Gordon K. Livingston

Interlaboratory Comparison of the Dicentric Chromosome Assay for Radiation Biodosimetry in Mass Casualty Events

Abstract Wilkins, R. C., Romm, H., Kao, T-C., Awa, A. A., Yoshida, M. A., Livingston, G. K., Jenkins, M. S., Oestreicher, U., Pellmar, T. C. and Prasanna, P. G. S. Interlaboratory Comparison of the Dicentric Chromosome Assay for Radiation Biodosimetry in Mass Casualty Events. Radiat. Res. 169, 551–560 (2008). This interlaboratory comparison validates the dicentric chromosome assay for assessing radiation dose in mass casualty accidents and identifies the advantages and limitations of an international biodosimetry network. The assays validity and accuracy were determined among five laboratories following the International Organization for Standardization guidelines. Blood samples irradiated at the Armed Forces Radiobiology Research Institute were shipped to all laboratories, which constructed individual radiation calibration curves and assessed the dose to dose-blinded samples. Each laboratory constructed a dose–effect calibration curve for the yield of dicentrics for 60Co γ rays in the 0 to 5-Gy range, using the maximum likelihood linear-quadratic model, Y = c + αD + βD2. For all laboratories, the estimated coefficients of the fitted curves were within the 99.7% confidence intervals (CIs), but the observed dicentric yields differed. When each laboratory assessed radiation doses to four dose-blinded blood samples by comparing the observed dicentric yield with the laboratorys own calibration curve, the estimates were accurate in all laboratories at all doses. For all laboratories, actual doses were within the 99.75% CI for the assessed dose. Across the dose range, the error in the estimated doses, compared to the physical doses, ranged from 15% underestimation to 15% overestimation.

International study of factors affecting human chromosome translocations

Chromosome translocations in peripheral blood lymphocytes of normal, healthy humans increase with age, but the effects of gender, race, and cigarette smoking on background translocation yields have not been examined systematically. Further, the shape of the relationship between age and translocation frequency (TF) has not been definitively determined. We collected existing data from 16 laboratories in North America, Europe, and Asia on TFs measured in peripheral blood lymphocytes by fluorescence in situ hybridization whole chromosome painting among 1933 individuals. In Poisson regression models, age, ranging from newborns (cord blood) to 85 years, was strongly associated with TF and this relationship showed significant upward curvature at older ages versus a linear relationship (p<0.001). Ever smokers had significantly higher TFs than non-smokers (rate ratio (RR)=1.19, 95% confidence interval (CI), 1.09-1.30) and smoking modified the effect of age on TFs with a steeper age-related increase among ever smokers compared to non-smokers (p<0.001). TFs did not differ by gender. Interpreting an independent effect of race was difficult owing to laboratory variation. Our study is three times larger than any pooled effort to date, confirming a suspected curvilinear relationship of TF with age. The significant effect of cigarette smoking has not been observed with previous pooled studies of TF in humans. Our data provide stable estimates of background TF by age, gender, race, and smoking status and suggest an acceleration of chromosome damage above age 60 and among those with a history of smoking cigarettes.

WHO 1st Consultation on the Development of a Global Biodosimetry Laboratories Network for Radiation Emergencies (BioDoseNet)

Abstract Blakely, W. F., Carr, Z., Chu, M. C-M., Dayal-Drager, R., Fujimoto, K., Hopmeir, M., Kulka, U., Lillis-Hearne, P., Livingston, G. K., Lloyd, D. C., Maznyk, N., Perez, M. D. R., Romm, H., Takashima, Y., Voisin, P., Wilkins, R. C. and Yoshida, M. A. WHO 1st Consultation on the Development of a Global Biodosimetry Laboratories Network for Radiation Emergencies (BioDoseNet). Radiat. Res. 171, 127–139 (2009). The World Health Organization (WHO) held a consultation meeting at WHO Headquarters, Geneva, Switzerland, December 17–18, 2007, to develop the framework for a global biodosimetry network. The WHO network is envisioned to enable dose assessment using multiple methods [cytogenetics, electron paramagnetic resonance (EPR), radionuclide bioassays, etc.]; however, the initial discussion focused on the cytogenetic bioassay (i.e., metaphase-spread dicentric assay). Few regional cytogenetic biodosimetry networks have been established so far. The roles and resources available from United Nations (UN) agencies that provide international cooperation in biological dosimetry after radiological emergencies were reviewed. In addition, extensive reliance on the use of the relevant International Standards Organization (ISO) standards was emphasized. The results of a WHO survey of global cytogenetic biological dosimetry capability were reported, and while the survey indicates robust global capability, there was also a clear lack of global leadership and coordination. The expert group, which had a concentrated focus on cytogenetic biodosimetry, formulated the general scope and concept of operations for the development of a WHO global biodosimetry laboratory network for radiation emergencies (BioDoseNet). Follow-on meetings are planned to further develop technical details for this network.

Biological Dosimetry by the Triage Dicentric Chromosome Assay: Potential Implications for Treatment of Acute Radiation Syndrome in Radiological Mass Casualties

Abstract Biological dosimetry is an essential tool for estimating radiation dose. The dicentric chromosome assay (DCA) is currently the tool of choice. Because the assay is labor-intensive and time-consuming, strategies are needed to increase throughput for use in radiation mass casualty incidents. One such strategy is to truncate metaphase spread analysis for triage dose estimates by scoring 50 or fewer metaphases, compared to a routine analysis of 500 to 1000 metaphases, and to increase throughput using a large group of scorers in a biodosimetry network. Previously, the National Institutes for Allergies and Infectious Diseases (NIAID) and the Armed Forces Radiobiology Research Institute (AFRRI) sponsored a double-blinded interlaboratory comparison among five established international cytogenetic biodosimetry laboratories to determine the variability in calibration curves and in dose measurements in unknown, irradiated samples. In the present study, we further analyzed the published data from this previous study to investigate how the number of metaphase spreads influences dose prediction accuracy and how this information could be of value in the triage and management of people at risk for the acute radiation syndrome (ARS). Although, as expected, accuracy decreased with lower numbers of metaphase spreads analyzed, predicted doses by the laboratories were in good agreement and were judged to be adequate to guide diagnosis and treatment of ARS. These results demonstrate that for rapid triage, a network of cytogenetic biodosimetry laboratories can accurately assess doses even with a lower number of scored metaphases.

Incidence of Sister Chromatid Exchange in Bone Marrow Cells of the Mouse Following Microwave Exposure

The measurement of sister chromatid exchange (SCE) constitutes the most sensitive indicator yet developed for detecting cytogenetic effects of mutagens and carcinogens. This sensitive assay was utilized to investigate potential mutagenic bioeffects of microwave (2450-MHz) radiation by comparing the incidence of SCE in bone marrow cells of sham control mice, standard control mice, and irradiated rice following a 28-day exposure to 20 mW/cm/sup 2/ incident power density. No statistically significant differences in the numbers of SCEs were detected between the exposed group and the control groups.

Interlaboratory Variation in Scoring Dicentric Chromosomes in a Case of Partial-Body X-Ray Exposure: Implications for Biodosimetry Networking and Cytogenetic “Triage Mode” Scoring

Abstract Ainsbury, E. A., Livingston, G. K., Abbott, M. G., Moquet, J. E., Hone, P. A., Jenkins, M. S., Christensen, D. M., Lloyd, D. C. and Rothkamm, K. Interlaboratory Variation in Scoring Dicentric Chromosomes in a Case of Partial-Body X-Ray Exposure: Implications for Biodosimetry Networking and Cytogenetic “Triage Mode” Scoring. The international radiation biodosimetry community has recently been engaged in activities focused on establishing cooperative networks for biodosimetric triage for radiation emergency scenarios involving mass casualties. To this end, there have been several recent publications in the literature regarding the potential for shared scoring in such an accident or incident. We present details from a medical irradiation case where two independently validated laboratories found very different yields of dicentric chromosome aberrations. The potential reasons for this disparity are discussed, and the actual reason is identified as being the partial-body nature of the radiation exposure combined with differing criteria for metaphase selection. In the context of the recent networking activity, this report is intended to highlight the fact that shared scoring may produce inconsistencies and that further validation of the scoring protocols and experimental techniques may be required before the networks are prepared to deal satisfactorily with a radiological or nuclear emergency. Also, the findings presented here clearly demonstrate the limitations of the dicentric assay for estimating radiation doses after partial-body exposures and bring into question the usefulness of rapid “triage mode” scoring in such exposure scenarios.

A Modified System for Analyzing Ionizing Radiation-Induced Chromosome Abnormalities

The analysis of dicentric chromosomes in human peripheral blood lymphocytes (PBLs) by Giemsa staining is the most established method for biological dosimetry. However, this method requires a well-trained person because of the difficulty in detecting aberrations rapidly and accurately. Here, we applied a fluorescence in situ hybridization (FISH) technique, using telomere and centromere peptide nucleic acid (PNA) probes, to solve the problem of biological dosimetry in radiation emergency medicine. A comparison by a well-trained observer found that FISH analysis of PBLs for the dose estimation was more accurate than the conventional Giemsa analysis, especially in samples irradiated at high doses. These results show that FISH analysis with centromeric/telomeric PNA probes could become the standard method for biological dosimetry in radiation emergency medicine.

Effect of Occupational Radiation Exposures on Chromosome Aberration Rates in Former Plutonium Workers

Abstract Livingston, G. K., Falk, R. B. and Schmid, E. Effect of Occupational Radiation Exposures on Chromosome Aberration Rates in Former Plutonium Workers. Radiat. Res. 166, 89–97 (2006). A fluorescence in situ hybridization (FISH) method was used to measure chromosome aberration rates in lymphocytes of 30 retired plutonium workers with combined internal and external radiation doses greater than 0.5 Sv along with 17 additional workers with predominantly external doses below 0.1 Sv. The former group was defined as high-dose and the latter as low-dose with respect to occupational radiation exposure. The two groups were compared to each other and also to 21 control subjects having no history of occupational radiation exposure. Radiation exposures to the high-dose group were primarily the result of internal depositions of plutonium and its radioactive decay products resulting from various work-related activities and accidents. The median external dose for the high-dose group was 280 mSv (range 10–730) compared to a median of 22 mSv (range 10–76) for the low-dose group. The median internal dose to the bone marrow for the high-dose group was 168 mSv (range 29–20,904) while that of the low-dose group was considered negligible. Over 200,000 metaphase cells were analyzed for chromosome aberrations by painting pairs 1, 4 and 12 in combination with a pancentromeric probe. Additionally, 136,000 binucleated lymphocytes were analyzed for micronuclei in parallel cultures to assess mitotic abnormalities arising from damaged chromosomes. The results showed that the frequency of structural aberrations affecting any of the painted chromosomes in the high-dose group correlated with the bone marrow dose but not with the external dose. In contrast, the frequency of micronuclei did not vary significantly between the study groups. The total translocation frequency per genome equivalent × 10−3 ± SE was 4.0 ± 0.6, 9.0 ± 1.1 and 17.0 ± 2.1 for the control, low-dose and high-dose groups, respectively. Statistical analysis of the data showed that the frequency of total translocations and S-cells correlated with the bone marrow dose, with P values of 0.005 and 0.004, respectively. In contrast, these two end points did not correlate with the external dose, with P values of 0.45 and 0.39, respectively. In conclusion, elevated rates of stable chromosome aberrations were found in lymphocytes of former workers decades after plutonium intakes, providing evidence that chronic irradiation of hematopoietic precursor cells in the bone marrow induces cytogenetically altered cells that persist in peripheral blood.

Evaluation of the annual Canadian biodosimetry network intercomparisons

Abstract Purpose: To evaluate the importance of annual intercomparisons for maintaining the capacity and capabilities of a well-established biodosimetry network in conjunction with assessing efficient and effective analysis methods for emergency response. Materials and methods: Annual intercomparisons were conducted between laboratories in the Canadian National Biological Dosimetry Response Plan. Intercomparisons were performed over a six-year period and comprised of the shipment of 10–12 irradiated, blinded blood samples for analysis by each of the participating laboratories. Dose estimates were determined by each laboratory using the dicentric chromosome assay (conventional and QuickScan scoring) and where possible the cytokinesis block micronucleus (CBMN) assay. Dose estimates were returned to the lead laboratory for evaluation and comparison. Results: Individual laboratories performed comparably from year to year with only slight fluctuations in performance. Dose estimates using the dicentric chromosome assay were accurate about 80% of the time and the QuickScan method for scoring the dicentric chromosome assay was proven to reduce the time of analysis without having a significant effect on the dose estimates. Although analysis with the CBMN assay was comparable to QuickScan scoring with respect to speed, the accuracy of the dose estimates was greatly reduced. Conclusions: Annual intercomparisons are necessary to maintain a network of laboratories for emergency response biodosimetry as they evoke confidence in their capabilities.

Role of dicentric analysis in an overarching biodosimetry strategy for use following a nuclear detonation in an urban environment.

AbstractIn the moments immediately following a nuclear detonation, casualties with a variety of injuries including trauma, burns, radiation exposure, and combined injuries would require immediate assistance. Accurate and timely radiation dose assessments, based on patient history and laboratory testing, are absolutely critical to support adequately the triage and treatment of those affected. This capability is also essential for ensuring the proper allocation of scarce resources and will support longitudinal evaluation of radiation-exposed individuals and populations. To maximize saving lives, casualties must be systematically triaged to determine what medical interventions are needed, the nature of those interventions, and who requires intervention immediately. In the National Strategy for Improving the Response and Recovery for an Improvised Nuclear Device (IND) Attack, the U.S. Department of Homeland Security recognized laboratory capacity for radiation biodosimetry as having a significant gap for performing mass radiation dose assessment. The anticipated demand for radiation biodosimetry exceeds its supply, and this gap is partly linked to the limited number and analytical complexity of laboratory methods for determining radiation doses within patients. The dicentric assay is a key component of a cytogenetic biodosimetry response asset, as it has the necessary sensitivity and specificity for assessing medically significant radiation doses. To address these shortfalls, the authors have developed a multimodal strategy to expand dicentric assay capacity. This strategy includes the development of an internet-based cytogenetics network that would address immediately the labor intensive burden of the dicentric chromosome assay by increasing the number of skilled personnel to conduct the analysis. An additional option that will require more time includes improving surge capabilities by combining resources available within the country’s 150 clinical cytogenetics laboratories. Key to this intermediate term effort is the fact that geneticists and technicians may be experts in matters related to identifying chromosomal abnormalities related to genetic disorders, but they are not familiar with dosimetry for which training and retraining will be required. Finally, long-term options are presented to improve capacity focus on ways to automate parts of the dicentric chromosome assay method.