Chromosome translocations are an established biomarker of cumulative exposure to external ionising radiation. Airline pilots are exposed to cosmic ionising radiation, but few flight crew studies have examined translocations in relation to flight experience.
We determined the frequency of translocations in the peripheral blood lymphocytes of 83 airline pilots and 50 comparison subjects (mean age 47 and 46 years, respectively). Translocations were scored in an average of 1039 cell equivalents (CE) per subject using fluorescence in situ hybridisation (FISH) whole chromo-some painting and expressed per 100 CE. Negative binomial regression models were used to assess the relationship between translocation frequency and exposure status and flight years, adjusting for age, diagnostic x ray procedures, and military flying.
There was no significant difference in the adjusted mean translocation frequency of pilots and comparison subjects (0.37 (SE 0.04) vs 0.38 (SE 0.06) translocations/100 CE, respectively). However, among pilots, the adjusted translocation frequency was significantly associated with flight years (p = 0.01) with rate ratios of 1.06 (95% CI 1.01 to 1.11) and 1.81 (95% CI 1.16 to 2.82) for a 1- and 10-year incremental increase in flight years, respectively. The adjusted rate ratio for pilots in the highest compared to the lowest quartile of flight years was 2.59 (95% CI 1.26 to 5.33).
This data suggests that pilots with long-term flying experience may be exposed to biologically significant doses of ionising radiation. Epidemiological studies with longer follow-up of larger cohorts of pilots with a wide range of radiation exposure levels are needed to clarify the relationship between cosmic radiation exposure and cancer risk.
We exposed human peripheral lymphocytes in vitro to 0.3 and 1 Gy of 60Co gamma rays to evaluate whether the ability and sensitivity to detect chromosomal aberrations by chromosome painting is independent or not to the specific paint probes. To detect structural aberrations (translocations), we painted chromosome spreads simultaneously with two whole-chromosome libraries for chromosomes 1, 2, 3, 4, 5, 6, 7, 11, 13, 16, and 18. To compare the rate of chromosome translocations detected by the different pairs of chromosomes, data were normalized according to the fraction of genome painted and evaluated by unconditional logistic regression. Our results show that any combination of paint probes can be used to score induced chromosomal aberrations. We observed that the amounts of translocations are dose dependent and quite homogeneous within each dose of radiation, independently of chromosomes painted. However, the use of small chromosome probes is not recommended because of the high number of cells to be analyzed due to the small amount of genome painted and because it is more difficult to detect translocations in small chromosomes.
The effects of maternal cigarette smoking during pregnancy on structural chromosome aberrations were evaluated in peripheral lymphocytes from 239 mothers and their 241 newborns to determine whether smoking during pregnancy, genetic susceptibility, and race are associated with chromosome aberrations including translocations. Demographic information and cigarette smoking data were obtained via questionnaire. There were 119 Caucasian Americans, 118 African Americans, and 2 Asian Americans. The average maternal age was 24.9 ± 5.8 (mean ± S.D.) years. Thirty-nine percent of the Caucasian Americans and 45.4% of the African Americans self-reported that they were active smokers during the index pregnancy. The average number of cigarettes smoked per day was 2.65 ± 5.75 and 1.37 ± 3.17 for Caucasian and African American mothers, respectively. Peripheral blood lymphocytes from the mother and from the fetal side of the placenta were evaluated for chromosome aberrations by whole chromosome painting and for genetic susceptibility using an in vitro bleomycin challenge assay. Spontaneous translocation frequencies in both maternal and newborn lymphocytes were not associated with cigarette smoking, socio-economic status, or education. The absence of a smoking effect may be attributable to the low level of cigarette usage in these subjects. The average bleomycin-induced damage in the maternal and newborn populations was 0.37 ± 0.27 and 0.15 ± 0.14 breaks per cell, respectively, a difference that was highly significant (p < 0.0001). In newborns there was a positive association between bleomycin sensitivity and the frequencies of aberrations as measured by chromosome painting: p ≤ 0.0007 for dicentrics and fragments, and p ≤ 0.002 for translocations. Caucasian American newborns demonstrated a significant association between dicentrics and fragments as measured by painting, and bleomycin sensitivity (p ≤ 0.0002), but no such association was observed for African American newborns. The results of this study indicate that while differences were observed between African Americans and Caucasian Americans, race does not appear to be a major contributor to chromosome damage in newborns or their mothers. However, peripheral lymphocytes in pregnant women are more susceptible to genetic damage than peripheral lymphocytes in newborns.
cigarette smoking; pregnancy; newborns; mothers; chromosome translocations; genomic susceptibility
From 1995 to 1996 about 15 people suspected of being overexposed to ionizing radiation were referred to the Institute for Nuclear Safety and Protection in Fontenay-aux-Roses, France, for investigation by chromosome aberration analysis. Biological estimates of accidental overexposure were first obtained by scoring radio-induced unstable structural chromosome aberrations (dicentrics, centric rings, and fragments) in peripheral blood lymphocytes. For dose estimates, the yield of these chromosomal aberrations observed in 500 metaphases was compared with the laboratory dose-response relationship established from human blood irradiated in vitro (gamma-rays, 60Co, 0.5 Gy/min). To extend the possibilities of detecting DNA damage from earlier exposures by visualizing stable chromosome aberrations, chromosome painting by fluorescence in situ hybridization (FISH painting) was developed using a cocktail of three composite whole human chromosome-specific DNA probes (numbers 2, 4, and 12). A laboratory calibration curve for scoring terminal and/or reciprocal translocations was established for the same radiation quality and dose rate as those used for conventional cytogenetics (gamma-rays, 60Co, 0.5 Gy/min). For dosimetry purposes, it was also important to verify whether FISH painting could be applied to each human blood sample assessed for conventional expertise. For each individual, 2000 metaphases were scored for the presence or absence of reciprocal and terminal translocations. We present here a comparison between the results obtained by the two technologies for each of the cases studied separately. We describe their similarities or differences and discuss the suitability of using FISH painting for routine expertise analysis.
The human lymphocyte is a premier cell for monitoring chromosome aneuploidy. The lymphocyte is easily obtained, can be studied before and after culture, and has been extensively investigated. Assays available for lymphocytes include the scoring of chromosome breaks (subjective and laborious), the analysis of chromosome abnormalities such as increase or decrease in number (versus normal background), dicentrics etc., and the micronucleus test (presumable end-state phenomena). We propose the monitoring of somatic chromosome translocations in human lymphocytes. Background data available from North America indicate that the frequency of de novo chromosome translocations in Halifax, Portland, Denver, and Atlanta is about 1.7 x 10(-3). The most common translocation arising in lymphocytes is between chromosomes 7 and 14 (with a frequency of 4 x 10(-4). All translocations occurring de novo in human lymphocytes tend to appear balanced with no evidence for loss or gain of chromosome material. Cytogenetic laboratories are processing lymphocytes daily. The resultant photographs and karyotypes are all scorable for de novo translocations. Suitable data on exposure to possible mutagenic agents could be collected in advance of these chromosome studies. This would provide a new method for monitoring chromosome changes in the population. The cost of monitoring lymphocyte chromosomes for somatic translocations would be small, since numerous laboratories study lymphocytes rountinely for clinical diagnostic purposes. There may be merit in availing ourselves of easily available data from a very available species: man.
Exposure to naphthalene, an IARC-classified possible carcinogen and polycyclic aromatic hydrocarbon (PAH), is widespread, though resulting health effects are poorly understood. Metabolites of naphthalene, 1- and 2-naphthol, are measurable in urine and are biomarkers of personal exposure. Chromosomal aberrations (CAs), including translocations, are established markers of cancer risk and a bio-dosimeter of clastogenic exposures. Although prenatal (maternal) PAH exposure predicts CAs in cord blood, few studies have examined CAs in school-age children and none has examined their association with metabolites of specific PAHs.
Using Whole Chromosome Paint Fluorescent in Situ Hybridization, we documented CAs including translocations, in 113 five year old urban minority children and examined their association with concurrent concentrations of PAH metabolites measured in urine.
We report that in lymphocytes, the occurrence and frequency of CAs including translocations are associated with levels of urinary 1- and 2-naphthol. When doubling the levels of urinary naphthols, gender-adjusted Odds Ratio (OR) for CAs are 1.63 (95%CI: 1.21, 2.19) and 1.44 (95%CI: 1.02, 2.04) for 1-and 2-naphthol respectively; and for translocations: OR=1.55 (95%CI: 1.11-2.17) and 1.92 (95%CI: 1.20-3.08) for 1- and 2-naphthol respectively.
Our results demonstrate that markers of exposure to naphthalene in children are associated with translocations in a dose related manner, and that naphthalene may be a clastogen.
Indoor exposure to elevated levels of naphthalene is prevalent in large regions of the world. This study is the first to present an association between a marker of naphthalene exposure and a pre-carcinogenic effect in humans.
Naphthalene; Chromosomal aberrations; Polycyclic aromatic hydrocarbons; Cohort study; Postnatal
The U.S. population has nearly one radiographic examination per person per year and concern about cancer risks associated with medical radiation has increased. Radiologic technologists were surveyed to determine whether their personal cumulative exposure to diagnostic x-rays was associated with increased frequencies of chromosome translocations, an established radiation biomarker and possible intermediary suggesting increased cancer risk. Within a large cohort of U. S. radiologic technologists, 150 provided a blood sample for whole chromosome painting and were interviewed about past x-ray examinations. The number and types of examinations reported were converted to a red bone marrow (RBM) dose score with units that approximated 1 mGy. The relationship between dose score and chromosome translocation frequency was assessed using Poisson regression. The estimated mean cumulative RBM radiation dose score was 49 (range 0 – 303). After adjustment for age, translocation frequencies significantly increased with increasing RBM dose score with an estimate of 0.004 translocations per 100 cell equivalents per score unit (95% confidence interval 0.002 to 0.007; P < 0.001). Removing extreme values or adjustment for gender, cigarette smoking, occupational radiation dose, allowing practice x-rays while training, work with radioisotopes, and radiotherapy for benign conditions did not affect the estimate. Cumulative radiation exposure from routine x-ray examinations was associated independently with increased chromosome damage, suggesting the possibility of elevated long-term health risks, including cancer. The slope estimate was consistent with expectation based on cytogenetic experience and atomic bomb survivor data.
Radiation exposure; diagnostic x-rays; chromosome translocations; FISH; risk factors
Non-random chromosomal translocations are frequently associated with a
variety of cancers, especially hematologic malignancies and childhood sarcomas
In addition to their diagnostic utility, chromosomal translocations are
increasingly being used in the clinic to guide therapeutic decisions. However,
the mechanisms which cause these translocations remain poorly understood.
Illegitimate V(D)J recombination, class switch recombination, homologous
recombination, non-homologous end joining, and genome fragile sites all have
potential roles in the production of non-random chromosomal translocations. In
addition, mutations in DNA repair pathways have been implicated in the
production of chromosomal translocations in humans, mice, and yeast. Although
initially quite surprising, the identification of these same oncogenic
chromosomal translocations in peripheral blood from healthy individuals strongly
suggests that the translocation is not sufficient to induce malignant
transformation, and that complementary mutations are required to produce a frank
Chromosomal translocation; hematologic malignancy; leukemia; lymphoma; non-homologous end-joining; DNA repair
Fluorescence in situ hybridization with chromosome-specific composite DNA probes (chromosome painting) is a reliable and efficient method for detecting structural chromosome aberrations. Painting is now being used to quantify chromosome damage in many human populations. In one such study we evaluated 91 unexposed people ranging in age from birth (cord bloods) to 79. We established a baseline frequency of stable aberrations that showed a highly significant curvilinear increase with age (p < 0.00001) that accounted for 70% of the variance among donors. The magnitude of this effect illustrates the importance of understanding the cytogenetic changes that occur with age, which is particularly important for quantifying the effects of prior adverse environmental, occupational, or accidental exposure. In this paper we use the data obtained in our previous study to characterize the distribution of stable aberrations by age and pack-years of cigarette smoking. We also provide estimates of the number of cell equivalents that need to be scored to detect a given increase in aberrations above the background level surveyed in this population.
Most mutagens and genotoxic carcinogens are efficient inducers of chromosomal alterations in exposed cells. Two important classes of aberrations, namely structural and numerical, are recognized and both types of aberrations are associated with congenital abnormalities and neoplasia in humans. These alterations can be easily detected and quantified in human peripheral blood lymphocytes. Conventional staining techniques can be used to detect these aberrations; this technique was used to estimate absorbed dose in the case of a radiation accident in Goiania, Brazil. A recently introduced fluorescent in situ hybridization technique (FISH) using DNA probes has increased the sensitivity and ease of detecting chromosome aberrations, especially stable chromosome aberrations. This technique allows, to some extent, the estimation of absorbed radiation dose from past exposures. Numerical aberrations can be directly estimated in metaphases by counting the number of FISH-painted chromosomes. Micronuclei are formed by lagging chromosome fragments or whole chromosomes during the anaphase stage of cell division. The nature of micronuclei as to whether they possess a centromere can be determined either by CREST staining (calcinosis, Raynoud's phenomenon, esophageal dysmotility, sclerodactyly, telangiectasia) or FISH with centromere-specific DNA probes. In several carcinogen-exposed populations, such as heavy smokers or people exposed to arsenic, aneuploidy appears to be more common than structural aberrations. In victims of radiation accidents, aneuploidy (hyperploidy) has been found to be common in addition to structural aberrations.
Genotoxic carcinogens in cigarette smoke interact with DNA, causing cytotoxicity. Cytogenetic damage therefore seems to be an excellent biomarker for determining the effect of exposure to chromosome-damaging agents in cigarette smoke.
To study the utility of measurement of frequency of satellite associations (SA) as a biomarker for chromosomal damage using cytogenetic assay in peripheral blood lymphocytes.
Materials and Methods:
This study was conducted on 30 smokers and 30 nonsmokers drawn from a rural population of South India. Smokers were divided into three groups of ten each based on their smoking index (SI) (group I: SI < 150, group II: SI 150–300, and group III: SI > 300) and the frequency of SAs was studied.
The frequency of SAs was significantly greater in smokers than in nonsmokers and the frequency of SAs among the smokers was also seen to increase with increase in SI.
The results of this study indicate that the genotoxic effect of cigarette smoke on chromosomes increases with smoking intensity. These findings can be used to support smoking cessation interventions.
Chromosome damage; genotoxicity; satellite associations; smoking index
To gain a better understanding of the mechanism of chromosomal translocations in cancer, we investigated the spatial proximity between CBFB and MYH11 genes involved in inv(16)(p13q22) found in acute myeloid leukemia patients. Previous studies have demonstrated a role for spatial genome organization in the formation of tumorigenic abnormalities. The non-random localization of chromosomes and, more specifically, of genes appears to play a role in the mechanism of chromosomal translocations. Here, two-color fluorescence in situ hybridization and confocal microscopy were used to measure the interphase distance between CBFB and MYH11 in hematopoietic stem cells, where inv(16)(p13q22) is believed to occur, leading to leukemia development. The measured distances in hematopoietic stem cells were compared to mesenchymal stem cells, peripheral blood lymphocytes and fibroblasts, as spatial genome organization is determined to be cell-type specific. Results indicate that CBFB and MYH11 are significantly closer in hematopoietic stem cells compared to all other cell types examined. Furthermore, the CBFB-MYH11 distance is significantly reduced compared to CBFB and a control locus in hematopoietic stem cells, although separation between CBFB and the control is ~70% of that between CBFB and MYH11 on metaphase chromosomes. Hematopoietic stem cells were also treated with fragile site-inducing chemicals since both genes contain translocation breakpoints within these regions. However, treatment with fragile site-inducing chemicals did not significantly affect the interphase distance. Consistent with previous studies, our results suggest that gene proximity may play a role in the formation of cancer-causing rearrangements, providing insight into the mechanism of chromosomal abnormalities in human tumors.
A complex familial chromosome translocation has been ascertained by combining classical cytogenetics and CISS (chromosomal in situ suppression). Cytogenetic analysis of a chorionic villus sample with G banding showed a 47,XX,-2, +der(2)t(2;22),+der(22)t(2;22) karyotype. Analysis of peripheral blood lymphocytes from the parents by G banding and CISS showed a more complex translocation in the father: 46,XY,-2,-11,-22, +der(2) t(2;11)(q13;q23), +der(11) t(11;22) (q23;q11.2), +der(22) t(2;22) (q13;q11.2). Definitive analysis of cultured amniotic fluid cells showed a double partial trisomy of chromosomes 11 and 22. The couple decided to continue the pregnancy. The fetal karyotype was confirmed at birth. Clinical abnormalities present in our patient were typical of an unbalanced 11;22 translocation. Our findings confirm that chromosome painting techniques allow a better characterisation of complex chromosome rearrangements which may be difficult to detect in G banded karyotypes.
Informative studies of cancer risks associated with medical radiation are difficult to conduct owing to low radiation doses, poor recall of diagnostic X rays, and long intervals before cancers occur. Chromosome aberrations have been associated with increased cancer risk and translocations are a known radiation biomarker. Seventy-nine U.S. radiologic technologists were selected for blood collection, and translocations were enumerated by whole chromosome painting. We developed a dose score to the red bone marrow for medical radiation exposure from X-ray examinations reported by the technologists that they received as patients. Using Poisson regression, we analyzed translocations in relation to the dose scores. Each dose score unit approximated 1 mGy. The estimated mean cumulative red bone marrow radiation dose score was 42 (range 1–265). After adjustment for age, occupational radiation, and radiotherapy for benign conditions, translocation frequencies significantly increased with increasing red bone marrow dose score with an estimate of 0.007 translocations per 100 CEs per score unit (95% CI, 0.002 to 0.013; P = 0.01). Chromosome damage has been linked with elevated cancer risk, and we found that cumulative radiation exposure from medical X-ray examinations was associated with increased numbers of chromosome translocations.
AIMS/BACKGROUND: To assess the presence of bcl-2/JH rearrangements in bone marrow and peripheral blood lymphocytes from patients affected by diseases other than malignant lymphomas. The t(14;18) (q32;q21) translocation, which juxtaposes the bcl-2 oncogene on chromosome 18 and the JH segment of the immunoglobulin heavy chain (IgH) genes on chromosome 14, is found frequently in follicular lymphomas. METHODS: A sensitive semi-nested polymerase chain reaction (PCR) was used to detect t(14;18) translocation in bone marrow aspirates and peripheral blood lymphocytes from 48 patients. In 137 additional individuals peripheral blood lymphocytes only were tested. RESULTS: Cells carrying bcl-2/JH rearrangements were detected in about a quarter of the bone marrow samples and half of the peripheral blood lymphocyte samples. In seven patients, t(14;18) positive cells were found in both the bone marrow and peripheral blood lymphocyte samples. The size of the PCR products and bcl-2/JH DNA sequence analysis showed that the same t(14;18) carrying clone was present in the bone marrow and the corresponding peripheral blood lymphocyte samples in three of these seven patients. Some patients had more than one bcl-2/JH rearrangement. There was no significant correlation between age and the translocation incidence. Cells carrying the t(14;18) translocation were present in peripheral blood lymphocyte samples with a similar incidence--between 47% and 52% in all age groups from 20 to 79 years. Patients older than 80 years had a lower (37%) but not significantly different incidence. CONCLUSIONS: These findings suggest that patients affected by non-lymphoid diseases may have several t(14;18) carrying cells and some of them undergo a clonal expansion. Whether individuals with t(14;18) positive cells are at a higher risk of lymphoid malignancies remains unanswered and further epidemiological studies are required.
Aarray painting is a technique that uses microarray technology to rapidly map chromosome translocation breakpoints. previous methods to map translocation breakpoints have used fluorescence in situ hybridization (FIsH) and have consequently been labor-intensive, time-consuming and restricted to the low breakpoint resolution imposed by the use of metaphase chromosomes. array painting combines the isolation of derivative chromosomes (chromosomes with translocations) and high-resolution microarray analysis to refine the genomic location of translocation breakpoints in a single experiment. In this protocol, we describe array painting by isolation of derivative chromosomes using a MoFlo flow sorter, amplification of these derivatives using whole-genome amplification and hybridization onto commercially available oligonucleotide microarrays. although the sorting of derivative chromosomes is a specialized procedure requiring sophisticated equipment, the amplification, labeling and hybridization of Dna is straightforward, robust and can be completed within 1 week. the protocol described produces good quality data; however, array painting is equally achievable using any combination of the available alternative methodologies for chromosome isolation, amplification and hybridization.
Microdissection and fluorescence in situ hybridisation (FISH) were used to elucidate the nature of a complex chromosome translocation, after GTG banding failed in the complete characterisation of the structural rearrangement between chromosomes 6 and 12. These chromosomes were painted with chromosome specific paints and one of the chromosome regions involved in the translocation was isolated by microdissection. Ten copies of the microdissected region were collected with microneedles from GTG banded metaphases, transferred to a collecting drop, and amplified by means of DOP-PCR. The PCR product was labelled with biotin-14-dATP and used as a FISH probe for hybridisation to normal metaphase chromosomes and metaphase chromosomes of the patients (microFISH). FISH with this chromosome region specific painting probe and with chromosome band specific probes enabled the characterisation of a complex chromosome rearrangement with five breakpoints in two chromosomes. This resulted in the following karyotype: 46,XY,t(6;12)(6pter--> 6q12::12q24.1-->12qter;12qter-->12q13.3:: 6q16.2-->6q26::12q13.3-->12q24.1::6q12--> 6q16.2::6q26-->6qter).
We discuss here the results from our studies demonstrating that simple translocations detected by chromosome painting can be used to reconstruct radiation doses for workers exposed within the dose limits and for individuals with past exposure. To be useful, a biomarker for exposure and risk assessment should employ an end point that is highly quantitative, stable over time, and relevant to human risk. Recent advances in chromosome staining using fluorescence in situ hybridization facilitate fast and reliable measurement of simple translocations, a type of DNA damage linked both to prior clastogenic exposure and to risk. In contrast to other biomarkers available, the frequency of simple translocations in individuals exposed to whole-body radiation is stable over time postexposure, has little interindividual variability, and can be measured accurately at low frequencies.
Dermatofibrosarcoma protuberans (DFSP) is a dermal sarcoma typically carrying a translocation between chromosomes 17 and 22 that generates functional platelet-derived growth factor B (PDGFB).
Patients and Methods
Two distinct phase II trials of imatinib (400 to 800 mg daily) in patients with locally advanced or metastatic DFSP were conducted and closed prematurely, one in Europe (European Organisation for Research and Treatment of Cancer [EORTC]) with 14-week progression-free rate as the primary end point and the other in North America (Southwest Oncology Group [SWOG]) with confirmed objective response rate as the primary end point. In the EORTC trial, confirmation of PDGFB rearrangement was required, and surgery was undertaken after 14 weeks if feasible. The SWOG study confirmed t(17;22) after enrollment.
Sixteen and eight patients were enrolled onto the EORTC and SWOG trials, respectively. Tumor size ranged from 1.2 to 49 cm. DFSP was located on head/neck, trunk, and limb in seven, 11, and six patients, respectively, and was classic, pigmented, and fibrosarcomatous DFSP in 13, one, and nine patients, respectively. Metastases were present in seven patients (lung involvement was present six patients). Eleven patients (4%) had partial response as best response, and four patients had progressive disease as best response. Median time to progression (TTP) was 1.7 years. Imatinib was stopped in 11 patients because of progression, one patient because of toxicity, and two patients after complete resection of disease. Median overall survival (OS) time has not been reached; 1-year OS rate was 87.5%.
Imatinib is active in DFSP harboring t(17;22) including fibrosarcomatous DFSP, with objective response rate approaching 50%. Response rates and TTP did not differ between patients taking 400 mg daily versus 400 mg twice a day.
A 6 month old boy presented with bilateral Wilms' tumour. Cytogenetic analysis of the lymphocytes from the patient showed a de novo balanced translocation t(5;6)(q21;q21), which was also present in the tumour material as the sole cytogenetic abnormality. To facilitate the identification of the translocation breakpoints, we have established a lymphoblastoid cell line (MA214L) from the patient which maintains the translocation in culture. We have used Genethon microsatellite markers as sequence tagged sites (STSs) to isolate yeast artificial chromosome (YAC) clones to 5q and 6q from human genomic libraries. Using fluorescence in situ hybridisation (FISH) on metaphase preparations of MA214L, we have physically defined the translocation breakpoints between YAC clones on each chromosome arm. The genetic distance separating the flanking YACs on 6q21 is 3 cM, while that on 5q21 is 4 cM. To date this is the first report of these chromosomal regions being implicated in Wilms' tumourigenesis.
We describe a method, termed reverse chromosome painting, which allows the rapid analysis of the content and breakpoints of aberrant chromosomes. The method involves the sorting of small numbers of the aberrant chromosome from short term blood culture preparations or cell lines by using bivariate flow karyotype analysis. The sorted chromosomes are amplified and biotin labelled enzymatically using a degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR), the product annealed to metaphase spreads from normal subjects, and hybridisation detected using fluorescence in situ hybridisation (FISH). We show the usefulness of this method for routine clinical cytogenetics by the analysis of cases involving an insertion, a deletion, a translocation, and two cases of a chromosome with additional material of unknown origin. The method has particular application for the rapid resolution of the origin of de novo unbalanced chromosome duplications.
A survey for the prevalence of chronic bronchitis in an industrial population in North India is reported. The prevalence of chronic bronchitis is 12.5 percent in 473 subjects between the ages of 17 and 64 years. The prevalence rate of chronic bronchitis is comparable to that observed in areas of low community air pollution in Europe and North America. There is no age-related rise in the frequency of respiratory symptoms. The consumption of tobacco in these subjects is low and is comparable to tobacco consumption of light smokers. The prevalence of chronic bronchitis in smokers is five times the prevalence in non-smokers and is similar to the values reported for light smokers in other surveys. These observations suggest that cigarette smoking is associated with the development of chronic bronchitis, and the differences in the prevalence rate of chronic bronchitis between this survey and other surveys conducted in Europe and North America are mainly due to differences in smoking habits. Air pollution has a minor effect only and ethnic differences do not appear to play any part. Forced expired volume in one second shows a negative correlation with age. It is lower in asymptomatic smokers than in non-smokers and is lower in chronic bronchitis than in controls.
After mitosis, mammalian chromosomes partially decondense to occupy distinct territories in the cell nucleus. Current models propose that territories are separated by an interchromatin domain, rich in soluble nuclear machinery, where only rare interchromosomal interactions can occur via extended chromatin loops. In contrast, recent evidence for chromatin mobility and high frequency of chromosome translocations are consistent with significant levels of chromosome intermingling, with important consequences for genome function and stability. Here we use a novel high-resolution in situ hybridization procedure that preserves chromatin nanostructure to show that chromosome territories intermingle significantly in the nucleus of human cells. The degree of intermingling between specific chromosome pairs in human lymphocytes correlates with the frequency of chromosome translocations in the same cell type, implying that double-strand breaks formed within areas of intermingling are more likely to participate in interchromosomal rearrangements. The presence of transcription factories in regions of intermingling and the effect of transcription impairment on the interactions between chromosomes shows that transcription-dependent interchromosomal associations shape chromosome organization in mammalian cells. These findings suggest that local chromatin conformation and gene transcription influence the extent with which chromosomes interact and affect their overall properties, with direct consequences for cell-type specific genome stability.
The authors apply a novel high-resolution in situ hybridization method that preserves chromatin nanostructure and show that chromosome territories intermingle significantly in the nucleus of human cells.
Groups of male B6C3F1 mice were exposed by inhalation to 0, 25, 50, 100 or 200 p.p.m. ethylene oxide (EO) for up to 48 weeks (6 hours/day, 5 days/week). Animals were sacrificed at 6, 12, 24 and 48 weeks after the start of the exposure for analyses of reciprocal translocations in peripheral blood lymphocytes and germ cells. The frequency of the total chromosomal aberrations in the peripheral blood lymphocytes was significantly increased at the 100 and 200 p.p.m. exposure concentrations at the 12-week time point, at 50, 100 and 200 p.p.m. at the 24-week time point and at all EO concentrations at the 48-week time point. The frequency of stable reciprocal translocations, which can be used as biomarkers, was increased (P < 0.05) at 100 and 200 p.p.m. at the 12-week time point, at 100 and 200 p.p.m. at the 24-week time point and at 50, 100 and 200 p.p.m. at the 48-week time point. No statistically significant increase could be observed in translocation frequencies at the 6-week time point in the peripheral blood lymphocytes. The exposure–response curves were non-linear when the frequencies of translocations were plotted against EO exposure durations or against EO exposure concentrations. There was no effect of exposure concentration rate on reciprocal translocation frequency. Reciprocal translocations induced in spermatogonial stem cells (observed at the sprematocyte stage) showed significant increases in translocation frequencies over controls at all EO concentrations at 48 weeks. However, increases were small and they did not occur in a dose-responsive manner. The statistically significant increase observed at 12 weeks in the spermatocytes was equivocal. This study provides low-level chronic exposure somatic cytogenetic data generated in mice that can be used to support the shape of the tumour dose–response in rodents and humans The germ cell cytogenetic data are discussed in terms of its relevance for a threshold response for genetic effects at low exposures.
We irradiated normal human lymphocytes and fibroblasts with 137Cs γ rays, 3.5 MeV α particles and 1 GeV/amu 56Fe ions and measured the subsequent formation of chromosome-type aberrations by mFISH at the first mitosis following irradiation. This was done for the purposes of characterizing the shape of dose-response relationships and determining the frequency distribution of various aberration types with respect to the parameters of dose, radiation quality and cell type. Salient results and conclusions include the following. For low-LET γ rays, lymphocytes showed a more robust dose response for overall damage and a higher degree of upward curvature compared to fibroblasts. For both sources of high-LET radiation, and for both cell types, the response for simple and complex exchanges was linear with dose. Independent of all three parameters considered, the most likely damage outcome was the formation of a simple exchange event involving two breaks. However, in terms of the breakpoints making up exchange events, the majority of damage registered following HZE particle irradiation was due to complex aberrations involving multiple chromosomes. This adds a decidedly nonlinear component to the overall breakpoint response, giving it a significant degree of positive curvature, which we interpret as being due to interaction between ionizations of the primary HZE particle track and long-range δ rays produced by other nearby tracks. While such track interaction had been previously theorized, to the best of our knowledge, it has never been demonstrated experimentally.