Critical level of radiation damage of root apical meristem and mechanisms for its recovery in <Emphasis Type="Italic">Pisum sativum</Emphasis> L.

Dose dependencies of growth and cytogenetic values have been built to determine that the critical level of root apical meristem damage induced by cute irradiation within the range from 2 to 20 Gy. Causal relationships between frequency of chromosome aberrations and death of tissue, organ, and organism have been analyzed. The critical level of damage in the stem apical meristem and root of seedlings was defined as 44–48% of aberrant anaphase. The exceeding of this level results in launch of a suicidal program in the meristem through induction of multiaberrant damages and interphase cell death. It is assumed that cell competition between clones of nonaberrant, aberrant with single damages, and multiaberrant cells plays an important role in mechanisms of recovery. The exceeding of a 50% level of aberrations results in total or partial recovery of root apical meristem by regeneration. Approximately 70% of chromosome aberrations are the critical index of root apical meristem damage which still allow its regeneration. However, these local regeneration processes are insufficient for recovery of morphogenesis and survival of seedlings.     

The dose dependence of cytogenetic damages and the adaptive response of mammalian cells under the action of ionizing radiation at low doses

The dose-effect dependence of cytogenetic damage after single dose irradiation in the dose range of 0.1-2 Gy and the adaptive response after double-dose irradiation were studied on Chinese hamster and human melanoma cells in culture. The non-linear dose dependencies were found for the induction of chromosome aberrations with decrease in cell radiosensitivity in the definite dose range. This decrease started at 10 and 20 cGy for melanoma and Chinese hamster cells respectively. The maximal adaptive response was induced at 1 cGy for melanoma cells and at 20 cGy for Chinese hamster cells. It can be supposed that the same inducible repair processes are responsible for non-linearity of dose-effect curves and induction of the adaptive response. These processes are similar in mechanisms and different in quantitative proportion for different cell types.     

Induction of chromosome aberrations and micronuclei in human peripheral blood lymphocytes at low dose of radiation

The chromosome damage induced by the doses of y-irradiation 6)Co in peripheral blood lymphocytes was studied using different cytogenetic assays. Isolated lymphocytes were exposed to 0.01-1.0 Gy, stimulated by PHA, and analysed for chromosome aberrations at 48 h postirradiation by metaphase method, at 49 h--by the anaphase method, at 58 h by micronucleus assay with cytochalasin B and, additionally, micronuclei were counted at 48 h on the slides prepared for the metaphase analysis without cytochalasin B. Despite of the quantitative differences in the amount of chromosome damage revealed by different methods all of them demonstrated complex nonlinear dose dependence of the frequency of aberrant cells and aberrations. At the dose range from 0.01 Gy to 0.05-0.07 Gy the cells had the highest radiosensitivity mainly due to chromatid-type aberration induction. With dose increasing the frequency of the aberrant cells and aberrations decreased significantly (in some cases to the control level). At the doses up to 0.5-0.7 Gy the dose-effect curves have become linear with the decreased slope compare to initial one (by factor of 5 to 10 for different criteria) reflecting the higher radioresistance of cells. These data confirm the idea that the direct linear extrapolation of high dose effect to low dose range--the procedure routinelly used to estimate genetic risk of low dose irradiation--cannot be effective and may lead to underestimation of chromosome damage produced by low radiation doses. Preferences and disadvantages of used cytogenetic assays and possible mechanisms of low ionising radiation doses action were discussed.     

Structural architectonics of the root apical meristem in connection with a quantitative evaluation of its radiation damage

The dose dependences of the frequency of chromosome aberrations in root meristems of pea germinants after their 48-h irradiation in a dose range from 4 to 8 Gy is nonlinear. The plateau of the curve indicates the activation of recovery processes. With the dose rise, the meristem size decreases, while the frequency of cellular inactivation, clustering disorders, and deformations of cell rows in the meristem and extension zone increase. However, the topology of cell rows at the 33% level of aberrations is mostly maintained. Conservation and recovery of the topology of cell rows are performed through repopulation and substitution of damaged cells and rows on its basis. The growth character of new cell rows in damaged tissue is intrusive. The promotion of aberrant cells to the extension zone is slowed down or blocked by the interruption of symplastic growth. In new cellular subpopulations, chromosome mutagenesis continues, and the effectiveness of recovery processes is in many respects defined by the competition between normal and aberrant cells and also their clones. The limits of the recovery potential of root apex are restricted by “critical mass” of the proliferating pool and the degree of damage to the extension zone. At a level of 50%, aberrant anatelophases meristem recovery triggers a more radical mechanism through regeneration, which leads to the complete substitution of apex tissues, including the extension zone.     

Cellular and tissue mechanisms of recovery processes in Hordeum distichum L. under irradiation

Ionizing and UV-B irradiation of barley seedlings increased the number of chromosome aberrations in vegetative and generative meristems and disturbances in microsporo- and microgametogenesis. Other irradiation-induced changes included enhanced cytomixis and an increase in the number of pathologies in meiosis and tetrads and in the polymorphism and disturbances in pollen grains. Damages induced by different irradiation types differ primarily quantitatively. The dynamics of occurrence of chromosome aberrations in the root meristem reciprocally depended on the irradiation dose. Damages induced by low UV doses were detected throughout the ontogeny of plants. Increased irradiation doses activated cytolytic processes during premeiotic interphase, meiosis, and tetrad formation, as well as after the end of gametogenesis. Exposure to maximum doses of UV radiation and moderate doses of γ-radiation restored the fertility of grain pollens. Microsporocytes, microspores, and pollen grain components undergo degeneration primarily via apoptosis. This type of cell death is apparently autonomous, being induced by the cell population itself. The results of this study confirmed the hypothesis on the positive role of cell competition in the recovery processes and adaptation of plants to mutagens.     

Structural architectonics of apical root meristems in coherence with the quantitative assessment of its damage by radiation

The dose dependencies of the aberrant anaphases frequency in the root meristem in 48 hours after irradiation in the range of doses of 4-10 Gy is characterized by threshold and plateau at 33% aberrant anaphase. The plateau indicates the activation of the recovery processes. Topology of cell rows in the primary meristem of the dose to 8 Gy are conserved and recovered damages. New cell rows are formed by local cell pools in the distal meristem, pericycle cells and subepidermy. It grows by intrusive character displacing the rows of damaged cells. Apparently the competition between clones of normal and aberrant cells plays the primary role in the mechanisms of recovery. Resulting to competition the promotion of aberrant cells to the extension zone is slowed down or blocked. So critical level of damage of the root apical meristem was defined about 50% of aberrant anaphase. Exceeding of this level leads to lethal consequence for meristem and it is accompanied by the inclusion of more radical process of restoration through regeneration. Regeneration leads to complete replacement of the apex tissues including the extension zone.     

The radioenhancement of two human head and neck squamous cell carcinomas by 2'-2' difluorodeoxycytidine (gemcitabine; dFdC) is mediated by an increase in radiation-induced residual chromosome aberrations but not residual DNA DSBs

PURPOSE: The present study aimed at investigating if 2'-2' difluorodeoxycytidine (dFdC) radioenhancement was mediated by an effect on induction and/or repair of radiation-induced DNA DSBs and chromosome aberrations in cells with different intrinsic radiosensitivity. METHODS: Confluent human head and neck squamous cell carcinoma cell lines designated SCC61 and SQD9 were treated with 5 microM dFdC for 3 or 24 h prior to irradiation. DNA DSBs induction and repair were analyzed by PFGE. Radiation-induced chromosome aberrations were examined with a FISH technique. RESULTS: In both cell lines, dFdC did not modify radiation-induced DNA DSBs in a dose range between 0 and 40 Gy. After a single dose of 40 Gy, dFdC affected neither the kinetic of repair nor the residual amount of DNA DSBs up to 4 h after irradiation. Whereas dFdC did not increase the induction of chromosome aberrations, after a single dose of 5 Gy, the percentage of aberrant cells and the number of aberrations per aberrant cells were significantly higher in combination with dFdC. CONCLUSION: Our data suggest that under experimental conditions yielding substantial radioenhancement, dFdC decreases the repair of genomic lesions inducing secondary chromosome breaks but has no effect on DNA DSBs repair as measured by PFGE.     

Vicia faba chromosome damage induced by low doses of gamma radiation

The rate of radiation damage to chromosomes by low doses of gamma rays (0.01-0.30 Gy) was studied in the root tips ofVicia faba. As criteria of the effect of ionizing radiation, the frequency of sister chromatid exchanges (SCEs), incidence of chromosomal aberrations and the number of micronuclei were evaluated and compared in irradiated cells. The results obtained confirmed that the analysis of SCEs did not represent an efficient indicator of radiation damage to chromosomes. On the contrary, the formation of chromosomal aberrations and micronuclei was effectively stimulated by low radiation doses, there being linear dose-effect relationships in the low doses region used.     

Effect of preliminary chronic irradiation and alpha-tocopherol on the frequency of chromosome aberrations in mouse bone marrow cells, induced by exposure to acute gamma-irradiation

The incidence of chromosome aberrations in bone marrow cells of femur did not exceed the spontaneous one in CBA mice exposed, during 70 days, to gamma-radiation at dose--rates of 33.7-35.8 nA/kg and cumulative dose of 2.75 Gy. A single acute exposure of intact animals to a dose of 2.98 Gy increased significantly the mutation level. Preirradiation with small doses increased the resistance of hereditary structures to sublethal radiation doses. Exogenous alpha-tocopherol (0.06 mg/20 g mass) protected the genetic apparatus of cells from total-body irradiation and was an additional factor decreasing the mutation level after acute exposure of mice at the background of long-term irradiation with small doses.     

Cytogenetic Effects of γ-Radiation in Onion (<Emphasis Type="Italic">Allium cepa</Emphasis> L.) Seedlings

The effect of γ-radiation on the cytogenetic parameters of root meristem cells of onion seedlings was studied in laboratory experiments (Allium-test). An increase in the overall frequency of chromosomal aberrations and micronucleus frequencies in seedling cells at low γ-radiation doses (≤0.1 Gy) was detected for the first time. At a maximum absorbed dose of 13 Gy, chromosomal aberrations were detected in the majority of cells in the anaphase and telophase stages of the cell cycle, and the number of cells with multiple aberrations increased. The main contribution to the overall frequency of chromosomal aberrations, in addition to multiple aberrations, is made by the bridge-type aberrations, fragments, and lagging chromosomes. The data obtained allow using the cytogenetic indices of Allium cepa seedlings to assess the biological effects of lowdose γ-radiation.     

Meiotic non-disjunction induced by fission neutrons relative to X-rays observed in mouse secondary spermatocytes. II. Dose-effect relationships after treatment of pachytene cells

(C57B1/Cne X C3H/Cne)F1 male mice were irradiated with single acute doses of 0.4 MeV neutrons (from 0.11 to 0.72 Gy) or 250 kV X-rays (from 0.25 to 3 Gy) and sacrificed 5 days later. Chromosome preparations of secondary spermatocytes, irradiated at the stage of pachytene, were analysed and the incidence of hyper-haploidies and chromosome fragments was recorded. Data on numerical aberrations were fitted by highly significant linear relationships for both types of radiation. A relative biological effectiveness (RBE) value of 5.65 was estimated by the ratio between the slopes of the two regression lines. The same linear fitting was applied to frequencies of cells with fragments, even if in this case other types of functions could not be excluded. An RBE value was estimated in the same way as for numerical aberrations and yielded a comparable figure of 5.23. A significant correlation was also found between the incidence of numerical and structural aberrations, which points to the chromosome itself as the prevalent target for radiation-induced non-disjunction (ND). In addition, the highly significant linearity of the dose-effect relationship observed for the induction of aneuploidies suggests, as the simplest hypothesis, a single-hit mechanism of radiation action, possibly through pre-non-disjunctional damage to the centromeric region, rather than an indirect induction of segregational difficulties after primarily induced chromatid interchanges.     

The blocking of the proliferation of the human endothelial cells in culture by beta-irradiation from internal and external sources of the radiation. S-block is induced by the 3H2O beta-particles

Recently we shown that low doses (0.12-0.46 Gy) of (methyl-3H)-thymidine incorporated into human endothelial cells induce the accumulation cells in G2-phase of the cell cycle. The temperate doses of (1-6 Gy) gamma-rays 137Cs were less effective in the induction of the G2-block estimated by flow cytometry analysis of DNA content and in the induction of the chromosome aberrations (bridges and fragments in anaphase). The aim of this study was the comparative investigation of efficiency of beta-rays emitted 3H from 3H-thymidine and 3H2O by several of the cellular parameters. Here we shown that at the equal conditions of the incubation of the cells in medium with 3H2O induced the accumulation cells in S-phase without decreasing of the mitotic activity and without increasing of the chromosome aberrations level. Unlike from 3H2O the incubation of the cells with 3H-thymidine induced the accumulation cells in G2-phase with decrease of the mitotic activity and with increase of the chromosome aberrations level. Concurrent treatment cells with 3H-thymidine and thymidine abrogate these cellular effects of the 3H-thymidine. Inhibitor ATM-kinase caffeine abrogate as G2-block as S-phase block. These results suggest that the low-dose beta-radiation activates S-phase and G2-phase checkpoints requiring ATM-mediated signal transduction pathway. The factors, which impact on the efficiency of the internal and of the external sources of the irradiation, depend on theirs disposition in relation to radiosensitive target--DNA was discussed.     

The effect of continuous gamma irradiation on formation of sister chromatid exchanges (SCEs) and chromosomal aberrations in meristematic cells ofVicia faba

Germinated seeds ofVicia faba were continuously irradiated at low dose rate of gamma rays (0.05 Gy h-¹) up to a total accumulated dose of 2 Gy. The FPG (fluorescence plus Giemsa) technique of differential chromatid staining was used to monitor the frequency of sister chromatid exchanges (SCEs) in irradiated root tip meristem cells. The results of the experiments have demonstrated that SCE frequency is raised by continuous gamma irradiation only in plant cells containing BrdU in the chromosomal DNA. No effect concerning SCE formation was recorded at continuous irradiation of meristematic cells of Vicia faba with native, i. e. BrdU-nonsubstituted, DNA. In contrast to SCEs, a significant increase was found in the yield of chromosomal aberrations in all variants of irradiation.     

Dynamics of the induced chromosomal instability in welsh onion (Allium fistulosum L.): gamma irradiation of the seeds of different storage periods

The chromosome aberrations in root meristem cells of welsh onion (Allium fistulosum L.) seeds after gamma-irradiation (5 and 10 Gy) of different-aged seeds (7, 19, 31, 43 and 55 months of storage) were studied. The irradiation dose of 5 Gy significantly increased the frequency of aberrant anaphases (FAA) for 31- and 43-months seeds; the dose of 10 Gy significantly increased the FAA in seeds of all age groups. The irradiation of young (7 months) seeds resulted in decreasing of the fraction of bridges to the control level of the old (55-months) seeds for the dose of 5 Gy and below the control level of the old seeds--for the dose of 10 Gy. Some peculiarities of cytogenetic parameters of genome instability and the germinating capacity of the seeds made it possible to suppose that the third year of storage is a critical period for the welsh onion seeds.     

Comparison of recovery from potential mitotic abnormality in mitotically quiescent lens cells after X, neutron, and 56Fe irradiations

After exposure to various doses of 250 kVp X radiation, 0.85 Me V fission spectrum neutrons, or 600 MeV/A iron (Fe) particles, mitotically quiescent rat lens cells showed no visible evidence of radiation injury. However, following the mitogenic stimulus of wounding, mitotic abnormalities became evident when responding cells entered mitosis. Latent damage and recovery therefrom were monitored at 3, 7, 14, and 28 days after irradiation. Following doses of 1 to 10 Gy of X radiation, the recovery rate, indicated by a decrease in abnormalities with time, was proportional to dose, and the dose-effect slope decreased exponentially with time. Virtually no recovery occurred during the 28 days after 1.25 to 2.25 Gy of fission neutron radiation. After doses of 0.5 to 3.0 Gy of Fe particles, an increased expression of mitotic damage or recovery than recovery occurred. As a consequence of the differing patterns in time for expression of damage or recovery following X rays and the high-LET radiations, the relative biological effectiveness (RBE) increased from 3.6 to 16 for neutrons and from 2 to 10 for Fe particles over the 28-day observation period.     

Elimination of X-ray-induced chromosomal aberrations in the progeny of female mice

Female NMRI mice were irradiated with various doses of X-rays and induced chromosome aberrations were scored in MII oocytes (Dosage: 0.222, 0.666, 2 and 6 Gy). After irradiation with 2 Gy, early zygotes were examined in the 2-cell stage; additional dominant lethals were counted and surviving embryos were examined after 13.5 days of pregnancy. 87.2% of the MII oocytes showed structural chromosomal aberrations after irradiation with 2 Gy. Surviving embryos, however, failed to show any increase in the aberration rate. This result points to (almost) complete elimination of genetically damaged oocytes and zygotes already before birth. In addition to the structural aberrations, aneuploidies were induced. Most of them, however, were hypoploidies. Hence, the study confirmed the well-known susceptibility of oocytes around the time of fertilization for induced chromosome loss. Induced hyperploidies, however, were very rare. Evidence for induction of meiotic non-disjunction was weak. In surviving embryos, no increase in numerical aberrations, either hypoploid or hyperploid was discovered. The significance of these data for the prediction of chromosomal damage due to to ionizing radiation in humans is discussed. Recent risk estimates of UNSCEAR and other agencies represent very cautious upper levels.     

Effects of gamma-irradiation on the infectivity and chromosome aberration of Clonorchis sinensis

Effects of gamma irradiation on the worm survival and chromosomal aberration of Clonorchis sinensis were studied. The metacercariae irradiated with various amounts of gamma radiation (ranging from 5 Gy to 50 Gy) were fed to rats, and the effects were compared with those of non-irradiated controls. Recovery rates of adult worms in irradiated groups were reduced gradually as increasing of the irradiation doses. No worm was recovered from rats which were fed with 50 Gy irradiated metacercariae. The chromosome number was 2n = 56 in all worms from all experimental groups. However, the groups irradiated with 20 Gy, 25 Gy or 30 Gy showed variations in the chromosome number, depending on different cells in the same individual. Radiation doses used in this study did not appear to induce chromosome aberrations, however, irradiation with 30 Gy showed slightly reduced chromosome size.     

Cytogenetic effect of plant tissue culture medium with certain growth substances onAllium sativum L. meristem root tip cells

The effect of plant tissue culture medium with different concentrations and combinations of growth regulators (kinetin, indol-3-ylacetic acid, 2,4-dichlorophenoxyacetic acid) was evaluated on mitosis ofAllium sativum meristem root tip cells. Different combinations of growth regulators at low concentrations had no effect on induction of mitotic aberrations or inhibition of mitotic activity. Inhibition of mitotic activity, a tendency to chromosome stickiness and clumping and a slight increase in the frequency of mitotic aberrations were observed at higher concentrations. It may be proposed that plant tissue culture media have no direct effect on induction of mitotic aberrations in plant tissue culturesin vitro.     

Radiation-induced chromosomal aberrations in human lymphocytes. I. Dependence on the dose of gamma-rays and an anomaly at low doses

Cultures of human lymphocytes obtained from blood of healthy adult donors were irradiated with different doses of ⁶⁰Co γ-rays and the irradiated cells were analysed in metaphase 50 h after irradiation. The effect (total yield of abberations of chromosome type, or total yield of exchange type abberations) produced by the lowest dose (5 rad) appears to be statistically significant in a sample of 1500 cells. In the usual dose range (25–400 rad), both parabolic and linear-quadratic equations give a satisfactory fit of experimental data (dicentrics, fragments, or all aberrations of chromosome type). Low doses of γ-rays, however, produced more aberrations than expected, if one extrapolates dose-effect curves from higher doses. Both relations should be considered, therefore, merely as empirical equations. Dicentrics show at low doses (10–30 rad) a plateau which appears to be statistically significant. Some indications are obtained that the total number of chromosome-type aberrations is a more reliable criterion of cytogenetic damage than the usually accepted yeild of dicientrics and rings.     

Effect of low-dose radiation on repair of DNA and chromosome damage

In this report results of studies on the effect of different doses of low LET (linear energy transfer) radiations on the unscheduled DNA synthesis (UDS) and DNA polymerase activity as well as the induction of adaptive response in bone marrow cells (BMC) by low dose radiation were presented. It was found that whole-body irradiation (WBI) with X-ray doses above 0.5 Gy caused a dose-dependent depression of both UD5 and DNA polymerase activity, while low dose radiation below 250 mGy could stimulate the DNA repair synthesis and the enzyme activity. WBI of mice with low doses of X-rays in the range of 2-100 mGy at a dose rate of 57.3 mGy per minute induced an adaptive response in the BMC expressed as a reduction of chromosome aberrations following a second exposure to a larger dose (0.65 mGy). It was demonstrated that the magnitude of the adaptive response seemed to be inversely related to the induction dose. The possibility of induction of adaptive response in GO phase of the cell cycle and the possibility of a second induction of the adaptive response were discussed.