Dicentric yields induced by gamma-radiation and chromosome arm number in primates.

To evaluate the effect of the chromosome arm number on the yield of dicentric chromosomes, frequencies of gamma-ray-induced chromosome aberrations were examined with peripheral lymphocytes from three different primate species, Saimiri sciureus (arm number, 77), Macaca fascicularis (arm number, 83) and Nycticebus coucang (arm number, 99). Irradiated blood samples were cultured by the same standard technique as that commonly used for human lymphocytes. The yields of dicentrics and dicentrics plus rings at doses of 100, 200 and 300 rad of gamma-irradiation were not significantly different among the three species, in spite of the difference in the chromosome arm number. Furthermore, dose-response relationships for these species were consistent with that for man. Statistical analysis indicated that the expected dicentric yields calculated from the arm number model were significantly different from the observed yields at 200 and 300 rad doses (P less than 0.01). From these results it can be pointed out that there is no correlation between the yield of dicentrics and the effective chromosome arm number, and that the chromosomal radiosensitivity of these primates is essentially the same as that of man, at least in the lymphocyte system.     

Blood lymphocyte culture system: quantitative analysis of X-ray-induced chromosome aberrations in man, muntjac and cattle

Peripheral blood lymphocytes of 3 mammalian species, man, muntjac and cattle, which have various amounts of DNA and divergent karyotypes, were exposed to 100-400 rad of X-rays, and frequencies of dicentrics and other aberrations were analysed at first post-irradiation metaphases. During experiments, various preparative or physical and biological factors that could influence the yield of chromosome aberrations were taken into account. The frequency of dicentrics scored at first post-irradiation metaphases showed best fit to both linear and quadratic dose-response curves, y = a + bD and y = bD + cD2 with a high correlation coefficient of 0.98 (P less than 0.001). The frequency of dicentrics obtained at different post-irradiation fixation times did not show significant variation, indicating a homogeneous sensitivity of peripheral lymphocytes to X-irradiation. BrdU incorporation following X-irradiation showed no increase in the frequency of chromosome aberrations. The frequency of dicentrics in man, muntjac and cattle showed a close correlation with their DNA content, but no meaningful correlation was found between the yield of dicentrics and the chromosome arm number or the nuclear volume. The ratio of dicentric yields, 1.00:0.67:1.04 obtained in man, muntjac and cattle were comparable to the ratio of their DNA contents, 1.00: 0.65: 1.07. The base-line frequency of SCEs was similar in the 3 species and no significant variation in SCE frequency was noticed even after administration of 400 rad of X-rays.     

Enhanced frequency of chromosome aberrations in workers occupationally exposed to diagnostic X-rays

To estimate the level of radiation exposure of personnel handling diagnostic X-ray machines, the yield of chromosomal aberrations was analysed in peripheral blood lymphocyte cultures. These occupationally exposed individuals showed higher frequencies of dicentrics as well as acentrics than normal controls. Absorbed radiation doses calculated by extrapolating reference in vitro dose-response curve for dicentrics ranged between 0.13 and 0.17 Gy. This implies exposure beyond the permissible limit of 0.05 Gy/year for the whole body. However, no obvious trend of increased aberrations as a function of either duration of employment or age was noticed. The increase in the aberration yields in this personnel underscores the need of adopting measures to avoid or minimise such overexposure.     

Quantitative analyses of the induction of chromosome aberrations and sister-chromatid exchanges in human lymphocytes exposed to gamma-rays and mitomycin-C in combination

Most chemicals are S-dependent and are potent inducers of SCE, but do not produce chromosome-type aberrations in the first metaphases after exposure. Ionizing radiation, which is an S-independent agent, produces chromosome-type aberrations, especially dicentrics and rings, but inefficiently produces chromatid-type aberrations. A series of experiments has been performed to investigate whether cytogenetic damage induced by ionizing radiation (gamma-rays) might be assessed separately from that induced by the alkylating chemical, mitomycin C (MMC), when human lymphocytes were exposed to these 2 agents in combination. Whole-blood cultures of human lymphocytes in G0 phase were exposed to gamma-rays and MMC in combination or separately. Cytogenetic analyses were done for both chromosome aberrations (CA), analyzed in cultures incubated for 56 h without BrdUrd, and sister-chromatid exchanges (SCEs) in cultures incubated for 72 h with BrdUrd. The frequency of chromosome-type aberrations (dicentrics and rings) increased with increasing doses of gamma-rays from 0.5 to 4.0 Gy. The dose-response relationships were the same with or without concomitant treatment with MMC (10(-6) M). Although the SCE frequency increased with increasing doses of MMC, the increase was nearly the same as when cells were treated with both MMC and gamma-rays (2 Gy). There was no interaction between MMC and gamma-rays concerning these 2 endpoints.     

Aberrations induced by fission neutrons in human peripheral lymphocytes

Analysis of dose-response relationship was carried out for chromosome aberrations produced in human peripheral lymphocytes by fission neutrons at doses of 25, 50, 100 or 200 rad.Statistical treatment showed experimental data to be fitted by a regression curve described by the mathematical model Y = a+bD. A linear relation to dose characterized both one-break and two-break aberration yields. Numerical values of coefficients are reported for yields of dicentrics, chromosome fragments, minutes, aberrant cells, total number of aberrations, and total breakage.Based on chromosome fragments and aberrant cells, relative biological efficiency (RBE) value derived for fission neutrons relative to 180 kV X-rays for chromosome fragments was 2.53, and for aberrant cells it was 2.80.     

The biological activity of hydrogen peroxide. I. Induction of chromosome-type aberrations susceptible to inhibition by scavengers of hydroxyl radicals in human embryonic fibroblasts

The cytogenetic effect of hydrogen peroxide (H2O2) was investigated in human embryonic fibroblasts. Chromosome-type aberrations were found together with chromatid-type aberrations in metaphase cells harvested 24 h after a single 10-min treatment with 10(-5)-10(-3) M H2O2 in 0.9% NaCl solution. The chromosome-type aberrations were observed to be predominantly dicentrics and deletions. Both types of aberration showed a dose-response relationship to the dose of H2O2 over the range of 10(-5)-1.5 X 10(-4) M H2O2. The intercellular distribution of dicentrics showed a Poisson distribution. Centric and acentric rings and abnormal monocentrics were a minor fraction of the chromosome-type aberrations. The chromatid-type aberrations observed, such as breaks, exchanges and gaps, showed no dose-response relationship. The frequency of isochromatid breaks was higher than that of chromatid breaks and approximately 70% of the isochromatid breaks were found in the centromeric or pericentromeric region. The intercellular distribution of chromatid exchanges showed an over-dispersed distribution. The generation of aberrations by H2O2 was effectively suppressed by catalase and several scavengers of hydroxyl radicals (.OH) such as ethanol, dimethyl sulfoxide (DMSO) and mannitol. This result suggest that .OH plays an essential role in the generation of the chromosome aberrations by H2O2.     

Cell type-specific quantitative predictions of radiation-induced chromosome aberrations: a computer model approach

A quantitative computer model was applied to simulate the three-dimensional (3D) spatial organization of chromatin in human cell nuclei under defined conditions of virtual irradiation to explore the implications of spatial organization on chromosome aberrations. To calibrate the virtual irradiation algorithm, a dose-dependent spectrum of radiation-induced chromosome aberrations such as dicentrics, translocations and centric rings was calculated for low-LET radiation doses ranging from 0.5 to 5 Gy. This was compared with the results from experimental studies. While the dose-response curves calculated from model simulations agree well with experimental dose-response curves for dicentrics and translocations, centric rings are significantly more frequent in the model simulation than in experiments despite taking into account exclusive arm territories in the applied Spherical 1 Mbp Chromatin Domain (SCD) computer model explicitly. Taking into account the non-random positioning of chromosome territories observed in lymphocyte cell nuclei (a so-called gene density-correlated arrangement of chromosome territories), aberration frequencies were calculated with the calibrated irradiation algorithm to investigate the impact of chromosome territory neighborhood effects (proximity effects). The absolute frequencies of pairwise exchanges agree well with those found in an experimental study. In conclusion, the results obtained using the computer model approach presented here based on only a few adjustable parameters correlated well with those of experimental studies of chromosome aberration frequencies. Thus the model may be a useful tool in radiation-induced cancer risk estimates in combination with epidemiological studies.     

Models of chromosome aberration induction: an example based on radiation track structure

A few examples of models of chromosome aberration induction are summarised and discussed on the basis of the three main theories of aberration formation, that is "breakage-and-reunion", "exchange" and "one-hit". A model and code developed at the Universities of Milan and Pavia is then presented in detail. The model provides dose-response curves for different aberration types (dicentrics, translocations, rings, complex exchanges and deletions) induced in human lymphocytes by gamma rays, protons and alpha particles of different energies, both as monochromatic fields and as mixed fields. The main assumptions are that only clustered - and thus severe - DNA breaks ("Complex Lesions", CL) can participate in the production of aberrations, and that only break free ends in neighbouring chromosome territories can interact and form exchanges. The yields of CLs induced by the various radiation types of interest are taken from a previous modelling work. These lesions are distributed within a sphere representing the cell nucleus according to the radiation track structure, e.g. randomly for gamma rays and along straight lines for light ions. Interphase chromosome territories are explicitly simulated and configurations are obtained in which each chromosome occupies an intranuclear domain with volume proportional to its DNA content. In order to allow direct comparisons with experimental data, small fragments can be neglected since usually they cannot be detected in experiments. The presence of a background level of aberrations is also taken into account. The results of the simulations are in good agreement with experimental dose-response curves available in the literature, that provides a validation of the model both in terms of the adopted assumptions and in terms of the simulation techniques. To address the question of "true" incompleteness, simulations were also run in which all fragments were assumed to be visible.     

两例事故受照者染色体畸变分析及生物剂量估算

应用外周血淋巴细胞染色体畸变形和CB微核分析方法,对2000年河南许昌“3.06”^60Co辐射事故1例受照（A）和2000年河南开封“6.26”辐射事故一例过量放射性核素内污染致γ射线照射受照（B）的生物剂量进行估算。结果表明,“A”和“B”两例受照依据双＋环率估算的剂量分别为1.44Gy和0.15Gy,CB微核率估算的剂量分别为1.43Gy和0.22Gy,与用物理方法估算的剂量比较接近,亦与放射损伤的临床诊断一致。泊松分布检验证实,“A”偏离泊松分布,受到不均匀照射。提示染色体畸变和CB微核分析是非常可靠的生物计量估算方法。     

Chromosome aberration frequencies produced by a 70-MeV proton beam

The effectiveness of a 70-MeV proton beam in the induction of chromosome aberrations was studied. We employed peripheral lymphocytes and analyzed the frequencies of dicentrics and rings after irradiation at doses ranging from 0.1 to 8.0 Gy at various depths within a Lucite phantom. The frequency of chromosome aberrations after irradiation with an unmodulated proton beam at 5 mm showed a dose-response relationship similar to that of 60Co gamma rays. However, irradiation at greater depths with the spread-out Bragg peak induced higher aberration frequencies at doses lower than those with gamma rays. Furthermore, the distribution curve of chromosome aberration frequencies as a function of depth was found to be slightly different from the physically measured depth-dose curve. With the spread-out Bragg peak the biological effects were more marked at greater depths, resulting in a distribution of relative biological effectiveness values. The results obtained from chromosome aberration analysis may not be related directly to those for the relationship between dose and cell killing. Slight differences in values for relative biological effectiveness due to the change of dose and site of proton beam irradiation may not be important for practical proton beam therapy, but may be important in the prevention of late radiation injuries.     

Maximum likelihood estimation for cytogenetic dose-response curves

In vitro dose-response curves are used to describe the relation between chromosome aberrations and radiation dose for human lymphocytes. The lymphocytes are exposed to low-LET radiation, and the resulting dicentric chromosome aberrations follow the Poisson distribution. The expected yield depends on both the magnitude and the temporal distribution of the dose. A general dose-response model that describes this relation has been presented by Kellerer and Rossi (1972, Current Topics on Radiation Research Quarterly 8, 85-158; 1978, Radiation Research 75, 471-488) using the theory of dual radiation action. Two special cases of practical interest are split-dose and continuous exposure experiments, and the resulting dose-time-response models are intrinsically nonlinear in the parameters. A general-purpose maximum likelihood estimation procedure is described, and estimation for the nonlinear models is illustrated with numerical examples from both experimental designs. Poisson regression analysis is used for estimation, hypothesis testing, and regression diagnostics. Results are discussed in the context of exposure assessment procedures for both acute and chronic human radiation exposure.     

Radiation-induced chromosome aberrations in lymphocytes from man and crab-eating monkey: The dose-response relationships at low doses

To obtain information on the relation beteween yiels of chrmosome aberrations and dose at low-dose levels, experiments were conducted with 5, 10, 20, 30 and 50 rad of ¹³⁷Cs γ-rays, on lymphocytes from man and crab-eating monkey (Macaca fascicularis). The dose-response relationship for dicentrics was obtained from the combined data of these low-dose experiments with those of our previous onse at high doses (100–400 rad) When the difference between observed yields and those expected from the linear-quadratic model were computed, the dose-response curve had a good fit for man, but not for the monkey. The linear regression lines between 0 and 30 rad were calculated, because the expected values of α/β for man and monkey would be about 100 and 60 rad.The human date gave a satisfactory fit to a linear model, i.e., a linear increase in aberration frequency with dose, whereas this was not so for those of the monkey. Furtyhermose, there was some suggestive evidence for the existence of plateau in dicentrics yields between 10 and 30 rad for the monkey and between 20 and 30 rad for human lymphocytes, but more data would be needed to verify this suggestion, particularly for human lymphocytes.     

Comparison of cytogenic response of human lymphocytes to in vivo and in vitro exposure to low doses of gamma-rays. Unstable chromosomal exchanges detected by FPG techniques

On peripheral lymphocytes of eight cancer patients undergone whole-body therapeutic irradiation (at daily dose of 10 cGy up to total dose of 50 cGy of 60Co gamma-rays) the dose-response of unstable chromosome exchanges (dicentrics and centric rings) was studied. This dose response fitted well linear function. The lower slope of dose-response curve was found for in vivo irradiated lymphocytes as compared to the dose response curve obtained for in vitro irradiated lymphocytes of the same patients. This finding seems to provide evidence that in case of protracted irradiation of individuals an absorbed dose could be underestimated if for biological dosimetry an in vitro dose response curve for unstable chromosome aberrations is used as referent one.     

Genotoxic effects of radiotherapy and chemotherapy on the circulating lymphocytes of breast cancer patients. I. Chromosome aberrations induced in vivo

Unstable chromosome aberrations were scored in peripheral blood lymphocytes (PBL) serially collected from 21 breast cancer patients before and after radiotherapy (RT), chemotherapy (CT) and combined treatments. Local radiotherapy as treatment for mammary cancer induced unstable chromosome aberrations in peripheral blood lymphocytes. Only a fraction of these lymphocytes were exposed to irradiation during treatment and the chromosomal damage observed in PBL was equivalent to that induced by irradiation in vitro with 2 Gy at high dose rate, i.e., about 4% of the total dose delivered locally. Chemotherapy alone did not induce such anomalies. Apart from the observed interindividual variations in either the level or the fate of dicentrics with time, different features of chromosome damage were found when chemotherapy was given before or after local cobaltotherapy: secondary chemotherapy did not alter the frequency and the overdispersed distribution of dicentrics observed after first-line radiotherapy; in contrast, when CT was given before radiotherapy, a lower dicentric frequency was scored, the distribution of dicentrics was not always found to be overdispersed and there was a time-dependent decrease in dicentrics after in vivo exposure.     

Analysis of chromosome aberrations in human peripheral lymphocytes induced by 5.4 keV x-rays

Irradiation of human lymphocytes by x-rays has been seen, in past studies, to produce increasing frequencies of chromosome aberrations at lower x-ray energies. However, in one earlier irradiation experiment with chromium x-rays, the relative biological effectiveness (RBE) did not appear to be larger than that of hard x-rays, especially at higher doses. A possible reason for this unexpected result may have been the irradiation and culture conditions. We have, therefore, in the present study used a technique that has been developed in our laboratory to ensure uniformity of irradiation within lymphocytes and to avoid artefacts due to the cell cycle kinetics. Monolayers of 3-h-stimulated lymphocytes were exposed to 5.4 keV x-rays. A linear-quadratic dose-response was found for dicentrics. The comparison to an earlier finding with 220 kV x-rays shows the expected result of the RBE of the 5.4 keV x-rays to be above that of 220 kV x-rays. The intercellular distribution of dicentrics did not differ significantly from a Poisson distribution. Received: 17 January 1997 / Accepted in revised form: 17 July 1997     

Cytogenetic studies of blood lymphocytes of cosmonauts after long-ter, space flights

An analysis was performed of unstable chromosomal aberrations in peripheral blood of 36 cosmonauts after long-term space missions on "Mir" orbital station. 25 cosmonauts were examined before their flights to score spontaneous yields of cytogenetical damage. In all cases the doses absorbed by crews during space flights did not exceed permissible levels of irradiation, adopted for cosmonauts. The frequencies of chromosomal-type aberrations after space missions were found to increase significantly compared to the pre-flight levels. The yields of dicentrics and centric rings on the average were as high as 0.12 +/- 0.02 and 0.47 +/- 0.06% before and after the 1st flight, 0.18 +/- 0.05 and 0.71 +/- 0.11% before and after the 2nd flight respectively. During the inter-flight periods, usually lasted 1.5-2 years, the yields of chromosome damage lowered, but did not reach their spontaneous values. After each next flight the yields of chromosome aberrations increased again. The cytogenetical damage detected in cosmonauts' peripheral blood lymphocytes after chronic action of low doses of space radiation points out a possible increase in risks of stochastic effects in distant future for crews after long-term space missions.     

A comparative study of the frequencies of radiation-induced chromosome aberrations in somatic and germ cells of the rhesus monkey (Macaca mulatta)

Frequencies of radiation-induced chromosome aberrations in spermatogonia, peripheral blood lymphocytes and bone-marrow cells of the rhesus monkey (Macaca mulatta) and in human blood lymphocytes, were determined at different exposures of X-rays. The dose-response curve for the induction of reciprocal translocations in spermatogonia suggested a “hump” at about 200 rad. The absolute frequencies of chromosome aberrations in somatic and germ cells of the rhesus monkey were low in comparison with most other mammalian species and the ratio between aberrations in the two tissues was 25 to 1 at the 100 rad level. Although the numbers of “effective chromosome arms” in man and rhesus monkey are similar (81 vs. 83), the rhesus monkey showed a lower rate of induction of dicentrics in blood lymphocytes than man at all doses, reaching statistical significance at the 300 rad level.     

The induction of chromosome aberrations during the course of radiation therapy for morbus Hodgkin

In a patient with Morbus Hodgkin, structural aberrations of the chromosome type in peripheral lymphocytes were analyzed during radiation therapy (accumulated target dose 44.6 Gy: 22 fractions of 1.8 Gy each and 2 fractions of 2.5 Gy each at the end of the therapy). The blood was sampled about 5 min after a fraction and/or 24, 48, or 72 h thereafter. The frequency of dicentric chromosomes:acentric fragments:centric ring chromosomes is 37:14:1 throughout the therapy. Independent of the time of blood sampling after a fraction, the distributions of dicentrics and acentrics are overdisperse and represent negative binomial distributions. The yields from these aberrations, as determined during the course of radiotherapy, are best fitted to a linear-quadratic function with a negative quadratic term. The two dose-effect curves (blood sampling about 5 min and 24 to 72 h after a fraction) of dicentrics and acentrics do not differ significantly. Up to an accumulated target dose of about 20 Gy the percentages of cells with chromosome aberrations increase to about 48 to 65% and, at this level, remain constant until the end of therapy.     

The mouse splenocyte assay, an in vivo/in vitro system for biological monitoring: studies with X-rays, fission neutrons and bleomycin.

A modified mouse splenocyte culture system was standardized after testing different mitogens (i.e., phytohemagglutinin (PHA), concanavalin A (Con A)). The mitotic index was determined for comparison between different mitogens. Following selection of appropriate mitogen (PHA 16, Flow), a series of experiments were conducted to evaluate the application of a cytokinesis-block for scoring micronuclei and assays for chromosomal aberrations produced by treatment in G0 and G2 for the purposes of biological dosimetry following in vivo and/or in vitro exposure to X-rays, fission neutrons and bleomycin. In the X-irradiation studies, the frequencies of micronuclei and chromosomal aberrations (i.e., dicentrics and rings) increased in a dose-dependent manner. These data could be fitted to a linear-quadratic model. No difference was observed between irradiation in vivo and in vitro, suggesting that measurement of dicentrics and micronuclei in vitro after X-irradiation can be used as an in vivo dosimeter. Following in vivo irradiation with 1 MeV fission neutrons and in vitro culturing of mouse splenocytes, linear dose-response curves were obtained for induction of micronuclei and chromosomal aberrations. The lethal effects of neutrons were shown to be significantly greater than for a similar dose of X-rays. The relative biological effectiveness (RBE) was 6-8 in a dose range of 0.25-3 Gy for radiation-induced asymmetrical exchanges (dicentrics and rings), and about 8 for micronuclei in a dose range of 0.25-2 Gy. Furthermore, the induction of chromosomal aberrations by bleomycin was investigated in mouse G0 splenocytes (in vitro) and compared with X-ray data. Following bleomycin treatment (2 h) a similar pattern of dose-response curve was obtained as with X-rays. In this context a bleomycin rad equivalent of 20 micrograms/ml = 0.50 Gy was estimated.     

Distribution of the various radiation-induced chromosomal rearrangements in relation to the dose and sampling time

The quantitative analysis of the chromosome rearrangements detected in 2128 R-banded metaphases, obtained from gamma-irradiated human lymphocytes after 48 to 96 h in culture is reported. Depending on the culture time, and possibly on the dose of radiation (from 1 to 3 Gy), the most frequent type of rearrangement was either dicentrics or reciprocal translocations. In first generation mitoses, the frequency of cells without rearrangement ranged from 0.66 to 0.18, and the mean number of rearranged chromosomes per cell from 0.79 to 3.28. The dose-response curve follows a quadratic function for dicentric aberration yields, but not for other rearrangements.