Alan A. Edwards

935 MHz cellular phone radiation. An in vitro study of genotoxicity in human lymphocytes.

Purpose: The possibility of genotoxicity of radiofrequency radiation (RFR) applied alone or in combination with x-rays was investigated in vitro using several assays on human lymphocytes. The chosen specific absorption rate (SAR) values are near the upper limit of actual energy absorption in localized tissue when persons use some cellular telephones. The purpose of the combined exposures was to examine whether RFR might act epigenetically by reducing the fidelity of repair of DNA damage caused by a well-characterized and established mutagen. Methods: Blood specimens from 14 donors were exposed continuously for 24 h to a Global System for Mobile Communications (GSM) basic 935 MHz signal. The signal was applied at two SAR; 1 and 2 W/Kg, alone or combined with a 1-min exposure to 1.0 Gy of 250 kVp x-rays given immediately before or after the RFR. The assays employed were the alkaline comet technique to detect DNA strand breakage, metaphase analyses to detect unstable chromosomal aberrations and sister chromatid exchanges, micronuclei in cytokinesis-blocked binucleate lymphocytes and the nuclear division index to detect alterations in the speed of in vitro cell cycling. Results: By comparison with appropriate sham-exposed and control samples, no effect of RFR alone could be found for any of the assay endpoints. In addition RFR did not modify any measured effects of the x-radiation. Conclusions: This study has used several standard in vitro tests for chromosomal and DNA damage in Go human lymphocytes exposed in vitro to a combination of x-rays and RFR. It has comprehensively examined whether a 24-h continuous exposure to a 935 MHz GSM basic signal delivering SAR of 1 or 2 W/Kg is genotoxic per se or whether, it can influence the genotoxicity of the well-established clastogenic agent; x-radiation. Within the experimental parameters of the study in all instances no effect from the RFR signal was observed.

Intercomparison of translocation and dicentric frequencies between laboratories in a follow-up of the radiological accident in Estonia

Purpose : To perform an interlaboratory comparison of FISH chromosome painting and to study the time-course of translocations and dicentrics in three accident victims exposed to radiation. Also, to use the data in the validation of the FISH technique as a retrospective dosimeter. Materials and methods : Twelve blood samples were collected during 4 years from three subjects exposed to radiation in an accident in Estonia in 1994 involving γ -radiation from a 137Cs source. Two of the subjects were exposed during approximately 7 h, both receiving a protracted dose of about 1 Gy and also localized exposure. The third subject received a protracted whole-body dose of 2.7Gy during 4 weeks as well as a short-term partialbody dose. Preparations from 48-h metaphase cultures were painted by the FISH technique using routine methods and probe cocktails in four laboratories. Samples from each subject were analysed in two different laboratories that used different combinations of whole chromosome probes. The PAINT nomenclature was applied when recording chromosome aberrations. Results : The intercomparison of FISH analysis data showed reasonable similarities between laboratories, the largest discrepancy being 21% in the frequency of two-way translocations in subject 3. Half-time calculations, based on combined data sets from two laboratories, showed that dicentrics decreased rapidly with half-times of approximately 2 years. In all cases, the initial dicentric yields were lower than the initial translocation yields. During the 4-year follow-up, the frequencies of all translocations in cells containing only simple rearrangements fell on average to about 65% of their initial value. Two-way translocations were slightly more persistent than all translocations. The average halftime was about 8 years for two-way translocations and around 6 years for all translocations. Cells containing complex rearrangements were few in number and they disappeared with time. In general, the inclusion of complex cells caused a more rapid fall in aberration yield. Conclusions : In general, the results imply that relatively consistent scoring data were obtained with different chromosome painting protocols. They also support the idea that the reduction of translocations with time is associated with partial-body irradiation.

The 60Co gamma ray dose-response for chromosomal aberrations in human lymphocytes analysed by FISH; applicability to biological dosimetry.

PURPOSE To investigate the in vitro dose-response for 60Co irradiated human lymphocytes assayed by FISH, and to consider how this may be applied to retrospective dosimetry. METHOD Blood was irradiated with doses in the range 0.25-4.0 Gy. Cultured lymphocytes were scored for all stable and unstable aberrations involving painted chromosomes 2, 3 and 5 and, in addition, all unstable aberrations in the counterstained chromosomes. A pancentromeric probe was included. RESULTS The relative numbers of painted and full genome dicentrics agreed well with the Lucas hypothesis for calculating genome equivalence. The involvement of each painted chromosome in exchanges agreed with their relative arm lengths. The dose-response relationship fitted well to the linear quadratic model; Y=(0.9 x 10(-2))D+(6.5 x 10(-2))D2 where D is the dose in Gy for the incidence Y, of all one plus two-way translocations in all cells corrected for genome equivalence. Complex rearrangements also became more frequent with increasing dose. A correlation was noted between the distributions of dicentrics and translocations among the cells and this was entirely due to complexes. CONCLUSIONS For retrospective dosimetry it is recommended to use an in vitro dose-response for apparently simple translocations in stable (Cs) cells. To date, acute linear yield coefficients from FISH data carry statistical uncertainties too large for useful application to retrospective dosimetry of persons exposed to chronic or low doses. As an interim measure it is suggested that one may derive a linear term from full genome dicentrics corrected by a factor representing the translocation to dicentric ratio.

The Relationship between Colony-forming Ability and Chromosomal Aberrations Induced in Human T-lymphocytes after γ-Irradiation

Using aliquots from the same samples of irradiated normal human blood, T-lymphocyte survival was measured by (a) 14-day colony growth, (b) the proportion of chromosomally normal first post-irradiation metaphases and (c) the proportion of cells not suffering interphase death and delay within 48 h after irradiation. Combining (b) and (c) gives a prediction of the proportion of cytogenetically undamaged cells which reach metaphase after 48 h. These cells should be capable of producing viable colonies under suitable culture conditions. A comparison of these values with those observed in (a) shows a reasonable agreement in accordance with the hypothesis that gross chromosomal damage and cell reproductive death are closely related.

Retrospective Biodosimetry among United States Radiologic Technologists

Abstract Parveen, B., Preston, D. L., Doody, M. M., Hauptmann, M., Kampa, D., Alexander, B. H., Petibone, D., Simon, S. L., Weinstock, R. M., Bouville, A., Yong, L. C., Freedman, D. M., Mabuchi, K., Linet, M. S., Edwards, A. A., Tucker, J. D. and Sigurdson, A. J. Retrospective Biodosimetry among United States Radiologic Technologists. Radiat. Res. 167, 727–734 (2007). Measurement of chromosome translocations in peripheral blood lymphocytes has been used to quantify prior exposure to ionizing radiation, including for workers exposed to low, chronic doses. We assessed translocation frequencies in a subset of U.S. radiologic technologists to substantiate ionizing radiation dose estimates developed for 110,418 technologists who worked between 1916 and 1984. From 3,441 cohort members known to have begun working before 1950, we selected a sample of 152, stratified by estimated cumulative dose, oversampling from higher-dose categories and excluding persons with a prior cancer diagnosis, a personal or family history of chromosomal instability disorders, or a current history of smoking. Estimates of film-badge dose ranged from less than 10 cSv to more than 30 cSv. Blood samples, obtained in 2004, were analyzed by fluorescence in situ hybridization (FISH) whole chromosome painting by simultaneously labeling chromosomes 1, 2 and 4 in red and 3, 5 and 6 in green. Translocations were scored in 1800 well-spread metaphase cells and expressed per 100 cell equivalents (CE) per person. Linear Poisson regression models with allowance for overdispersion were used to assess the relationship between estimated occupational red bone marrow absorbed dose in cGy and translocation frequency, adjusted for age, gender and estimated red bone marrow absorbed dose score from personal diagnostic procedures. We observed 0.09 excess translocations per 100 CE per cGy red bone marrow dose (95% CI: −0.01, 0.2; P = 0.07), which is similar to the expected estimate based on previous cytogenetic studies (0.05 excess translocations per 100 CE per cGy). Despite uncertainty in the estimates of occupational red bone marrow absorbed doses, we found good general agreement between the doses and translocation frequencies, lending support to the credibility of the dose assessment for this large cohort of U.S. radiologic technologists.

A Study to Verify a Reported Excess of Chromosomal Aberrations in Blood Lymphocytes of Namibian Uranium Miners

Abstract Lloyd, D. C., Lucas, J. N., Edwards, A. A., Deng, W., Valente, E., Hone, P. A. and Moquet, J. E. A Study to Verify a Reported Excess of Chromosomal Aberrations in Blood Lymphocytes of Namibian Uranium Miners. Radiat. Res. 155, 807–815 (2001). This report describes a study to verify an earlier report of excess chromosomal damage in the blood lymphocytes of uranium miners. Coded blood samples from 10 miners and 10 controls were analyzed conventionally for unstable aberrations and by FISH for translocations. Conventional analysis, scoring 1000 metaphases per subject, showed no significant difference between miners and controls in the frequencies of chromosome- and chromatid-type aberrations. Investigators at two laboratories undertook FISH analyses, each scoring 4000 metaphases per subject. When the data from each laboratory were examined separately, one found slightly more translocations in the miners while the other found fewer. In neither case was the difference significant at the 95% level of confidence. Combining the data likewise showed no significant excess of damage in the miners. This applied to simple one- and two-way translocations and to cells with complex exchanges. There was no correlation between levels of translocations and total lifetime doses from occupational and/or background irradiation. A borderline significant excess of rogue cells was found in the miners. This may be a chance observation, as these rare, highly abnormal cells are considered to be unrelated to radiation exposure and are probably due to a virus. The overall conclusion is that the frequency of chromosomal damage in the miners did not exceed that in the controls. Therefore, the result of the earlier study was not confirmed.

The Induction of Micronuclei in Human Lymphocytes by in Vitro Irradiation with Alpha Particles from Plutonium-239

Human peripheral blood lymphocytes were irradiated for 24 h in vitro with plutonium-239 citrate. The dose response for micronucleus induction using the cytochalasin B blocking technique fitted well to a linear relationship. Comparison with published data from the same laboratory using X-irradiation, indicated an RBE of 3.6 for alpha particles at low doses.

ROUTINE DIAGNOSTIC X-RAY EXAMINATIONS AND INCREASED FREQUENCY OF CHROMOSOME TRANSLOCATIONS AMONG U. S. RADIOLOGIC TECHNOLOGISTS

The U.S. population has nearly one radiographic examination per person per year, and concern about cancer risks associated with medical radiation has increased. Radiologic technologists were surveyed to determine whether their personal cumulative exposure to diagnostic X-rays was associated with increased frequencies of chromosome translocations, an established radiation biomarker and possible intermediary suggesting increased cancer risk. Within a large cohort of U.S. radiologic technologists, 150 provided a blood sample for whole chromosome painting and were interviewed about past X-ray examinations. The number and types of examinations reported were converted to a red bone marrow (RBM) dose score with units that approximated 1 mGy. The relationship between dose score and chromosome translocation frequency was assessed using Poisson regression. The estimated mean cumulative RBM radiation dose score was 49 (range, 0-303). After adjustment for age, translocation frequencies significantly increased with increasing RBM dose score with an estimate of 0.004 translocations per 100 cell equivalents per score unit (95% confidence interval, 0.002-0.007; P < 0.001). Removing extreme values or adjustment for gender, cigarette smoking, occupational radiation dose, allowing practice X-rays while training, work with radioisotopes, and radiotherapy for benign conditions did not affect the estimate. Cumulative radiation exposure from routine X-ray examinations was associated independently with increased chromosome damage, suggesting the possibility of elevated long-term health risks, including cancer. The slope estimate was consistent with expectation based on cytogenetic experience and atomic bomb survivor data.

Increased Frequency of Chromosome Translocations Associated with Diagnostic X-Ray Examinations

Abstract Bhatti, P., Doody, M. M., Preston, D. L., Kampa, D., Ron, E., Weinstock, R. W., Simon, S., Edwards, A. A. and Sigurdson, A. J. Increased Frequency of Chromosome Translocations Associated with Diagnostic X-Ray Examinations. Radiat. Res. 170, 149–155 (2008). Informative studies of cancer risks associated with medical radiation are difficult to conduct owing to low radiation doses, poor recall of diagnostic X rays, and long intervals before cancers occur. Chromosome aberrations have been associated with increased cancer risk and translocations are a known radiation biomarker. Seventy-nine U.S. radiologic technologists were selected for blood collection, and translocations were enumerated by whole chromosome painting. We developed a dose score to the red bone marrow for medical radiation exposure from X-ray examinations reported by the technologists that they received as patients. Using Poisson regression, we analyzed translocations in relation to the dose scores. Each dose score unit approximated 1 mGy. The estimated mean cumulative red bone marrow radiation dose score was 42 (range 1–265). After adjustment for age, occupational radiation, and radiotherapy for benign conditions, translocation frequencies significantly increased with increasing red bone marrow dose score with an estimate of 0.007 translocations per 100 CEs per score unit (95% CI, 0.002 to 0.013; P = 0.01). Chromosome damage has been linked with elevated cancer risk, and we found that cumulative radiation exposure from medical X-ray examinations was associated with increased numbers of chromosome translocations.

Choosing Metaphases for Biological Dosimetry by Fluorescence In Situ Hybridization (FISH)

Abstract Edwards, A., Maznik, N., Moquet, J., Hone, P., Vinnikov, V., Lloyd, D. and Cox, R. Choosing Metaphases for Biological Dosimetry by Fluorescence In Situ Hybridization (FISH). Radiat. Res. 157, 469–471 (2002). Data are presented for a subset of lymphocytes characterized by FISH as missing painted chromosomal material. These lymphocytes occur in both control and irradiated subjects. These cells have a much greater frequency of one-way translocations than cells in which all of the painted chromosomal material is present. Their presence contributes to interindividual variability in control translocation yields. These cells do not appear to be more prevalent in persons exposed to high radiation doses. It is suggested that their exclusion when selecting cells for analysis may improve the sensitivity of FISH as a biological dosimeter at low doses. Mechanisms for the production of these one-way translocations in vivo are also discussed, with a proposal that their variable frequency in individuals may be consistent with exposure to chemical clastogens.