Nori Nakamura

Genetic alterations in thyroid tumor progression: association with p53 gene mutations.

To identify the genetic events that must be involved in thyroid tumor progression, we initially investigated p53 gene alterations in 10 papillary adenocarcinomas, 4 follicular adenocarcinomas, and 8 undifferentiated carcinomas. Base substitutional mutations in exons 5 to 8 and loss of heterozygosity (LOH) of the p53 gene were not detected in papillary or follicular adenocarcinomas. However, 7 of 8 undifferentiated carcinomas were carrying base substitutional mutations, and LOH was detected in 3 of 5 informative cases. Furthermore, to verify that the p53 gene alterations are truly involved in tumor progression, DNA from individual foci of the four undifferentiated carcinomas coexisting with a differentiated focus and from one follicular adenocarcinoma with an undifferentiated focus was analyzed by direct sequencing and polymerase‐chain‐reaction‐restriction‐fragment‐length polymorphism (PCR‐RFLP). Base substitutional mutations in the p53 gene from exons 5 to 8 were identified exclusively in the undifferentiated foci, but not in the differentiated foci. LOH was observed in 3 of 4 informative undifferentiated foci. In one of these positive cases, LOH was observed in both papillary adenocarcinoma and undifferentiated carcinoma. However, a p53 gene mutation at codon 248 was detected in the undifferentiated carcinoma but not in the papillary adenocarcinoma. The results imply that LOH occurs first in papillary adenocarcinoma followed by a p53 mutation during the transition from papillary adenocarcinoma to undifferentiated carcinoma. Maintenance of LOH during tumor progression excludes the possibility that these different histological foci are derived from different origins and represents molecular evidence that undifferentiated carcinoma is very likely derived from preexisting papillary adenocarcinoma. Furthermore, these results strongly suggest that the mutated p53 gene plays a crucial role in de‐differentiation during the progression of thyroid tumors.

Mutation frequency in human blood cells increases with age

Using either the colony formation assay or flow cytometry, it is feasible to measure the frequency of rare mutant lymphocytes or erythrocytes in human peripheral blood. Accordingly, we have investigated the mutant cell frequencies of the hypoxanthine-guanine phosphoribosyltransferase and T-cell receptor genes in T lymphocytes and of the glycophorin A gene in erythrocytes of several hundred persons aged 0-96 years. The mutant frequency of every one of these genes increased significantly with age. A simple accumulation of mutations in hematopoietic stem cells over time may explain the age-dependent increase in the frequency of glycophorin A mutants. In contrast, a balance between mutant cell generation and loss should be taken into account for the mechanism of the increase of T-cell mutations.

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.

A close correlation between electron spin resonance (ESR) dosimetry from tooth enamel and cytogenetic dosimetry from lymphocytes of Hiroshima atomic-bomb survivors

PURPOSE To estimate gamma-ray doses received by Hiroshima atomic bomb survivors using electron spin resonance (ESR) of tooth enamel and to compare the results with cytogenetic data. MATERIALS AND METHODS Tooth enamel ESR was performed for 100 teeth donated from 69 Hiroshima survivors, and conventional cytogenetic examinations were conducted for 61 of the tooth donors. To evaluate possible contamination from dental X-ray exposure, which primarily affects the tooths buccal surface, each tooth was divided into buccal and lingual parts for subsequent independent enamel isolation and ESR measurement. RESULTS Almost 20 teeth showed considerably larger buccal doses than lingual doses, but most of these discrepant teeth were incisors and canines. The results are probably attributable to solar light exposure. In contrast, the buccal and lingual doses found in molars were similar. Conventional translocation data of lymphocytes and ESR-estimated doses of 40 donors of molars showed the dose-response for translocations to be almost the same as that expected from in vitro gamma-ray irradiation experiments. CONCLUSIONS Both tooth enamel ESR and lymphocyte cytogenetics are useful measures for individual biodosimetry of acute radiation exposure, even half a century after the exposure occurred.

Stable Chromosome Aberrations in Atomic Bomb Survivors: Results from 25 Years of Investigation

Abstract Kodama, Y., Pawel, D., Nakamura, N., Preston, D., Honda, T., Itoh, M., Nakano, M., Ohtaki, K., Funamoto, S. and Awa, A. A. Stable Chromosome Aberrations in Atomic Bomb Survivors: Results from 25 Years of Investigation. Radiat. Res. 156, 337–346 (2001). Frequencies of stable chromosome aberrations from more than 3,000 atomic bomb survivors were used to examine the nature of the radiation dose response. The end point was the proportion of cells with at least one translocation or inversion detected in Giemsa-stained cultures of approximately 100 lymphocytes per person. The statistical methods allow for both imprecision of individual dose estimates and extra-binomial variation. A highly significant and nonlinear dose response was seen. The shape of the dose response was concave upward for doses below 1.5 Sv but exhibited some leveling off at higher doses. This curvature was similar for the two cities, with a crossover dose (i.e. the ratio of the linear coefficient to the quadratic coefficient) of 1.7 Sv (95% CI 0.9, 4). The low-dose slopes for the two cities differed significantly: 6.6% per Sv (95% CI 5.5, 8.4) in Hiroshima and 3.7% (95% CI 2.6, 4.9) in Nagasaki. This difference was reduced considerably, but not eliminated, when the comparison was limited to people who were exposed in houses or tenements. Nagasaki survivors exposed in factories, as well as people in either city who were outside with little or no shielding, had a lower dose response than those exposed in houses. This suggests that doses for Nagasaki factory worker survivors may be overestimated by the DS86, apparently by about 60%. Even though factory workers constitute about 20% of Nagasaki survivors with dose estimates in the range of 0.5 to 2 Sv, calculations indicate that the dosimetry problems for these people have little impact on cancer risk estimates for Nagasaki.

Frequency of mutant T lymphocytes defective in the expression of the T-cell antigen receptor gene among radiation-exposed people

The frequency of mutant T lymphocytes defective in T-cell receptor gene (alpha or beta) expression was measured using the 2-color flow cytometric technique. Results for a total of 203 atomic bomb survivors, 78 of whom were proximally exposed (DS86 doses of greater than or equal to 1.5 Gy) and 125 of whom were distally exposed (DS86 dose of less than 0.005 Gy), showed that the mutant frequency was significantly higher in males than in females. No significant dose effects were observed. In contrast, a significant increase of mutant frequency was observed for 6 patients treated with Thorotrast, a contrast medium containing thorium-232 formerly used for radioligands. In addition, thyroid disease patients treated with 131I showed a dose-related increase of mutant frequency. It was suggested that the present T-cell receptor mutation assay has a unique characteristic as a biological dosimeter for measurement of recent exposures to genotoxic agents.

Strong static magnetic field and the induction of mutations through elevated production of reactive oxygen species in Escherichia coli soxR.

Purpose : Although strong static magnetic fields (SMF) are supposed to have the potential to affect biological systems, the effects have not been evaluated sufficiently. Experiments should be performed with a powerful SMF-generating apparatus to evaluate the biological effects of SMF. Materials and methods : An Escherichia coli mutation assay was used to assess the mutagenic effects of strong SMF. Various mutant strains of E. coli were exposed to up to 9 Tesla (T) for 24 h and the frequencies of rifampicin-resistant mutations were then determined. The expression of the soxS::lacZ fusion gene was assessed by measurement of β-galactosidase activity. Results : The results for survival or mutation were obtained with wild-type E. coli strain GC4468 and its derivatives defective in DNA repair enzymes or redox-regulating enzymes were all negative. On the other hand, the mutation frequency was significantly increased by the SMF exposure in soxR and sodAsodB mutants, which are defective in defence mechanisms against oxidative stress. Furthermore, the expression of superoxide-inducible soxS::lacZ fusion gene was stimulated 1.4- and 1.8-fold in E. coli when exposed to 5 and 9 T, respectively. Conclusions : These results indicate that strong SMF induce mutations through elevated production of intracellular superoxide radicals in E. coli.

Absence of Correlations between Radiosensitivities of Human T-Lymphocytes in G0 and Skin Fibroblasts in Log Phase

Dose-survival curves were obtained for matched samples of peripheral T-lymphocytes and skin fibroblasts from a total of 22 patients who underwent various surgical procedures using loss of colony-forming ability as the end point. The results showed that the mean D10 (dose required to kill 90% of cells) +/- SD was 3.58 +/- 0.21 Gy for T-lymphocytes irradiated in G0 and 3.19 +/- 0.37 Gy for skin fibroblasts irradiated in log phase. The coefficients of variation were found to be 6 and 11%, respectively. Contrary to the expectation, regression analysis of D10 values for the two types of cells revealed no significant correlations. The absence of correlation most probably derives from the fact that the apparent interindividual variability of dose-survival curves is caused primarily by random experimental fluctuations at least in the case of lymphocytes. Possible reasons for the greater variability observed in the fibroblast assay are discussed.

Human Fetuses do not Register Chromosome Damage Inflicted by Radiation Exposure in Lymphoid Precursor Cells except for a Small but Significant Effect at Low Doses

Abstract Ohtaki, K., Kodama, Y., Nakano, M., Itoh, M., Awa, A. A., Cologne, J. and Nakamura, N. Human Fetuses do not Register Chromosome Damage Inflicted by Radiation Exposure in Lymphoid Precursor Cells except for a Small but Significant Effect at Low Doses. Radiat. Res. 161, 373–379 (2004). Human fetuses are thought to be highly sensitive to radiation exposure because diagnostic low-dose X rays have been suggested to increase the risk of childhood leukemia. However, animal studies generally have not demonstrated a high radiosensitivity of fetuses, and the underlying causes for the discrepancy remain unidentified. We examined atomic bomb survivors exposed in utero for translocation frequencies in blood lymphocytes at 40 years of age. Contrary to our expectation of a greater radiosensitivity in fetuses than in adults, the frequency did not increase with dose except for a small increase (less than 1%) at doses below 0.1 Sv, which was statistically significant. We interpret the results as indicating that fetal lymphoid precursor cells comprise two subpopulations. One is small in number, sensitive to the induction of both translocations and cell killing, but rapidly diminishing above 50 mSv. The other is the major fraction but is insensitive to registering damage expressed as chromosome aberrations. Our results provide a biological basis for resolving the long-standing controversy that a substantial risk of childhood leukemia is implicated in human fetuses exposed to low-dose X rays whereas animal studies involving mainly high-dose exposures generally do not confirm it.

Early-onset breast cancer in A-bomb survivors

1 Benowitz NL, Pharmacologic aspects of cigarette smoking and nicotine addiction. N Engl J Med 1988; 319: 1318-30. 2 Wooten MR, Khangure MS, Murphy MJ. Intracerebral haemorrhage and vasculitis related to ephedrine abuse. Ann Neurol 1983; 3: 337-40. 3 Kaye BR, Fainstst M. Cerebral vasculitis associated with cocaine abuse. JAMA 1987; 258: 2104-06. 4 Glick R, Hoying J, Cerullo L, Perlman S. Phenylpropanolamine: an over-the-counter drug causing central nervous system vasculitis and intracerebral haemorrhage. Case report and review. Neurosurgery 1987; 20: 969-74. 5 Maouad J, Fernandez F, Barrillon A, Gerbaux A, Gay J. Diffuse or segmental narrowing (spasm) of the coronary arteries during smoking demonstrated on angiography. Am J Cardiol 1984; 53: 354-55.