ABSTRACT
Sister chromatid exchanges (SCE) were studied in chromosomes of peripheral blood lymphocytes taken from 19 normal individuals of both sexes and cultured in the presence of 5-bromodeoxyuridine (BUdR). BUdR was added to the culture in final concentration of 15 or 30 microgram/ml for the last 28 hours of cultivation. The chromosome preparations were stained with the fluorescent dye Hoechst-33258, followed by Giemsa staining. At BUdR concentration of 15 microgram/ml the mean number of SCE was significantly less (8.9 per metaphase), that at the concentration of 30 microgram/ml (12.3 per metaphase). The incidence of SCE among 19 individuals varied from 6.5 to 13.7. The mean numbers of SCE in five individuals were significantly different from the mean number calculated for the whole group of 19 subjects. There were no differences between the sexes in the incidence of SCE. The distribution of SCE between chromosomes was found to be proportional to the chromosome length and to the amount of DNA. However, the chromosomes of groups E, F and G were highly significantly under-represented in the number of exchanges.
Subject(s)
Chromatids , Chromosomes, Human , Lymphocytes/ultrastructure , Adult , Bromodeoxyuridine , Chromatids/analysis , DNA/analysis , Female , Humans , MaleABSTRACT
The incorporation of 3H-deoxycytidine (3H-Cdr) in the presence of thymidine (Tdr) into cultured human blood lymphocytes has been studied. The analysis of the label in interphase nuclei as well as in chromosomes at metaphase was carried out. The labeling was much higher when 3H-Cdr (0.5 to 1.0 C/ml, 2--4 x 10(-5) mM) was added to the cultures simultaneously with Tdr (4 x 10(-1) mM). This observation is considered as an indication that in the presence of high doses of Tdr exogeneous Cdr is utilized to synthesize cytosine of DNA rather than thymidine. During the first hours after its addition, the bulk of 3H-Cdr is eliminated from the culture medium. At 12 hrs of the incubation, the medium seems to be free of the nucleoside as shown particularly from the single chromatid localization of the label in chromosomes of the second mitosis. The incorporation into lymphocytes of 3H-Tdr administered in the same dose under the same conditions was registered for the whole period of observation (24 hrs). The data obtained are discussed in relation to lymphocyte catabolism of exogeneous nucleosides.
Subject(s)
Bromodeoxycytidine/metabolism , Chromatids/drug effects , Crossing Over, Genetic/drug effects , Cytosine Nucleotides/biosynthesis , DNA/biosynthesis , Deoxycytidine/analogs & derivatives , Nucleic Acid Precursors/metabolism , Cells, Cultured , Humans , Interphase , Lymphocytes/metabolism , Lymphocytes/ultrastructure , Metaphase , Thymidine/metabolism , Time Factors , TritiumABSTRACT
In the cultured in vitro human blood lymphocytes treated with 5-bromodeoxycytidine (BrdC) plus thymidine (dT) in concentrations of 0.05 mM and 0.4 mM, respectively, metaphases of the second division, containing chromosomes with differentially stained sister chromatids appear after a 30 hours treatment, attaining 85 per cent at 36 hours. Under the treatment with BrdC or BrdU alone, the similar percentage of metaphases is observed at 28 hours. Similar results are obtained if, at 16--18 hours, the culture medium, containing BrdC plus dT is either replaced with a medium free of both the precursors, or supplied once more vith (0.05 mM) or dC(0.05 mM). The The observed slowing movement of cells through out the cell cycle, due to the treatment with BrdC plus dT, is explained by the inhibitory effect of dT on DNA synthesis which is manifested as BrdC catabolism. It is concluded that BrdC administered together with the cell cycle-slowing doses of thymidine is predominantly incorporated into cytosine of DNA.
Subject(s)
Bromodeoxycytidine/pharmacology , Chromatids , DNA/biosynthesis , Deoxycytidine/analogs & derivatives , Thymidine/pharmacology , Chromosomes, Human/ultrastructure , Depression, Chemical , Humans , Lymphocytes/ultrastructureABSTRACT
Cell movement through the mitotic cycle and sister chromatid exchanges (SCE) were studied in human blood lymphocytes cultured in the presence of 5-bromodeoxycytidine (BrdC, 0.05 mM) plus thymidine (dT 0.4, 0.8, and 1.0 mM). In controls, lymphocytes were cultivated in the presence of 5-bromodeoxyuridine (BrdU, 0.05 mM) and deoxycytidine (0.1 mM), or BrdC alone. All nucleosides were added to the cultures 28 hours prior to fixation and were maintained in the medium for 16 hours. As determined from percentage of metaphases of 1st to 3rd divisions, BrdC did not release from thymidine block. This fact leads us to conclude that BrdC in contrast to deoxycytidine does not serve as a cytosine precursor. No significant differences in the frequency of SCE and their distribution among chromosomes were found between cultures treated with BrdC and with BrdU.
Subject(s)
Bromine/metabolism , Bromodeoxycytidine/metabolism , Chromatids/metabolism , Cytosine Nucleotides/metabolism , DNA/metabolism , Deoxycytidine/analogs & derivatives , Nucleic Acid Precursors/metabolism , Thymine/metabolism , Cells, Cultured , Humans , In Vitro Techniques , Lymphocytes/metabolism , MetaphaseABSTRACT
The frequency of sister chromatid exchanges (SCE) was studied in cultivated blood lymphocytes of three normal individuals under elimination of DNA synthesis inhibiting action of 5-bromodeoxyuridine (BrdUrd 0.05 mM) with deoxycytidine (Cdr 0.1 and 0.01 mM). The frequency of SCE was significantly increased in the presence of 0.1 mM Cdr. In parallel with SCE frequency, Cdr elevated the percentage of metaphases of the second division. The increase of SCE in the presence of Cdr may be connected with normalization of the DNA replication under its action.