female genotype in manifestation of dominant lethal mutations induced by thiophosphamide in the spermatids of male rats]. / Rol' genotipa samok v proiavlenii dominantnykh letal'nykh mutatsii, indutsirovannykh tiofosfamidom v spermatidakh samtsov myshei.

Hybrid male mice FICD2 were injected intraperitoneally with thioTEPA at a dose of 1.5 mg/kg or 2.0 mg/kg. During the second week after the treatment males were mated with females of 101/HY, C57BL/6JY and CBA/LacY strains, and embryoic mortality was studied on 3.5 day of pregnancy cytologically and on 15-17 days -- by the dominant lethal method. ThioTEPA did not affect the rate of fertilization. The dominant lethals frequencies differed significantly in females 101 (67%), B6 (59%) and CBA (43%). It is suggested that the differences observed are due to different activity of repair systems in oocytes of females of these strains.

[Fecundity and embryonal mortality of four inbred mouse lines, BALB/c, B10.CW, A/Sn, CC57W and their hybrids]. / Plodovitost' i émbrional'naia smertnost' chetyrekh inbrednykh liniï mysheï BALB/c, BUO.CW, A/Sn, CC57W i ikh gibridov

The object of this investigation was the potential fecundity of four inbred strains of mice, viz. BALB/c, B10.SW, CC57W, A/Sn and of their different hybrid combinations. The inbred strains studied had different normal ovulation numbers varying from 9,2 to 11,9 and different death-rate of embryos before (10,99-39,49%) and after A/Sn, B10.CW and CC57W were practically equal to one another, but significantly larger than in the strain BALB/c. Interlinear crosses resulted in a considerable decrease of the total embryonic death rate, while the normal ovulation number did not undergo any changes. The number of live embryos in simple hybrids did not differ significantly from that in the maternal inbred strains. Therefore the heterozygosity of embryos did not affect significantly the potential fecundity of females. The number of surviving embryos per female increased in the cross between the simple hybrids (BALB/cXB10.CW)X(CC57WXA/Sn) to 8.9 +/- 0,37. This heterosis was the result of the total death-rate of embryos down to 14,89%. As it is shown by the comparison of the potential fecundity of pregnant females carrying homo- and heterozygous embryos to that of pregnant hybrid females, the rate of survival of embryos depends more on the genotype of the mother, than on that of the embryos.

[Cytogenetic effect of mitomycin C and cytosine arabinoside on mice of different genotypes]. / Tsitogeneticheskii éffekt mitomitsina C i tsitozin-arabinozida u myshei raznogo genotipa

Cytogenetic effect of mitomycin C (MC) and cytosine arabinoside (CA) on bone marrow cells of male mice of the strains 101/HY, C57BL/6Y C,3H/SnY and of the (C3HX101) F1 hybrids was studied. The frequencies of cells with chromosome aberrations after the treatment with MC at a 5 mg/kg dose were 54,4%; 41,8%; 40,4% and 26,8% in 101H, B6, C3H/Sn mice and in the F1 hybrids (C3HX101) respectively. The frequencies of cells with chromosome aberrations after the treatment with CA at a 500 mg/kg dose were 25,2%; 17,8%; 10,8% and the 101/H, B6, C3H/Sn mice and in the F1 hybrids (C3HX101) respectively. Both mutagens induced the greatest number of chromosome aberrations in the 101/H strain and the smallest number in the F1 hybrid (C3HX101). A positive correlation was established between the levels of induced and spontaneous chromosome lesions.

[Fecundity and embryonal mortality of four inbred strains of mice, BALB/c, B10.CW, A/Sn, CC57W and their hybrids]. / Plodovitost' i émbrional'naia smertnost' chetyrekh inbrednykh linii myshei BALB/c, B10.CW, A/Sn, CC57W i ikh gibridov

The object of this investigation was the potential fecundity of four inbred strains of mice, viz. BALB/c, B10.SW, CC57W, A/Sn and of their different hybrid combinations. The inbred strains studied had different normal ovulation numbers varying from 9,2 to 11,9 and different death-rate of embryos before (10,99-39,49%) and after (9,05-22,47%) the implantation. The numbers of live embryos per female in the strains A/Sn, B10.CW and CC57W were practically equal to one another, but significantly larger than in the strain BALB/c. Interlinear crosses resulted in a considerable decrease of the total embryonic death-rate, while the normal ovulation number did not undergo any changes. The number of live embryos in simple hybrids did not differ significantly from that in the maternal inbred strains. Therefore the heterozygosity of embryos did not affect significantly the potential fecundity of females. The number of surviving embryos per female increased in the cross between the simple hybrids (BALB/cXB10.CW) X (CC57WXA/Sn) to 8,9 +/- 0,37. This heterosis was the result of the total death-rate of embryos down to 14,89%. As it is shown by the comparison of the potential fecundity of pregnant females carrying homo- and heterozygous embryos to that of pregnant hybrid females, the rate of survival of embryos depends more on the genotype of the mother, than on that of the embryos.