Kinetics of formation of exchanges and rejoining of breaks in human G0 and G2 lymphocytes after low-LET radiation.

PURPOSE: To study the kinetics of chromosomal break rejoining and exchange formation in human G0 and G2 lymphocytes after low-LET irradiation using the premature chromosome condensation (PCC) technique. METHODS: For the G0 experiments, isolated human lymphocytes were irradiated (1, 3 and 8 Gy) and immediate and delayed fusions (1, 3, 5, 8 and 24 h) were performed with CHO-cells, and 48-h cultures were set up simultaneously. Interphase and metaphase human chromosomes were hybridized with dual colour combination probes for chromosomes 2 and 4. For the G2 experiments, isolated lymphocytes from the same donor were cultured for 69 h, irradiated with 3 Gy 60Co gamma rays and fused with CHO cells immediately and after 1, 2 and 3 h. Chromatid breaks and exchanges were scored after Giemsa staining. RESULTS: In both G0 and G2 cells, exchanges were already observed at the first fusion and, at lower doses, their frequencies remained relatively stable over time. The exchange-type aberrations seen in G0 cells were mostly incomplete, especially in the first fusions, while the exchanges seen in metaphase were mostly complete. The frequency of complete exchanges increased by time during incubation, while the frequency of incomplete exchanges showed more variation and a general decline by time in the G0-PCC experiment. The number of breaks declined rapidly by time both in G0 and G2. The break rejoining rate was higher at 8 Gy than in the two lower doses in the G0-PCC. The overall aberration frequencies in G0 and G2 cells studied by PCC were higher than in the corresponding metaphase. The initial number of breaks in G2 was higher than in G0 and the break rejoining rate in the G2 was two-fold. CONCLUSIONS: The formation of complete exchanges in G0 is a delayed process. The incomplete exchanges appear unstable in G0. The decreasing proportion of incomplete exchanges and simultaneous increase in complete exchanges in G0-PCC indicates that formation of exchanges is not coordinated in time.

Cytogenetic effects of softwood kraft pulp bleaching effluents and methanesulfonyl chloride in Chinese hamster ovary cells.

The genotoxicity of effluents collected from a conventional 5-stage softwood kraft pulp bleaching process was studied in Chinese hamster ovary (CHO) cells in vitro. Spent liquor from the first chlorination stage (C/D), where elemental chlorine and chlorine dioxide had been used in equal proportions, was shown to induce a dose-dependent increase in sister chromatid exchanges (SCEs) without metabolic activation (4-h treatment), with a maximum increase of 1.6 times over the control level at 204 microliters/ml; this dose also induced 15.5- and 20.5-fold increases in cells with chromatid-type chromosomal aberrations after 4-h and a 20-h treatment, respectively. Another C/D stage spent liquor from a process where the ratio of elemental chlorine and chlorine dioxide had been 9:1 produced a 40.5-fold elevation of cells with chromatid-type aberrations at 204 microliters/ml (20-h treatment). This sample clearly increased chromosomal aberrations also when tested as a concentrate (4-h treatment), which showed that the observed clastogenicity was not unspecifically due to the relatively large volumes used in the treatments with the unconcentrated liquors. In general, the use of rat liver S9 mix reduced the genotoxicity of the spent liquors. The results agree with earlier findings on the Salmonella mutagenicity of the same C/D samples: both the prokaryotic and eukaryotic assays showed a reduction in genotoxicity when the amount of elemental chlorine in the bleaching process was reduced. An effluent sample collected from the alkaline stage of the process was not clastogenic with or without metabolic activation. Methanesulfonyl chloride, a new compound identified in bleaching plant air, was found to be induce chromosomal aberrations in the presence of S9 mix.

Sister-chromatid exchanges induced by vinyl esters and respective carboxylic acids in cultured human lymphocytes.

Vinyl acetate--an efficient inducer of sister-chromatid exchanges (SCEs)--is known to be hydrolyzed in mammalian cells into acetic acid and acetaldehyde, the latter being the likely metabolite responsible for the SCE induction. As similar hydrolysis to acetaldehyde and to a carboxylic acid is also expected for other vinyl esters, five such compounds--vinyl formate, vinyl chloroformate, vinyl propionate, vinyl crotonate and vinyl-2-ethylhexanoate--and five carboxylic acids--formic acid, acetic acid, propionic acid, crotonic acid and 2-ethylhexanoic acid--were tested for their ability to induce SCEs in cultured (72 h) human lymphocytes with a 48-h treatment, starting at 24 h after culture initiation. Vinyl formate, vinyl propionate and vinyl crotonate induced a clear dose-dependent increase in the number of SCEs/cell at concentrations of 0.125-0.5 mM and vinyl chloroformate at 0.063-1 mM, i.e., at roughly the same concentration range as vinyl acetate and acetaldehyde. Vinyl-2-ethylhexanoate required slightly higher concentrations (0.25-4 mM) for SCE induction. All of the carboxylic acids tested also elevated SCEs, but only slightly. Formic acid and crotonic acid produced some SCE increase at a concentration of 10 mM, acetic acid at 5 and 10 mM and propionic acid at 2.5 mM. 2-Ethylhexanoic acid induced SCEs at a lower concentration range (0.63-2.5 mM) than the other acids. The positive concentrations of the first three carboxylic acids lowered the pH of the culture medium immediately after the treatment by 0.5-1.0 pH unit (lowest observed pH 6.53). The pH differences from the control cultures became smaller in measurements done 24 h and 48 h after the beginning of treatment. Propionic acid and 2-ethylhexanoic acid affected medium pH only slightly (maximum drop 0.2 pH units) at the concentrations that induced SCEs. The results lend support to the idea that the efficient SCE induction observed with the vinyl esters results from the formation of acetaldehyde, with carboxylic acids--with the possible exception of 2-ethylhexanoic acid--playing no significant role. The slight SCE induction obtained with the carboxylic acids cannot be explained by lowered pH alone.