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Formation of chromatid-type aberrations with procarbazine hydrochloride in postmeiotic male germ cells of mice
366 tan Isotope Research Institute, Tokyo, and 2 Teikyo University, Tokyo (Japan) Examination of DNA lesion by the alkaline elution technique. II. DNA lesion induced by hydrogen peroxide DNA single-strand breaks (SSBs) induced by hydrogen peroxide (H202) in the chromatin of mouse leukemic cells (L5178Y) were examined employing an 'alkaline elution' technique. An obvious relationship was obtained between the concentration of H202 and the amount of single-stranded DNA fragments. The amount of fragmented DNA increased along the time course of H202 treatment at low concentration (0.5 /~g/ml). However, this relation was reversed when raising the concentration of H202 up to 10 #g/ml. HzOz-induced DNA SSB was inhibited after pretreatment of cells with catalase within 5 min at 0 °C or 25 °C. DNA SSB repair was evident after removal of H202 and cell incubation for 5 min at 37 o C although not evident after incubation for 20 min at 0°C. The repair was not inhibited by 2 mM caffeine or 0.2 mM cyclohexamide.
17 Kanaya, H., and Y. Takeda, Hatano Research Institute, Food and Drug Safety Center, Hadano, Kanagawa (Japan)
18 Kasai, H., H. Hayami and S. Nishimura, National Cancer Center Research Institute, Tokyo (Japan) Hydroxylation of deoxyguanosine at the C-8 position by mutagens and carcinogens The C-8 position of deoxyguanosine (dGuo) was hydroxylated by ascorbic acid in the presence of oxygen (02) in 0.1 M phosphate buffer (pH 6.8) at 37°C. Addition of hydrogen peroxide (H202) remarkably enhanced this hydroxylation. The Udenfriend system (ascorbic acid, Fe n, ethylenediaminetetraacetic acid (EDTA) and O 2) was also effective for hydroxylation of dGuo in high yield. Guanine residues in DNA were also hydroxylated by ascorbic acid. Other reducing agents, such as hydroxylamine, hydrazine, dihydroxymaleic acid, sodium bisulfite and acetol, were also effective for the hydroxylation reaction, as were metal-EDTA complexes (Fe ll-, Snn-, Ti m-, CuLEDTA). Dihydroxymaleic acid showed considerable mutagenic activity to TA100 (about 3000 revertants/mg). An OH radical seemed to be involved in this hydroxylation reaction in most of the above hydroxylating systems, but another reaction mechanism may also be involved, particularly when dGuo is hydroxylated by ascorbic acid alone or ascorbic acid plus H202. The possible biological significance of the hydroxylation of guanine residues in DNA in relation to mutagenesis and carcinogenesis is discussed.
Formation of mutagens and nitroso compounds from cardiovascular drugs by reaction with nitrite The formation of mutagens and nitroso compounds from 18 cardiovascular drugs by drugnitrite interaction was examined by the screening procedure developed in our laboratory (Takeda and Kanaya (1982) Chem. Pharm. Bull., 30, 3399-3404). On the reaction of 50 mM drug and 500 mM nitrite at pH 3.0-3.4 and 37 o C for 4 h, formation of bacterial mutagens (S. typhimurium TA98 and TA100) was observed for clonidine, bamethan, hydralazine and bethanidine. The formation of nitroso compounds was observed for ajmaline, alprenolol, dilazep, trimetazidine, bamethan and propranolol.
19 Katoh, M., and S. Iwahara, Hatano Research Institute, Food and Drug Safety Center (FDSC), Kanagawa (Japan) Formation of chromatid-type aberrations with procarbazine hydrochloride in postmeiotic male germ cells of mice To study the relationship between the types of chromosome aberrations and the rates of heritable translocations induced in postmeiotic male germ cells of mice after treatment with mutagens, we investigated whether procarbazine hydrochloride
367 induces mainly the chromosome type or chromatid type in postmeiotic male germ cells of mice. Male mice were injected intraperitoneally with 800 m g / k g of procarbazine hydrochloride and each male was then serially mated with 2 virgin females per week for 6 weeks. Chromosome aberrations were scored at the paternal chromosome sets in the first cleavage metaphases after fertilization. Type of chromosome aberrations induced in postmeiotic stages were mainly chromatid type. From these results and previous reports (Katoh et al., 1980, 1981, 1982, 1983; Leonard and Deknudt, 1968; Sotomayor and Cumming, 1975; Lang and Adler, 1977; Adler, 1980), we suggest that the frequency of heritable translocations caused by mutagens such as procarbazine hydrochloride and mitomycin C which induce chromatid-type aberrations is lower than that by mutagens such as methyl methanesulfonate, cyclophosphamide and X-ray which induce chromosome-type aberrations in postmeiotic stages.
20 Kishi, K., and A. Tonomura, Tokyo Medical and Dental University, Yushima, Tokyo (Japan) Cytogenetic effects of sodium fluoride
Sodium fluoride (NaF) is widely used for the prevention of dental caries at various concentrations. The clastogenic effect of NaF has been tested by the use of several cytogenetic assay systems, but the findings on its genotoxicity are not consistent. In this study, the effects of N a F on chromosomes, unscheduled DNA synthesis (UDS) and sister-chromatid exchanges (SCEs) were investigated using cultured human lymphocytes. For clastogenicity testing, cells were treated for 24 h in various concentrations of NaF. At least two donors were tested for each concentration and more than 10 000 cells were totally observed. The frequencies of chromosome aberrations were 0.78 _+ 0.72, 0.88 + 0.56, 0.77 + 0.45, 5.95 + 5.35, 57.76 _ 31.46, 108.00 _ 59.40, 80.00 _ 53.70 and 40.00 _ 5.66 per 100 cells for concentrations of 0, 0.25, 0.5, 1.0, 2.0, 4.0, 6.0 and 8.0 mM, respectively. Considerable differences among individuals were observed, but
there was no significant difference between sexes. Sodium fluoride treatment had remarkable effects on the induction of isochromatid gaps and chromosome breaks (NUpds). At various concentrations of NaF ranging from 1.0 to 4.0 mM, no increase in UDS and SCE was observed.
21 Kitamori, S., R. Nakagawa, M. Kuromoto and H. Tokiwa, Fukuoka Environmental Research Center, Fukuoka 818-01 (Japan) Identification of nitroarenes in diesel exhaust particulates
The direct-acting mutagens in diesel particulate extracts were identified. Small particles collected had electron microscopically the size range from 0.1 to 0.3 /~m, and were extracted with toluenehexane-dichloromethane by ultrasonification for 30 min. Nitroarenes in the extracts were analyzed using Sephadex LH20 column chromatography, HPLC, and ECD-gas chromatography. Among nitroarenes identified, it was concluded that the major mutagens are in all probability 1,6- and 1,8-dinitropyrene. 1-Nitropyrene and 3-nitrofluoranthene were also detected. The dinitropyrenes detected contributed 43% of the entire activity of the crude extracts for strain TA98, whereas 1-nitropyrene (or 3-nitrofluoranthene) was responsible for less than 10% of the activity.
22 Kiuchi, H., T. Inoue, T. Kada, O. Makino 1, T. Shibata 1 and T. Ando 1, National Institute of Genetics, Mishima, and 1 Riken Institute, Wakoshi (Japan) Effect of an antimutagenic metal compound, cobaltous chloride, on E. coli RecA protein in vitro
Cobaltous chloride, which exhibits a potent antimutagenic activity in various mutation assay sys-
17 Kanaya, H., and Y. Takeda, Hatano Research Institute, Food and Drug Safety Center, Hadano, Kanagawa (Japan)
18 Kasai, H., H. Hayami and S. Nishimura, National Cancer Center Research Institute, Tokyo (Japan) Hydroxylation of deoxyguanosine at the C-8 position by mutagens and carcinogens The C-8 position of deoxyguanosine (dGuo) was hydroxylated by ascorbic acid in the presence of oxygen (02) in 0.1 M phosphate buffer (pH 6.8) at 37°C. Addition of hydrogen peroxide (H202) remarkably enhanced this hydroxylation. The Udenfriend system (ascorbic acid, Fe n, ethylenediaminetetraacetic acid (EDTA) and O 2) was also effective for hydroxylation of dGuo in high yield. Guanine residues in DNA were also hydroxylated by ascorbic acid. Other reducing agents, such as hydroxylamine, hydrazine, dihydroxymaleic acid, sodium bisulfite and acetol, were also effective for the hydroxylation reaction, as were metal-EDTA complexes (Fe ll-, Snn-, Ti m-, CuLEDTA). Dihydroxymaleic acid showed considerable mutagenic activity to TA100 (about 3000 revertants/mg). An OH radical seemed to be involved in this hydroxylation reaction in most of the above hydroxylating systems, but another reaction mechanism may also be involved, particularly when dGuo is hydroxylated by ascorbic acid alone or ascorbic acid plus H202. The possible biological significance of the hydroxylation of guanine residues in DNA in relation to mutagenesis and carcinogenesis is discussed.
Formation of mutagens and nitroso compounds from cardiovascular drugs by reaction with nitrite The formation of mutagens and nitroso compounds from 18 cardiovascular drugs by drugnitrite interaction was examined by the screening procedure developed in our laboratory (Takeda and Kanaya (1982) Chem. Pharm. Bull., 30, 3399-3404). On the reaction of 50 mM drug and 500 mM nitrite at pH 3.0-3.4 and 37 o C for 4 h, formation of bacterial mutagens (S. typhimurium TA98 and TA100) was observed for clonidine, bamethan, hydralazine and bethanidine. The formation of nitroso compounds was observed for ajmaline, alprenolol, dilazep, trimetazidine, bamethan and propranolol.
19 Katoh, M., and S. Iwahara, Hatano Research Institute, Food and Drug Safety Center (FDSC), Kanagawa (Japan) Formation of chromatid-type aberrations with procarbazine hydrochloride in postmeiotic male germ cells of mice To study the relationship between the types of chromosome aberrations and the rates of heritable translocations induced in postmeiotic male germ cells of mice after treatment with mutagens, we investigated whether procarbazine hydrochloride
367 induces mainly the chromosome type or chromatid type in postmeiotic male germ cells of mice. Male mice were injected intraperitoneally with 800 m g / k g of procarbazine hydrochloride and each male was then serially mated with 2 virgin females per week for 6 weeks. Chromosome aberrations were scored at the paternal chromosome sets in the first cleavage metaphases after fertilization. Type of chromosome aberrations induced in postmeiotic stages were mainly chromatid type. From these results and previous reports (Katoh et al., 1980, 1981, 1982, 1983; Leonard and Deknudt, 1968; Sotomayor and Cumming, 1975; Lang and Adler, 1977; Adler, 1980), we suggest that the frequency of heritable translocations caused by mutagens such as procarbazine hydrochloride and mitomycin C which induce chromatid-type aberrations is lower than that by mutagens such as methyl methanesulfonate, cyclophosphamide and X-ray which induce chromosome-type aberrations in postmeiotic stages.
20 Kishi, K., and A. Tonomura, Tokyo Medical and Dental University, Yushima, Tokyo (Japan) Cytogenetic effects of sodium fluoride
Sodium fluoride (NaF) is widely used for the prevention of dental caries at various concentrations. The clastogenic effect of NaF has been tested by the use of several cytogenetic assay systems, but the findings on its genotoxicity are not consistent. In this study, the effects of N a F on chromosomes, unscheduled DNA synthesis (UDS) and sister-chromatid exchanges (SCEs) were investigated using cultured human lymphocytes. For clastogenicity testing, cells were treated for 24 h in various concentrations of NaF. At least two donors were tested for each concentration and more than 10 000 cells were totally observed. The frequencies of chromosome aberrations were 0.78 _+ 0.72, 0.88 + 0.56, 0.77 + 0.45, 5.95 + 5.35, 57.76 _ 31.46, 108.00 _ 59.40, 80.00 _ 53.70 and 40.00 _ 5.66 per 100 cells for concentrations of 0, 0.25, 0.5, 1.0, 2.0, 4.0, 6.0 and 8.0 mM, respectively. Considerable differences among individuals were observed, but
there was no significant difference between sexes. Sodium fluoride treatment had remarkable effects on the induction of isochromatid gaps and chromosome breaks (NUpds). At various concentrations of NaF ranging from 1.0 to 4.0 mM, no increase in UDS and SCE was observed.
21 Kitamori, S., R. Nakagawa, M. Kuromoto and H. Tokiwa, Fukuoka Environmental Research Center, Fukuoka 818-01 (Japan) Identification of nitroarenes in diesel exhaust particulates
The direct-acting mutagens in diesel particulate extracts were identified. Small particles collected had electron microscopically the size range from 0.1 to 0.3 /~m, and were extracted with toluenehexane-dichloromethane by ultrasonification for 30 min. Nitroarenes in the extracts were analyzed using Sephadex LH20 column chromatography, HPLC, and ECD-gas chromatography. Among nitroarenes identified, it was concluded that the major mutagens are in all probability 1,6- and 1,8-dinitropyrene. 1-Nitropyrene and 3-nitrofluoranthene were also detected. The dinitropyrenes detected contributed 43% of the entire activity of the crude extracts for strain TA98, whereas 1-nitropyrene (or 3-nitrofluoranthene) was responsible for less than 10% of the activity.
22 Kiuchi, H., T. Inoue, T. Kada, O. Makino 1, T. Shibata 1 and T. Ando 1, National Institute of Genetics, Mishima, and 1 Riken Institute, Wakoshi (Japan) Effect of an antimutagenic metal compound, cobaltous chloride, on E. coli RecA protein in vitro
Cobaltous chloride, which exhibits a potent antimutagenic activity in various mutation assay sys-