Trans-generational effects of radiation exposure from the Chernobyl Nuclear
Plant Accident: A review of studies using mutation markers of repeat DNA
sequences
FURITSU Katsumi
Department of Genetics, Hyogo Collage of Medicine,
Mukogawa-cho 1-1, Nishinomiya, Hyogo 663-8131, Japan

Introduction
The assessment of the possible radiation effects on the children who were conceived after the
Chernobyl accident is needed in order to estimate the long-term health effects of the accident, including
the effects on future generations. If germline cells were even exposed at the stage before conception, thehealth effect of children could be “trans-generational”. Additionally, the health effects of children due toexposure to radiation might also be caused by exposure at successive stages after conception including theembryonic and fetal stages.
We already have definite evidence of the trans-generational effect of radiation from experimental
animals. However, it has not yet been demonstrated in humans. The Radiation Effect Research Foundation(RERF) reported, “Extensive studies of the children of survivors of the atomic bombings in Hiroshima andNagasaki have thus far yielded no statistically significant increases in genetic effects compared to control
populations.” They have done an on-going epidemiological study collecting mortality from all causes, andhave performed molecular studies of DNA, as well as conducting a clinical health study investigating thefrequency and prevalence of adult-onset life-style-related diseases.1) Under such circumstances, we have
only the genetic risk assessment of radiation exposure, which was estimated from the data of spontaneous
mutation rates in human Mendelian diseases and chronic multi-factorial diseases, and the data of theradiation-induced germline mutations of experimental mice.2) Studying the possible trans-generational
effects induced by radiation exposure from the Chernobyl nuclear power plant accident is important inorder to discuss the reassessment of the genetic risk of radiation. People who were exposed to theradiation from the Chernobyl accident form another large population, other than the atomic survivors ofHiroshima and Nagasaki where the radiation exposure was quite different.
This review focuses on the studies using mutation markers of repeat DNA sequences which isrelevant to the radiation exposures that resulted from the Chernobyl accident.
Repeat DNA sequences as markers for detection of “trans-generational effects”
The choice of mutation markers is one of the critical issues in the studies of the trans-generational
effects of mutagens including radiation. The markers should be sensitive and be able to effectively detectmutations, and the system should be capable of obtaining quantitative results of the dose-effect.
Spontaneous mutation rates of protein-coding genes are at most in the order of 10-5-10-6 per generation.
Trying to detect trans-generational effects using markers of these coding genes requires very large numberof samples. Therefore, it is not realistic to expect “significant results” using these markers in a population
which is actually available for study.
There are considerable numbers of sites with repeat DNA sequences, multiple repeats of a certain
sequence of bases (a repeat unit), in the genomes of various living things including human beings. The
different numbers of repeat units at a site in the genomes of individuals of the same species may lead to- 165 -

polymorphic phenotypes. Most of the repeat sequences are in non-coding regions. These are sites, which
can spontaneously mutate in high frequencies per generation. These sites can be used as markers to detect
the genetic effects of mutagens in a limited-size population. In studies of the trans-generational effects ofradiation, the results of research using markers of hypervariable minisatellites and microsatellites havebeen reported in human studies, as well as the results of experiments using markers of Expanded Simple
Tandem Repeat (ESTR) in mice have also been reported.
Hypervariable minisatellites in humans are composed of varying numbers of GC-rich core repeats of
10 - 100 base pairs (bp). The core repeats within a stretch of minisatellite are related but are not identical.
The total number of base pairs in a minisatellite is up to a few tens of thousands. There are more than ten
thousand minisatellite loci in the human genome and most of them are situated close to telomeres. Someminisatellites exhibit spontaneous instability in germ cells with mutation frequencies as high as 10-1 - 10-2
per gamete, however such high mutation rates can not be seen in somatic cell lines. Recombination andgene conversion at minisatellite loci during meiosis result in complex changes in the loci, while
recombination and slippage during mitosis lead to minisatellite instability of somatic cells. These different
mechanisms for mutations are considered to be one of the reasons for such high instability found only in
the germline.3)
Microsatellites in humans are composed of repeat units of less than 10 bp and the total length of a
microsatellite is usually less than 100 bp. There are more than a hundred thousand microsatellites in thehuman genome and they are dispersed widely throughout the genome. The spontaneous mutation ratevaries in each locus and is 10-2 - 10-4 per generation. It is considered that replication slippage leads tomicrosatellite mutations. A higher mutation rate was reported in paternal loci compared to maternal loci.
Larger numbers of mitotic germline cell divisions in spermatogenesis than in oogenesis might be one
possible reason for the difference in mutation rates between paternal and maternal germlines.4)
ESTRs of mice were at first cloned for their sequence homology to human minisatellites and they
were called “minisatellites of mice” at first. However, their repeat units are only 5 - 6 bp and there is novariation of repeat units within a locus. Because of these structural differences between human and mouseminisatellites, these repeat sequences in mice were renamed ESTRs.5) Replication slippage is thought tocause the instability of ESTR although other mechanisms are possible.3)
Human experimental somatic cells showed a dose-dependent increase in mutation rates atminisatellite and microsatellite loci, when they were exposed to radiation.6) As for human germ cells, noincrease was detected in the minisatellite mutation rate of sperm from volunteer patients who were given
radiotherapy for seminoma.7) There is no similar research examining the radiation effects on the humanmicrosatellite mutations of germline cells using samples from volunteer patients.
Induction of germline mutations of ESTRs in male mice by acute gamma irradiation was reported
independently by two research groups.8, 9) The observed mutation frequency induced by irradiation was
two orders of magnitude higher than the frequency which could be expected from the assumption that the
DNA double strand break at the ESTR locus would cause the mutation. This suggests that ESTR locimight not be a direct target of radiation, but rather radiation might induce an indirect mechanism of some
sort that would increase genetic instability in a cell.3) Further research is needed to clarify this mechanism.
As for the most sensitive stage during spermatogenesis, different results were reported from these two
research groups. Niwa’s group reported that the late spermatid stage of post-meiosis was most sensitive toradiation with regard to ESTR mutations.9) This result was compatible with the findings previously
obtained from the experiments of the specific recessive loci in mice germline. On the contrary, Dubrova’s- 166 -

group reported that the pre-meiotic stages of spermatogenesis were sensitive.10)
Research on children living in contaminated areas of Chernobyl
In 1996, ten years after the Chernobyl accident, Dubrova et al. reported the results of studies on
residents in the Chernobyl contaminated areas in Belarus, which showed that “the frequency of mutation
was found to be twice as high in the exposed families as in the control group.”11) This research drew theattention of the scientific community as it was the first “evidence” that radiation could increase germline
mutations in human beings. In 1997, they published another report which included additional subjects andminisatellite probes to reinforce the results of the first report.12) However, these reports were criticizedbecause they used samples of Caucasian families in the UK as the non-exposed control group which wereof different ethnic origins than the Belarusian exposed families.13) Therefore, they conducted further
research on the residents in the contaminated areas in Ukraine. In this study, they compared the children
who were conceived before and after the accident, so that they could investigate the exposed subjects andunexposed controls from the same ethnic background. They analyzed the paternal and maternal mutation
rates using eight single-locus hypervariable minisatellite probes and reported that “a statisticallysignificant 1.6-fold increase in mutation rate was found in the germline of exposed fathers, whereas thematernal germline mutation rate in the exposed families was not elevated.”14)
Dubrova et al. also investigated the populations around the Semipalatinsk nuclear test site in
Kazakhstan, as well as exposed populations in rural villages along the Techa River, which were
contaminated by radioactive releases from the Mayak plutonium separation combine. They used the sameeight hypervariable minisatellite probes as mutation markers and again reported elevated minisatellite
mutation rates in the exposed populations.15, 16)
However, the results reported from Dubrova’s group were contrary to the findings of other groups
which had also investigated human populations using minisatellite markers. Significant elevations of theminisatellite mutation rates were not detected in the reports of the study using sperm samples of volunteer
patients whose testicles had been irradiated by radiotherapy for treatment of seminoma,7) in the research offamilies of atomic bomb survivors in Hiroshima and Nagasaki,17-19) or in the study of the families ofChernobyl clean-up workers who were called “liquidators”.20-22) The reasons for this discrepancy are as yet
unknown. Only the research group of Dubrova investigated the exposed residents in the contaminatedareas. The residents in the contaminated areas were chronically exposed to low-dose-rates of radiation,
while atomic bomb survivors and the Chernobyl liquidators were mainly exposed to high-dose-rates ofradiation for certain limited periods. These differences in type of radiation exposure might be the reason
for the discrepancy. In the case of the residents in the contaminated areas, exposure might not be limited toa specific stage of germline maturation, but could be a successive irradiation throughout the whole processof germline maturation and it could even include the stages after conception.
One of the backgrounds of these studies using minisatellite markers to detect the radiation effects onhuman germline was from previous studies using experimental mice. Mutation rates of ESTRs (these lociwere formerly called “minisatellites of mice”) of the experimental mice were significantly increased byacute gamma irradiation.8, 9) As mentioned above, because of the structural differences between humanminisatellites and “minisatellites of mice”, the latter were renamed ESTRs and different mechanisms are
considered to cause mutations at these loci in mice.5) Radiation may possibly cause changes that increase
the genetic instability in cells, which could lead to mutations of ESTRs in mice. Considering possible
different mechanisms causing mutations, the experimental model of ESTR in mice cannot be directly
applied to solve questions related to human minisatellite mutations. Further research for the mechanism of- 167 -

human minisatellite mutations is necessary before minisatellites will be used more widely as markers todetect and monitor the trans-generational effects of radiation in human populations.
Studies of the children of Chernobyl liquidators
As for the children of Chernobyl liquidators, results of studies using minisatellites20-22) or
microsatellites22, 23) as mutation markers have been reported. There is no study as yet which has
demonstrated “significant effects” with regard to germline mutations in liquidators.
The exposure period was much shorter in liquidators as compared to residents in contaminated areas;in most cases it was a period of up to several months. The radiation dose of individual liquidators varied
widely depending on location and occupation. (However, the information on individual radiation dose wasquite limited and usually external exposure alone was recorded.) In the studies of liquidators, only thepaternal exposure was analyzed because the liquidators were largely men. Children conceived before theaccident or residents in the non-contaminated areas of the same country were used as the non-exposed
control groups.
Kiuru et al. studied the offspring of Estonian liquidators using the mutation markers of eight
hypervariable minisatellite probes which Dubrova had also used in their study of residents. The
minisatellite mutation rate was nonsignificantly increased among children born after the accident (Odds
ratio: 1.33, 95% CI: 0.80-2.20). The rate was higher in the children born to fathers with recorded doses of
0.2 Sv or above (Odds ratio: 3.00, 95% CI: 0.97-9.30) relative to their siblings born before the accident,
although this increase was not statistically significant.21)
Livshits et al. reported that children of Chernobyl workers in Ukraine did not show elevated rates ofmutations in minisatellite alleles. They used the five hypervariable minisatellites as mutation markers
which had also been used by Dubrova. Their control group was made up of residents in non-contaminatedareas in Ukraine. Their results showed as well that the mutation rate of the residents in the contaminatedareas in Belarus (4.94%) was higher than the mutation rate of their controls (3.72%) that were residents of
non-contaminated areas in Ukraine. This was reasonable as it suggested an elevated mutation rate in theresidents in the contaminated areas as compared to those in non-contaminated areas; both groups were ofsimilar ethnic background.20)
The study using microsatellite mutations as markers to investigate the trans-generational effects ofradiation in humans presents a more recent challenge than studies using minisatellites. The protocol forexperiments using microsatellite markers is relatively simple: amplifying a target site including a
microsatellite locus with a pair of primers, one of which is fluorescence-labeled, and analyzing thedifferences in length of PCR (Polymerase Chain Reaction) products using an automatic genetic analyzer.
One of the advantages of this method is higher sensitivity which can detect even a difference in the lengthof a single base in microsatellite loci. However in the experiments using minisatellite markers, thesensitivity is less than this, with bands of different lengths of DNA fragments being analyzed in Southern
Blotting.
However, as is mentioned above, the spontaneous mutation rate of microsatellite loci is one order lessthan that of minisatellite loci. Furitsu and Ryo et al. used as many as 72 microsatellite loci as mutation
markers when they investigated a population of a limited number of liquidator families in Belarus. These
72 microsatellites loci were selected from the loci which had been reported in the literature as having
higher spontaneous mutation rates. Analyzing the mutation of 40 Y-linked mincrosatellite loci, a higher
mutation rate (2.9 x 10-3) was detected in the families of liquidators in Belarus as compared to the- 168 -

mutation rate of the unexposed control group (2.1 x 10-3), although the increase was not statisticallysignificant.23)
There is not yet sufficient basic experimental data on the sensitivity to radiation with regard to
microsatellite mutations in mammalian germlines. Further research on experimental mice and humanpopulations with different radiation doses as well as various types of exposure is necessary.

Studies on living things other than human beings in Chernobyl-contaminated areas
There have been studies of mutations in DNA repeat sequences in barn swallows and wheat plants in
the Chernobyl-contaminated areas. In the study of barn swallows, an increased germline mutation rate wasobserved at the two hypervariable microsatellite loci in populations breeding in Chernobyl- contaminatedareas in Ukraine. European populations of barn swallows migrate across the Sahara desert, wintering insouthern Africa. Ellegren et al. captured adult and nestling barn swallows (as “families”) in Ukraine and
Italy in 1991 and 1996. The mutation rate in the two microsatellite loci was 7.2% (number of meiosis
investigated: 138) for the “families” in the contaminated areas. This rate was significantly higher than themutation rates of control groups in the non-contaminated areas in Ukraine (3.8%, meiosis: 131) and Italy(2.0%, meiosis: 2,002).24)
In the study of wheat plants, two genetically identified populations derived from the same
homogeneous parental line were compared. One population was grown in heavily contaminated plots ofland near the Chernobyl nuclear power plant, the other was sown in a clean (<1 Ci/km2) control area 30
km away with comparable agrochemical characteristics. They analyzed the 13 microsatellite loci in theoffspring plants and estimated that the spontaneous mutation rate was 1.03 x 10-3 per locus (95% IC: 0.44
- 2.03 x 10-3), whereas the mutation rate in the exposed group was 6.63 x 10-3 (95% IC: 4.28 - 9.70 x10-3).25) They also analyzed the complex pattern of microsatellite mutations in the germline of wheat andconcluded that a simple model of replication slippage could not account for mutation events at these lociin wheat.26)
A study on the variation in the hypervariable portion of the mitochondrial control-region haplotypes
in a population of the bank vole in northern Ukraine has been carried out, although it is not a study on therepeat sequences of nuclear DNA. Specimens of the bank vole exhibit the highest internal doses ofradiocesium among small mammals inhabiting the most contaminated sites in Chernobyl. The rate ofvariation in the Chernobyl population was higher than in the control group living in less contaminatedsites. This suggested that exposure to ionizing radiation in the contaminated environments increased thematernal mutation rate and has consequently increased mitochondrial DNA diversity.27) However, thisincreased diversity might be a function of asymmetric migration from a genetically diverse source
population existing before the Chernobyl accident. The longitudinal genetic monitoring project is also
on-going.28)
Plants and small animals usually alternate generations more rapidly than humans and they cannot
intentionally avoid the environmental contamination. Therefore, it is very important to monitor the genetic
changes of these living things in the contaminated areas to estimate the long-term consequences of theChernobyl accident in the whole ecosystem.
Conclusion
In the studies of the children of Chernobyl victims so far, only Dubrova’s study showed a significant
increase in the mutation rate of repeat sequences of DNA, through research done on the residents in the- 169 -

contaminated areas using the markers of minisatellite loci. Other studies investigating the Chernobyl
liquidators did not show any significant increase in the mutation rate even using the same hypervariable
minisatellite probes as mutation markers and the same experimental method as Dubrova’s group. This
discrepancy might be due to the different types of exposure to radiation between residents and liquidators.
Further investigation is required to solve the question: to what extent the increase in mutations in
repeat sequences would reflect in actual human health through its phenotype. Most of the repeat sequencesin the human genome are in non-coding regions. However, some of them are known to cause diseases, orincrease the severity of diseases, as they are in coding or modulator regions in DNA, or near them. The
sensitivity to radiation in the repeat sequences, which relate to certain diseases, should be further
examined and discussed concretely with respect to each site. If some radiation-induced mechanism, whichleads to increased genetic instability in cells, would affect various regions in the genome other than inregions with repeat sequences, it could also influence the coding or modulator regions and induce anumber of deleterious expressions of genes which might lead to the ill health of individuals.
Further research is needed on the mechanism and sensitivity for the induction of mutations throughradiation exposure in DNA repeat sequences in the human germline, in each stage of germ cell maturation
and also in the stages after conception.
As for the studies using microsatellite loci as mutation markers, the studies on barn swallows andwheat plants showed a significant increase in mutation rates, while studies on liquidators have so far failed
to show a significant increase in mutation rates. The residents in the contaminated areas have not yet been
investigated using the mutation markers of microsatellites. Basic data using microsatellite loci inexperimental mice should be obtained. Further research using microsatellite markers with other humanpopulations exposed to radiation in various ways should also be carried out.
In the studies which demonstrated significant increases of mutation rates in DNA repeat sequences,
the observed mutation rate is much higher than the rate which could be expected from the assumption thatthe locus itself is the direct target of radiation and a double strand break at the locus can lead to mutation.
This suggests that the loci might not have been directly targeted by radiation, but radiation might haveinduced some sort of mechanism to increase the genetic instability of a cell. The mechanisms, possibly
including an epigenetic process, might be at work here and they could conceivably be common
mechanisms for other biological effects of radiation, especially at low doses and through chronic
exposure.
As mentioned above, there are still many unsolved problems regarding radiation effects on mutationsof DNA repeat sequences. However, these results, including the significantly elevated mutation rate inmice ESTR, suggest that the genetic instability is actually increased by radiation to the germline.
Therefore, it should not be ignored when we assess the risk of trans-generational effects of radiation. It isespecially important in the genetic risk assessment of low dose and chronic exposure to radiation.
It is critical to carry out a long-range scientific monitoring of the trans-generational effect of the
populations exposed to radiation from the Chernobyl accident. For that purpose, it is vital to establish aninternational research system in cooperation with the researchers in the affected countries.
Not only the environmental contamination by the Chernobyl accident but also other social andeconomic factors have affected the health of people in the Chernobyl areas. This complicated situation
makes it more difficult to investigate the radiation health effects from the Chernobyl accident. It is noteasy to study the radiation health effect separately from other risk factors. The exact radiation dose
estimation is critical in assessing the health effects. However, estimating the exact individual radiation- 170 -

dose from both external and internal exposures can be problematic as it depends on multiple factors suchas individuals’ working conditions, life styles and so on. Therefore, analyzing health effects according todoses received by individuals who may be ill is not a simple process.
Many problems are still unsolved, even though more than 20 years have passed since the Chernobyl
accident. Challenges in the scientific assessment of the various radiation effects of the accident, includingtrans-generational effects, remain with us. However, we should not just wait for the results of further
research, which could provide us with scientific information about radiation, mutations and their relation
to health effects in people, without taking any action right now. We must take action, if we seriously
consider the wellbeing of future generations and remedies for the Chernobyl victims as well as the
problem of nuclear energy in our society.
Table 1 and 2 are the summaries of reports so far on “Trans-generational effects” of humanpopulations using mutation markers of minisatellites and microsatellite, respectively.

Table 1. Reports on the “Trans-generational effects” of human populations using mutation markers
of minisatellites
A. Chernobyl Nuclear Power Plant Accident(1) Dubrova et al. (1996) [11]z Residents in the heavily contaminated rural areas of the Mogilev¾ Exposed groupprovince in Belarus: 79 families with children born between
February and September 1994; both parents had resided
continuously in the contaminated areas.
z Caucasian families in the UK: 105 families; sex was matched with
¾ Non-exposed control
the offspring in the exposed group.
group¾

Estimated radiation dose¾

Minisatellite probes¾

Main results¾

Radiation Effects(2) Dubrova et al. (1997)[12]¾ Exposed group¾

Non-exposed control
group

z The level of Cs137 contamination was 1-15Ci/km2．Individual
radiation dose for external and internal chronic exposure to Cs137
was estimated to be less than 5mSv/year.
z A multi-locus probe: 33.15(MS1, MS31), single-locus probes:
MS32, CEB1．
z The overall mutation rate was 1.97-fold as high in the exposed groupas it was in the control group.
z Number of mutations / Total number of bands in offspring: exposedgroup [49/1615], control group[23/1491]
z Mutation rate was correlated with the level of Cs 137 surface
contamination.
z Mutation spectrum (ratio between male and female germline
mutations, gain or loss of repeat units, size distribution) was similar
in both groups.
z Positivez Residents in the heavily contaminated rural areas of the Mogilev
province in Belarus: 127 families with children born between
February and September 1994 (male: 60, female: 67) ; both parents
had resided continuously in the contaminated areas.（They added
samples to the previous study reported in 1996.）
z Caucasian families in the UK: 120 families; sex ratio of children wasmatched with the offspring in the exposed group (male: 53, female:57).（They added samples to the previous study reported in 1996.）- 171 -

¾

Estimated radiation dose¾

Minisatellite probes¾

Main results¾

Radiation effects

z The mean dose over all families for parental external and internal
chronic exposure to Cs137 was 27.6±3.3mSv.
z Multi-locus probes: 33.15, 33.6, single-locus probes: MS32, CEB1,
B6.7, CEB15, CEB25, CEB36.
z A 1.87-fold increase in the mutation rate was found in the exposedgroup.
z Number of mutations / Total number of bands in offspring: exposedgroup [136/6616], control group [56/5099]
z Mutation rate was correlated with the estimated radiation dose of theparents.
z Mutation spectrum (ratio between male and female germline
mutations, gain or loss of repeat units, size distribution) was similar
in both groups.
z Positive(3) Dubrova et al.(2002)[14]¾

Exposed group¾

Non-exposed control
group

¾

Estimated radiation dose¾

Minisatellite probes¾

Main results¾

Radiation effectsz Ukrainian families inhabiting in the contaminated areas with 240
children conceived after the Chernobyl accident and born between
1987 and 1996; number of families [171+27], 27 families withchildren conceived before and after the accident.
z Ukrainian families inhabiting in the contaminated areas with 98
children conceived before the Chernobyl accident and born between
1976 and 1986; number of families [54+27], 27 families withchildren conceived before and after the accident. Ethnic background,maternal age, parental occupations and number of smokers ofparents were similar for both exposed and control groups. The
paternal age in the exposed group exceeded that for the control
group.
z The level of Cs137 contamination was >2Ci/km2 and estimated dosefrom chromosome aberrations was 0.2-0.4Gy．
z Single-locus probes: B6.7, CEB1, CEB15, CEB25, CEB36, MS1,
MS31, MS32．
z A 1.56-fold increase in the paternal mutation rate was found in the
exposed families.
z Number of mutations / Total number of bands in offspring:∗ Paternal mutation: exposed group [112/178], control group[29/706]∗ Maternal mutation: exposed group [25/1727], control group[10/701]z Paternal: positive, maternal:negative(4) Livshits et al. (2001)[20]¾

Exposed group¾

Non-exposed control
group
Estimated radiation dose¾

z Liquidators families from Kiev in Ukraine; fathers had worked on
the Chernobyl site during the period 1986-1987; 161 families with183 children; 88 children were conceived while the fathers were
working on the Chernobyl site or within 2 months after the fathers
stopped working on the site and 95 children were conceived at least
4 months after the fathers had stopped working on the site.
z Families from southern Ukraine (non-contaminated areas); 163
families with 163 children.
z 0.048 - 1.2 Sv．Data on individual dose estimate were provided for
only 28% of liquidators.

- 172 -

¾

Minisatellite probes¾

Main results¾

Radiation effectsz Single-locus probes: B6.7, CEB1, CEB15, CEB25, CEB36, CEB42,
CEB72．
z Only paternal mutations were analyzed and mutation rates per locus
in the children of liquidators did not differ significantly from thecontrol group．
z Number of mutations / Total number of bands in offspring (analyzing
the result of paternal mutations using the seven single-locus probes):
exposed group [53/1154], control group [51/1036].
z Analyzing the result of mutation rates using the five single-locus
probes, which were common to Dubrova’s group:∗ The mutation rate of the families in the contaminated areas ofBelarus (4.94%, Dubrova’s data) was 1.33-fold higher than themutation rate of the families from non-contaminated areas in
Ukraine (3.72%, Livshits’s data).
∗ The mutation rate of the families from non-contaminated areas in
Ukraine was 1.35-fold higher than the mutation rate of Caucasianfamilies in the UK (2.75%, Dubrova’s data).
z Negative(5) Kiuru et al. (2003)[21]¾

Exposed group¾¾

Non-exposed control
group
Estimated dose of
exposed group
Minisatellite probes¾

Main results¾

Radiation effects¾

z Estonian liquidator families; father had been worked at Chernobyl
over a median period of 3 months between 1986 and 1991; 147
families with 155 children born within 33 months after fathers were
exposed at Chernobyl (post-Chernobyl children).
z The same 147 liquidator families with 148 children born before theChernobyl accident (pre-Chernobyl children).
z Mean radiation dose was 0.11±0.06 Sv with less than 1.4% of thecohort receiving more than 0.25 Sv.
z Single-locus probes: B6.7, CEB1, CEB15, CEB25, CEB36, MS1,
MS31, MS32．
z Mutation rate was higher in post-Chernobyl children than in
pre-Chernobyl children but the elevation was not statisticallysignificant (Odds ratio: 1.33, 95% CI: 0.80-2.20).
z Number of mutations / Total number of bands in offspring: exposedgroup [52/1238], control group [42/1182]
z Post-Chernobyl children of fathers with radiation dose of ≧0.2Sv
had a threefold higher mutation rate (Odds ratio: 3.00, 95% CI:
0.97-9.30).
z Negative (There was some indication in the liquidator exposed group≧0.2Sv.)(6) Slebos et al. (2004)[22]¾

Exposed group¾

Non-exposed control
group

z Liquidator families from Kiev and Chernigov in Ukraine; father had
served in the cleanup operation of the Chernobyl accident between
1986 and 1990; 75 families with 75 children conceived after the startof the cleanup operation (within them, 39 families with 39 children
did not have children before the start of the cleanup operation).
z 41 liquidators families from Kiev and Chernigov in Ukraine, fromthe same registry as the exposed group, with 41 children conceived
before the start of the cleanup operation (within them, 5 families
with 5 children did not have children after the start of the cleanup
operation). Both “before” (control) and “after” (exposed) children in- 173 -

¾¾¾

Estimated radiation dose
Minisatellite probes
Main results¾

Radiation effects

a family were conceived by the same mother.
z Median accumulated dose was 152mSv.
z Multi-locus minisatellite probes: 33.5, 33.15
z No statistically significant difference in mutation rates between
exposed and control groups was observed. (Small sample size
limited statistical power.)z Number of mutations / Total number of bands in offspring: exposedgroup [9/82], control group [9/472]
z Negative (Small sample size limited statistical power.)

B. Nuclear tests(7) Dubrova et al. (2002)[15]¾

Exposed group¾

Non-exposed control
group

¾
¾

Estimated radiation dose
Minisatellite probes¾

Main results¾

Radiation effectsz Three-generations of families inhabiting the rural areas of theBeskaragai district of Kazakhstan around the Semipalatinsk nuclear
test site: those in the first generation [P0] were born between 1920
and 1950, those in the second generation [F1] were born between
1951 and 1974 and those in the third generation [F2] were childrenof F1. 40 families with 135 of F1 and 97 of F2.
z Three-generations of families from the geographically similar
non-contaminated rural area of Kazakhstan; matched with theexposed group by ethnicity, year of birth, parental age, occupation,
whether or not they were smokers. 28 families with 83 of F1 and 65
of F2.
z >1Svz Single-locus probes: B6.7, CEB1, CEB15, CEB25, CEB36, MS1,
MS31, MS32．
z A statistically significant 1.8-fold increase in mutation rate wasfound in the P0 generation and a less marked 1.5-fold increase wasfound in the F1 generation.
z Positive

C. Mayak plutonium separation combine(8) Dubrova et al. (2006)[16]z Populations in rural villages along the Techa River, which were
¾ Exposed groupcontaminated by radioactive releases from the Mayak plutonium
separation combine; children conceived between 1950 and 1972;both parental exposure [53 families with 101 children], paternal
exposure [10 families with 18 children] and maternal exposure [22families with 40 children].
z
53 families of non-irradiated parents with 110 children: 49 families¾ Non-exposed control
from the non-contaminated areas with children conceived between
group1946 and 1977, and 4 families from the village along Techa River
with all children born between 1950 and 1952 before the discharge;
sex ratio, paternal age, parental occupation and number of smokers
were matched with the exposed group, but maternal age in thecontrol group was higher than that of the exposed group.
¾ Estimated radiation dose z Mean paternal dose was l02±12mSv and mean maternal dose was
86±9mSv. Over 80% of the total parental dose was attributed to theinternal exposure of the radionuclide.
z Single-locus probes: B6.7, CEB1, CEB15, CEB25, CEB36, MS1,
¾ Minisatellite probes
MS31, MS32．

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¾

Main results¾

Radiation effectsz A statistically significant 1.67-fold increase in mutation rate wasfound in the paternal mutations, whereas maternal mutation rate was
not elevated in the exposed families.
z Number of mutations / Total number of bands in offspring:∗ Paternal mutation: exposed group [42/861], control group[31/1044]∗ Maternal mutation: exposed group [6/980], control group [10/885]
z Paternal: positive, maternal: negative

D. Atomic bombings(9) Kodaira, Satoh et al. (1995, 1996, 2004)[17,18,19]
z Atomic bomb survivor families in Hiroshima and Nagasaki in which¾ Exposed groupone or both parents received A-bomb radiation of > 0.01Sv (gonadal
dose). Most of the children were born more than 10 years after thebombing. Within a total of 48 families with 61 children, 1 child hadparents both of whom were exposed, 29 children had exposure only
through the father and 31 children had exposure only through themother.
z
Atomic bomb survivor families in Hiroshima and Nagasaki in which¾ Non-exposed control
one or both parents were exposed to less than 0.01Sv or the families
groupwhich were not in Hiroshima or Nagasaki at the time of thebombing; 49 families with 58 children.
¾ Estimated radiation dose z More than 75% of the exposed parents had an estimated dose of>1Sv (using a neutron RBE value of 10).
z Single-locus minisatellite probes: B6.7, CEB1, CEB15, CEB25,¾ Minisatellite probes
CEB36, MS1, MS31, MS32.
z No evidence that the mutation clustered in individual offspring was¾ Main results
found.
z Number of mutations / Total number of bands in offspring:∗ Paternal mutation: exposed group [11/240], control group [33/709]∗ Maternal mutation: exposed group [2/256], control group [6/694]
z Negative¾ Radiation effects
E. Radiotherapy(10) May, Tamaki et al. [7](2000)
z Three seminoma patients aged 33-49 years donated semen samples¾ Exposed group
after radiotherapy. The post-treatment samples represented sperm
derived from cells irradiated at different stages of spermatogenesis,
the sperms having been taken for several times on different days
after irradiation.
z
Sperm samples donated from the same three patients before having¾ Non-exposed control
radiotherapy.
group¾ Estimated radiation dose z Total testicular X-ray doses were 0.75, 0.82 and 0.38 Gy for eachpatient.
z Single-locus minisatellite probes : B6.7, CEB1.
¾ Minisatellite probesz No significant difference in pre- and post-irradiation mutation rates¾ Main results
was observed at any stages.
z
Negative¾ Radiation effects- 175 -

Table 2. Reports on the “Trans-generational effects” of human populations using mutation markers
of microsatellites
A. Chernobyl nuclear power plant accident(1) Slebos et al. (2004)[22]z Liquidator families from Kiev and Chernigov in Ukraine; father had¾ Exposed groupserved in the cleanup operation of the Chernobyl accident between
1986 and 1990; 75 families with 75 children conceived after the startof the cleanup operation (within them, 39 families with 39 children
did not have children before the start of the cleanup operation).
z 41 liquidator families from Kiev and Chernigov in Ukraine, from the¾ Non-exposed control
same registry as the exposed group, with 41 children conceived
group
before the start of the cleanup operation (within them, 5 families
with 5 children, that did not have children after the start of thecleanup operation). Both “before” (control) and “after” (exposed)
children in a family were conceived by the same mother.
¾ Estimated radiation dose z Median accumulated dose was 152mSv.
z Autosomal loci: 4, X-linked locus: 1¾ Microsatellite lociz More paternal mutations were seen in “after” children than in¾ Main results“before” children, although differences were minimal.
z Total number of mutations at the 5 microsatellite loci / Total numberof microsatellite loci examined: exposed group [6/325], control
group [2/174]
z D7S1482 demonstrated germline hypermutability.
z Negative (Small sample size limited statistical power.)¾ Radiation effects(2) Furitsu, Ryo et al. (2005)[23]z Belarusian liquidator families, in which either or both of the parents¾ Exposed groupwere involved in cleanup operations in the contaminated areas ofChernobyl between April 1986 and July 1987 during the periods
from several weeks to several months; Total number of families
examined was 64 with 73 children; among them, 61 children, whose
fathers were exposed before conception, were analyzed.
z Belarusian families living in the non-contaminated areas in Belarus,¾ Non-exposed control
in which none of the parents were involved in cleanup operations ingroup
the contaminated areas; sex ratio of the children were matched to the
exposed group; 66 families with 69 children.
¾ Estimated radiation dose z Very little information of the recorded individual radiation dose wasavailable for this cohort. UNSCEAR (2000) reported that the meaneffective dose was 39mSv for the liquidators in Belarus during
1986-1987 according to the national registry.
z Autosomal loci: 31, Y-linked loci: 40, X-linked locus: 1.
¾ Microsatellite lociz A higher mutation rate (2.9 x 10-3) of Y-linked loci was detected in¾ Main results
the families of liquidators as compared to that of the control group(2.1 x 10-3), although the increase was not statistically significant.
z Number of mutations / Total number of microsatellite loci examined:∗ 40 Y-linked loci: exposed group [4/1392], control group [3/1458]∗ 31autosomal loci: exposed group [11/1852], control group [18/2108]∗ 1 X-linked locus: no mutations were detected in either exposed orcontrol groupsz
Negative¾ Radiation effects- 176 -

(3) Satoh et al. (1996)[18]¾

Exposed group¾

Non-exposed control
group

¾

Estimated radiation dose¾¾

Microsatellite loci
Main results¾

Radiation effectsz Atomic bomb survivor families in Hiroshima and Nagasaki in which
one or both parents received A-bomb radiation of > 0.01Sv (gonadal
dose). Most of the children were born more than 10 years after thebombing. Within the totally 50 families with 64 children, all children
were exposed through either the mother or the father except one
child where both parents had been exposed.
z Atomic bomb survivor families in Hiroshima and Nagasaki in which
one or both parents were exposed to less than 0.01Sv or the families
that were not in Hiroshima or Nagasaki at the time of the bombing;
50 families with 60 children.
z Mean doses for gametes: maternal 1.7Sv, paternal 2.1Sv (using aneutron RBE value of 20).
z Autosomal loci: 3, X-linked loci: 2.
z There was no significant difference in the mean mutation rate
between the children of the exposed and control groups.
z Total number of mutations in the 5 microsatellite loci / Total numberof microsatellite loci examined: exposed group [0/307], control
group [4/809]
z Negative

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