Action of ftorafur on the chromosomes of normal and tumorous cells of the Djzungarian hamster in vitro]

The chromosome-breaking effect of antitumor agent ftorafur (0,1 mg/ml) was studied on cell cultures of normal and transformed Djungarian hamster's fibroblasts. After 24 hours of exposition with the drug 79% of aberrant malignant cells was registered, while the chromosomes of normal cells remained undamaged. After ftorafur's action for 2--12 hours and the following removal of the agent the level of aberrant metaphase among normal cells rose to 11--15%. Chromatid breaks were the predominant type of drug-induced aberrations. Thse results confirm the idea that ftorafur blocks the cell cycle of normal fibroblasts and that it's necessary to wash the drug off some period before fuxation in order to reveal its chromosome-breaking effect in metaphase cells. The striking distinctions in susceptibility of normal and malignant cells to the mutagenic effect of ftorafur is partly due to the distrubances of the cell cycle caused by this drug.     

Effect of ftorafur and 5-fluorouracil on the chromosomes of human tumor cell cultures]

The effect of two antitumour drugs, ftorafur (Ft) and 5-fluorouracil (5-FU) on chromosomes of human tumour cells (strain CA-1) was studied in vitro. Since no data on the karyotype of this tumour strain had been published, the chromosome set of the model was investigated at first. Significant quantitative and structural divergence from the normal human male karyotype were observed. Steam line cells contained 47-49 chromosomes, including 9 permanent markers. No Y-chromosome was revealed. Ft and 5-FU hardly injured the chromosomes of CA-1 cells; the level of aberrant metaphases reached 94%. Chromatid deletions and gaps formed the major part of drug-induced cytogenetic abnormalities.     

A comparison of aberration distribution and cell-cycle progression in cells treated with bleomycin with those exposed to X-rays

The extent of cell-cycle delay and the frequency of aberrant metaphases induced by bleomycin (BLM) and X-rays have been compared at doses which produce similar frequencies of chromosome aberrations by the 2 clastogenic agents (BLM, 40 micrograms/ml and X-rays, 2 Gy) in muntjac lymphocytes. The frequency of aberrant metaphases was low in BLM-treated cells; however, the number of aberrations per metaphase was higher than in cells exposed to X-rays. Thus in contrast to their uniform sensitivity to X-rays, the lymphocytes showed differential sensitivity to BLM. This might be due to differences among the cells in their uptake of BLM and/or its action on the nuclear membrane-DNA complex. In spite of the total number of chromosome aberrations being similar to that induced by X-rays, BLM did not induce a significant delay in cell-cycle progression as observed in the case of X-rays. A possible explanation could be that the DNA damages being limited to fewer cells than in the case of X-irradiation, the BLM-treated cultures had more normal cells allowing faster progression and/or unlike X-rays BLM may not be causing other cellular damages in addition to DNA breaks.     

The induction of chromosome aberrations in human purified peripheral blood lymphocytes following in vitro exposure to selenium

Human lymphocyte cultures were treated with increasing concentrations (8.0 X 10(-8) M to 8.0 X 10(-5) M) of sodium selenite and selenomethionine 24 h after stimulation with phytohemagglutinin and were scored for chromosomal aberrations at 48 h. The yield of abnormal metaphases was dependent on the dose and the form of selenium used. At 8.0 X 10(-5) M the proportion of aberrant cells reached 53.5% and 43.0% for selenite and selenomethionine, respectively. The selenium-induced chromosomal aberrations were primarily of the chromatid type and included breaks and fragments. Chromosomal exchanges were less frequent and included triradials and quadriradials. These results confirm that selenium is clastogenic for cultured human lymphocytes.     

Spontane und chloramphenicolinduzierte Chromosomenmutationen und biochemische Befunde bei zwei Fällen mit Glutathionreduktasemangel (NAD(P)H: Glutathione oxidoreductase,E.C. 1.6.4.2.)

Summary In one proband with pancytopenia and in his mother with normal blood picture the pattern of the erythrocyte enzymes revealed a glutathione reductase deficiency with a missing GR-II-band in the electropherogram. The incidence of spontaneous aberrations in the chromosomes of both individuals (52–58% and 7.2–8% metaphases with chromosomal damage) was significantly higher than in normal controls (0–3%). After 0.5 mg chloramphenicol per ml culture medium was applied 24 hours before fixation the number of damaged metaphases further increased to about 81% and 19–21%, respectively. Possible reasons for this phenomenon are discussed.     

Chromosomal aberrations and sister-chromatid exchanges in Chinese hamster cells treated in vitro with hexavalent chromium compounds.

Chinese hamster cells (CHO line) were treated in vitro for 30--39 h with hexavalent chromium compounds (K2Cr2O7 and Na2CrO7), at concentrations ranging from 0.1 to 1.0 microgram of Cr6+ per ml, in medium containing BUdr. Chromosomal aberrations and sister-chromatid exchanges were scored on BUdr-labelled 2nd division metaphases, collected at the end of treatment and stained with Giemsa. Treatment with mitomycin C 0.009--0.030 microgram/ml) was carried out as a control for the responsiveness of the cell system to chromosomal damage. Both chromium compounds induced marked mitotic delays. Chromosomal aberrations were increased about 10-fold by exposure to Cr6+ (1.0 microgram/ml). The principal aberrations observed were single chromatid gaps, breaks and interchanges, whose frequencies increased proportionally to the concentration of chromium. Dicentric chromosomes, isochromatid breaks, chromosome and chromatid rings were also induced. The frequenyc of sister-chromatid exchanges was hardly doubled 30 h after exposure to Cr6+ at 0.3 microgram/ml, whereas it was trebled 39 h after treatment, in the cells whose division cycle had been slowed down by chromium.     

Chromosomal changes induced by chrysotile fibres or benzo-3,4-pyrene in rat pleural mesothelial cells

The induction of chromosomal aberrations in rat pleural mesothelial cells (RPMC) following in vitro treatment with chrysotile fibres has been demonstrated. The production of chromosomal aberrations was also observed after treatment of the cells with benzo-3,4-pyrene (BP). The yield of abnormal metaphases was dose-dependent and reached 58% at a BP dose of 2 micrograms/ml. Chrysotile fibres at 7 micrograms/ml induced 21% abnormal metaphases and the frequency decreased with further increases in fibre concentration. Their decline is possibly related to a lethal effect. Chrysotile-induced chromosomal aberrations were primarily of the chromatid type and included breaks and fragments. BP induced chromosome exchanges which were not seen following chrysotile treatment. Minutes and double minutes were detected in BP-treated RPMC and occasionally found after chrysotile application. These results confirm that chrysotile fibres are clastogenic for some cultured cells and demonstrate that the fibres induce chromosome damage in target RPMC.     

In vivo radioprotection by alpha-TMG: preliminary studies

alpha-TMG is a novel water-soluble derivative of Vitamin E that has shown excellent antioxidant activity. The parent compound has demonstrated protection against radiation induced chromosomal damage in vivo. Hence, the preliminary experiments to determine the radioprotective activity of alpha-TMG were carried out in adult Swiss albino mice. Acute toxicity of the drug was studied taking 24h, 72 h and 30 day mortality after a single intraperitoneal injection of 500-2000 mg/kg body weight of the drug. The drug LD(50) for 24h and 72 h/30 day survival were found to be 1120 and 1000 mg/kg body weight, respectively. The optimum time of drug administration and drug dose-dependent effect on in vivo radiation protection of bone marrow chromosomes was studied in mice. Injection of 600 mg/kg of the drug 15 min before or within 5, 15 or 30min after 3Gy whole body gamma radiation resulted in a significant decrease in the aberrant metaphases percent at 24h post-irradiation; the maximum effect was seen when the drug was given immediately after irradiation. Injection of 200-800 mg/kg TMG within 5 min of irradiation with 3 Gy produced a significant dose-dependent reduction in the radiation induced percent aberrant metaphases and in the frequency of micronucleated erythrocytes at 24h after exposure, with a corresponding decrease in the different types of aberrations. The optimum dose for protection without drug toxicity was 600 mg/kg body weight. At this dose, TMG produced 70 and >60% reduction in the radiation induced percent aberrant metaphases and micronucleated erythrocytes, respectively. The high water solubility and effectiveness when administered post-irradiation favor TMG as a likely candidate for protection in case of accidental exposures.     

In vivo radioprotection by 5-aminosalicylic acid

The radioprotective effect of 5-aminosalicylic acid (5ASA) was investigated in mouse bone marrow. The present study was aimed at investigating the radioprotective effect of pre-irradiation treatment with 5ASA against a range of whole-body lethal (8-11 Gy) and sublethal (1-4 Gy) doses of gamma-radiation (RT) in adult Swiss albino mice. Protection against lethal irradiation was evaluated from 30-day mouse survival and against sublethal doses was assessed from chromosomal aberrations in the bone marrow 24 h after irradiation. An intraperitoneal injection of 5ASA at a dose of 25mg/kg body weight (b. wt.) 30 min before lethal RT increased survival, giving a dose modification factor (DMF) of 1.08. Injection of 5ASA (25 mg/kg b. wt.) 60 or 30 min before or within 15 min after 3 Gy whole body RT resulted in a significant decrease in the radiation-induced aberrant metaphases, at 24 h post-irradiation. Maximum effect was seen when the drug was administered 30 min before irradiation. 5ASA (25 mg/kg b. wt.) significantly reduced the number of aberrant metaphases and the different types of aberrations at all the radiation doses (1-4 Gy) tested, giving a DMFs of 1.43 for number of aberrant metaphases. 5ASA pretreatment also significantly enhanced the endogenous spleen colonies in mouse exposed to 11 Gy RT. Pretreatment with 5ASA, protected plasmid DNA (pGEM-7Zf) against breakage induced by RT and Fenton reactants. Using nanosecond pulse radiolysis technique, the bimolecular rate constant of the reaction of 5ASA with hydroxyl radical was found to be 6.7x10(9)M(-1)s(-1). The p53 and p21 protein levels of bone marrow and spleen were evaluated to identify the specific molecular mechanisms. Both p53 and p21 increased 24h after 6 Gy irradiation, while treatment with 5ASA inhibited this RT-induced increase. Therefore, the present data suggest that 5ASA pretreatment decreases death caused by RT-induced gastrointestinal and hemopoeitic syndromes. The proposed mechanism of radioprotection by 5ASA is through the inhibition of damage to DNA, lipids, and proteins; and prevention of RT-induced increased expression of p53 and p21.     

The effects of increasing dosages of X-radiation on the chromosomes of the central mudminnow, Umbra limi (Kirtland) (Salmoniformes: Umbridae)

Fish subjected to 350 R, 660 R and 990 R of X-radiation showed chromosomal aberrations such as chromatid breaks and gaps, and chromatid exchanges between several chromosomes. The frequency of aberrations/metaphase increased with radiation dosage. Likewise, the percentage of aberrant cells increased with increased irradiation. The countable metaphases fish was lower for higher doses of radiation. At lower doses single chromatid breaks accounted for most of the aberrations whereas complex aberrations involving the breakage and exchange of fragments between several chromosomes were more frequent in fish subjected to 990 R. Gill tissue yielded three times as many countable metaphases as did spleen tissue.     

Anticlastogenic effect of Vitamin C on cisplatin induced chromosome aberrations in human lymphocyte cultures.

Vitamin C (ascorbic acid) is an antioxidant that can scavenge free radicals and protect cellular macromolecules, including DNA, from oxidative damage induced by different agents. The protective effect of Vitamin C on cisplatin induced chromosome aberrations has been determined in the human peripheral lymphocyte chromosome aberration test in vitro. The results of treatments with Vitamin C indicated that it statistically significantly decreases the number of chromosome aberrations and number of metaphases with aberrations induced with cisplatin, but it can not completely protect cells from damage. The test concentrations of Vitamin C (10 and 100 microg/ml) had a limited antimutagen effect on cisplatin (0.5 microg/ml), which can cause genetic damage through free radical mechanisms. The antimutagen effect included the anticlastogenic effect of Vitamin C and its ability to decrease the number of aneuploid mitoses. Vitamin C showed the most efficient anticlastogenic effect during simultaneous treatment with cisplatin. Also, Vitamin C reduced cell toxicity of cisplatin during simultaneous treatment.     

Chromosome aberration assays for the study of cyclophosphamide and Bacillus thuringiensis in Oxya chinensis (Orthoptera: Acrididae)

Chromosome aberrations induced by an anti-neoplastic drug, cyclophosphamide (CP) and a bioinsecticide, Bacillus thuringiensis (B.t.) were examined using grasshoppers as an animal model, with injection as the route of exposure. Oxya chinensis (Thunberg), having a small number (2n male symbol =23) of large-sized chromosomes in males, was used for this purpose. The fifth instar nymphs were treated with various concentrations of CP (2, 5 and 10 mg/ml) and B.t. (0.55, 1.83 and 5.50 IU/ml) by injection into the abdomen, using physiological saline and distilled water as negative controls, respectively. The chromosomal preparations were made from the spermatogonia of the specimen testis at different intervals after dosing (24 and 48 h). The effect of the high dose of CP (10 mg/ml) in O. chinensis was also analyzed at the 42-h time point. The chromosome aberrations observed were mainly chromatid and chromosome breaks. CP induced a dose- and time-dependents increase in the number of chromosome aberrations (CAs) per cell and in the percentage of aberrant cells. The strongest effect was seen when grasshoppers were injected with the highest dose and cells were analyzed at the 48-h time point. The results show that CP induced a significant increase in the frequency of CAs in testicular cells of O. chinensis with the three doses employed, compared to the negative control. Our results suggest that there exists in the grasshopper an enzyme system analogous to liver-S9 fraction, and that CP may be used as a positive control in genotoxicity test in this species. In addition, the evaluation of the chromosome aberrations induced by B.t. in the grasshoppers' testicular cells showed that B.t. may induce chromosome aberrations, mainly chromatid and chromosome breaks, in spermatogonia. By statistical analysis, B.t. showed significant dose-effect relationships and it may be mutagenic in this species. Recent research has focused on the development of biological insecticides to protect cereal crops against damage by insect species, such as beetles and grasshoppers. The present studies may contribute to our knowledge of entomological genotoxicity in grasshoppers and provide reference for the research on the mechanism of B.t. toxicity.     

Anticlastogenic effect of Vitamin C on cisplatin induced chromosome aberrations in human lymphocyte cultures

Vitamin C (ascorbic acid) is an antioxidant that can scavenge free radicals and protect cellular macromolecules, including DNA, from oxidative damage induced by different agents. The protective effect of Vitamin C on cisplatin induced chromosome aberrations has been determined in the human peripheral lymphocyte chromosome aberration test in vitro. The results of treatments with Vitamin C indicated that it statistically significantly decreases the number of chromosome aberrations and number of metaphases with aberrations induced with cisplatin, but it can not completely protect cells from damage. The test concentrations of Vitamin C (10 and 100 μg/ml) had a limited antimutagen effect on cisplatin (0.5 μg/ml), which can cause genetic damage through free radical mechanisms. The antimutagen effect included the anticlastogenic effect of Vitamin C and its ability to decrease the number of aneuploid mitoses. Vitamin C showed the most efficient anticlastogenic effect during simultaneous treatment with cisplatin. Also, Vitamin C reduced cell toxicity of cisplatin during simultaneous treatment.     

The "protective" effect of gamma-irradiation of cells in metaphase of mitosis following V-irradiation during the S period]

20,1% cells with chromosomes aberrations were obtained after UV-irradiation of embryonal fibroblasts of mice at the S-stage in vitro at a decreasing dose of 40erg/mm2. Subsequent gamma-irradiation at the metaphase of the first mitosis at a 5 krad dose led to a statistically significant decrease of the frequency of aberrant cells observed in the same mitosis down to 11,7%. The frequency of spontaneous aberrations did not change during the first few minutes after gamma-irradiation of intact cells at the metaphase. The "protective" effect of gamma-rays can not be explained either by unequal changes of the duration of mitotic stages for aberrant and normal cells, or by sticking of chromosome fragments or by breaks of bridges at the anaphase. The death of cells "under irradiation" also appears to be a hardly probable case of the effect observed. It is assumed that the decrease of the aberrations frequency is the result of predicted earlier modification of the processes of realization of potential chromosome damages into visible aberrations at the metaphase.     

Mutagenicity and clastogenicity of teniposide (VM-26) in L5178Y/TK +/- -3.7.2C mouse lymphoma cells

The antitumor drug teniposide (VM-26) is a potent inducer of DNA breaks (Long et al., Cancer Res., (1985) 45, 3106), but it is only weakly mutagenic at the hprt locus in CHO cells (Singh and Gupta, Cancer Res., (1983) 43, 577). In the present study, the mutagenic and clastogenic activities of teniposide were evaluated in L5178Y/TK +/- -3.7.2C mouse lymphoma cells. Although teniposide is a weak mutagen at the hprt locus, it is a potent mutagen at the tk locus, with as little as 0.5 ng/ml producing 220 TK mutants/10(6) survivors at 96% survival (background = 100/10(6) survivors). This same dose of teniposide induced 38 aberrations per 100 metaphases (background = 7/100 cells). At 7 ng/ml, teniposide induced approximately 2700 TK mutants/10(6) survivors at approximately 10% survival. At the highest dose sampled for aberration analysis (5 ng/ml), teniposide induced 44 aberrations/100 cells. Most of the aberrations were chromosomal rather than chromatid events. As expected for a compound acting primarily by a clastogenic mechanism, most of the TK mutants were small colonies. Thus, teniposide is a potent clastogen, and it is a potent mutagen at the tk locus but not at the hprt locus. These results support the hypothesis that the location of the target gene affects the ability of the assay to detect both intragenic events and events causing functional multilocus effects. Thus, a heterozygous locus (like tk) but not a functionally hemizygous locus (like hprt) may permit the detection of mutagens that act primarily by a clastogenic mechanism. Because teniposide induces topoisomerase II-associated DNA breaks, and because there is evidence that teniposide may not interact directly with DNA, we discuss the possibility that the potent clastogenic/mutagenic activity of teniposide may be mediated by topoisomerase II.     

Chromosome aberration assays for the study of cyclophosphamide and Bacillus thuringiensis in Oxya chinensis (Orthoptera: Acrididae)

Chromosome aberrations induced by an anti-neoplastic drug, cyclophosphamide (CP) and a bioinsecticide, Bacillus thuringiensis (B.t.) were examined using grasshoppers as an animal model, with injection as the route of exposure. Oxya chinensis (Thunberg), having a small number (2n♂=23) of large-sized chromosomes in males, was used for this purpose. The fifth instar nymphs were treated with various concentrations of CP (2, 5 and 10 mg/ml) and B.t. (0.55, 1.83 and 5.50 IU/ml) by injection into the abdomen, using physiological saline and distilled water as negative controls, respectively. The chromosomal preparations were made from the spermatogonia of the specimen testis at different intervals after dosing (24 and 48 h). The effect of the high dose of CP (10 mg/ml) in O. chinensis was also analyzed at the 42-h time point. The chromosome aberrations observed were mainly chromatid and chromosome breaks. CP induced a dose- and time-dependents increase in the number of chromosome aberrations (CAs) per cell and in the percentage of aberrant cells. The strongest effect was seen when grasshoppers were injected with the highest dose and cells were analyzed at the 48-h time point. The results show that CP induced a significant increase in the frequency of CAs in testicular cells of O. chinensis with the three doses employed, compared to the negative control. Our results suggest that there exists in the grasshopper an enzyme system analogous to liver-S9 fraction, and that CP may be used as a positive control in genotoxicity test in this species. In addition, the evaluation of the chromosome aberrations induced by B.t. in the grasshoppers’ testicular cells showed that B.t. may induce chromosome aberrations, mainly chromatid and chromosome breaks, in spermatogonia. By statistical analysis, B.t. showed significant dose–effect relationships and it may be mutagenic in this species. Recent research has focused on the development of biological insecticides to protect cereal crops against damage by insect species, such as beetles and grasshoppers. The present studies may contribute to our knowledge of entomological genotoxicity in grasshoppers and provide reference for the research on the mechanism of B.t. toxicity.     

Trans-generational radiation-induced chromosomal instability in the female enhances the action of chemical mutagens

Genomic instability can be produced by ionising radiation, so-called radiation-induced genomic instability, and chemical mutagens. Radiation-induced genomic instability occurs in both germinal and somatic cells and also in the offspring of irradiated individuals, and it is characterised by genetic changes including chromosomal rearrangements. The majority of studies of trans-generational, radiation-induced genomic instability have been described in the male germ line, whereas the authors who have chosen the female as a model are scarce. The aim of this work is to find out the radiation-induced effects in the foetal offspring of X-ray-treated female rats and, at the same time, the possible impact of this radiation-induced genomic instability on the action of a chemical mutagen. In order to achieve both goals, the quantity and quality of chromosomal damage were analysed.

In order to detect trans-generational genomic instability, a total of 4806 metaphases from foetal tissues from the foetal offspring of X-irradiated female rats (5 Gy, acute dose) were analysed. The study's results showed that there is radiation-induced genomic instability: the number of aberrant metaphases and the breaks per total metaphases studied increased and were found to be statistically significant (p ≤ 0.05), with regard to the control group.

In order to identify how this trans-generational, radiation-induced chromosomal instability could influence the chromosomal behaviour of the offspring of irradiated rat females in front of a chemical agent (aphidicolin), a total of 2481 metaphases were studied. The observed results showed that there is an enhancement of the action of the chemical agent: chromosomal breaks per aberrant metaphases show significant differences (p ≤ 0.05) in the X-ray- and aphidicolin-treated group as regards the aphidicolin-treated group.

In conclusion, our findings indicate that there is trans-generational, radiation-induced chromosomal instability in the foetal cells from X-ray-treated female rats and that this RIGI enhances the chromosomal damage caused by the chemical agent aphidicolin.     

Unusual dose--response of chromosome abberrations induced in human lymphocytes by very low dose exposures to tritium

Leukocyte cultures of human peripheral blood were chronically exposed for 48 h to tritiated water and [3H]thymidine using a wide range of tritium doses, and aberrations in lymphocyte chromosomes at the first metaphases were examined. In the experimental conditions, the types of aberrations induced by radiation emitted from both tritiated water and [3H]thymidine were mostly chromatid types, such as chromatid gaps and deletions. The dose--response relations for chromatid breaks per cell exhibited unusual dose-dependency in both cases. It was demonstrated that at higher dose range the yields of chromatid breaks increased linearly with dose, while those at lower dose range were significantly higher than would be expected by a downward extrapolation from the linear relation. Partial-hit or partial-target kinetic events appears at very low dose exposure.     

Comparative analysis of DNA breakage,chromosomal aberrations and apoptosis induced by the anti-herpes purine nucleoside analogues aciclovir,ganciclovir and penciclovir

Nucleoside analogues have been used in antiviral therapy and suicide cancer gene therapy. Therefore, it is of importance to compare their potential cytotoxic and genotoxic action. Using metabolically competent CHO cells expressing the thymidine kinase gene of herpes simplex virus type 1 (CHO-HSVtk cells) as a model system, the induction of DNA breaks was compared with the induction of structural chromosomal aberrations and apoptosis/necrosis after exposure to the anti-herpes nucleoside analogues aciclovir (ACV), ganciclovir (GCV) and penciclovir (PCV). After continuous treatment of CHO-HSVtk cells with the drugs, LD(10) in a colony-forming assay was 50, 0.5 and 1 microM for ACV, GCV and PCV, respectively, with GCV to be the most potent agent as determined at a given dose level. There was a remarkable difference in the activity of the agents to kill HSVtk expressing and non-expressing cells: the difference in cellular sensitivity of HSVtk(+) versus HSVtk(-) cells at LD(10) level was 7-fold for ACV, 60-fold for GCV and 400-fold for PCV. The drugs were shown to be strong inducers of apoptosis that was analysed as to concentration- and time-dependence; they induced to only very low extent necrosis. The agents were also highly potent in the induction of DNA single-strand breaks (SSBs) and double-strand breaks (DSBs) (as measured by single cell gel electrophoresis (SCGE)) and chromosomal aberrations. Although PCV induced DNA DSBs with a kinetics and frequency similar to that of GCV, it caused mostly condensation defects instead of "typical" structural chromosomal aberrations. For the drugs used, the frequency of apoptotic cells and the induction of abnormal mitoses appear to be related indicating genotoxic effects induced by the agents to be involved in cell killing due to apoptosis.     

Relationship of DNA repair to chromosome aberrations, sister-chromatid exchanges and survival during liquid-holding recovery in X-irradiated mammalian cells

The repair of X-ray induced DNA single strand breaks and DNA—protein cross-links was investigated in stationary phase, contact-inhibited mouse cells by the alkaline-elution technique. Approx. 90% of X-ray induced single strand breaks were rejoined during the first hour of repair, whereas most of the remaining breaks were rejoined more slowly during the next 5 h. At early repair times, the number of residual non-rejoined sungle strand breaks was approx. proportional to the X-ray dose. DNA—protein cross-links were removed at a slower rate (T1/2 approx. 10–12 h). Cells were held in stationary growth for various periods of time after irradiation before subculture at low density to score for colony survival (potentially lethal damage repair), chromosome aberrations in the first mitosis, and sister-chromatid exchanges in the second mitosis. Both cell killing and the frequency of chromosome aberrations decreased during the first several hours of recovery, reaching a minimum level by 6 h; this decrease correlated temporally with the repair of the slowly rejoining DNA-strand breaks. Relatively few sister-chromatid exchanges were observed when the cells were subcultured immediately after X-ray. The exchange frequency rose to maximum levels after a 4-h recovery interval, and returned to control levels after 12 h of recovery. The possible relationship of DNA repair to these changes in survival, chromosome aberrations, and sister-chromatid exchanges during liquid-holding recovery is discussed.