RERF Report No.9-15

Paper published in Radiation Research§“Irradiation at different fetal stages results in different translocation frequencies
in adult mouse thyroid cells”

Kanya Hamasaki, Reid D. Landes, Asao Noda, Nori Nakamura, Yoshiaki Kodama
Radiat Res 2016 (October); 186(4):360-6(doi: 10.1667/RR14385.1)

Study Findings
We examined chromosome aberration frequencies in thyroid tissues of adult mice
irradiated during the fetal stage and found that radiation effects varied depending on thefetal stage at the time of irradiation. While mice irradiated as 6.5-day fetuses (beforeorganogenesis) had a relatively low chromosome aberration frequency (5.8×10-3), adultmice irradiated as 15.5-day fetuses (during the fetal stage) showed frequencies that were
nearly the same as that of their mothers’ (25.3×10-3). Taking into account findings fromprevious studies, we interpreted these results as an indication that fetal tissue stem cells
record radiation damage primarily when exposed after locating in their niche*; before
locating in their niche, however, radiation-damaged cells may undergo negative selection
and be unable to locate in the niche, and therefore the effects of radiation damage may
not remain.
Niche*: Microenvironment in which tissue stem cells can function.
Explanations
Based on findings from epidemiological studies (case-control studies) starting in the1950s, fetuses have been considered highly sensitive to cancer risks from radiation.
However, RERF’s study using chromosome aberration frequency in lymphocytes as amarker revealed that few effects of radiation were observed among in-utero survivors.
The present study investigating chromosome aberrations in tissues of mice irradiated as
fetuses may provide a clue for understanding these apparently contradictory observation
results.
1. Study Purpose
To verify differences in fetal irradiation-related chromosome aberrations among
tissues of different types, we previously studied hematopoietic cells in mice andmammary gland epithelial cells in rats. In this study, we examined chromosome
aberration frequencies in mouse thyroid epithelial cells. We also conducted experiments
using irradiation at different stages of fetal development to determine whether or notchromosome aberration frequencies following fetal irradiation differ by when irradiation
occurs.
2. Study Methods
Pregnant female mice were irradiated with 2 Gy of X-rays at day 6.5 or 15.5 afterconception. After the neonatal mice reached adulthood (at least eight weeks of age), their
thyroid glands and those of their mothers were removed. We cultured thyroid epithelial
cells for five or six days and prepared chromosome samples using conventional methods.
To detect stable chromosome aberrations, such as translocation, occurring in thyroid cells
due to fetal irradiation, we employed fluorescence in situ hybridization (FISH),* whichlabels chromosomes 1 and 3 in green and red, respectively. We then observed slides oflabeled chromosomes and counted the number of translocations and other stable
chromosome aberrations. In a portion of the mice, spleen T lymphocytes were also
cultured and assessed for chromosome analysis.

RERF Report No.9-15*FISH (fluorescence in situ hybridization) is a staining technique. To determine thepresence or absence of structural abnormality in a target chromosome (chromosomes 1and 3 in this study), DNA is extracted from a library of the target chromosome andlabeled with fluorescence to prepare a probe. (A library contains DNA extracted fromindividual chromosomes and amplified in Escherichia coli.) Following preparation of
single-strand DNA from double-strand DNA of the target chromosome sample(denaturalization), a reaction is prompted with the aforementioned probe(hybridization), making it possible to label a specific DNA sequence artificially.
3. Study Results(1) Adult mice irradiated with 2 Gy of X-rays as 15.5-day fetuses had a highertranslocation frequency (30/1155 or 25.3×10-3) in thyroid cells than did non-irradiatedadult controls (0/1007 or 0.1×10-3), a figure that was nearly the same as thatexperienced by irradiated mothers and other adult females (43/1244 or 33.6×10-3). These
results are consistent with previously observed results in fetally irradiated rat
mammary cells.
(2) When fetuses were exposed to 2 Gy at an earlier stage of development (day 6.5) before
thyroid organogenesis, however, the resulting translocation frequency was much lower(3/502 or 5.8×10-3) than that in the mice irradiated at day 15.5 (30/1155 or 25.3×10-3).
These results reveal that fetal radiation sensitivity differs by timing of irradiation, andthat the effects of thyroid irradiation prior to thyroid organogenesis are insignificant.
On the other hand, few chromosome aberrations were observed in hematopoietic tissues
regardless of the timing of irradiation (it is understood that hematopoietic stem cells
locate in their niches only after birth).
Study SignificanceThis study revealed that chromosome aberration frequencies in mouse thyroid cells
due to fetal irradiation differed by timing of irradiation (prior to organogenesis/during thefetal stage). Such differences may be partly attributed to whether or not irradiated tissue
stem cells have located in their niche. If tissue stem cells are irradiated before locating inthis way, aberrant cells cannot locate in their niche; when such cells are irradiated after
locating in their niches, however, stem cells may repair DNA damage with the aim ofsurvival, and thus cause mis-repairs of DNA that lead to chromosome aberrations. Thisstudy’s results will contribute to understanding of the mechanisms behind cancer risks
due to fetal radiation exposure.
The Radiation Effects Research Foundation has studied A-bomb survivors and their offspring inHiroshima and Nagasaki for around 70 years. RERF’s research achievements are considered theprincipal scientific basis for radiation risk assessment by the United Nations Scientific Committee
on the Effects of Atomic Radiation (UNSCEAR) and for recommendations regarding radiation
protection standards by the International Commission on Radiological Protection (ICRP). RERF
expresses its profound gratitude to the A-bomb survivors and survivors’ offspring for theircooperation in our studies.

§Radiation Research, which is an official monthly journal of the Radiation Research Society, publishes
original peer-reviewed papers and review articles on radiation effects and related issues in the fields ofphysics, chemistry, biology, and medicine. (Impact factor in 2015: 2.67)