DNA strand break and rejoining in cultured human fibroblasts exposed to fast neutrons or gamma rays

The production and rejoining of DNA single-strand and double-strand breaks have been monitored in monolayer cultures of proliferating human skin fibroblasts by means of sensitive techniques. Cells were irradiated with low doses of either 60Co gamma-rays or 14.6 MeV neutrons at 0 degrees C (0-5 Gy for measurement of single-strand breaks by alkaline elution and 0-50 Gy for double-strand breaks measured by neutral elution). The yield of single-strand breaks induced by neutrons was 30 per cent of that produced by the same dose of gamma-rays; whilst in the induction of double-strand breaks neutrons were 1.6 times as effective as gamma-rays. Upon post-irradiation incubation of cells at 37 degrees C, neutron-induced single-strand and double-strand breaks were rejoined with a similar time-course to gamma-induced breaks. Rejoining followed biphasic kinetics; of the single-strand breaks, 50 per cent disappeared within 2 min after gamma-rays and 6-10 min after neutrons. Fifty per cent of the double-strand breaks disappeared within 10 min, after gamma-rays and neutrons. Cells derived from patients suffering from ataxia-telangiectasia showed the same capacity for repair of single- and double-strand breaks induced by 14.6 MeV neutrons, as cells established from normal donors. The comparison of neutrons and gamma-rays in the induction of DNA breaks did not explain the elevated r.b.e. on high LET radiation. However, a study of the variation in the spectrum of lesions induced by different radiation sources will probably contribute to the clarification of the relative importance of other radio products.     

The induction and repair of DNA damage and its influence on cell death in primary human fibroblasts exposed to UV-A or UV-C irradiation

Irradiation with UV-A of normal human fibroblasts in phosphate-buffered saline induced cell death, measured as lack of colony-forming ability. A specially filtered sunlamp, emitting wavelengths greater than 330 nm, was used as UV-A source. After UV-A irradiation, single-strand breaks (alkali-labile bonds) could be detected in DNA; these lesions were rapidly repaired. The induction of these single-strand breaks was almost eliminated when irradiation was performed in the presence of catalase. However, catalase, when present during UV-A irradiation, did not reduce cell death of the fibroblasts. Excision repair, monitored as unscheduled DNA synthesis, was induced strongly by irradiation with UV-C (predominantly 254 nm), but could not be detected after UV-A irradiation. Moreover, very little accumulation of incision breaks during post-irradiation incubation with hydroxyurea and 1-beta-D-arabinofuranosylcytosine (araC) was detected after UV-A. This is consistent with the low amount of pyrimidine dimers (measured as UV-endonuclease susceptible sites) induced by UV-A. Xeroderma pigmentosum fibroblasts of complementation group A, which are extremely sensitive to UV-C irradiation, showed the same sensitivity to UV-A as normal fibroblasts. The results indicate that lethality by UV-A wavelengths greater than 330 nm is caused by lesions other than single-strand breaks (alkali-labile bonds) and pyrimidine dimers.     

Toxicity and DNA damage induced by 1-nitropyrene and its derivatives in Chinese hamster lung fibroblasts

1-Nitropyrene and its chemically synthesised derivatives were investigated for their cytotoxicity and ability to induce DNA-strand breaks in Chinese hamster lung fibroblasts. Both 1-nitrosopyrene (0.25-60 micrograms/ml) and 1-aminopyrene (0.25-25 micrograms/ml) were cytotoxic, and induced the formation of DNA lesions, which were measured as DNA single-strand breaks after sedimentation in alkaline sucrose-density gradients. Higher doses of 1-aminopyrene (25-60 micrograms/ml) inhibited the formation of DNA single-strand breaks. 1-Nitropyrene was not toxic (0.25-60 micrograms/ml) and induced low levels of detectable DNA strand breaks, whilst N-acetyl-1-aminopyrene was inactive. The post-mitochondrial supernatant fraction of Aroclor-induced rat-liver containing 4 mM NADPH (S9 mix) did not promote the activation of 1-nitropyrene. In fact DNA strand breaks induced by either 1-nitropyrene or 1-nitrosopyrene was abolished in the presence of S9 mix. The 1-nitropyrene reduced intermediate, N-hydroxy-1-aminopyrene was synthesised by the reduction of 1-nitrosopyrene with ascorbic acid. In the presence of ascorbic acid, 1-nitrosopyrene caused a 5-fold increase in the number of DNA single-strand breaks when compared to cells treated with 1-nitrosopyrene alone. The results are discussed in terms of the metabolic activation of 1-nitropyrene and 1-aminopyrene in Chinese hamster lung cells.     

Effect of 3-aminobenzamide on DNA strand-break rejoining and cytotoxicity in CHO cells treated with hydrogen peroxide

The influence of the nuclear ADP-ribosyltransferase inhibitor 3-aminobenzamide on the DNA strand-break rejoining kinetics and cytotoxicity in Chinese hamster ovary cells following H2O2 treatment was investigated. For the DNA damage studies, cells were treated on ice with H2O2 (0-20 microM) for 1 h in serum-free medium, after which the H2O2 was removed and the cells were allowed to repair their damage in complete medium at 37 degrees C in the presence or absence of 3-aminobenzamide (5 mM) for periods up to 2 h. The DNA strand breaks remaining as a function of time were then estimated by alkaline elution. A linear relationship between the H2O2 concentration and the initial level of DNA single-strand breaks (zero time allowed for repair) was observed. No double-strand breaks or DNA-protein cross-links were detected at these doses. The rejoining of single-strand breaks after H2O2 (20 microM) alone was characterized by a single exponential process with a t1/2 of approx. 5 min. However, in the presence of 3-aminobenzamide, rejoining was much slower and biphasic, with t1/2 of approx. 10 and 36 min. The inhibitory action of 3-aminobenzamide was concentration-dependent and completely reversible in that, when the 3-aminobenzamide was removed from the treated cultures, the strand-break rejoining kinetics rapidly returned to the t1/2 of 5 min typical of H2O2 alone. Considerably higher concentrations of H2O2 (up to 600 microM) were required for cell killing compared to the DNA damage studies. Cell killing by H2O2 alone was characterized by a shoulderless, exponential survival curve (D0 = 880 microM). The cytotoxicity was potentiated when the cells were treated with 3-aminobenzamide (5 mM) for 1 h after the H2O2 treatment; the survival curve with 3-aminobenzamide also assumed a biphasic character (D0 of 212 microM and 520 microM). These results are consistent with the theory that OH.-induced single-strand breaks do not normally represent lethal lesions to the cell because of their rapid, efficient repair. However, interference with these repair processes (in this case by 3-aminobenzamide) can alter this relationship, possibly allowing lesion fixation.     

XRCC1 and DNA strand break repair

DNA single-strand breaks can arise indirectly, as normal intermediates of DNA base excision repair, or directly from damage to deoxyribose. Because single-strand breaks are induced by endogenous reactive molecules such as reactive oxygen species, these lesions pose a continuous threat to genetic integrity. XRCC1 protein plays a major role in facilitating the repair of single-strand breaks in mammalian cells, via an ability to interact with multiple enzymatic components of repair reactions. Here, the protein-protein interactions facilitated by XRCC1, and the repair processes in which these interactions operate, are reviewed. Models for the repair of single-strand breaks during base excision repair and at direct breaks are presented.     

Differential repair of 1-beta-D-arabinofuranosylcytosine-detectable sites in DNA of human fibroblasts exposed to ultraviolet light and 4-nitroquinoline 1-oxide

The extent of DNA excision repair was determined in dermal fibroblast strains from clinically normal and xeroderma pigmentosum (XP; complementation group A) human donors after single or combined exposures to 254-nm ultraviolet light and 4-nitroquinoline 1-oxide (4NQO). The repair was monitored by incubation of the treated cultures in the presence of 1-beta-D-arabinofuranosylcytosine (araC), a potent inhibitor of long-patch excision repair, followed by quantitation of araC-accumulated DNA single-strand breaks (representing repair events) by velocity sedimentation analysis in alkaline sucrose gradients. The amount of repair in normal fibroblast strains increased as a function of UV fluence and reached a plateau at 15 J/m2; strand breaks were not detected when these same cultures were irradiated with as much as 60 J/m2 UV and incubated in the absence of araC, implying that an initial (incision) step is rate-limiting in the repair of UV damage. In normal fibroblasts (i) the incidence of araC-detectable lesions removed during fixed intervals following exposure to 4NQO (4 microM; 30 min) was approximately 2.5 times greater than that seen following irradiation with repair-saturating fluences (greater than or equal to 15 J/m2) of UV-rays; and (ii) the amount of repair in cultures treated simultaneously with 4NQO (0.5-6 microM; 30 min) and a repair-saturating fluence of UV (20 J/m2) was found to approach the sum of that arising from exposure to each separately. The XP cells (XP12BE) exhibited a deficiency in the removal of araC-detectable DNA lesions following exposure to either of the carcinogens. Since araC is known to inhibit the repair of alkali-stable 4NQO-DNA adducts (i.e., lesions assumed to be removed by the UV-like excision pathway) but not that of alkali-labile sites (i.e., DNA lesions operated on by the X-ray-like repair pathway), our results strongly imply that the multistep excision-repair pathway operative on UV photoproducts in human fibroblasts differs from that responsible for removing alkali-stable (araC-detectable) 4NQO adducts by at least one step, presumably the rate-limiting incision reaction mediated by a lesion-recognizing endonuclease.     

Induction of chromosome damage by Neurospora endonuclease in repair-inhibited quiescent normal human fibroblasts

The purpose of these experiments was to determine the role of double-strand breaks in chromosome aberration formations. Quiescent normal human fibroblasts were treated with 3 μM nitrogen mustard and then allowed to repair their DNA damage for 24 h prior to cell fusion and induction of premature chromosome condensation. The extent of chromosome damage was determined in the G1 prematurely condensed chromosomes (G1 PCC). The presence of cytosine arabinoside and hydroxyurea during the repair period in order to accumulate single-strand DNA breaks resulted in an increase in the chromosome-break frequency. Treatment of these repair-inhibited cells with single-strand-specific neurospora endonuclease during fusion to change single-strand lesions into double-strand breajs resulted in a doubling of the aberration frequency. These results support the notion that double-strand breaks are important in chromosome-aberration formation.     

Induction and repair of DNA damage in gamma-irradiated human lymphoblasts: irradiation in the presence and absence of misonidazole

The effects of oxygen and misonidazole on the induction of DNA lesions were examined in human TK6 lymphoblasts irradiated with 60Co gamma rays. We have investigated both the formation and subsequent repair of two classes of DNA damage, single-strand breaks and lesions recognized by the gamma endonuclease activity in a cell-free extract of Micrococcus luteus. Relative to irradiation under hypoxia, single-strand break yields were increased by the presence of either oxygen or misonidazole at the time of irradiation. In contrast, M. luteus enzyme-sensitive site yields were unaffected by the presence of either oxygen or misonidazole. No significant differences in single-strand break or enzyme-sensitive site repair kinetics were observed for lesions induced under any of the irradiation conditions employed. These results confirm the sensitizing effects of oxygen and oxygen-mimetic drugs on the induction of single-strand breaks but provide no support for their ability to enhance the induction of enzyme-sensitive sites.     

Hypochlorous acid potentiates hydrogen peroxide-mediated DNA-strand breaks in human mononuclear leucocytes.

In this study the formation of DNA single-strand breaks in MNL in close proximity to activated phagocytes, or in contact with added H2O2 and/or HOCl, were evaluated. Neutrophils activated by phorbol myristate acetate (PMA), induced DNA-strand breaks in neighboring lymphocytes which increased after 1-2 h incubation in a repair medium. These DNA-strand breaks could be prevented by the addition of catalase or substitution of the neutrophils with cells from a patient with chronic granulomatous disease. Inclusion of the myeloperoxidase (MPO) inhibitor, sodium azide (NaN3), to the system was associated with less damage after 1-2 h incubation and a faster repair rate. Exposure of MNL to added reagent H2O2 (12-100 microM) was also accompanied by DNA damage. Addition of reagent HOCl (3-25 microM) did not induce any DNA-strand breaks. However, when combined with H2O2 (12.5 microM), HOCl increased H2O2-mediated DNA damage and compromised the repair process. Interactions between the phagocyte-derived reactive oxidants H2O2 and HOCl are probably involved in the etiology of inflammation-related cancer.     

Hydrogen peroxide-induced DNA damage in bovine lens epithelial cells

The present investigation was undertaken to determine the types and extent of DNA damage resulting from incubation of primary cultures of bovine lens epithelial cells with hydrogen peroxide. Significant numbers of DNA single-strand breaks were detected by alkaline elution after exposure to as little as 25 microM H2O2 for 5 min at 37 degrees C. The extent of single-strand breakage was concentration dependent and linear from 25 to 200 microM H2O2. The observed single-strand breaks appear primarily due to the action of the hydroxyl radical via a Fenton reaction as both an iron chelator, 1,10-phenanthroline and OH. scavengers, including DMSO, KI and glycerol, significantly inhibited the DNA-damaging effect of H2O2. Diethyldithiocarbamate, an inhibitor of superoxide dismutase, further potentiated the DNA-damaging effects of H2O2, presumably by increasing the steady-state concentration of Fe2+. DNA-protein cross-linking was not observed. In addition, significant levels of 5,6-saturated thymine residues or pyrimidine dimers were not detected after modification of the alkaline elution methodology to allow the use of either E. coli endonuclease III or bacteriophage T4 endonuclease V, respectively. No double-strand breaks were detected after incubation of epithelial cell cultures with H2O2 concentrations of up to 400 microM for 10 min and subsequent neutral filter elution. Since, in vivo, the lens epithelium contains populations of both quiescent and dividing cells, the degree of susceptibility to oxidative damage was also studied in actively growing and plateau-phase cultures. Reduced levels of single-strand breakage were observed when plateau-phase cultures were compared to actively growing cells. In contrast, essentially no differences in repair rates were noted at equitoxic doses of H2O2. The above results suggest that lens epithelial cells may be particularly sensitive to oxidative damage and thus are a good model system in which to study the effects of oxidative stress.     

Repair of MMS-induced DNA double-strand breaks in haploid cells of Saccharomyces cerevisiae, which requires the presence of a duplicate genome.

Summary The formation and repair of double-strand breaks induced in DNA by MMS was studied in haploid wild type and MMS-sensitive rad6 mutant strains of Saccharomyces cerevisiae with the use of the neutral and alkaline sucrose sedimentation technique. A similar decrease in average molecular weight of double-stranded DNA from 5–6x10⁸ to 1–0.7x10⁸ daltons was observed following treatment with 0.5% MMS in wild type and mutant strains. Incubation of cells after MMS treatment in a fresh drug-free growing medium resulted in repair of double-strand breaks in the wild type strain, but only in the exponential phase of growth. No repair of double-strand breaks was found when cells of the wild type strain were synchronized in G-1 phase by treatment with factor, although DNA single-strand breaks were still efficiently repaired. Mutant rad6 which has a very low ability to repair MMS-induced single-strand breaks, did not repair double-strand breaks regardless of the phase of growth.These results suggest that (1) repair of double-strand breaks requires the ability for single-strand breaks repair, (2) rejoining of double-strand breaks requires the availability of two homologous DNA molecules, this strongly supports the recombinational model of DNA repair.     

Less repair of pyrimidine dimers and single-strand breaks in genes by scid cells.

Severe combined immunodeficient (Scid) mice have a mutation in the catalytic subunit of the DNA binding protein kinase that is involved in repair of double-strand breaks in DNA. To determine if the protein also influences repair of single-strand breaks, we examined the ability of Scid cells to repair lesions introduced by ultraviolet light and gamma-ray irradiation. DNA repair was measured both in total genomic DNA and in specific genes from murine Scid and wildtype fibroblast cell lines. The removal of pyrimidine dimers and repair of strand breaks in genes was measured using quantitative Southern blot analyses. After ultraviolet irradiation, there was no significant difference in the repair of photoproducts in bulk DNA between Scid and wildtype cells, as measured by cellular survival and unscheduled DNA synthesis. However, deficient repair was evident in genes, where Scid cells had 25-50% less repair in the c-myc and dihydrofolate reductase genes. After gamma-irradiation, Scid fibroblasts had 20-35% less repair of DNA breaks in immunoglobulin kappa and heavy constant genes than wildtype cells. The data suggest that intact DNA-PK enzyme is needed for the efficient operation of cellular repair of pyrimidine dimers and single-strand breaks in genes, as well as in its established role in rejoining double-strand breaks.     

Characterization of intracellular DNA strand breaks induced by neocarzinostatin in Escherichia coli cells.

DNA strand breaks induced by Neocarzinostatin in Escherichia coli cells have been characterized. Radioactively labeled phage lambda DNA was introduced into lysogenic host bacteria allowing the phage DNA to circularize into superhelical molecules. After drug treatment DNA single- and double-strand breaks were measured independently after neutral sucrose gradient sedimentation. The presence of alkali-labile lesions was measured in parallel in alkaline sucrose gradients. The cell envelope provided an efficient protection towards the drug, since no strand breaks were detected unless the cells were made permeable with toluene or with hypotonic Tris buffer. In permeable cells, no double strand breaks could be detected, even at high NCS concentration (100 micrograms/ml). Induction of single-strand breaks leveled off after 15 min at 20 degrees C in the presence of 2 mM mercaptoethanol. Exposure to 0.3N NaOH doubled the number of strand breaks. No enzymatic repair of the breaks could be observed.     

Nature of DNA lesions induced in human hepatoma cells, human colonic cells and human embryonic lung fibroblasts by the antiretroviral drug 3'-azido-3'-deoxythymidine

This study tried to clarify the question if nuclear genotoxicity played a role in 3'-azido-3'-deoxythymidine (AZT) toxicity. We investigated cytotoxic and DNA-damaging effects of AZT on human hepatoma HepG2 and human colonic CaCo-2 cells as well as on human diploid lung fibroblasts HEL. The amount of induced DNA damage was measured by standard alkaline single cell gel electrophoresis (SCGE). The nature of induced DNA lesions was evaluated (1) by modified SCGE, which includes treatment of lysed cells with DNA repair enzymes Endo III and Fpg and enables to recognize oxidized bases of DNA, and (2) by SCGE processed in parallel at pH 13.0 (standard technique) and pH 12.1, which enables to recognize alkali labile DNA lesions and direct DNA strand breaks. Cytotoxicity of AZT was evaluated by the trypan blue exclusion technique. Our findings showed that 3-h treatment of cells with AZT decreased the viability of all cell lines studied. SCGE performed in the presence of DNA repair enzymes proved that AZT induced oxidative lesions to DNA in all cell types. In hepatoma HepG2 cells and embryonic lung fibroblasts HEL the majority of AZT-induced DNA strand breaks were pH-independent, i.e. they were identified at both pH values (12.1 and 13.0). These DNA lesions represented direct DNA breaks. In colonic Caco-2 cells DNA lesions were converted to DNA strand breaks particularly under strong alkaline conditions (pH>13.0), which is characteristic for alkali-labile sites of DNA. DNA strand break rejoining was investigated by the standard comet assay technique during 48 h of post-AZT-treatment in HepG2 and Caco-2 cells. The kinetics of DNA rejoining, considered an indicator of DNA repair, revealed that AZT-induced DNA breaks were repaired in both cell types slowly, though HepG2 cells seemed to be more repair proficient with respect to AZT-induced DNA lesions.     

Chromatid damage induced by fluorescent light during G2 phase in normal and Gardner syndrome fibroblasts. Interpretation in terms of deficient DNA repair

Skin fibroblasts from Gardner syndrome (GS) compared with those from normal donors showed a significantly higher incidence of chromatid gaps and breaks following exposure to low-intensity, cool-white fluorescent light during G2 phase of the cell cycle. Considerable evidence supports the concept that chromatid gaps and breaks seen directly after exposure to DNA-damaging agents represent unrepaired DNA single- and double-strand breaks respectively. The changes in incidence of chromatid aberrations with time after light exposure are consistent with the sequence of events known to follow DNA damage and repair. Initially, the incidence of light-induced chromatid gaps was equivalent in GS and normal fibroblasts. In the normal cells, the chromatid gaps disappeared by 1 h post-exposure, presumably as a result of efficient repair of DNA single-strand breaks. In contrast, the incidence of gaps increased in GS cells by 0.5 h followed by a decrease at 1 h and concomitant increase in chromatid breaks. It appears from these findings that the increased incidence of chromatid damage in GS fibroblasts results from deficient repair of DNA single-strand breaks which arise from incomplete nucleotide excision of DNA damage during G2 phase.     

Induction and repair of double- and single-strand DNA breaks in bacteriophage lambda superinfecting Escherichia coli

Induction and repair of double- and single-strand DNA breaks have been measured after decays of 125I and 3H incorporated into the DNA and after external irradiation with 4 MeV electrons. For the decay experiments, cells of wild type Escherichia coli K-12 were superinfected with bacteriophage lambda DNA labelled with 5'-(125I)iodo-2'-deoxyuridine or with (methyl-3H)thymidine and frozen in liquid nitrogen. Aliquots were thawed at intervals and lysed at neutral pH, and the phage DNA was assayed for double- and single-strand breakage by neutral sucrose gradient centrifugation. The gradients used allowed measurements of both kinds of breaks in the same gradient. Decays of 125I induced 0.39 single-strand breaks per double-strand break. No repair of either break type could be detected. Each 3H disintegration caused 0.20 single-strand breaks and very few double-strand breaks. The single-strand breaks were rapidly rejoined after the cells were thawed. For irradiation with 4 MeV electrons, cells of wild type E. coli K-12 were superinfected with phage lambda and suspended in growth medium. Irradiation induced 42 single-strand breaks per double-strand break. The rates of break induction were 6.75 x 10(-14) (double-strand breaks) and 2.82 x 10(-12) (single-strand breaks) per rad and per dalton. The single-strand breaks were rapidly repaired upon incubation whereas the double-strand breaks seemed to remain unrepaired. It is concluded that double-strand breaks in superinfecting bacteriophage lambda DNA are repaired to a very small extent, if at all.     

Structure of repaired sites in human DNA synthesized in the presence of inhibitors of DNA polymerases alpha and beta in human fibroblasts

Excision repair of ultraviolet damage in human fibroblasts was partially inhibited by drugs that block DNA polymerases alpha or beta (cytosine arabinoside, aphidicolin and dideoxythymidine) causing a reduction in unscheduled synthesis and an accumulation of single-strand breaks. The strand breaks accumulated in the presence of aphidicolin could be resealed within 30 min after removal of the drug, but those accumulated by cytosine arabinoside took many hours. Digestion of repaired DNA with exonuclease III or S1 nuclease revealed that even the highest concentration of polymerase inhibitors, singly or in combination, that produced maximal accumulation of single-strand breaks only blocked 37-86% of repair sites. Use of single-strand break frequencies to measure the number of repair events can therefore be in error by as much as a factor of 3. The blocked patches with free 3'OH termini were, on average, 22% of normal length, corresponding to between 6 and 17 bases (assuming a normal patch of 25-75 bases in length). Patches that remained unsealed in vivo were also resistant to sealing by T4 ligase in vitro. The data are more consistent with a mechanism of repair in which long single-strand gaps are first made by excision enzymes and subsequently filled in by DNA polymerase alpha. Strand displacement or nick translation mechanisms seem unlikely.     

Changes of 8-OH-dG levels in DNA and its base excision repair activity in rat lungs after inhalation exposure to hexavalent chromium

The co-genotoxic effects of cadmium are well recognized and it is assumed that most of these effects are due to the inhibition of DNA repair. We used the comet assay to analyze the effect of low, non-toxic concentrations of CdCl2 on DNA damage and repair-induced in Chinese hamster ovary (CHO) cells by UV-radiation, by methyl methanesulfonate (MMS) and by N-methyl-N-nitrosourea (MNU). The UV-induced DNA lesions revealed by the comet assay are single-strand breaks which are the intermediates formed during nucleotide excision repair (NER). In cells exposed to UV-irradiation alone the formation of DNA strand breaks was rapid, followed by a fast rejoining phase during the first 60 min after irradiation. In UV-irradiated cells pre-exposed to CdCl2, the formation of DNA strand breaks was significantly slower, indicating that cadmium inhibited DNA damage recognition and/or excision. Methyl methanesulfonate and N-methyl-N-nitrosourea directly alkylate nitrogen and oxygen atoms of DNA bases. The lesions revealed by the comet assay are mainly breaks at apurinic/apyrimidinic (AP) sites and breaks formed as intermediates during base excision repair (BER). In MMS treated cells the initial level of DNA strand breaks did not change during the first hour of recovery; thereafter repair was detected. In cells pre-exposed to CdCl2 the MMS-induced DNA strand breaks accumulated during the first 2h of recovery, indicating that AP sites and/or DNA strand breaks were formed but that further steps of BER were blocked. In MNU treated cells the maximal level of DNA strand breaks was detected immediately after the treatment and the breaks were repaired rapidly. In CdCl2 pre-treated cells the formation of MNU-induced DNA single-strand breaks was not affected, while the repair was slower, indicating inhibition of polymerization and/or the ligation step of BER. Cadmium thus affects the repair of UV-, MMS- and MNU-induced DNA damage, providing further evidence, that inhibition of DNA repair is an important mechanism of cadmium induced mutagenicity and carcinogenicity.     

Effect of age on endogenous DNA single-strand breakage, strand break induction and repair in the adult housefly, Musca domestica

It has been suggested that genomic alterations involving DNA damage and the ability to repair such damage play an important role in cellular senescence. In this study, endogenous DNA single-strand breaks, the susceptibility of DNA to induced strand breakage and the capacity to repair these breaks were compared in postmitotic cells from young (3-day-old) and old (23-day-old) houseflies. DNA single-strand breaks did not accumulate during normal aging in the housefly. However, cells of the old flies exhibited a greater sensitivity to single-strand breakage induced by gamma-radiation and UV light. The capacity to repair these exogenously induced single-strand breaks declined with age. Results do not support the view that DNA single-strand breaks are a causal factor in aging in the housefly. An age-related increase in the susceptibility to undergo single-strand breakage suggests alterations in chromatin during the aging process.