Prevalence of adult-onset multifactorial disease among offspring of atomic bomb survivors.

The first study to examine whether parental radiation exposure leads to increased heritable risk of common adult-onset multifactorial diseases (i.e., hypertension, diabetes mellitus, hypercholesterolemia, ischemic heart disease, and stroke) was conducted among 11,951 participants in the clinical examination program out of a potential of 24,673 mail survey subjects who were offspring of survivors born from May 1946 through December 1984. Logistic regression analyses demonstrated no evidence of an association between the prevalence of multifactorial diseases in the offspring and parental radiation exposure, after adjusting for age, city, gender and various risk factors. The odds ratio (OR) for a paternal dose of 1 Gy was 0.91 [95% confidence interval (CI) 0.81-1.01, P = 0.08], and that for a maternal dose of 1 Gy was 0.98 (95% CI 0.86-1.10, P = 0.71). There was no apparent effect of parental age at exposure or of elapsed time between parental exposure and birth, but male offspring had a low odds ratio (OR = 0.76 at 1 Gy) for paternal exposure, but cautious interpretation is needed for this finding. The clinical assessment of nearly 12,000 offspring of A-bomb survivors who have reached a median age of about 50 years provided no evidence for an increased prevalence of adult-onset multifactorial diseases in relation to parental radiation exposure.

Liver cancer in atomic-bomb survivors: histological characteristics and relationships to radiation and hepatitis B and C viruses.

Histological features of primary liver cancer among atomic-bomb survivors and their relationship to hepatitis B (HBV) and C viral (HCV) infections are of special interest because of the increased risk of liver cancer in persons exposed to ionizing radiation and the high and increasing liver cancer rates in Japan and elsewhere. We conducted a pathology review of liver cancers occurring from 1958 to 1987 among subjects in the 120,321 member cohort of 1945 Hiroshima and Nagasaki residents. A panel of pathologists classified tumor histological types and defined accompanying cirrhotic changes of the liver. Archival tissue samples were assessed for HBV using pathology stains and PCR. Reverse transcriptase (RT) PCR was used to determine HCV status. We used unconditional logistic regression to compare 302 hepatocellular carcinoma (HCC) cases to 53 cholangiocarcinoma (CC) cases, adjusting for age, year of diagnosis, sex and viral status. Cirrhotic changes occurred significantly more often among HCC than CC cases (76% in HCC and 6% in CC). Compared to CC cases, HCC cases were 10.9 times more likely to be HBV-positive (95% confidence interval: 2.1-83.2) and 4.3 times more likely to be HCV-positive (95% confidence interval: 1.1-20.5). No significant differences were found between HCC and CC cases in radiation exposures. The predominance of HCC in the atomic-bomb survivors follows the background liver cancer pattern in Japan. Our findings suggest that HBV and HCV are involved in the pathogenesis of HCC with or without cirrhosis and are significantly less important in that of CC.

FISH examination of lymphocytes from Mayak workers for assessment of translocation induction rate under chronic radiation exposures.

PURPOSE: To estimate the translocation-induction rate under chronic exposure conditions by measuring chromosome aberration frequencies in lymphocytes from Mayak nuclear workers using fluorescence in situ hybridization (FISH). MATERIALS AND METHODS: Lymphocytes were examined from 27 nuclear workers at the Mayak Production Association and two control individuals using FISH with probes for chromosomes 1, 2 and 4. Official doses derived from worker film-badge records varied from 0 to 8.50 Gy. RESULTS: The mean (+/-SD) genome-equivalent translocation frequency (F(G)) was 2.30 (+/-0.75)% in the zero-dose group (n = 7), and Poisson regression analysis provided the best-fit equation of F(G)(%) = 2.96(+/-0.39) + 0.69(+/-0.14)D + 0.12(+/-0.05)A, where D is the film-badge-derived dose (Gy), and A is age centred at 67 years. The induction rate would increase to nearly 1% Gy(-1) if the radiation dose to bone marrow, one of the major organs for lymphocytes and where their precursor cells reside, is considered. CONCLUSION: The estimated induction rate in vivo appeared substantially smaller than linear coefficients estimated from various in vitro studies.

Temporal changes in liver cancer incidence rates in Japan: accounting for death certificate inaccuracies and improving diagnostic techniques.

Primary liver cancer (PLC) rates have risen dramatically during the past few decades in some regions, particularly in Japan, where PLC is now the third major cause of cancer death. PLC is one of the most difficult tumors to diagnose correctly, because (i) the liver is a frequent site of cancer metastasis and (ii) death from PLC is often attributed to cirrhosis or chronic hepatitis. Also, because the disease is often rapidly fatal, a large proportion of liver cancer cases are identified based on death certificates alone without confirmation by clinical records. Thus, worldwide differences in published incidence rates for this disease reflect regional or national differences in both the accuracy of death certificates and the sensitivity of diagnostic methods. By comparing death certificate causes of death with those based on pathology review, we were able to adjust 1958--1994 incidence rates for a large Japanese cohort for these errors. Although the death certificate false-positive error rate declined, the false-negative error rate remained high throughout the study. The introduction of improved liver cancer diagnostic methods in Japan in the early 1980s was associated with a sharp increase in PLC incidence. We conclude that errors in death certificate causes of death and changes in liver cancer diagnostic techniques have had an important impact on the reported incidence of this disease. Taking these factors into account, rates of hepatocellular carcinoma rose between 2.4- and 4.3-fold in our Japanese cohort from 1960 to 1985, peaked about 1993 and declined thereafter. Incidence rates of cholangiocarcinoma remained stable through 1987.

Impact of comparison group on cohort dose response regression: an example using risk estimation in atomic-bomb survivors.

Cohort-based dose-response analyses can be biased if based on a comparison group that is not comparable to the exposed persons with respect to uncontrolled factors related to disease incidence or mortality. When data exist over a range of doses including the very low dose region, internal regression standardized analyses based on the regression intercept derived from the exposed subcohort alone can provide risk estimates that are not subject to such comparison-group bias. In the Life Span Study cohort of atomic-bomb survivors, persons with dose estimates of zero comprise a broader geographic distribution than that of persons with non-zero dose estimates. Because there is geographic variation in mortality rates, the zero-dose persons might bias background rate estimates thereby affecting inference about radiation risk. This is illustrated using mortality due to all causes. Restricting the comparison group to certain geographically defined subcohorts resulted in as much as a 6% increase or 8% decrease in the risk estimate. This bias can be corrected using an SMR-type estimate in the regression model, allowing retention of the comparison group in the analysis if it is needed for stability or precision in estimating age, time, and sex effects. Consideration of heterogeneity in comparison groups is particularly important in dose-response studies focused on low doses at which the response may be comparable in magnitude to such heterogeneity.

Chromosomal instability in BRCA1- or BRCA2-defective human cancer cells detected by spontaneous micronucleus assay.

The BRCA1 and BRCA2 gene products are believed to play an important part in the onset and/or development of many sporadic mammary cancers. Recently, it has been reported that these two proteins contribute to a centrosome function which is believed to help maintain the integrity of the chromosome segregation process. This may mean a reduced level of the BRCA1 or BRCA2 protein in mammary cells will occasionally lead to nondisjunctional chromosomal loss or gain. We now report that spontaneous micronuclei arising from chromosome(s) which fail to be incorporated into the relevant daughter nuclei during mitosis tend to occur more frequently in BRCA1- or BRCA2-defective human cancer cells than in BRCA-positive cancer cells. Some cases of mammary carcinogenesis may therefore stem from the loss of integrity of chromosome segregation in cells which have a reduced capacity to express either BRCA1 or BRCA2.

Longevity of atomic-bomb survivors.

BACKGROUND: Conflicting claims have been made regarding biological and health consequences of exposure to low doses of radiation. Studies have suggested that certain low-dose exposed atomic-bomb survivors live longer than their peers. Earlier studies in other radiation-exposed populations demonstrated life shortening from mortality from cancer but lacked dosimetry and relied on comparison groups which may introduce bias because of lack of comparability. We have re-examined the effect of radiation on life expectancy in one cohort of survivors of the atomic bombings of Hiroshima and Nagasaki, Japan. METHODS: We did a prospective cohort study of 120,321 survivors. The study encompasses 45 years of mortality follow-up with radiation-dose estimates available for most cohort members. We calculated relative mortality rates and survival distribution using internal comparison (cohort-based estimation of background mortality). FINDINGS: Median life expectancy decreased with increasing radiation dose at a rate of about 1.3 years per Gy, but declined more rapidly at high doses. Median loss of life among cohort members with estimated doses below 1 Gy was about 2 months, but among the small number of cohort members with estimated doses of 1 Gy or more it was 2.6 years. Median loss of life among all individuals with greater-than-zero dose estimates was about 4 months. INTERPRETATION: These results are important in light of the recent finding that radiation significantly increases mortality rates for causes other than cancer. The results do not support claims that survivors exposed to certain doses of radiation live longer than comparable unexposed individuals. Because the cohort was intentionally constructed to contain a higher proportion of high-dose atomic-bomb survivors, average loss of life among all exposed atomic-bomb survivors would be less than the 4 months found for the study cohort.

Modelling haemopoietic stem cell division by analysis of mutant red cells.

Data describing the number of human red cells mutated at the glycophorin A locus, measured flow cytometrically, are reported for 752 adults and 49 neonates. The variance increases with age more rapidly than the approximately linear increase in mean. It is postulated that this discrepancy is explained by the known property of asymmetric stem cell division, so that the division of a single mutant stem cell may result in zero, one or two progeny stem cells. A mathematical analysis allows description of this process with three parameters: stem cell number, mean division rate and mutation rate per division. The values of these parameters can not be deduced from the data presented here. However, estimates of either stem cell number or mutation rate from other sources enable deduction of the two other parameters. The mean number of divisions per stem cell per lifetime was estimated to be about 70. This analysis therefore implies that the rate at which blood cell telomeres shorten with age acts as a direct measure of stem cell turnover. Furthermore, it is argued that this low figure implies that mutations occurring during early life, including organogenesis, are relatively important in initiating stem cell-derived malignancy. Finally, the number of human stem cell divisions per lifetime is similar to shorter-lived mammals, suggesting this number is important in the ageing process.

Effects of radiation on incidence of primary liver cancer among atomic bomb survivors.

We describe the radiation risk for primary liver cancers between 1958 and 1987 in a cohort of atomic bomb survivors in Hiroshima and Nagasaki, Japan. The analysis is based on a comprehensive pathology review of known or suspected liver neoplasms that generated 518 incident, first primary cases, mostly hepatocellular carcinoma. Excess relative risk from atomic bomb radiation was linear: 0.81 per sievert weighted liver dose (95% CI [0.32, 1.43]; P < 0.001). Males and females had similar relative risk so that, given a threefold higher background incidence in males, the radiation-related excess incidence was substantially higher in males. Excess risk peaked for those with age at exposure in the early 20s; there was essentially no excess risk in those exposed before age 10 or after age 45. Whether this was due to a difference in sensitivity or possible confounding by other factors could not be addressed retrospectively in the full cohort. A paucity of cholangiocarcinoma and hemangiosarcoma cases suggested that they are not significantly associated with whole-body radiation exposure, as they are with the internal alpha-particle-emitting radiological contrast medium Thorotrast. Because most of the radiation-related excess cases occurred among males, it is important to ascertain what factors put men at greater risk of radiation-related liver cancer.

Statistical issues in biological radiation dosimetry for risk assessment using stable chromosome aberrations.

Biological dosimeters are useful for epidemiologic risk assessment in populations exposed to catastrophic nuclear events and as a means of validating physical dosimetry in radiation workers. Application requires knowledge of the magnitude of uncertainty in the biological dose estimates and an understanding of potential statistical pitfalls arising from their use. This paper describes the statistical aspects of biological dosimetry in general and presents a detailed analysis in the specific case of dosimetry for risk assessment using stable chromosome aberration frequency. Biological dose estimates may be obtained from a dose-response curve, but negative estimates can result and adjustment must be made for regression bias due to imprecise estimation when the estimates are used in regression analyses. Posterior-mean estimates, derived as the mean of the distribution of true doses compatible with a given value of the biological endpoint, have several desirable properties: they are nonnegative, less sensitive to extreme skewness in the true dose distribution, and implicitly adjusted to avoid regression bias. The methods necessitate approximating the true-dose distribution in the population in which biological dosimetry is being applied, which calls for careful consideration of this distribution through other information. An important question addressed here is to what extent the methods are robust to misspecification of this distribution, because in many applications of biological dosimetry it cannot be characterized well. The findings suggest that dosimetry based solely on stable chromosome aberration frequency may be useful for population-based risk assessment.

Lifespan of human memory T-cells in the absence of T-cell receptor expression.

To evaluate the intrinsic lifespan of human memory T-cells in the absence of T-cell receptor signaling, we used radiation-induced mutant CD4+ T-cells lacking surface expression of TCR/CD3 complex as an in vivo cell marker. We analyzed the long-term kinetics of TCR/CD3 - mutant T-cells among CD4+ CD45RA+ naive and CD4+ CD45RA- memory T-cell fractions in peripheral blood of gynecological cancer patients receiving radiotherapy. Both the proportion and number of these mutant T-cells decayed exponentially with time following radiotherapy. The estimated half-life of mutant memory T-cells was 2 to 3 years and did not differ from that of mutant naive T-cells. These results indicate that the lifespan of mature CD4+ T-cells is limited regardless of their memory or naive phenotype in the absence of TCR/CD3 expression. This finding may suggest that continued T-cell receptor signaling is required for lifetime maintenance of human memory T-cells.

Risk factors for primary breast cancer in Japan: 8-year follow-up of atomic bomb survivors.

BACKGROUND: Findings from the Life Span Study (LSS) of the health effects of exposure to atomic bomb radiation have documented a strong dose-response relation between radiation exposure and breast cancer incidence. PURPOSE: We analyzed data from the LSS cohort to identify nonradiation risk factors for breast cancer and to determine whether these factors were independent of the effects of radiation on breast cancer occurrence. METHODS: Breast cancer incidence was ascertained among a cohort of 22,200 residents of Hiroshima and Nagasaki, Japan, who had completed a mail survey between 1979 and 1981 to study nonradiation risk factors for disease. During the subsequent follow-up period (average 8.31 years), 161 cases of primary breast cancer were identified through population-based tumor registries in the two cities. RESULTS: The risk of breast cancer was inversely related to age at menarche and weakly positive in relation to age at menopause and years of menstruation. A significant negative association of full-term pregnancy against breast cancer was observed, although the number of pregnancies beyond the first was not related to the rate of breast cancer in the cohort. Women having their first full-term pregnancy before age 30 were at decreased risk of breast cancer relative to older women, but there was no trend. A nonsignificant, positive trend in risk was associated with increasing weight and body mass (kg/m2). The risk of breast cancer among women with a history of estrogen use was 1.64 (95% confidence interval 1.02-2.64) and with diabetes 2.06 (95% confidence interval 1.27-3.34). It was not possible to distinguish among additive and multiplicative models of the joint association of radiation dose and various non-radiation-related exposures (age at menarche, full-term pregnancy, female hormone preparations) identified in this analysis. CONCLUSIONS: Nonradiation risk factors for breast cancer among Japanese atomic bomb survivors were consistent with those identified among other populations of women, although the prevalence of common risk factors was low. Reproductive factors and hormone use appear to act independently of radiation exposure on the risk of breast cancer among this population.

Somatic cell mutations at the glycophorin A locus in erythrocytes of atomic bomb survivors: implications for radiation carcinogenesis.

To clarify the relationship between somatic cell mutations and radiation exposure, the frequency of hemizygous mutant erythrocytes at the glycophorin A (GPA) locus was measured by flow cytometry for 1,226 heterozygous atomic bomb (A-bomb) survivors in Hiroshima and Nagasaki. For statistical analysis, both GPA mutant frequency and radiation dose were log-transformed to normalize skewed distributions of these variables. The GPA mutant frequency increased slightly but significantly with age at testing and with the number of cigarettes smoked. Also, mutant frequency was significantly higher in males than in females even with adjustment for smoking and was higher in Hiroshima than in Nagasaki. These characteristics of background GPA mutant frequency are qualitatively similar to those of background solid cancer incidence or mortality obtained from previous epidemiological studies of survivors. An analysis of the mutant frequency dose response using a descriptive model showed that the doubling dose is about 1.20 Sv [95% confidence interval (CI): 0.95-1.56], whereas the minimum dose for detecting a significant increase in mutant frequency is about 0.24 Sv (95% CI: 0.041-0.51). No significant effects of sex, city or age at the time of exposure on the dose response were detected. Interestingly, the doubling dose of the GPA mutant frequency was similar to that of solid cancer incidence in A-bomb survivors. This observation is in line with the hypothesis that radiation-induced somatic cell mutations are the major cause of excess cancer risk after radiation exposure. Furthermore, the dose response was significantly higher in persons previously or subsequently diagnosed with cancer than in cancer-free individuals. This may suggest an earlier onset of cancer due to elevated mutant frequency or a higher radiation sensitivity in the cancer group, although the possibility of dosimetry errors should be considered. The findings obtained in the present study suggest that the GPA mutant frequency may reflect the cancer risk among people exposed to radiation.

Monitoring exposure to atomic bomb radiation by somatic mutation.

Atomic bomb survivors are a population suitable for studying the relationship between somatic mutation and cancer risk because their exposure doses are relatively well known and their dose responses in terms of cancer risk have also been thoroughly studied. An analysis has been made of erythrocyte glycophorin A (GPA) gene mutations in 1,226 atomic bomb survivors in Hiroshima and Nagasaki. The GPA mutation frequency (Mf) increased slightly but significantly with age at the time of measurement and with the number of cigarettes smoked. After adjustment for the effect of smoking, the Mf was significantly higher in males than in females and higher in Hiroshima than in Nagasaki. All of these characteristics of the background GPA Mf were in accord with those of solid tumor incidence obtained from an earlier epidemiological study of A-bomb survivors. Analysis of the dose effect on Mf revealed the doubling dose to be about 1.20 Sv and the minimum dose for detection of a significant increase to be about 0.24 Sv. No significant dose effect for difference in sex, city, or age at the time of bombing was observed. Interestingly, the doubling dose for the GPA Mf approximated that for solid cancer incidence (1.59 Sv). And the minimum dose for detection was not inconsistent with the data for solid cancer incidence. The dose effect was significantly higher in those diagnosed with cancer before or after measurement than in those without a history of cancer. These findings are consistent with the hypothesis that somatic mutations are the main cause of excess cancer risk from radiation exposure.

Optimal case-control matching in practice.

We illustrate modern matching techniques and discuss practical issues in defining the closeness of matching for retrospective case-control designs (in which the pool of subjects already exists when the study commences). We empirically compare matching on a balancing score, analogous to the propensity score for treated/control matching, with matching on a weighted distance measure. Although both methods in principle produce balance between cases and controls in the marginal distributions of the matching covariates, the weighted distance measure provides better balance in practice because the balancing score can be poorly estimated. We emphasize the use of optimal matching based on efficient network algorithms. An illustration is based on the design of a case-control study of hepatitis B virus infection as a possible confounder and/or effect modifier of radiation-related primary liver cancer in atomic bomb survivors.

Effect of radiation and cigarette smoking on expression of FUdR-inducible common fragile sites in human peripheral lymphocytes.

In vitro X-irradiation of human peripheral blood lymphocytes increased the frequencies of fluorodeoxyuridine-induced fragile sites in a dose-related manner. However, the cells from 30 atomic bomb survivors exposed to either high or low radiation doses 47 years earlier showed no demonstrable difference in fragile site expression, indicating that fragile site induction was ephemeral in nature. When fragile sites were analyzed on the basis of tobacco smoking habits, an elevated number was observed in the smokers. The results confirm that fragile sites can be affected by recent exposure to exogenous agents, but the effect is probably of limited duration, based on the atomic bomb survivor experience.

Nonparametric regression analysis of data from the Ames mutagenicity assay.

The Ames assay has received widespread attention from statisticians because of its popularity and importance to risk assessment. However, investigators have yet to routinely apply modern regression methods that have been available for more than a decade. We study yet another approach, the application of nonparametric regression techniques, not as the ultimate solution but rather as a framework within which to address some of the shortcomings of other methods. But nonparametric regression is itself prone to difficulties when applied to Ames assay data, as we show through the use of two examples and some simulation studies. We argue that there remains a great need for further development of statistical methods suitable to the Ames assay. It is hoped that such work can be stimulated and guided by greater collaboration between statisticians and laboratory investigators.

Application of generalized estimating equations to a study of in vitro radiation sensitivity.

We describe an application of the generalized estimating equation (GEE) method (Liang and Zeger, 1986, Biometrika 73, 13-22) for regression analysis of correlated Poisson data from a split-plot design with a small number of experimental units. As an alternative to the use of an arbitrarily chosen working correlation matrix, we demonstrate the use of GEE with a reasonable model for the true covariance structure among repeated observations within individuals. We show that, under such a split-plot design with large clusters, the asymptotic relative efficiency of GEE with simple (independence or exchangeable) working correlation matrices is rather low. We conclude by summarizing issues and needs for further work concerning efficiency of the GEE parameter estimates in practice.

Radiosensitivity of atomic bomb survivors as determined with a micronucleus assay.

If A-bomb survivors include a disproportionately large number of either radioresistant or radiosensitive persons, the surviving population would provide a biased estimate of the true risk of radiogenic cancer. To test this hypothesis, the in vitro X-ray sensitivities of peripheral blood lymphocytes obtained from 937 A-bomb survivors were measured with a cytokinesis-blocking micronucleus assay. Background frequencies (no irradiation in vitro) of micronuclei show a wide distribution. Frequencies in both males and females tend to increase with increasing donor age. Frequencies in females are significantly higher than those in males. Donor age decreases the sensitivity of lymphocytes to in vitro X-ray exposure at a rate of about 0.001 micronuclei per cell per year per gray. There is no effect of donors' sex on in vitro radiation sensitivity. Atomic bomb radiation and cigarette smoking had no significant effect on background and X-ray-induced micronuclei frequencies. Thus there is no difference in radiosensitivity of peripheral blood lymphocytes between proximally and distally exposed survivors.