The micronucleus assay in radiation accidents

The cytokinesis-block micronucleus assay in peripheral blood lymphocytes is a standardised and validated technique for biodosimetry. Automated scoring of micronuclei allows large scale applications as in population triage in case of radiation accidents or malevolent use of radioactive sources. The dose detection limit (95% confidence) of the micronucleus assay for individual dose assessment is restricted to 0.2 Gy but can be decreased to 0.1 Gy by scoring centromeres in micronuclei using fluorescence in situ hybridization (FISH). In the past the micronucleus assay was applied for a number of large scale biomonitoring studies of nuclear power plant workers and hospital workers. Baseline micronucleus frequencies depend strongly on age and gender. The assay was also already used for biodosimetry of radiation accidents. In a multiple endpoint biodosimetry study for dose assessment of a worker exposed accidentally in 2003 to X-rays, a good agreement was obtained between dose estimates resulting from the micronucleus assay, the scoring of dicentrics and translocations. Automated scoring of micronuclei in combination with centromere signals, allowing systematic biodosimetry of exposed populations, remains a challenge for the future

The micronucleus assay as a biological dosimeter of in vivo ionising radiation exposure

Biological dosimetry, based on the analysis of micronuclei (MN) in the cytokinesis-block micronucleus (CBMN) assay can be used as an alternative method for scoring dicentric chromosomes in the field of radiation protection. Biological dosimetry or Biodosimetry, is mainly performed, in addition to physical dosimetry, with the aim of individual dose assessment. Many studies have shown that the number of radiation-induced MN is strongly correlated with dose and quality of radiation. The CBMN assay has become, in the last years, a thoroughly validated and standardised technique to evaluate in vivo radiation exposure of occupational, medical and accidentally exposed individuals. Compared to the gold standard, the dicentric assay, the CBMN assay has the important advantage of allowing economical, easy and quick analysis. The main disadvantage of the CBMN assay is related to the variable micronucleus ( MN) background frequency, by which only in vivo exposures in excess of 0.2-0.3 Gy X-rays can be detected. In the last years, several improvements have been achieved, with the ultimate goals (i) of further increasing the sensitivity of the CBMN assay for low-dose detection by combining the assay with a fluorescence in situ hybridisation centromere staining technique, (ii) of increasing the specificity of the test for radiation by scoring nucleoplasmic bridges in binucleated cells and (iii) of making the assay optimally suitable for rapid automated analysis of a large number of samples, viz. in case of a large-scale radiation accident. The development of a combined automated MN-centromere scoring procedure remains a challenge for the future, as it will allow systematic biomonitoring of radiation workers exposed to low-dose radiation

Low doses of ionizing radiation induce immune-stimulatory responses in isolated human primary monocytes

The health effects arising from exposure to low doses of ionizing radiation are of particular concern, mainly due to the increased application of diagnostic and therapeutic X-ray modalities. The mechanisms behind the cell and tissue responses to low doses remain to be elucidated. Accumulating evidence suggests that low doses of ionizing radiation induce activation of the immune response; however, the processes involved have yet to be adequately investigated. Monocytes are key players in the induction of an immune response. Within the context of this study, we investigated the activation of toll-like receptors (TLRs), mitogen-activated protein kinases (MAPKs) and NF-B signaling in isolated human primary monocytes in response to low doses (0.05 and 0.1 Gy) and a high dose (1 Gy) of ionizing radiation. Using quantitative RT-PCR and ELISA techniques, our results showed a positive regulation of TLR signaling in response to low doses but a less significant response at high doses. This activation was demonstrated via the activation of TLR signaling molecules (HMGB1, TLR4, TLR9, MyD88 and IRAK1). Furthermore, and in contrast to the high dose, the low doses showed increased phosphorylation levels of the protein IB, and therefore positive signaling of the NF-B pathway. This result denotes pro-survival and pro-inflammatory responses. Additionally, MAPKs were activated in response to 0.05 Gy, while 0.1 and 1 Gy showed a downregulatory trend that may be related to activation of the PF4 gene. On the other hand, there was highly significant involvement of activated p53 and damaged genes in response to high but not low doses. In conclusion, this study addressed the need to re-evaluate health risks arising from exposure to low doses of ionizing radiation, particularly in view of the accumulating evidence reporting inflammatory and oncogenic consequences from these exposures

A semi-automated FISH-based micronucleus-centromere assay for biomonitoring of hospital workers exposed to low doses of ionizing radiation

The aim of the present study was to perform cytogenetic analysis by means of a semi-automated micronucleus-centromere assay in lymphocytes from medical radiation workers. Two groups of workers receiving the highest occupational doses were selected: 10 nuclear medicine technicians and 10 interventional radiologists/cardiologists. Centromere-negative micronucleus (MNCM-) data, obtained from these two groups of medical radiation workers were compared with those obtained in matched controls. The blood samples of the matched controls were additionally used to construct a 'low-dose' (0-100 mGy) MNCM-dose-response curve to evaluate the sensitivity and suitability of the micronucleus-centromere assay as an 'effect' biomarker in medical surveillance programs. The physical dosimetry data of the 3 years preceding the blood sampling, based on single or double dosimetry practices, were collected for the interpretation of the micronucleus data. The in vitro radiation results showed that for small sized groups, semi-automated scoring of MNCM-enables the detection of a dose of 50 mGy. The comparison of MNCM-yields in medical radiation workers and control individuals showed enhanced MNCM-scores in the medical radiation workers group (P=0.15). The highest MNCM-scores were obtained in the interventional radiologists/cardiologists group, and these scores were significantly higher compared with those obtained from the matched control group (P=0.05). The higher MNCM-scores observed in interventional radiologists/cardiologists compared with nuclear medicine technicians were not in agreement with the personal dosimetry records in both groups, which may point to the limitation of 'double dosimetry' procedures used in interventional radiology/cardiology. In conclusion, the data obtained in the present study supports the importance of cytogenetic analysis, in addition to physical dosimetry, as a routine biomonitoring method in medical radiation workers receiving the highest occupational radiation burdens

Novel X-ray imaging technology enables significant patient dose reduction in interventional cardiology while maintaining diagnostic image quality

Objectives: The purpose of this study was to quantify the reduction in patient radiation dose during coronary angiography (CA) by a new X-ray technology, and to assess its impact on diagnostic image quality. Background: Recently, a novel X-ray imaging technology has become available for interventional cardiology, using advanced image processing and an optimized acquisition chain for radiation dose reduction. Methods: 70 adult patients were randomly assigned to a reference X-ray system or the novel X-ray system. Patient demographics were registered and exposure parameters were recorded for each radiation event. Clinical image quality was assessed for both patient groups. Results: With the same angiographic technique and a comparable patient population, the new imaging technology was associated with a 75% reduction in total kerma-area product (KAP) value (decrease from 47 Gycm(2) to 12 Gycm(2), P<0.001). Clinical image quality showed an equivalent detail and contrast for both imaging systems. On the other hand, the subjective appreciation of noise was more apparent in images of the new image processing system, acquired at lower doses, compared to the reference system. However, the higher noise content did not affect the overall image quality score, which was adequate for diagnosis in both systems. Conclusions: For the first time, we present a new X-ray imaging technology, combining advanced noise reduction algorithms and an optimized acquisition chain, which reduces patient radiation dose in CA drastically (75%), while maintaining diagnostic image quality. Use of this technology may further improve the radiation safety of cardiac angiography and interventions

Early biomarkers related to secondary primary cancer risk in radiotherapy treated prostate cancer patients: IMRT versus IMAT

AbstractPurposeTo investigate whether rotational techniques (Volumetric Modulated Arc Therapy – VMAT) are associated with a higher risk for secondary primary malignancies compared to step-and-shoot Intensity Modulated Radiation Therapy (ss-IMRT). To this end, radiation therapy (RT) induced DNA double-strand-breaks and the resulting chromosomal damage were assessed in peripheral blood T-lymphocytes of prostate cancer (PCa) patients applying γH2AX foci and G0 micronucleus (MN) assays.Methods and materialsThe study comprised 33PCa patients. A blood sample was taken before start of therapy and after the 1st and 3rd RT fraction to determine respectively the RT-induced γH2AX foci and MN. The equivalent total body dose (DETB) was calculated based on treatment planning data.ResultsA linear dose response was obtained for γH2AX foci yields versus DETB while MN showed a linear-quadratic dose response. Patients treated with large volume (LV) VMAT show a significantly higher level of induced γH2AX foci and MN compared to IMRT and small volume (SV) VMAT (p<0.01). Assuming a linear-quadratic relationship, a satisfactory correlation was found between both endpoints (R2 0.86).ConclusionsBiomarker responses were governed by dose and irradiated volume of normal tissues. No significant differences between IMRT and rotational therapy inherent to the technique itself were observed

The radiosensitizing effect of Ku70/80 knockdown in MCF10A cells irradiated with X-rays and p(66)+Be(40) neutrons

Background: A better understanding of the underlying mechanisms of DNA repair after low-and high-LET radiations represents a research priority aimed at improving the outcome of clinical radiotherapy. To date however, our knowledge regarding the importance of DNA DSB repair proteins and mechanisms in the response of human cells to high-LET radiation, is far from being complete. Methods: We investigated the radiosensitizing effect after interfering with the DNA repair capacity in a human mammary epithelial cell line (MCF10A) by lentiviral-mediated RNA interference (RNAi) of the Ku70 protein, a key-element of the non-homologous end-joining (NHEJ) pathway. Following irradiation of control and Ku-deficient cell lines with either 6 MV X-rays or p(66)+Be(40) neutrons, cellular radiosensitivity testing was performed using a crystal violet cell proliferation assay. Chromosomal radiosensitivity was evaluated using the micronucleus (MN) assay. Results: RNAi of Ku70 caused downregulation of both the Ku70 and the Ku80 proteins. This downregulation sensitized cells to both X-rays and neutrons. Comparable dose modifying factors (DMFs) for X-rays and neutrons of 1.62 and 1.52 respectively were obtained with the cell proliferation assay, which points to the similar involvement of the Ku heterodimer in the cellular response to both types of radiation beams. After using the MN assay to evaluate chromosomal radiosensitivity, the obtained DMFs for X-ray doses of 2 and 4 Gy were 2.95 and 2.66 respectively. After neutron irradiation, the DMFs for doses of 1 and 2 Gy were 3.36 and 2.82 respectively. The fact that DMFs are in the same range for X-rays and neutrons confirms a similar importance of the NHEJ pathway and the Ku heterodimer for repairing DNA damage induced by both X-rays and p(66)+Be(40) neutrons. Conclusions: Interfering with the NHEJ pathway enhanced the radiosensitivity of human MCF10A cells to low-LET Xrays and high-LET neutrons, pointing to the importance of the Ku heterodimer for repairing damage induced by both types of radiation. Further research using other high-LET radiation sources is however needed to unravel the involvement of DNA double strand break repair pathways and proteins in the cellular response of human cells to high-LET radiation

Factors modifying the risk for developing acute skin toxicity after whole-breast intensity modulated radiotherapy

Background: After breast-conserving radiation therapy most patients experience acute skin toxicity to some degree. This may impair patients' quality of life, cause pain and discomfort. In this study, we investigated treatment and patient-related factors, including genetic polymorphisms, that can modify the risk for severe radiation-induced skin toxicity in breast cancer patients. Methods: We studied 377 patients treated at Ghent University Hospital and at ST.-Elisabeth Clinic and Maternity in Namur, with adjuvant intensity modulated radiotherapy (IMRT) after breast-conserving surgery for breast cancer. Women were treated in a prone or supine position with normofractionated (25 x 2 Gy) or hypofractionated (15 x 2.67 Gy) IMRT alone or in combination with other adjuvant therapies. Patient-and treatment-related factors and genetic markers in regulatory regions of radioresponsive genes and in LIG3, MLH1 and XRCC3 genes were considered as variables. Acute dermatitis was scored using the CTCAEv3.0 scoring system. Desquamation was scored separately on a 3-point scale (0-none, 1-dry, 2-moist). Results: Two-hundred and twenty patients (58%) developed G2+ dermatitis whereas moist desquamation occurred in 56 patients (15%). Normofractionation (both p = D (p = 0.001 and p = 0.043) and concurrent hormone therapy (p = 0.001 and p = 0.037) were significantly associated with occurrence of acute dermatitis and moist desquamation, respectively. Additional factors associated with an increased risk of acute dermatitis were the genetic variation in MLH1 rs1800734 (p=0.008), smoking during RT (p = 0.010) and supine IMRT (p = 0.004). Patients receiving trastuzumab showed decreased risk of acute dermatitis (p < 0.001). Conclusions: The normofractionation schedule, supine IMRT, concomitant hormone treatment and patient related factors (high BMI, large breast, smoking during treatment and the genetic variation in MLH1 rs1800734) were associated with increased acute skin toxicity in patients receiving radiation therapy after breast-conserving surgery. Trastuzumab seemed to be protective

Intensity modulated radiotherapy induces pro-inflammatory and pro-survival responses in prostate cancer patients

Intensity modulated radiotherapy (IMRT) is one of the modern conformal radiotherapies that is widely used within the context of cancer patient treatment. It uses multiple radiation beams targeted to the tumor, however, large volumes of the body receive low doses of irradiation. Using γ-H2AX and global genome expression analysis, we studied the biological responses induced by low doses of ionizing radiation in prostate cancer patients following IMRT. By means of different bioinformatics analyses, we report that IMRT induced an inflammatory response via the induction of viral, adaptive, and innate immune signaling. In response to growth factors and immune-stimulatory signaling, positive regulation in the progression of cell cycle and DNA replication were induced. This denotes pro-inflammatory and pro-survival responses. Furthermore, double strand DNA breaks were induced in every patient 30 min after the treatment and remaining DNA repair and damage signaling continued after 18-24 h. Nine genes belonging to inflammatory responses (TLR3, SH2D1A and IL18), cell cycle progression (ORC4, SMC2 and CCDC99) and DNA damage and repair (RAD17, SMC6 and MRE11A) were confirmed by quantitative RT-PCR. This study emphasizes that the risk assessment of health effects from the out-of-field low doses during IMRT should be of concern, as these may increase the risk of secondary cancers and/or systemic inflammation