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1.
Two conflicting phenomena, the bystander effect and the adaptive response, are important in determining biological responses at low doses of radiation and have the potential to have an impact on the shape of the dose-response relationship. Using the Columbia University charged-particle microbeam and the highly sensitive AL cell mutagenic assay, we reported previously that nonirradiated cells acquired mutagenesis through direct contact with cells whose nuclei had previously been traversed with either a single or 20 alpha particles each. Here we show that pretreatment of cells with a low dose of X rays 4 h before alpha-particle irradiation significantly decreased this bystander mutagenic response. Furthermore, bystander cells showed an increase in sensitivity after a subsequent challenging dose of X rays. Results from the present study address some of the pressing issues regarding both the actual target size and the radiation dose response and can improve on our current understanding of radiation risk assessment.  相似文献   

2.
Ionizing radiation is a known human carcinogen that can induce a variety of biological effects depending on the physical nature, duration, doses and dose-rates of exposure. However, the magnitude of health risks at low doses and dose-rates (below 100mSv and/or 0.1mSvmin(-1)) remains controversial due to a lack of direct human evidence. It is anticipated that significant insights will emerge from the integration of epidemiological and biological research, made possible by molecular epidemiology studies incorporating biomarkers and bioassays. A number of these have been used to investigate exposure, effects and susceptibility to ionizing radiation, albeit often at higher doses and dose rates, with each reflecting time-limited cellular or physiological alterations. This review summarises the multidisciplinary work undertaken in the framework of the European project DoReMi (Low Dose Research towards Multidisciplinary Integration) to identify the most appropriate biomarkers for use in population studies. In addition to logistical and ethical considerations for conducting large-scale epidemiological studies, we discuss the relevance of their use for assessing the effects of low dose ionizing radiation exposure at the cellular and physiological level. We also propose a temporal classification of biomarkers that may be relevant for molecular epidemiology studies which need to take into account the time elapsed since exposure. Finally, the integration of biology with epidemiology requires careful planning and enhanced discussions between the epidemiology, biology and dosimetry communities in order to determine the most important questions to be addressed in light of pragmatic considerations including the appropriate population to be investigated (occupationally, environmentally or medically exposed), and study design. The consideration of the logistics of biological sample collection, processing and storing and the choice of biomarker or bioassay, as well as awareness of potential confounding factors, are also essential.  相似文献   

3.
This paper (1) presents an analysis of published data on the molecular nature of spontaneously arising and radiation-induced mutations in mammalian somatic cell systems and (2) examines whether the molecular nature and mechanisms of origin of radiation-induced mutations, in mammalian in vivo and in vitro systems, as currently understood, are consistent with expectations based on the biophysical and microdosimetric properties of ionizing radiation. Depending on the test system (CHO cells, human T lymphocytes and human lymphoid cell line TK6), 80-97% of spontaneous HPRT mutations show normal Southern patterns; the remainder is due to gross changes, predominantly partial (intragenic) deletions. Total gene deletions at the HPRT locus are rare except in the TK6 cell line. At the APRT locus in CHO cells, 80-97% of spontaneous mutations are due to base-pair changes, the remainder being, mostly, partial deletions. The latter can extend upstream in the 5' direction but not beyond the APRT gene in the 3' direction. At the human HLA-A locus (T lymphocytes), the percentage of mutations with normal Southern patterns is lower than that for HPRT, and in the range of 50-60%. At the HLA-A locus, mitotic recombination contributes substantially to the mutation spectrum (approximately 30% of mutations recovered) and this is likely to be true of the TK locus in the TK6 cell line as well. With a few exceptions, most of the radiation-induced mutations show altered Southern patterns and are consistent with their being deletions and/or other gross changes (HPRT, 70-90% (CHO); 50-85% (TK6); 50-75% (T lymphocytes); TK, 60-80% (TK6); HLA-A, 80% (T lymphocytes); DHFR, 100% (CHO]. The exceptions are APRT mutations in CHO cells (16-20% of mutants with deletions or other changes) and HPRT mutations in T lymphocytes from A-bomb survivors (15-25%); the latter finding is consistent with the occurrence of in vivo selection against HPRT mutant cells. In cases of HPRT intragenic deletions analyzed (CHO cells and V79 Chinese hamster cells), there is evidence for a non-random distribution of breakpoints. The spontaneous mutation frequencies vary widely, from about 0.04/10(6) cells (sickle cell mutations at the human HBB locus) to 30.8/10(6) cells (HLA-A mutations in T lymphocytes) and are dependent on the locus, the system employed and a number of other factors. Those for the other loci fall between these limits.(ABSTRACT TRUNCATED AT 400 WORDS)  相似文献   

4.
5.
Heteroduplex-induced mutagenesis in mammalian cells   总被引:3,自引:0,他引:3  
We have shown previously that heteroduplexes containing single-stranded loops are repaired efficiently in monkey cells, but not always correctly: 2% of the repair products acquired mutations within a 350 base-pair target (Weiss, U. and Wilson, J.H., Proc. Natl. Acad. Sci. USA 87:1123-1126, 1987). The structures of the mutant genomes, which are described here, are consistent with an error-prone repair system. The spectrum of mutations includes about 25% point mutations and 75% rearrangements, which consist of deletions, duplications, and substitutions. The mutations are clustered in the vicinity of single-stranded loops in the original heteroduplex. The high frequency of mutation, their clustering, and the positions of rearrangement endpoints suggest that the mutations were generated during repair of the heteroduplexes.  相似文献   

6.
L Ghisolfi  AC Keates  X Hu  DK Lee  CJ Li 《PloS one》2012,7(8):e43628
The cancer stem cell (CSC) model posits the presence of a small number of CSCs in the heterogeneous cancer cell population that are ultimately responsible for tumor initiation, as well as cancer recurrence and metastasis. CSCs have been isolated from a variety of human cancers and are able to generate a hierarchical and heterogeneous cancer cell population. CSCs are also resistant to conventional chemo- and radio-therapies. Here we report that ionizing radiation can induce stem cell-like properties in heterogeneous cancer cells. Exposure of non-stem cancer cells to ionizing radiation enhanced spherogenesis, and this was accompanied by upregulation of the pluripotency genes Sox2 and Oct3/4. Knockdown of Sox2 or Oct3/4 inhibited radiation-induced spherogenesis and increased cellular sensitivity to radiation. These data demonstrate that ionizing radiation can activate stemness pathways in heterogeneous cancer cells, resulting in the enrichment of a CSC subpopulation with higher resistance to radiotherapy.  相似文献   

7.
8.
Exposure to ionizing radiation can result in delayed effects that can be detected in the progeny of an irradiated cell multiple generations after the initial exposure. These effects are described under the rubric of radiation-induced genomic instability and encompass multiple genotoxic endpoints. We have developed a green fluorescence protein (GFP)-based assay and demonstrated that ionizing radiation induces genomic instability in human RKO-derived cells and in human hamster hybrid GM10115 cells, manifested as increased homologous recombination (HR). Up to 10% of cells cultured after irradiation produce mixed GFP(+/-) colonies indicative of delayed HR or, in the case of RKO-derived cells, mutation and deletion. Consistent with prior studies, delayed chromosomal instability correlated with delayed reproductive cell death. In contrast, cells displaying delayed HR showed no evidence of delayed reproductive cell death, and there was no correlation between delayed chromosomal instability and delayed HR, indicating that these forms of genome instability arise by distinct mechanisms. Because delayed hyperrecombination can be induced at doses of ionizing radiation that are not associated with significantly reduced cell viability, these data may have important implications for assessment of radiation risk and understanding the mechanisms of radiation carcinogenesis.  相似文献   

9.
Molecular analysis of mutagenesis in mammalian cells   总被引:1,自引:0,他引:1  
Mammalian cells are constantly facing various types of mutagens. However, due to the high complexity of the cell genome, the molecular analysis of mutagenesis has not yet been possible. Therefore, we have used simian virus 40 (SV40) as a biological and molecular probe to characterize mutagenesis at the nucleotide level. By using a reversion assay from a temperature-sensitive phenotype towards a wild-type phenotype, we have analysed mutagenesis induced by u.v.-light and by apurinic sites (Ap sites). We report here experiments allowing us to quantify and to compare the mutagenic efficiency of various DNA lesions measured on the SV40 genome. The Ap sites are very mutagenic in this type of assay. The molecular analysis of u.v.-induced mutagenesis reveals that mutations correspond to single base-pair substitutions always located opposite Py-Py lesions. The mutations are almost equally distributed between transition and transversion types, and between the 5' and the 3' side of the Py-Py targets. These results demonstrate for the first time in animal cells the existence of targeted mutations induced by u.v.-light. We propose therefore, the use of SV40 as an efficient biological and molecular probe for assaying mutagenic pathways in mammalian cells.  相似文献   

10.
Aristolochic acids I and II (AA-I, AA-II) are found in all Aristolochia species. Ingestion of these acids either in the form of herbal remedies or as contaminated wheat flour causes a dose-dependent chronic kidney failure characterized by renal tubulointerstitial fibrosis. In ∼50% of these cases, the condition is accompanied by an upper urinary tract malignancy. The disease is now termed aristolochic acid nephropathy (AAN). AA-I is largely responsible for the nephrotoxicity while both AA-I and AA-II are genotoxic. DNA adducts derived from AA-I and AA-II have been isolated from renal tissues of patients suffering from AAN. We describe the total synthesis, de novo, of the dA and dG adducts derived from AA-II, their incorporation site-specifically into DNA oligomers and the splicing of these modified oligomers into a plasmid construct followed by transfection into mouse embryonic fibroblasts. Analysis of the plasmid progeny revealed that both adducts blocked replication but were still partly processed by DNA polymerase(s). Although the majority of coding events involved insertion of correct nucleotides, substantial misincorporation of bases also was noted. The dA adduct is significantly more mutagenic than the dG adduct; both adducts give rise, almost exclusively, to misincorporation of dA, which leads to AL-II-dA→T and AL-II-dG→T transversions.  相似文献   

11.
Using fluorescence in situ hybridization with human band-specific DNA probes we examined the effect of ionizing radiation on the intra-nuclear localization of the heterochromatic region 9q12-->q13 and the euchromatic region 8p11.2 of similar sized chromosomes 9 and 8 respectively in confluent (G1) primary human fibroblasts. Microscopic analysis of the interphase nuclei revealed colocalization of the homologous heterochromatic regions from chromosome 9 in a proportion of cells directly after exposure to 4 Gy X-rays. The percentage of cells with paired chromosomes 9 gradually decreased to control levels during a period of one hour. No significant changes in localization were observed for chromosome 8. Using 2-D image analysis, radial and inter-homologue distances were measured for both chromosome bands. In unexposed cells, a random distribution of the chromosomes over the interphase nucleus was found. Directly after irradiation, the average inter-homologue distance decreased for chromosome 9 without alterations in radial distribution. The percentage of cells with inter-homologue distance <3 micro m increased from 11% in control cells to 25% in irradiated cells. In contrast, irradiation did not result in significant changes in the inter-homologue distance for chromosome 8. Colocalization of the heterochromatic regions of homologous chromosomes 9 was not observed in cells irradiated on ice. This observation, together with the time dependency of the colocalization, suggests an underlying active cellular process. The biological relevance of the observed homologous pairing remains unclear. It might be related to a homology dependent repair process of ionizing radiation induced DNA damage that is specific for heterochromatin. However, also other more general cellular responses to radiation-induced stress or change in chromatin organization might be responsible for the observed pairing of heterochromatic regions.  相似文献   

12.
13.
14.
Spontaneous mutant frequency in the male germline increases with age, thereby increasing the risk of siring offspring with genetic disorders. In the present study we investigated the effect of age on ionizing radiation-induced male germline mutagenesis. lacI transgenic mice were treated with ionizing radiation at 4-, 15- and 26-month-old, and mutant frequencies were determined for pachytene spermatocytes and round spermatids at 15 days or 49 days after ionizing radiation treatment. Cells collected 15 days after treatment were derivatives of irradiated differentiating spermatogenic cells while cells collected 49 days later were derivatives of spermatogonial stem cells. The results showed that (1) spontaneous mutant frequency increased in spermatogenic cells recovered from nonirradiated old mice (26-months-old), particularly in the round spermatids; (2) mutant frequencies were significantly increased in round spermatids obtained from middle-aged mice (15-months-old) and old age mice (26-months-old) at 15 and 49 days after irradiation compared to the sham-treated old mice; and (3) pachytene spermatocytes obtained from 15- or 26-month-old mice displayed a significantly increased mutant frequency at 15 days post irradiation. This study indicates that age modulates the mutagenic response to ionizing radiation in the male germline.  相似文献   

15.
DNA-chain elongation rates, determined by sedimentation analysis, were found to be similar in control and ataxia-telangiectasia lymphoblastoid cells. A γ-radiation dose of 6 Gray, which had previously been shown to have a marked inhibitory effect on initiation of DNA replication, had no appreciable effect on elongation rates in either cell type. Elongation rates were also determined at 20 Gray of γ-rays by pulsing cells with [3H]thymidine prior to irradiation to avoid anomalous sedimentation behaviour. At this radiation dose elongation was almost completely inhibited in control cells while little or no inhibition was observed in ataxia-telangiectasia cells. Deoxyribonucleoside triphosphate pool equilibration times were not altered at either dose.  相似文献   

16.
17.
The relationship between bromodeoxyuridine (BrdUrd) mutagenesis in mammalian cells and the effects of BrdUrd on deoxyribonucleoside triphosphate pools was analyzed. It was found that the exposure of Syrian hamster melanoma cells to mutagenic concentrations of BrdUrd resulted in the formation of a large bromodeoxyuridine triphosphate (BrdUTP) pool, which remained at a high level for several days. In contrast, the size of the deoxycytidine triphosphate (dCTP) pool dropped rapidly after the addition of BrdUrd, reached a minimum at about 6 h, and then expanded gradually to nearly its original level over the next 3 days. The addition of lower concentrations of BrdUrd, which had less of a mutagenic effect, resulted in the formation of a smaller BrdUTP pool and a slightly smaller drop in the dCTP pool. When a high concentration of deoxycytidine was added at the same time as a normally mutagenic concentration of BrdUrd, the drop in the dCTP pool was prevented, as was BrdUrd mutagenesis. In all of these experiments, mutagenesis was related to the ratio of BrdUTP to dCTP in the cells. In addition, it was shown that mutagenesis occurred primarily during the first 24 h of BrdUrd exposure, when the BrdUTP/dCTP ratio was at its highest level. It appears that there is a critical ratio of BrdUTP to dCTP that must be attained for high levels of mutagenesis to occur and that the extent of mutagenesis is related to the ratio of the BrdUrd and dCTP pools.  相似文献   

18.
19.
Estimates of genetic risks of radiation exposure of humans are traditionally expressed as expected increases in the frequencies of genetic diseases (single-gene, chromosomal and multifactorial) over and above those of naturally-occurring ones in the population. An important assumption in expressing risks in this manner is that gonadal radiation exposures can cause an increase in the frequency of mutations and that this would result in an increase in the frequency of genetic diseases under study. However, despite compelling evidence for radiation-induced mutations in experimental systems, no increases in the frequencies of genetic diseases of concern or other adverse effects (i.e., those which are not formally classified as genetic diseases), have been found in human studies involving parents who have sustained radiation exposures. The known differences between spontaneous mutations that underlie naturally-occurring single-gene diseases and radiation-induced mutations studied in experimental systems now permit us to address and resolve these issues to some extent. The fact that spontaneous mutations (among which are point mutations and DNA deletions generally restricted to the gene) originate through a number of different mechanisms and that the latter are intimately related to the DNA organization of the genes, are now well-documented. Further, spontaneous mutations include those that cause diseases through loss of function as well as gain of function of genes. In contrast, most radiation-induced mutations studied in experimental systems (although identified through the phenotypes of the marker genes) are predominantly multigene deletions which cause loss of function; the recoverability of an induced deletion in a livebirth seems dependent on whether the gene and the genomic region in which it is located can tolerate heterozygosity for the deletion and yet be compatible with viability. In retrospect, the successful mutation test systems (such as the mouse specific locus test) used in radiation studies have involved genes which are non-essential for survival and are also located in genomic regions, likewise non-essential for survival. In contrast, most of the human genes at which induced mutations have been looked for, do not seem to have these attributes. The inference therefore is that the failure to find induced germline mutations in humans is not due to the resistance of human genes to induced mutations but due to the structural and functional constraints associated with their recoverability in livebirths. Since the risk of inducible genetic diseases in humans is estimated using rates of "recovered" mutations in mice, there is a need to introduce appropriate correction factors to bridge the gap between these rates and the rates at which mutations causing diseases are potentially recoverable in humans. Since the whole genome is the "target" for radiation-induced genetic damage, the failure to find increases in the frequencies of specific single-gene diseases of societal concern does not imply that there are no genetic risks of radiation exposures: the problem lies in delineating the phenotypes of recoverable genetic damage that are recognizable in livebirths. Data from studies of naturally-occurring microdeletion syndromes in humans and those from mouse radiation studies are instructive in this regard. They (i) support the view that growth retardation, mental retardation and multisystem developmental abnormalities are likely to be among the quantitatively more important adverse effects of radiation-induced genetic damage than mutations in a few selected genes and (ii) underscore the need to expand the focus in risk estimation from known genetic diseases (as has been the case thus far) to include these induced adverse developmental effects although most of these are not formally classified as "genetic diseases". (ABSTRACT TRUNCATED)  相似文献   

20.
Mammalian cells were after irradiation suspended in melted agarose, and casted on microscope slides. The slides were after gelling at 0°C immersed in a neutral detergent solution which lysed the cells. A weak electric field (5 V/cm) was then applied over the gel for 5 minutes. The DNA in the gel was stained with the fluorescent dye acridine orange and gives a green emission in a microscope photometer. DNA had migrated towards the anode and this migration was more pronounced in irradiated than in control cells. The differences in migration pattern were quantitatively measured. The lower detection limit was below 0.5 Gy and a plateau in the dose-effect curve was reached at about 3 Gy. In repair experiments residual DNA damage could be observed after postirradiation incubation for 60 minutes.The advantages of the method is: no radioactive labelling and only a few number of cells is required.  相似文献   

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