Mutagen-induced DNA Damage Research Articles

In Vitro Genotoxic and Antigenotoxic Effects of Ten Novel Synthesized 4-Thiazolidinone Derivatives.

Heterocyclic compounds are found in a variety of drug molecules, and bioactive natural products. 4-Thiazolidinones (4-TZDs), which represent an important class of heterocyclic compounds, are of great interest today with their diverse bioactivities. In this study, ten novel 4-TZD derivatives (C1-C10) were synthesized, characterized by spectroscopic techniques, and their genotoxic, and antigenotoxic properties were investigated in vitro using the Ames Salmonella/microsome mutagenicity assay in the concentration range of 0.2-1.0 mM/plate. The results revealed that none of the compounds were mutagenic on the three different Salmonella typhimurium strains up to the highest concentration tested. Furthermore, in our study, C1, C4, C6, and C9 showed significant, ranging from moderate to strong, antigenotoxic effects against mutagen-induced DNA damage at relatively higher doses. Among these, C4 had the best potential to inhibit the number of revertant colonies induced by 9-aminoacridine (9-AA), with a maximum inhibition rate of 47.9 % for 1.0 mM/plate. As a result, preliminary knowledge about the safety of the use of ten novel synthesized 4-TZD compounds likely to exhibit many bioactivities was obtained in this study. In addition, the significant in vitro antimutagenic activity of some derivatives increases the importance of studies for the development of new pharmacological agents for cancer prevention.

Increased Mutagen Sensitivity and DNA Damage in Pulmonary Arterial Hypertension.

Pulmonary arterial hypertension (PAH) is a serious lung condition characterized by vascular remodeling in the precapillary pulmonary arterioles. We and others have demonstrated chromosomal abnormalities and increased DNA damage in PAH lung vascular cells, but their timing and role in disease pathogenesis is unknown. We hypothesized that if DNA damage predates PAH, it might be an intrinsic cell property that is present outside the diseased lung. We measured DNA damage, mutagen sensitivity, and reactive oxygen species (ROS) in lung and blood cells from patients with Group 1 PAH, their relatives, and unrelated control subjects. Baseline DNA damage was significantly elevated in PAH, both in pulmonary artery endothelial cells (P < 0.05) and peripheral blood mononuclear cells (PBMC) (P < 0.001). Remarkably, PBMC from unaffected relatives showed similar increases, indicating this is not related to PAH treatments. ROS levels were also higher (P < 0.01). DNA damage correlated with ROS production and was suppressed by antioxidants (P < 0.001). PBMC from patients and relatives also showed markedly increased sensitivity to two chemotherapeutic drugs, bleomycin and etoposide (P < 0.001). Results were consistent across idiopathic, heritable, and associated PAH groups. Levels of baseline and mutagen-induced DNA damage are intrinsically higher in PAH cells. Similar results in PBMC from unaffected relatives suggest this may be a genetically determined trait that predates disease onset and may act as a risk factor contributing to lung vascular remodeling following endothelial cell injury. Further studies are required to fully characterize mutagen sensitivity, which could have important implications for clinical management.

Impaired DNA Damage Repair Capacity is Associated with an Increased Risk of Esophageal Adenocarcinoma: A Case Control Study

Background: Inherited suboptimal DNA repair capacity in peripheral blood lymphocytes (PBLs) can be unmasked by mutagen challenge and has been associated with susceptibility to cancer. Purpose: To use comet assay to assess the esophageal adenocarcinoma (EAC) risk in relation to mutageninduced DNA damage in PBLs. Materials and methods: In a case-control study, the baseline, benzo[a]pyrene diol epoxide (BPDE)-induced, and γ radiation-induced DNA damage were quantified by the Olive tail moment (TM) in PBLs from 172 Caucasian EAC patients and 154 healthy controls who were frequency matched on age and gender. Logistic regression analysis was used to calculate odds ratios (OR) and 95% confidence intervals (CI) to estimate EAC risk in relation to DNA damage. Results: EAC patients tended to have higher DNA damage than controls, as measured by baseline, net BPDEand net γ radiation-induced TM, but the difference was statistically significant only for net BPDE-induced DNA damage (0.88 ± 0.94 vs. 0.62 ± 0.77, P=0.031). Using the 75th percentile TM in the controls as cutoff point, we found that high levels of net BPDE- and γ radiation-induced DNA damage were associated with significantly increased risks of EAC, with adjusted ORs of 2.15 (95% CI, 1.13-4.10) and 2.27 (95% CI, 1.24–4.16), respectively. EAC risks were further increased among individuals with both net mutagen-induced DNA damages and exposure to gastroesophageal reflux disease or smoking, known risk factors for EAC. Conclusion: Our results suggest impaired repair capacity of mutagen-induced DNA damage in PBLs assessed by comet assay may be a risk factor of EAC.

In silico study of effects of polymorphisms on biophysical chemical properties of oxidized N-terminal domain of X-ray cross-complementing group 1 protein

Base excision repair (BER) is the major pathway involved in removal of endogenous and mutagen-induced DNA damage. The X-ray cross-complementing group 1 protein (XRCC1), which participates in BER, is a scaffolding protein. The oxidized XRCC1 N-terminal domain (NTD) forms additional interactions with DNA polymerase β (Pol β). Any change in the residues of a protein (XRCC1, XRCC4, etc.) may alter its stability and function. Many coding regions of genes have single nucleotide polymorphisms (SNPs) that change the conformation of their products, and they are probably involved in some diseases. The R7L and R107H mutations are located in the XRCC1-NTD. In the present study, biophysical chemical properties of oxidized XRCC1-NTD (wild type or mutants) were investigated at different temperatures (290, 295, 298, 301, 304, 309, 310, 311, and 312 K) in water using in silico molecular mechanic computational methods. Comparison of the average calculated potential energies of oxidized XRCC1-NTD reveals that the R7L mutation increases stability, but the R107H and R7L&R107H mutations are destabilizing. Therefore, mutant types of this protein (R107H or R7L&R107H) may not function correctly. Furthermore, quantitative structure-activity relationship (QSAR) of oxidized XRCC1-NTD and docking assay showed that the R7L mutation is advantageous but the R107H and R7L&R107H mutations are disadvantageous for XRCC1-NTD, and in the latter cases it cannot interact with Pol β as well as the wild type does. Hence, DNA repair may be defective. Also, using the equation dE = ∂Ε/(∂Τ)V·dT + ∂Ε/(∂V)T·dV, it was determined that the best temperature for normal activity of oxidized XRCC1-NTD is exactly the natural body temperature (310 K).

Risk Assessment of Esophageal Adenocarcinoma Using γ-H2AX Assay

Mutagen-induced DNA damage as measured in peripheral blood lymphocytes (PBL) has been associated with increased risks of cancers. The formation of γ-H2AX is an early cellular response to DNA double-strand breaks (DSB). We hypothesize that higher level of radiation-induced γ-H2AX in PBLs may be associated with an increased risk of esophageal adenocarcinoma. Laser scanning cytometer-based immunocytochemical method was used to measure baseline and irradiation-induced γ-H2AX levels in PBLs from 211 patients with esophageal adenocarcinoma and 211 healthy controls. The ratio of induced γ-H2AX level to baseline level was used to evaluate individual susceptibility to DSBs. Relative risks for esophageal adenocarcinoma associated with γ-H2AX were assessed by multivariable logistic regression analysis. Radiation-induced γ-H2AX level and the γ-H2AX ratio were significantly higher in cases than in controls. Dichotomized at the median in controls, a significantly increased risk for esophageal adenocarcinoma was observed in association with high γ-H2AX ratio [OR = 2.94; 95% confidence interval (CI), 1.83-4.72]. Quartile analyses showed significant dose-response associations between higher γ-H2AX ratio and increased risk of esophageal adenocarcinoma (Ptrend, 1.64E-06). In addition, joint effect between γ-H2AX ratio and smoking was observed: smokers who had high γ-H2AX ratio exhibited the highest risk of esophageal adenocarcinoma (OR = 5.53; 95% CI, 2.71-11.25) compared with never smokers with low γ-H2AX ratio. Radiation-induced DNA damage assessed by γ-H2AX ratio is associated with an increased risk of esophageal adenocarcinoma. γ-H2AX assay is a new and robust method to measure DSB damage in PBLs, which can be used to assess mutagen sensitivity and esophageal adenocarcinoma risk.

Spanish honeys protect against food mutagen-induced DNA damage

Honey contains a variety of polyphenols and represents a good source of antioxidants, while the human diet often contains compounds that can cause DNA damage. The present study investigated the protective effect of three commercial honey samples of different floral origin (rosemary, heather and heterofloral) from Madrid Autonomic Community (Spain) as well as an artificial honey on DNA damage induced by dietary mutagens, using a human hepatoma cell line (HepG2) as in vitro model system and evaluation by the alkaline single-cell gel electrophoresis or comet assay. Rosemary, heather and heterofloral honeys protected against DNA strand breaks induced by N-nitrosopyrrolidine (NPYR), benzo(a)pyrene (BaP) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), but none of the honey samples tested prevented DNA strand breaks induced by N-nitrosodimethylamine (NDMA). Heterofloral and heather (unifloral) honeys with higher phenolic content were most effective in protecting HepG2 cells against DNA damage induced by food mutagens. Heterofloral honey was more protective against NPYR and BaP, while heather honey was more protective against PhIP. Artificial honey did not show a protective effect against DNA damage induced by any of the food mutagens tested, indicating that the protective effects of honeys could not be due to their sugar components. The results suggest that the protective effect of three kinds of Spanish honey of different floral origin could be attributed in part to the phenolics present in the samples. Honeys with higher phenolic content, i.e. heather and heterofloral honeys, were most effective in protecting against food mutagen-induced DNA damage in HepG2 cells. In addition, a possible synergistic effect between other minor honey components could also be involved.

Comet‐FISH with rDNA probes for the analysis of mutagen‐induced DNA damage in plant cells

We used comet-fluorescence in situ hybridization (FISH) in the model plant species Crepis capillaris following exposure of seedlings to maleic hydrazide (MH). FISH with 5S and 25S rDNA probes was applied to comets obtained under alkaline conditions to establish whether these DNA regions were preferentially involved in comet tail formation. MH treatment induced significant fragmentation of nuclear DNA and of rDNA loci. A 24-h post-treatment recovery period allowed a partial reversibility of MH-induced damage on nuclear and rDNA regions. Analyses of FISH signals demonstrated that rDNA sequences were always involved in tail formation and that 5S rDNA was more frequently present in the tail than 25S rDNA, regardless of treatment. The involvement of 25S rDNA in nucleolus formation and differences in chromatin structure between the two loci may explain the different susceptibility of the 25S and 5S rDNA regions to migrate into the tail. This work is the first report on the application of FISH to comet preparations from plants to analyze the distribution and repair of DNA damage within specific genomic regions after mutagenic treatment. Moreover, our work suggests that comet-FISH in plants may be a useful tool for environmental monitoring assessment.

The pol3-t Hyperrecombination Phenotype and DNA Damage‐Induced Recombination in Saccharomyces cerevisiae Is RAD50 Dependent

The DNA polymerase δ (POL3/CDC2) allele pol3-t of Saccharomyces cerevisiae has previously been shown to be sensitive to methylmethanesulfonate (MMS) and has been proposed to be involved in base excision repair. Our results, however, show that the pol3-t mutation is synergistic for MMS sensitivity with MAG1, a known base excision repair gene, but it is epistatic with rad50Δ, suggesting that POL3 may be involved not only in base excision repair but also in a RAD50 dependent function. We further studied the interaction of pol3-t with rad50Δ by examining their effect on spontaneous, MMS-, UV-, and ionizing radiation-induced intrachromosomal recombination. We found that rad50Δ completely abolishes the elevated spontaneous frequency of intrachromosomal recombination in the pol3-t mutant and significantly decreases UV- and MMS-induced recombination in both POL3 and pol3-t strains. Interestingly, rad50Δ had no effect on γ-ray-induced recombination in both backgrounds between 0 and 50 Gy. Finally, the deletion of RAD50 had no effect on the elevated frequency of homologous integration conferred by the pol3-t mutation. RAD50 is possibly involved in resolution of replication forks that are stalled by mutagen-induced external DNA damage, or internal DNA damage produced by growing the pol3-t mutant at the restrictive temperature.

Attenuation of acridine mutagen ICR-191 — DNA interactions and DNA damage by the mutagen interceptor chlorophyllin

We have investigated the ability of chlorophyllin (CHL) to interact with acridine mutagen ICR-191 (2-methoxy-6-chloro-9-(3-(2-chloroethyl)aminopropylamino)acridine) and also its ability to decrease binding of ICR-191 to DNA in a simple three-component competition system: CHL-ICR–DNA. Our data indicate a strong association of ICR-191 with CHL, stronger even than the association of ICR-191 with DNA. Calculations based on the measured affinity data show that a two- to three-fold excess of CHL reduces by about two-fold the concentration of the mutagen-DNA complex. We also exposed human leukemic HL-60 cells to ICR-191 in the absence and presence of CHL and measured the mutagen-induced DNA damage. The extent of DNA damage was assessed by analysis of histone H2AX phosphorylation. While ICR-191 induced significant increase in expression of phosphorylated H2AX (γH2AX), particularly in DNA replicating cells, this increase was totally abolished in the cells treated with ICR-191 in the presence of CHL.

Deficiency of cell cycle checkpoints and DNA repair system predispose individuals to esophageal cancer

Cell cycle checkpoints and DNA repair capacity are critical for the maintenance of genome integrity. We hypothesized that, in comparison to healthy controls, esophageal cancer patients might have a higher frequency of deficiencies in cell cycle checkpoints and/or DNA repair system. Using flow cytometry and comet assay, we assessed the γ-radiation-induced S phase and G2-M phase accumulation, and benzo( a)pyrene-diol-epoxide (BPDE)- and γ-radiation-induced DNA damage, in peripheral blood lymphocytes of 99 newly diagnosed esophageal cancer patients and 112 age-, gender-, and ethnicity-matched healthy controls. The mean γ-radiation-induced cell accumulation at G2-M phase was significantly lower in esophageal cancer patients than the control subjects (case versus control: 5.27% ± 5.11% versus. 7.06% ± 5.04%, P = 0.013). The less G2-M phase cell accumulation resulted in a significant increased risk for esophageal cancer with an odds ratio of 2.08 (95% confidence interval 1.15–3.77). After normalization to baseline S fraction, the radiation-induced increment in the 4N/2N ratio was also significantly lower in esophageal cancer patients than in controls (case versus control: 0.76% versus 1.04%, P = 0.0039). The less increment in the radiation-induced 4N/2N ratio was associated with 2.24(95% confidence interval 1.22–4.11)-fold increase of esophageal cancer risk. We also compared the mutagen-induced DNA damage level among individuals with different S or G2-M phase cell accumulation. We found that the less G2-M phase accumulation was associated with both high BPDE induced and γ-radiation-induced DNA damage in the healthy controls ( P for trend = 0.023 and 0.015, respectively). Similar pattern was observed for S phase accumulation ( P for trend = 0.033 and 0.022, respectively). However, such association was not seen in esophageal cancer patients. This study provides the first molecular epidemiologic evidence linking increased esophageal cancer risk with defects in cell-cycle checkpoints and DNA repair capacity.

Comet-assay in combination with PNA-FISH detects mutagen-induced DNA damage and specific repeat sequences in the damaged DNA of transformed cells

The Comet-assay was applied to three transformed cell lines (HT1080, CCRF-CEM line and CHO) which were treated with the cytostatics bleomycin (BLM) or mitomycin C (MMC). In addition, PNA probes for the telomere repeat (TTAGGG)(n) were used for detection of telomeric DNA sequences in the damaged DNA. Data were compared with previously obtained results from peripheral leukocytes. The amount of migrating DNA increased in all cell types in a dose-dependent manner after BLM exposure. CHO cells reacted sensitively at low doses of the mutagen, and leukocytes had the highest dose-related effect up to 25 IU/ml which, however, did not further increase. A rather linear dose response characterized the HT1080 cells, the effect was lowest for the CCRF-CEM cells. While MMC at lower doses increased the percentage of migrating DNA in a dose-dependent manner, the higher doses induced shorter comets, on average, than the lower ones in all cell lines. With PNA-Comet-FISH obvious differences were found between the studied cell lines with respect to quantitative head/tail distribution of telomeric signals after BLM exposure. A large number of signal spots of various sizes were found in CHO cells, very small signals could be detected in the comets of both neoplasia cell lines. Dose-dependence of telomeres in the tail was most pro-nounced in CCRF-CEM and normal leukocytes, less in HT1080. The steepest dose-related increase of telomeric signals in the tail was found in CHO cells. The ratio between the migrated DNA and the telomeric signals in the tail varied distinctly between the examined cell types from 3:1 to 1:1. Taken together, Comet-FISH can detect mutagenic effects on specific DNA sequences. This may be of high practical value if amplified DNA sequences will be addressed by those examinations in future.

Dietary Carotenoids and Genetic Instability Modify Bladder Cancer Risk

In vitro and in vivo studies have shown that carotenoid supplementation is associated with decreased DNA damage, but the role of dietary carotenoids in cancer risk remains controversial because epidemiologic studies have yielded conflicting results. Limited data exist regarding the role of dietary carotenoids in the context of constitutional genetic instability in cancer risk. This case-control study estimated dietary carotenoid intake [microg/(kJ . d)] from a FFQ for 423 patients with bladder cancer and 467 healthy controls, and quantified baseline and benzo[a]pyrene diol epoxide (BPDE)- and gamma-radiation-induced DNA damage in the peripheral blood lymphocytes using the comet assay. Overall, intake of total carotenoids was lower (P < 0.01) for bladder cancer cases (mean +/- SD: 1273.4 +/- 688.9) compared with healthy controls (1501.3 +/- 791.5). When categorized into quartiles, there was an inverse association between increasing levels of carotenoid intake and bladder cancer risk with greatest protective effect [odds ratio (OR) = 0.56, 95% CI, 0.37-0.85] in the quartile with the highest level of intake. Baseline and mutagen-induced DNA damage was significantly higher in cases than in controls; when analyzed jointly with carotenoid intake, high DNA damage and low carotenoid intake were associated with the highest risk. For example, with high baseline DNA damage and low total carotenoid intake, the OR was 3.08 (95% CI, 1.64-5.77); with high baseline DNA damage and high total carotenoid intake, the risk was somewhat attenuated (OR = 2.49, 95% CI, 1.28-4.84). The risk was decreased further for low baseline DNA damage and low total carotenoid intake (OR = 2.18; 95% CI, 1.13-4.22). This study provides evidence of a preventive role for carotenoids in bladder cancer, and these data may have important implications for cancer prevention, especially for individuals susceptible to DNA damage.

The Fanconi Anaemia Gene FANCC Promotes Homologous Recombination and Error-Prone DNA Repair

The Fanconi anemia (FA) protein FANCC is essential for chromosome stability in vertebrate cells, a feature underscored by the extreme sensitivity of FANCC-deficient cells to agents that crosslink DNA. However, it is not known how this FA protein facilitates the repair of both endogenously acquired and mutagen-induced DNA damage. Here, we use the model vertebrate cell line DT40 to address this question. We discover that apart from functioning in homologous recombination, FANCC also promotes the mutational repair of endogenously generated abasic sites. Moreover in these vertebrate cells, the efficient repair of crosslinks requires the combined functions of FANCC, translesion synthesis, and homologous recombination. These studies reveal that the FA proteins cooperate with key mutagenesis and repair processes that enable replication of damaged DNA.

Differential genotoxicity of ethyl methanesulphonate, N-ethyl- N-nitrosourea and maleic hydrazide in tobacco seedlings based on data of the Comet assay and two recombination assays

The purpose of this study was to determine if mutagen-induced DNA damage is correlated with the frequency of induced recombination events. The alkylating agents ethyl methanesulphonate (EMS) and N-ethyl- N-nitrosourea (ENU), and the plant growth regulator and herbicide maleic hydrazide (MH) were compared in tobacco seedlings for their ability to induce DNA damage measured by the Comet assay, and recombination activity measured by the GUS gene reactivation assay, and by the somatic twin sectors assay. While EMS and ENU induced a dose-dependent increase in DNA damage in leaf nuclei, MH had no significant effect. By contrast, MH induced a 6-fold higher frequency of homologous recombination as expressed by the GUS assay and a 2.8-fold higher frequency of somatic twin sectors than after EMS treatments.

The comet assay: genotoxic damage or nuclear fragmentation?

The single cell gel electrophoresis (SCGE) or comet assay is based on the assumption that comet images result from genotoxic damage that ultimately generate DNA single- or double-strand breaks. A criticism of the assay is that some or all of the comet images may be the result of apoptosis-mediated nuclear fragmentation. The objective of this study was to determine if mutagen-induced DNA damage leading to strand breakage observed in the SCGE assay was repairable or was due to nonrepairable nuclear fragmentation. Chinese hamster ovary cells were treated with ethylmethanesulfonate, 2-acetoxyacetylaminofluorene, or H(2)O(2). These mutagens induce genetic damage by different molecular mechanisms. One group of SCGE slides was prepared immediately after treatment, while parallel treated cultures were repeatedly washed and allowed to undergo liquid holding recovery for DNA repair. It was hypothesized that cells with genotoxic damage can repair their genomic DNA, while apoptotic cells cannot reverse nuclear fragmentation. We found a significant decrease in the tail moments of nuclei from mutagen-treated cells after 4 hr of liquid holding. However, this measurement may represent only those cells capable of repair. Apoptotic cells may continue DNA fragmentation during the recovery time and this DNA may become so diffuse that the nuclei disappear after electrophoresis. To overcome this possible artifact, images of nuclei were captured before and after alkaline electrophoresis. Constellations of nuclei were located on SCGE slides by their coordinates on the microscope stage. We found that no nuclei were lost due to apoptotic nuclear fragmentation and DNA migration. Even the so-called "hedgehog" comet images with extreme DNA damage were not lost during liquid holding. These data support the conclusion that mutagen-induced DNA damage is the principal cause of the damage measured in the comet assay.

The influence of temperature during alkaline treatment and electrophoresis on results obtained with the comet assay

The alkaline comet assay (single-cell gel electrophoresis) is becoming established as a genotoxicity test with manyfold applications in vitro and in vivo. While the underlying principles are identical, various modifications of the method are in use which clearly affect the sensitivity and resolving power of the assay. One variable of potential importance that has been disregarded until now is temperature during alkaline treatment and electrophoresis. We therefore performed comet assay experiments with human blood and V79 Chinese hamster cells using two different temperatures (4 and 20°C, i.e. room temperature) during alkaline treatment and electrophoresis. DNA damage was induced by the two standard mutagens gamma irradiation and methyl methanesulfonate (MMS). The results clearly indicate significant differences in the detection of background and mutagen-induced DNA damage at these two temperatures. Under otherwise identical test conditions (including the duration of alkaline treatment and electrophoresis), increased temperature during alkaline treatment and electrophoresis strongly enhances DNA migration. Our findings suggest that the comet assay should be performed under strictly controlled and reproducible temperature conditions. In any case the temperature during alkaline treatment and electrophoresis should be stated in a publication to allow for a critical evaluation of results obtained with the comet assay.

The effect of arsenic and cadmium on the persistence of mutagen-induced DNA damage in human cells

The effect of arsenic and cadmium on the persistence of mutagen-induced DNA damage in human cells

Quantification of DNA damage and repair in amino acid auxotrophs and UV-sensitive mutants of Aspergillus nidulans using an ELISA

An ELISA used to investigate DNA repair in mammalian cells has been adapted to investigate mutagen-induced DNA damage and repair in protoplasts of Aspergillus nidulans. The assay shows a reduced rate of repair of DNA damage in methionine and arginine auxotrophs ( methG and argB), which were shown previously to be hypersensitive to UV radiation and chemical mutagens. The assay also showed a considerably reduced ability to repair mutagen-induced damage in the uv-sensitive mutants uvsB and uvsH. The increased sensitivity of amino acids auxotrophs to mutagens is, therefore, correlated with a reduced capacity to repair mutagen-induced DNA damage.

Induction of Micronuclei by HTLV-I Tax: A Cellular Assay for Function

Cellular chromosomal damage is ubiquitously seen in HTLV-I-transformed lymphocytes. It is also characteristic of cells that have been exposed to mutagens. A sensitive measurement for mutagen-induced DNA damage is the formation of micronuclei in treated cells. Because current evidence suggests that HTLV-I Tax is etiologically linked to transformation, we tested for its activity in inducing micronuclei. We show here that transfection into cells of a Tax-producing plasmid rapidly induced the formation of micronuclei. This effect cooperated with that of a mutagen (mitomycin C) and was correlated with the inherent trans-activation capacity of Tax. These findings suggest that a commonly used mutagen assay could be a quick biological test for putatively oncogenic proteins.

Repair of sister-chromatid exchange-inducing lesions in mutagen-treated cultures of human whole blood and purified fresh or frozen lymphocytes

Repair of mutagen-induced lesions that result in sister-chromatid exchanges was evaluated in 10 normal individuals. The mutagens used were mitomycin C (MMC), 4-nitroquinoline 1-oxide (4NQO), and N-methyl- N′ nitro- N-nitrosoguanidine (MNNG). Cultures of whole blood, freshly purified lymphocytes, or purified lymphocytes cryopreserved for 6 months were analyzed after the mutagen treatments. All 3 mutagens induced reparable damage as evaluated by comparison of sister-chromatid exchanges between cultures that were given time to repair induced damage before 5-bromo-2′-deoxyuridine (BrdUrd)was added to the culture medium with those where BrdUrd was added immediately after the administration of the mutagens (MMC or 4NQO) or at culture initiation (MNNG). Repair of mutagen-induced DNA damage was detected in all 3 culture types; thus cryopreservation did not appear to alter the capacity of lymphocytes to repair mutagen-induced lesions. Quantitative differences in apparent repair capabilities were observed among individuals. Variability also existed among the different culture types within an individual, suggesting that caution should be exercised in interpreting these apparent differences.