The automated FADU-assay, a potential high-throughput in vitro method for early screening of DNA breakage

Moreno‐Villanueva, María

doi:10.14573/altex.2011.4.295

Cited by 42 publications

(44 citation statements)

References 38 publications

(40 reference statements)

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“…We have previously published a modified and automated version of the FADU assay [1],[11],[12]. Compared with the original FADU assay [2], the automated FADU assay shows comparable sensitivity, yet increased robustness and throughput, and decreased operator time.…”

Section: Discussionmentioning

confidence: 99%

Mathematical modelling of the automated FADU assay for the quantification of DNA strand breaks and their repair in human peripheral mononuclear blood cells

et al. 2014

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BackgroundCells continuously undergo DNA damage from exogenous agents like irradiation or genotoxic chemicals or from endogenous radicals produced by normal cellular metabolic activities. DNA strand breaks are one of the most common genotoxic lesions and they can also arise as intermediates of DNA repair activity. Unrepaired DNA damage can lead to genomic instability, which can massively compromise the health status of organisms. Therefore it is important to measure and quantify DNA damage and its repair.ResultsWe have previously published an automated method for measuring DNA strand breaks based on fluorimetric detection of alkaline DNA unwinding [1], and here we present a mathematical model of the FADU assay, which enables to an analytic expression for the relation between measured fluorescence and the number of strand breaks.ConclusionsAssessment of the formation and also the repair of DNA strand breaks is a crucial functional parameter to investigate genotoxicity in living cells. A reliable and convenient method to quantify DNA strand breakage is therefore of significant importance for a wide variety of scientific fields, e.g. toxicology, pharmacology, epidemiology and medical sciences.

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Section: Discussionmentioning

confidence: 99%

Mathematical modelling of the automated FADU assay for the quantification of DNA strand breaks and their repair in human peripheral mononuclear blood cells

et al. 2014

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“…The discrepancy between previous and the current study may be due to different assays for analysis of endogenous SSB levels. We employed the FADU assay due to reported lower coefficient of variation compared with the alkaline comet assay used in previous studies (Forchhammer et al , 2010; Moreno-Villanueva et al , 2011). Compared with the FADU assay, the alkaline comet assay requires more manual handling, which is potentially more stressful to frail cells from old study subjects.…”

Section: Discussionmentioning

confidence: 99%

“…Consequently, there may be a greater risk of overestimating the endogenous level of SSBs when using the alkaline comet assay on cells from old individuals. On the other hand, the FADU assay has proven to have high reproducibility on PBMCs (Moreno-Villanueva et al , 2009, 2011). A statistically significant age-related decline was observed in DSBR and γ-H2AX response in the age span investigated (Figs 2B and 3B; Table 2).…”

Section: Discussionmentioning

confidence: 99%

Age and gender effects on DNA strand break repair in peripheral blood mononuclear cells

et al. 2012

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Summary Exogenous and endogenous damage to DNA is constantly challenging the stability of our genome. This DNA damage increase the frequency of errors in DNA replication, thus causing point mutations or chromosomal rearrangements and has been implicated in aging, cancer, and neurodegenerative diseases. Therefore, efficient DNA repair is vital for the maintenance of genome stability. The general notion has been that DNA repair capacity decreases with age although there are conflicting results. Here, we focused on potential age-associated changes in DNA damage response and the capacities of repairing DNA single-strand breaks (SSBs) and double-strand breaks (DSBs) in human peripheral blood mononuclear cells (PBMCs). Of these lesions, DSBs are the least frequent but the most dangerous for cells. We have measured the level of endogenous SSBs, SSB repair capacity, γ-H2AX response, and DSB repair capacity in a study population consisting of 216 individuals from a population-based sample of twins aged 40– 77 years. Age in this range did not seem to have any effect on the SSB parameters. However, γ-H2AX response and DSB repair capacity decreased with increasing age, although the associations did not reach statistical significance after adjustment for batch effect across multiple experiments. No gender differences were observed for any of the parameters analyzed. Our findings suggest that in PBMCs, the repair of SSBs is maintained until old age, whereas the response to and the repair of DSBs decrease.

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“…An automated version of the Fluorimetric Detection of Alkaline DNA Unwinding (FADU) method [39,40] was used for analysing formation and repair of DNA strand breaks in living cells [41]. This method is characterized by high reproducibility and high throughput.…”

Section: Methods Of Studymentioning

confidence: 99%

Effects of Psychotherapy on DNA Strand Break Accumulation Originating from Traumatic Stress

Morath¹,

Moreno‐Villanueva

Hamuni³

et al. 2014

Psychother Psychosom

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Background: Previous research reveals an association between traumatic stress and an increased risk for numerous diseases, including cancer. At the molecular level, stress may increase carcinogenesis via increased DNA damage and impaired DNA repair mechanisms. We assessed DNA breakage in peripheral blood mononuclear cells from individuals with post-traumatic stress disorder (PTSD) and measured the cellular capacity to repair single-strand breaks after exposure to ionizing X-radiation. We also investigated the effect of psychotherapy on both DNA breakage and DNA repair. Methods: In a first study we investigated DNA breakage and repair in 34 individuals with PTSD and 31 controls. Controls were subdivided into 11 trauma-exposed subjects and 20 individuals without trauma exposure. In a second study, we analysed the effect of psychotherapy (Narrative Exposure Therapy) on DNA breakage and repair. Thirty-eight individuals with PTSD were randomly assigned to either a treatment or a waitlist control condition. Follow-up was performed 4 months and 1 year after therapy. Results: In study 1 we found higher levels of basal DNA breakage in individuals with PTSD and trauma-exposed subjects than in controls, indicating that traumatic stress is associated with DNA breakage. However, single-strand break repair was unimpaired in individuals with PTSD. In study 2, we found that psychotherapy reversed not only PTSD symptoms, but also DNA strand break accumulation. Conclusion: Our results show - for the first time in vivo - an association between traumatic stress and DNA breakage; they also demonstrate changes at the molecular level, i.e., the integrity of DNA, after psychotherapeutic interventions.

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The automated FADU-assay, a potential high-throughput in vitro method for early screening of DNA breakage

Cited by 42 publications

References 38 publications

Mathematical modelling of the automated FADU assay for the quantification of DNA strand breaks and their repair in human peripheral mononuclear blood cells

Mathematical modelling of the automated FADU assay for the quantification of DNA strand breaks and their repair in human peripheral mononuclear blood cells

Age and gender effects on DNA strand break repair in peripheral blood mononuclear cells

Effects of Psychotherapy on DNA Strand Break Accumulation Originating from Traumatic Stress

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