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Micronucleus test as an indicator of the adaptive response in human lymphocytes from 5 blood donors. The non-adapted cultures (2 Gy) were compared with the adapted ones (5 cGy + 2 Gy). Points are means ± SD of at least three independent experiments with 2−3 slides in each and 500 binucleated cells scored per slide. Asterisks mark statistically significant differences between pairs: * p < 0.05, ** p < 0.01, with t-test.
Source publication
Adaptive response (AR), an enhanced resistance to a high dose of ionising radiation acquired after pretreatment with a very low dose, was estimated in normal human lymphocytes. The question posed was whether the extent of radioadaptation, assessed by micronucleus test, would be related to the rate of DNA double-strand break (DSB) rejoining. Phytohe...
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Citations
... We use a label 'T' to specifically denote where the data is from tumor cell lines. All of the literature datasets are assumed or reported to be asynchronous, except Lliakis et al., which is synchronised in G1 cell cycle phase [51][52][53][54][55][56][57]. In all panels, lines with error bars representing SEM are simulated results from 70 repeats. ...
... We use a label 'T' to specifically denote where the data is from tumor cell lines. All of the literature datasets are assumed or reported to be asynchronous, except Lliakis et al., which is synchronised in G1 cell cycle phase[51][52][53][54][55][56][57]. In all panels, lines with error bars representing SEM are simulated results from 70 repeats. ...
Simple Summary
When cells are irradiated, their DNA can become damaged, this causes different types of repair processes to try and fix the DNA breaks. One of the most lethal types of DNA damage is double-strand breaks (DSBs). This work models the most used DSB repair process called Non-Homologous End Joining (NHEJ) and includes both its resection-independent and resection-dependent pathways. The models produced are benchmarked against experimental normal and deficient cell-types across a wide range of radiation qualities. We compare two approaches of modelling, the first is where the DSBs can repair in parallel and the second is where the DSBs repair is entwined. We find that it is necessary to consider both the resection-independent and resection-dependent pathways as entwined to produce a model which robustly matches experimental work. Through better modelling of NHEJ repair, it can improve our understanding of radiation response which has potential in biological optimisation for radiotherapy.
Abstract
Mechanistic in silico models can provide insight into biological mechanisms and highlight uncertainties for experimental investigation. Radiation-induced double-strand breaks (DSBs) are known to be toxic lesions if not repaired correctly. Non-homologous end joining (NHEJ) is the major DSB-repair pathway available throughout the cell cycle and, recently, has been hypothesised to consist of a fast and slow component in G0/G1. The slow component has been shown to be resection-dependent, requiring the nuclease Artemis to function. However, the pathway is not yet fully understood. This study compares two hypothesised models, simulating the action of individual repair proteins on DSB ends in a step-by-step manner, enabling the modelling of both wild-type and protein-deficient cell systems. Performance is benchmarked against experimental data from 21 cell lines and 18 radiation qualities. A model where resection-dependent and independent pathways are entirely separated can only reproduce experimental repair kinetics with additional restraints on end motion and protein recruitment. However, a model where the pathways are entwined was found to effectively fit without needing additional mechanisms. It has been shown that DaMaRiS is a useful tool when analysing the connections between resection-dependent and independent NHEJ repair pathways and robustly matches with experimental results from several sources.
Purpose:
Interindividual variability in normal tissue toxicity during radiation therapy is a limiting factor for successful treatment. Predicting the risk of developing acute reactions before initiation of radiation therapy may have the benefit of opting for altered radiation therapy regimens to achieve minimal adverse effects with improved tumor cure.
Methods and materials:
DNA double-strand break (DSB) induction and its repair kinetics in lymphocytes of head-and-neck cancer patients undergoing chemoradiation therapy was analyzed by counting γ-H2AX foci, neutral comet assay, and a modified version of neutral filter elution assay. Acute normal tissue reactions were assessed by Radiation Therapy Oncology Group criteria.
Results:
The correlation between residual DSBs and the severity of acute reactions demonstrated that residual γ-H2AX foci in head-and-neck cancer patients increased with the severity of oral mucositis and skin reaction.
Conclusions:
Our results suggest that γ-H2AX analysis may have predictive implications for identifying the overreactors to mucositis and skin reactions among head-and-neck cancer patients prior to initiation of radiation therapy.
Neutral filter elution assay is one of the methods used for detection of DNA double strand breaks (DSBs). However, it is laborious, expensive, and hazardous (radiolabeled precursors for DSB detection and scintillation counter for quantification), making it a less preferred method for DSB detection. In the present study, an attempt was made to improve the existing neutral filter elution assay by making use of fluorescent dye (PicoGreen) and microfiltration assembly for eluting the fragmented DNA, thereby reducing the cost and time required for the assay. We studied the effect of dye dilution, pH conditions, and cell number as a part of method standardization. X-ray dose-response and repair kinetics in lymphocytes as well as cell lines were studied for validating the sensitivity of the assay. A linear dose-response relationship for DSBs was observed at a cell number of 4×10(5)cells, a dye dilution of 500-fold, and at pH 10. Repair kinetics revealed a time-dependent repair of DSBs up to 360 min of posttreatment, indicating its usefulness in DSB repair studies. In conclusion, the present modified method is more efficient (in terms of cell number), cost effective, less time-consuming, and less hazardous compared to the existing method.
