Cytogenetic Dosimetry Research Articles

Extending culture time to improve Mitotic Index for cytogenetic dosimetry

Purpose To analyze the effects of extending lymphocyte cultivation time on the Mitotic Index, frequency of first-division cells, and dose estimation after irradiating blood samples with different doses of radiation. Materials and methods Blood samples from two healthy male volunteers were separately irradiated with three doses (3, 5, and 6 Gy) using a 60Co gamma source (average dose rate: 1.48 kGy.h−1) and cultivated in vitro for conventional (48 h) and extended (56, 68, and 72 h) amounts of time. Colcemid (0.01 µg.mL−1) was added at the beginning of the culture period. Cells were fixed, stained with fluorescence plus Giemsa (FPG), and analyzed under a light microscope. The effects of prolonged culture duration on the Mitotic Index (MI), frequency of first-division cells (M1 cells), and the First-Division Mitotic Index (FDMI) were investigated. The estimation of delivered doses was conducted using a conventional 48h-culture calibration curve. Results Overall, cells presented higher MI (up to 12-fold) with the extension of culture, while higher radiation doses led to lower MI values (up to 80% reduction at 48 h). Cells irradiated with higher doses (5 and 6 Gy) had the most significant increase (5- to 12-fold) of MI as the cultivation was prolonged. The frequency of M1 cells decreased with the prolongation of culture for all doses (up to 75% reduction), while irradiated cells presented higher frequencies of M1 cells than non-irradiated ones. FDMI increased for all irradiated cultures but most markedly in those irradiated with higher doses (up to 10-fold). The conventional 48h-culture calibration curve proved adequate for assessing the delivered dose based on dicentric frequency following a 72-hour culture. Conclusion Compared to the conventional 48-hour protocol, extending the culture length to 72 hours significantly increased the Mitotic Index and the number of first-division metaphases of irradiated lymphocytes, providing slides with a better scorable metaphase density. Extending the culture time to 72 hours, combined with FPG staining to score exclusively first-division metaphases, improved the counting of dicentric chromosomes. The methodology presented and discussed in this study can be a powerful tool for dicentric-based biodosimetry, especially when exposure to high radiation doses is involved.

Effect of changing the radiation dose range on the in vitro cytogenetic dose response to gamma-rays

Purpose To examine the distortion of the linear quadratic (LQ) model of in vitro cytogenetic dose response over an extended range of γ-ray doses by analyzing the available literature data, and to establish the dose ranges, in which the LQ dose response curve (DRC) can be most accurately fitted for biological dosimetry. Materials and methods Data on yields of dicentrics (Dic) or dicentrics plus centric rings (Dic + CR) induced in vitro in human lymphocytes by acute γ-rays were extracted from 108 open sources. The overall dose response dataset in the dose range up to 50 Gy was fitted to a fractional-rational (FR) model, which included a ‘basic’ LQ function in the numerator, and a reduction factor dependent on the square of the dose in the denominator. Cytogenetic dose response data obtained at Grigoriev Institute for Medical Radiology, Kharkiv, Ukraine (GIMRO) in the range 0.1 − 20.3 Gy acute γ-rays were fitted to the LQ model with the progressive changing minimum or maximum radiation dose. Results The overall dose response, as expected, followed the LQ function in the dose range ≤5 Gy, but in the extended dose range appeared to be S-shaped, with intensive saturation and a plateau at doses ≥22 Gy. Coefficients of the ‘basic’ LQ equation in FR model were very close to many published DRCs; calculated asymptote was 17. Fitting of the GIMRO dataset to the LQ model with the shift of the dose range showed the increase in linear coefficient with the increment of either minimum or maximum radiation dose, while the decline of the quadratic coefficient was regulated mostly by the increase of the highest dose. The best goodness of fit, assessed by lower χ2 values, occurred for dose ranges 0.1 − 1.0 Gy; 0.5 − 5.9 Gy; 1.0 − 7.8 Gy; 2.0 − 9.6 Gy, 3.9 − 16.4 Gy and 5.9 − 20.3 Gy. The ‘see-saw’ effect in changes of LQ coefficients was confirmed by re-fitting datasets published by other laboratories. Conclusions The classical LQ model with fixed coefficients appears to have limited applicability for cytogenetic dosimetry at radiation doses >5 Gy due to the saturation of the dose response. Different response of the LQ coefficients to the changes of the dose range must be considered during the DRC construction. Proper selection of minimum and maximum dose in calibration experiments makes it possible to improve the goodness of fit of the LQ DRC.

Persons chronically exposed to low doses of ionizing radiation: A cytogenetic dosimetry study

The dosimetry and control of exposure for individuals chronically exposed to ionizing radiation are important and complex issues. Assessment may be optimized by evaluating individual adaptation and radiosensitivity, but it is not possible for a single model to account for all relevant parameters. Our goal was to develop approaches for the calculation of doses for persons chronically exposed to ionizing radiation, taking their radiosensitivities into consideration. On the basis of ex vivo radiation of blood samples, dose-effect models were constructed for dose ranges 0.01–2.0 and 0.01–0.4 Gy, using different cytogenetic criteria. The frequencies of "dicentric chromosomes and rings" at low doses are too low to have predictive value. The different responses of subjects to radiation made it possible to categorize them according to their radiosensitivities and to generate separate dose-effect curves for radiosensitive, average, and radioresistant individuals, reducing the amount of error in retrospective dosimetry.

Радіоізотопи цезію і досвід цитогенетичної дозиметрії за аварійних ситуацій

An analysis of data from the literature and own experimental studies related to radiation accidents with 137Cs, the ways of its entry and behavior in the human body, cytogenetic indicators for exposure dosimetry, experience in assessing the dose of victims under conditions of external exposure, and the problem of cytogenetic dosimetry of internal exposure are presented.

Deep Neural Network-Based Automatic Dicentric Chromosome Detection Using a Model Pretrained on Common Objects.

Dicentric chromosome assay (DCA) is one of the cytogenetic dosimetry methods where the absorbed dose is estimated by counting the number of dicentric chromosomes, which is a major radiation-induced change in DNA. However, DCA is a time-consuming task and requires technical expertise. In this study, a neural network was applied for automating the DCA. We used YOLOv5, a one-stage detection algorithm, to mitigate these limitations by automating the estimation of the number of dicentric chromosomes in chromosome metaphase images. YOLOv5 was pretrained on common object datasets. For training, 887 augmented chromosome images were used. We evaluated the model using validation and test datasets with 380 and 300 images, respectively. With pretrained parameters, the trained model detected chromosomes in the images with a maximum F1 score of 0.94 and a mean average precision (mAP) of 0.961. Conversely, when the model was randomly initialized, the training performance decreased, with a maximum F1 score and mAP of 0.82 and 0.873%, respectively. These results confirm that the model could effectively detect dicentric chromosomes in an image. Consequently, automatic DCA is expected to be conducted based on deep learning for object detection, requiring a relatively small amount of chromosome data for training using the pretrained network.

Postirradiation temperature influences DSB repair and dicentric chromosome formation-potential impact for dicentric chromosome analysis in interlaboratory comparisons.

The objective was to investigate the influence of different pre-storage temperatures in the dicentric chromosome analysis (DCA) protocol (22°C vs. 37°C) by using γ-H2AX+53BP1 foci as a marker for deoxyribonucleic acid (DNA) double-strand break (DSB) damage induction and repair and the formation of dicentric chromosomes as a result of mis-repair. Repair of γ-H2AX+53BP1 DSB foci was absent in samples that were incubated for 2h at 22°C after exposure of 0.5 and 1.2Gy. When 0.5- and 1.2-Gy-exposed samples were incubated at 37°C for 2h, there was an average decline of 31 and 52% of DSB foci, respectively. This indicated that DNA repair occurred. There was a 27% decrease in dicentric chromosome yield at 1.2Gy and a 15% decrease at 3.5Gy after post-irradiation incubation for 2 h at 37°C relative to the observed dicentric frequencies at 22°C. Recommended to re-phase: our data suggested that there were more open DSBs after a 2-h incubation at 22°C, which contributed to more mis-repair and dicentric formation from the start of culture. Our findings are corroborated by publications showing that lesion interaction based on enzymatic activity is suppressed below 21°C. As such temperature variations can be a source of variation in DCA during interlaboratory comparison studies, we propose to establish a common guide for the standardisation of pre-culture conditions in cytogenetic dosimetry proficiency testing.

Establishment and validation of a calibration curve for dicentric chromosome induced by 6MV X-ray.

Radiation during radiotherapy and nuclear accidents is currently one of the biggest concerns for the international community. Biological dosimetry examines the amount of damage caused by radiation at the cellular level by quantifying a radiation biomarker. In particular, the dicentric chromosome assay is a biodosimetric technique that can quantify radiation damage by correlating radiation dose exposure with the frequency of dicentric chromosomes in the peripheral lymphocytes extracted from exposed individuals. This study aims to present of the reference dose-response calibration curve for biodosimetry laboratory of Mashhad University of Medical Sciences (north-east of Iran). In all, 40 samples of peripheral blood from four healthy volunteers were irradiated at doses of 0-5 Gray in a customised water phantom using a 6MV X-rays at dose rate of 2Gy/min from a linear accelerator. The irradiated samples were cultured and analysed according to the International Atomic Energy Agency Cytogenetic Dosimetry Protocol (2011) with some modifications. Linear-quadratic model curve fitting and further statistical analysis were done using Chromosome Aberration Calculation Software Version 2.0 and Dose Estimate (Version 5.2). The curve equation obtained was ${Y}_{dic}=0.0533{D}^2+0.0231D+0.0001$ and was in the range of other studies. Validation of the calibration curve was done by estimating the dose of blind samples.

Revision of cytogenetic dosimetry in the IAEA manual 2011 based on data about radio-sensitivity and dose-rate findings contributing.

In order to achieve the goal of rapid response, effective controland protection of life inlarge-scale radiation events, the IAEA Manual 2011 has been revised based on the data of radio-sensitivity, dose-rate findings. Analyze individual differences in radiation sensitivity using 60 Co radiation (0.27 Gy/min). Chromosomal aberrations with different irradiation dose rates were used to establish the biological dose curve and analyze the excess of the "dicentric + ring" caused by the dose rate at each dose point; DAPI-images and Metafer 4 were used to capture metaphase images and make further analysis. The data were collected in 2020, Dicentric + ring/100 Cells was 17.5-43.8, the average value was28.32 ± 6.98. The mean value of Dicentric + ring/100 Cells was 31.37 in males while 25.27 in females, there are significant differences (p < .01). The irradiation dose is dominant, At each dose point, the value of"(dicentric chromosome + centric rings)/cell" is proportional to "dose rate", that is, Y=kx + b, within the dose range of 1-5 Gy, "(dicentric chromosome + centric rings)/Cell" holds a quadratic linear relationship with dose rate, that is, y=ax2 + bx + c; The DAPI-images might give you more hints than those of conventional Giemsa-stain. The authors recommend that the IAEA Manual 2011 could be revised based on data of radio-sensitivity and dose-rate, which may contribute to the establishment of a unified dose-response calibration curve and stimulation of potential for automation in cytogenetic biodosimetry. (1) Individual differences of radiosensitivity are very large. (2) At each dose point, "(dicentric chromosome + centric rings)/cell" is proportional to "dose rate", that is, Y=kx + b. (3) "(dicentric chromosome + centric rings)/Cell" is a quadratic linear relationship with dose rate, that is, y=ax2 + bx + c. (4) We created a "Unity Standard Curve of Biological Dose Estimation". Creating a Unity Standard Curve of Biological Dose, under these circumstances, we can form a joint and rapid response to a nuclear and radiological accident.

Bayesian and Classical Solutions for Binomial Cytogenetic Dosimetry Problem

Publisher: School of Statistics, Renmin University of China, Journal: Journal of Data Science, Title: Bayesian and Classical Solutions for Binomial Cytogenetic Dosimetry Problem, Authors: M´arcia D. Branco, Heleno Bolfarine

Interim In vitro Dose-Response Curve for the Dicentric Biodosimeter Assay from a Philippine Radiotherapy Facility using a Linear Accelerator

Background. Accidental radiation exposure can occur anytime. Biodosimeters help in quantifying the absorbed dose of individuals who are not equipped with personal dosimeters during radiation exposure. The dicentric assay can quantify radiation damage by correlating radiation dose exposure with the frequency of dicentric chromosomes in the peripheral lymphocytes extracted from exposed individuals.&#x0D; Objective. The study aims to present the interim results of the reference dose-response curve for a Philippine radiotherapy facility constructed using a 6MV linear accelerator (ClinacX, Varian).&#x0D; Methods. Samples of peripheral blood from healthy volunteers were irradiated in a customized water phantom of doses 0.10 to 5.0 Gray using a linear accelerator. The irradiated samples were cultured and analyzed following the International Atomic Energy Agency Cytogenetic Dosimetry Protocol (2011) with modifications. Linear-quadratic model curve fitting and further statistical analysis were done using CABAS (Chromosome Aberration Calculation Software Version 2.0) and Dose Estimate (Version 5.2). Interim results of the samples were used to generate these curves.&#x0D; Results. The dose-response curve generated from the preliminary results were comparable to published dose response curves from international cytogenetic laboratories.&#x0D; Conclusion. The generated dose-response calibration curve will be useful for medical triage of the public and radiologic staff accidentally exposed to radiation during medical procedures or in the event of nuclear accidents.

Transfusion of γ-irradiated blood components to individuals does not compromise the cytogenetic dose assessment

Most blood components for transfusions are irradiated ex vivo to prevent transfusion-associated graft-versus-host disease (TA-GvHD); this irradiation can potentially affect the cytogenetic dose assessment of patients showing acute radiation syndrome (ARS) with bone marrow suppression or acute anaemia. Whole blood samples from five donors were irradiated with 0, 10 or 25 Gy γ-rays. The mitotic activity of each cultured blood sample was measured by calculating the mitotic index. A dicentric chromosome assay was used to evaluate the chromosomal aberrations and absorbed dose of blood lymphocytes. Mitogenic activity and scorable metaphase spreads were significantly decreased in the blood samples irradiated with 10 and 25 Gy (p < 0.001). Moreover, a significant increase in the mean scores of all types of chromosomal aberrations in the 10 Gy γ-irradiated samples was observed, with the estimated dose being 11.3 Gy (95% CI: 10.67–11.95 Gy); however, we were unable to estimate the exposure dose in the 25 Gy γ-irradiated samples due to a limited number of scorable metaphase spreads. The mitotic index of the 25 Gy γ-irradiated whole blood samples was significantly suppressed by more than 4-log fold. Thus, in the present study, we evaluated the effects of recommended radiation doses in γ-irradiated transplantation blood components using cytogenetic dosimetry. These results suggest that the partial transfusion of blood components to patients with ARS or acute anaemia did not compromise the estimation of the exposure dose using cytogenetic dosimetry.

CYTOGENETIC INDICATORS OF ACUTE RADIATION SICKNESS (THE CHORNOBYL EXPERIENCE).

The study was based on primary cytogenetic data obtained in May 1986 within examina-tion of the 30 clean-up workers («liquidators») having got stage I-III acute radiation sickness. Dose verificationwas performed using the cytogenetic dosimetry based on a culture of peripheral blood lymphocytes with metaphaseanalysis of chromosome aberrations. A new method of evaluating the results of patients' cytogenetic examination at the beginning of specifictherapy has been developed. Procedure was performed using a model of multiple linear regression (complex of cyto-genetic parameters) and provided a satisfactory diagnostic level (featuring a compliance with initially definedclinical and laboratory diagnoses). Overall frequency of the aberrant cells and radiation markers increased in high-er disease stages. There was a trend of the frequency growth of chromatid-type aberrations with increasing of radi-ation burden. Adequacy of the proposed method based on the regression analysis of cytogenetic results was con-firmed through the preservation of group differences in estimates of disease stage in subjects with verified diagnosis. Cytogenetic dosimetry in the scope of examination of persons exposed to ionizing radiation is an oblig-atory component of radiation sickness stage verification. The recommended method of cytogenetic data evaluationbefore and at the beginning of detoxification therapy provides a satisfactory level of diagnostics.

Rogue cell-like chromosomal aberrations in peripheral blood lymphocytes of interventional radiologists: A case study

We report two cases of interventional radiologists who had been exposed to radiation while performing fluoroscopically-guided interventional procedures (FGIPs), mainly transcatheter arterial chemoembolization, percutaneous catheter drainage, and percutaneous transhepatic biliary drainage procedures, for over 10 years. They had a unique multi-aberrant cell type with not only high numbers of dicentrics and/or centric rings but also excess acentric double minutes, similar to a rogue cell. As revealed in a self-administered questionnaire, they wore personal dosimeters and protective equipment at all times and used shielding devices during interventional fluoroscopy procedures. However, the exposed dose levels derived from cytogenetic dosimetry were much higher than the doses recorded on their personal dosimeters. A large number of unstable and stable chromosomal aberrations that were found in the peripheral blood lymphocytes of these interventional radiologists might be due to repeated and long-term exposure to ionizing radiation while performing FGIPs. Further investigations of chromosomal aberrations in interventional radiologists may improve the understanding of the long-term effects of radiation exposure on medical personnel.

Indicadores citogenéticos para la identificación de exposición a radiación ionizante en humanos

"La biodosimetría citogenética se aplica en la evaluación médica de las personas involucradas en situaciones radiológicas anormales, con el fin de evaluar las dosis recibidas, el peligro inminente para la salud y aplicar los tratamientos médicos más adecuados. Además, contribuye al esclarecimiento de sucesos cuando existen dudas respecto a los resultados de la dosimetría física por dosímetros defectuosos, no calibrados o ausentes. Es el método más preciso de dosimetría biológica, ya que existe una relación matemática que permite calcular la dosis, establecer el grado de homogeneidad de la exposición y, en caso de exposiciones no homogéneas, establecer la fracción del cuerpo irradiada y la dosis que recibió esa fracción mediante la cuantificación del número y tipos de aberraciones cromosómicas y de micronúcleos, y su distribución en los linfocitos de la sangre periférica. Para este análisis se establecen las relaciones dosis-efecto y un sistema automatizado para el cálculo de las dosis de radiación recibidas. Actualmente se está desarrollando un proyecto conjunto Universidad de Costa Rica – Hospital San Juan de Dios, con el objetivo de explorar los efectos cromosómicos de la radiación, en pacientes expuestos por razones terapéuticas y atendidos en este hospital. De igual modo, se hará la curva de calibración dosis – respuesta in vitro para rayos gama y se validará mediante la intercomparación con el Laboratorio de Dosimetría Citogenética del Centro para la Protección e Higiene de las Radiaciones de Cuba."

Use of biological dosimetry to confirm radiation exposure: Case study

The notification of a radiological accident/incident in Portugal is very rare. This study was conducted after a suspected occupational overexposure to ionizing radiation by a radiologist whose thermoluminescent dosimeter (TLD) had exceeded the value allowed by Portuguese legislation. After verifying the facilities a biological dosimetry study using the dicentric assay was performed using the dose-response curve for the Portuguese population. Cytogenetic dosimetry confirmed the TLD value and the worker's exposure to an accidental dose of ionizing radiation. The Portuguese Directorate-General for Health imposed health surveillance for the patient and he was excluded from further professional radiation exposure for 5 years.

THE ABILITY OF CYTOGENETIC ANALYSIS TO ESTIMATE RADIATION DOSES RETROSPECTIVELY IN CHERNOBYL ACCIDENT RECOVERY WORKER

Relevance. There is a need to evaluate doses retrospectively in Chernobyl nuclear power plant accident recovery workers without established external doses.Intention: To assess the ability of cytogenetic analysis to estimate external radiation doses retrospectively in Chernobyl nuclear power plant accident recovery workers exposed to low-dose radiation in the remote period.Methodology. Cytogenetic biological indication and biological dosimetry were performed in Chernobyl nuclear power plant accident recovery workers 27–30 years after their participation in recovery works. Stable and unstable chromosome aberrations were assessed in peripheral blood lymphocytes.Results and Discussion. The analysis of unstable chromosome aberrations showed that 45.1 % of Chernobyl nuclear power plant accident recovery workers had radiation markers more frequently than in the comparison group. Biological doses were retrospectively evaluated in 18 % of Chernobyl nuclear power plant accident recovery workers and ranged within 14–48 cGy. Specific anamnestic factors should be assessed while evaluating doses retrospectively.Conclusion. Analysis of stable chromosome aberrations allows retrospective dosimetry after low-dose exp sure if external dose is unavailable. Based on this, adverse health effects of low-dose exposure can be assessed and reimbursed in Chernobyl nuclear power plant accident recovery workers.

Dose response of prematurely condensed chromosome rings after gamma irradiation

Purpose: This paper presents a calibration curve for prematurely condensed chromosome rings (PCC-ring) which are scarce in cytogenetic dosimetry; here we analyzed the behavior of the PCC-ring frequency and PCC index after gamma irradiation in the dose interval from 0 to 20 Gy.Materials and methods: PCC-rings were induced in lymphocytes with Calyculin A. A total of 5788 PCC cells in G1, G2/M, and M/A stages were analyzed.Results: The best fitting curve between the frequency of PCC ring (Y) and the Dose (D) was obtained with the equation Y = (0.0308 ± 0.0012) D. The distribution of rings by cell followed a Poisson distribution. The PCC index was correlated with radiation dose and decreased in a dose-dependent manner.Conclusion: This PCC-ring dose–effect curve can be used in case of high dose accidental overexposure to gamma radiation, allowing a dose assessment in a reliable way in most of the simulated irradiation analyzed.

Laboratory Intercomparison of Cytogenetic Dosimetry Among 38 Laboratories in China.

A nationwide intercomparison exercise for estimating the irradiated dose was organized by the National Institute for Radiological Protection, Center for Disease Control and Prevention of China. Thirty-eight laboratories participated in this program. The main objective of this intercomparison exercise was to compare the participants’ ability of operation and dose assessment basing on the frequencies of dicentrics and centric rings. Whole blood samples were irradiated with different dosages of 60Co γ-rays. Each laboratory collected 2 blind samples and prepared the slides independently. All participants presented the estimated dose reports within 30 days. The doses assessed by the participants were acceptable within the reference dose of ±20%. The mean absolute difference of estimated dose relative to the reference dose was calculated, which reflected the overall accuracy of dose estimates for each laboratory. The overall estimation results of blind blood samples for intercomparison showed a good agreement with the reference dose for each sample, with nearly 75% of the participants producing acceptable results.

Simplified Bayesian method: application in cytogenetic biological dosimetry of mixed n\u2009+\u2009\u03b3 radiation fields

This article describes the application of a simplified Bayesian method for estimation of doses from a mixed field using cytogenetic biological dosimetry, taking as an example neutron and gamma radiation emitted from the MARIA nuclear research reactor in Poland. The Bayesian approach is a good alternative to the commonly used iterative method, which allows separate dose estimation. In the present paper, a computer program, which uses the iterative and simplified Bayesian methods to calculate mixed radiation doses, is introduced.

Dose Estimation Curves Following In Vitro X-ray Irradiation Using Blood From Four Healthy Korean Individuals.

Cytogenetic dosimetry is useful for evaluating the absorbed dose of ionizing radiation based on analysis of radiation-induced chromosomal aberrations. We created two types of in vitro dose-response calibration curves for dicentric chromosomes (DC) and translocations (TR) induced by X-ray irradiation, using an electron linear accelerator, which is the most frequently used medical device in radiotherapy. We irradiated samples from four healthy Korean individuals and compared the resultant curves between individuals. Aberration yields were studied in a total of 31,800 and 31,725 metaphases for DC and TR, respectively, obtained from 11 X-ray irradiation dose-points (0, 0.05, 0.1, 0.25, 0.5, 0.75, 1, 2, 3, 4, and 5 Gy). The dose-response relationship followed a linear-quadratic equation, Y=C+αD+βD2, with the coefficients C=0.0011 for DC and 0.0015 for TR, α=0.0119 for DC and 0.0048 for TR, and β=0.0617 for DC and 0.0237 for TR. Correlation coefficients between irradiation doses and chromosomal aberrations were 0.971 for DC and 0.6 for TR, indicating a very strong and a moderate correlation, respectively. This is the first study implementing cytogenetic dosimetry following exposure to ionizing X-radiation.