Fanconi's anemia lymphocytes: effect of caffeine, adenosine and niacinamide during G2 prophase

doi:10.1016/0027-5107(88)90241-2

Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis

Volume 199, Issue 1, May 1988, Pages 159-165

https://doi.org/10.1016/0027-5107(88)90241-2 Get rights and content

Abstract

In this investigation peripheral blood lymphocytes from 3 Fanconi's anemia (FA) patients, 2 FA heterozygotes and 4 normal subjects were treated with caffeine and/or adenosine, and/or niacinamide during G₂ prophase. Caffeine dramatically increased breakage levels in homozygote and heterozygote cells. Niacinamide and adenosine decreased the amount of chromosomal aberrations detected in FA homozygote and heterozygote lymphocytes treated and untreated with caffeine during G₂ prophase. Caffeine sensitivity of heterozygote lymphocytes is proposed as a new clinical test to explore heterozygosis in individuals of FA families.

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In the multinucleate cells induced in Allium cepa L. meristems, the nuclei surrounded by the largest cytoplasm environment complete replication earlier (advanced nuclei), but have a longer G2, than the others (delayed nuclei). Thus, all nuclei break down the nuclear envelope and start metaphase simultaneously. The present report shows that this synchronization relies on a checkpoint mechanism. When completion of replication was prevented in the delayed nuclei (due to in vivo 5-aminouracil feeding initiated when the advanced nuclei were already in G2), the metaphase was also further delayed in the advanced ones. In turn, some of the delayed nuclei overrode the G2 checkpoint (adaptation) and entered into mitosis with broken chromatids (Del Campo et al., 1997). Anoxic UVA (313 nm) irradiation apparently prevents the binding of regulatory proteins to Br-DNA. The present report shows that late replicating sequences are the targets of the checkpoint signal produced by the still replicating nuclei. This signal delays metaphase in the advanced nuclei, whose DNA is already fully replicated. Thus, when the already replicated sequences of late replicating DNA was modified in the advanced nuclei by bromosubstitution followed by anoxic UVA irradiation, they entered into mitosis without any delay, ignoring the inhibitory signals produced by the still replicating nuclei.
Fanconi anemia lymphocytes: Effect of DL-α-tocopherol (Vitamin E) on chromatid breaks and on G<inf>2</inf> repair efficiency
2001, Mutation Research - DNA Repair
The high frequency of chromosomal breaks in Fanconi anemia (FA) lymphocytes has been related to the increased oxidative damage shown by these cells.
The effect of 100 μM dl-α-tocopherol (Vitamin E) on the level of chromosomal damage in mitosis was studied in lymphocytes from five FA patients and from age matched controls, both under basal conditions and when G₂ repair was prevented by 2.5 mM caffeine (G₂ unrepaired damage). In addition, the effect of this antioxidant on G₂ duration and the efficiency of G₂ repair was also evaluated in the sample.
α-Tocopherol (AT) decreased the frequency of chromosomal damage (under basal and inhibited G₂ repair conditions) and the duration of G₂ in FA cells. This antioxidant protective effect, expressed as the decrease in chromatid breaks, was greater in FA cells (50.8%) than in controls (25%).
The efficiency of the G₂ repair process (G₂R rate) defined as the ratio between the percentage of chromatid breaks repaired in G₂ and the duration of this cell cycle phase was lesser in FA cells (10.6) than in controls (22.6). AT treatment slightly increased this G₂R rate, both in FA cells and controls.
These results suggest that an increased oxidative damage and a lower G₂ repair rate may be simultaneously involved in the high frequency of chromatid damage detected in FA cells.
Damage-resistant DNA synthesis in fanconi anemia cells treated with a DNA cross-linking agent
2000, Experimental Cell Research
Fanconi anemia (FA) is a recessive disorder associated with diverse congenital anomalies, progressive bone marrow failure, and a marked predisposition to develop cancer. At the cellular level, FA is characterized by a prolonged G₂ phase in proliferating cells and a marked hypersensitivity to both the cytotoxic and the clastogenic effects of agents which produce DNA interstrand cross-links. Treatment with these agents leads to even further prolongation of the G₂ phase in FA cells. We now show that FA cells, from four different complementation groups, fail to decrease their rates of replicative DNA synthesis, as do normal cells, following treatment with a DNA cross-linking agent. This may be responsible for the prolongation of the G2 phase seen in these cells, and suggests that the fundamental defect in response of FA cells to DNA cross-linking agents may be in the S phase, rather than the G₂ phase, of the cell cycle.
Arrest of S-phase progression is impaired in Fanconi anemia cells
2000, Experimental Cell Research
Fanconi anemia (FA) is an inherited cancer-susceptibility disorder, characterized by genomic instability, hypersensitivity to DNA cross-linking agents, and a prolonged G2 phase of the cell cycle. We observed a marked dose-dependent accumulation of FA cells in the G2 compartment after treatment with 4,5′,8-trimethylpsoralen (Me₃Pso) in combination with 365 nm irradiation. Using bivariate DNA distribution methodology, we determined the proportion of replicating and arresting S-phase cells and observed that, whereas normal cells arrested DNA replication in the presence of Me₃Pso cross-links and monoadducts, FA lymphoblasts failed to arrest DNA synthesis. Taken together, the above data suggest that, in response to damage induced by DNA cross-linking agents, the S-phase checkpoint is inefficient in FA cells. This would lead to accumulation of secondary lesions, such as single- and double-strand breaks and gaps. The prolonged time in G2 phase seen in FA cells therefore exists in order to allow the cells to remove lesions which accumulated during the preceding abnormal S phase.
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It has been shown that frameshift mutagenesis by 9-aminoacridine (9AA) in Salmonella typhimurium is significantly inhibited if glucose is present while cells are being treated in liquid defined medium. We suggested that this effect might be a result of glucose-provoked alterations of cAMP levels within the cell. We therefore sought to investigate the effects of exogenous cAMP on mutagenesis by 9-aminoacridine in both Salmonella typhimurium and Escherichia coli. Contrary to expectation, we found that frameshift mutagenesis was significantly depressed when high concentrations of cAMP were added to the defined medium during liquid treatment with 9AA. Other adenosine 5′-phosphates such as adenosine 5′-triphosphate (ATP), adenosine 5′-diphosphate (ADP) and adenosine 5′-monophosphate (AMP) added to the liquid medium during 9AA treatment also substantially decreased the reversion rate to prototrophy in both S. typhimurium and E. coli, as did adenosine itself. Further experiments showed that neither influx nor efflux of 9-aminoacridine molecules were greatly affected by adenosine compounds, and that although cAMP and adenosine exerted similar antimutagenic effects on 9AA-treated stationary phase cells, their effects on log phase cells were quite different. The antimutagenic effect of a representative adenosine compound (ATP) was found to persist for some time after stationary phase cells had been washed, with maximal mutability being regained only after about 3 h.
G<inf>2</inf> repair and aging: influence of donor age on chromosomal aberrations in human lymphocytes
1993, Mutation Research DNAging
The caffeine effects on chromosomal aberration frequency and mean G₂ duration were studied in human lymphocytes in vitro from three age groups of normal donors (I: 1–5 years old; II: 30–40 years old; III: 60–70 years old). Under control conditions, the three age group showed a similar frequency of chromosomal aberrations. All three age groups exhibited a linear dose response for aberrations with caffeine treatments. However, lymphocytes from aged individuals (groups II and III) showed higher chromosomal aberration frequencies and longer G₂ duration than cells from young individuals (group I). The caffeine effect in reducing G₂ length was rather similar in every age group. The reversion of caffeine effects by adenosine or niacinamide in lymphocytes from older individuals was higher than in cells from group I. The different caffeine effects and G₂ values between lymphocytes from old and young individuals are most likely due to a higher number of DNA lesions reaching G₂ phase and/or a decrease of the G₂ repair capability of lymphocytes from older individuals.

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Fanconi's anemia lymphocytes: effect of caffeine, adenosine and niacinamide during G2 prophase

Abstract

Mutation Res.

Mutation Res.

J. Mol. Biol.

Mutation Res.

Mutation Res.

Mutation Res.

Blood

Biophys. J.

Susceptibility of Fanconi's anemia fibroblast to chromosome damage by carcinogens

Nature (London)

Prenatal and post-natal diagnosis and carrier detection of Fanconi's anemia by a cytogenetic method

Pediatrics

DNA of HeLa cells during caffeine-promoted recovery from X-ray induced G2 arrest

J. Radiat. Biol.

Abnormal NAD+ levels in cells from patients with Fanconi's anemia

Nature (London)

Familial constitutional panmyelocytopathy, Fanconi's anemia (FA). I. Clinical aspects

Genes which increase chromosomal instability in somatic cells and predispose to cancer

Prog. Med. Genet.

Fanconi's anemia lymphocytes: effect of caffeine, adenosine and niacinamide during G₂ prophase

DNA of HeLa cells during caffeine-promoted recovery from X-ray induced G₂ arrest

Abnormal NAD⁺ levels in cells from patients with Fanconi's anemia