Cytogenetic monitoring of hospital staff exposed to ionizing radiation: optimize protocol considering DNA repair genes variability.

PURPOSE: Chronic occupational exposure to ionizing radiation (IR) induces a wide spectrum of DNA damages. The aim of this study was to assess the frequencies of micronucleus (MN), sister chromatid exchanges (SCE) and to evaluate their association with XRCC1 399 Arg/Gln and XRCC3 241 Thr/Met polymorphisms in Hospital staff occupationally exposed to IR. MATERIALS AND METHODS: A questionnaire followed by a cytogenetic analysis was concluded for each subject in our study. The exposed subjects were classified into two groups based on duration of employment (Group I < 15 years; Group II ≥15years). The genotypes of all individuals (subjects and controls) were determined by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). RESULTS: DNA damage frequencies were significantly greater in IR workers compared with controls (p < .05). However, no association arised between XRCC1 399 Arg/Gln and XRCC3 241 Thr/Met polymorphisms, on one hand, and the severity of DNA damages in the studied cohort of Tunisian population, on the other hand. CONCLUSION: Our data provide evidence for an obvious genotoxic effect associated with IR exposure and reinforce the high sensitivity of cytogenetic assays for biomonitoring of occupationally exposed populations. These results indicate that workers exposed to IR should have periodic monitoring, along their exposure. The variants, rs25487 and rs861539, of XRCC1 and XRCC3 genes have obvious functional effects. Paradoxically, these variants are not associated with the severity of damages, according to used assays, in the studied cohort of Tunisian population, unlike other studies.

Microwave frequency electromagnetic fields (EMFs) produce widespread neuropsychiatric effects including depression.

Non-thermal microwave/lower frequency electromagnetic fields (EMFs) act via voltage-gated calcium channel (VGCC) activation. Calcium channel blockers block EMF effects and several types of additional evidence confirm this mechanism. Low intensity microwave EMFs have been proposed to produce neuropsychiatric effects, sometimes called microwave syndrome, and the focus of this review is whether these are indeed well documented and consistent with the known mechanism(s) of action of such EMFs. VGCCs occur in very high densities throughout the nervous system and have near universal roles in release of neurotransmitters and neuroendocrine hormones. Soviet and Western literature shows that much of the impact of non-thermal microwave exposures in experimental animals occurs in the brain and peripheral nervous system, such that nervous system histology and function show diverse and substantial changes. These may be generated through roles of VGCC activation, producing excessive neurotransmitter/neuroendocrine release as well as oxidative/nitrosative stress and other responses. Excessive VGCC activity has been shown from genetic polymorphism studies to have roles in producing neuropsychiatric changes in humans. Two U.S. government reports from the 1970s to 1980s provide evidence for many neuropsychiatric effects of non-thermal microwave EMFs, based on occupational exposure studies. 18 more recent epidemiological studies, provide substantial evidence that microwave EMFs from cell/mobile phone base stations, excessive cell/mobile phone usage and from wireless smart meters can each produce similar patterns of neuropsychiatric effects, with several of these studies showing clear dose-response relationships. Lesser evidence from 6 additional studies suggests that short wave, radio station, occupational and digital TV antenna exposures may produce similar neuropsychiatric effects. Among the more commonly reported changes are sleep disturbance/insomnia, headache, depression/depressive symptoms, fatigue/tiredness, dysesthesia, concentration/attention dysfunction, memory changes, dizziness, irritability, loss of appetite/body weight, restlessness/anxiety, nausea, skin burning/tingling/dermographism and EEG changes. In summary, then, the mechanism of action of microwave EMFs, the role of the VGCCs in the brain, the impact of non-thermal EMFs on the brain, extensive epidemiological studies performed over the past 50 years, and five criteria testing for causality, all collectively show that various non-thermal microwave EMF exposures produce diverse neuropsychiatric effects.

Finding the genetic determinants of adverse reactions to radiotherapy.

Individual variation in radiosensitivity is thought to be at least partly determined by genetic factors. The remaining difference between individuals is caused by comorbidities, variation in treatment, body habitus and stochastic factors. Evidence for the heritability of radiosensitivity comes from rare genetic disorders and from cell-based studies. To what extent common and rare genetic variants might explain the genetic component of radiosensitivity has not been fully elucidated. If the genetic variants accounting for this heritability were to be determined, they could be incorporated into any future predictive statistical model of adverse reactions to radiotherapy. With the evolution of DNA sequencing and bioinformatics, radiogenomics has emerged as a new research field with the aim of finding the genetic determinants of adverse reactions to radiotherapy. Similar to the investigation of other complex genetic disease traits, early studies in radiogenomics involved candidate gene association studies--many plagued by false associations caused by low sample sizes and problematic experimental design. More recently, some promising genetic associations (e.g. with tumour necrosis factor) have emerged from large multi-institutional cohorts with built-in replication. At the same time, several small- to medium-sized genome-wide association studies (GWAS) have been or are about to be published. These studies will probably lead to an increasing number of genetic polymorphisms that may predict adverse reactions to radiotherapy. The future of the field is to create large patient cohorts for multiple cancer types, to validate the genetic loci and build reliable predictive models. For example, the REQUITE project involves multiple groups in Europe and North America. For further discovery studies, larger GWAS will be necessary to include rare sequence variants through next generation sequencing. Ultimately, radiogenomics seeks to predict which cancer patients will show radiosensitivity or radioresistance, so oncologists and surgeons can alter treatment accordingly to lower adverse reactions or increase the efficacy of radiotherapy.

Cytogenetic studies on newborns from high and normal level natural radiation areas of Kerala in southwest coast of India.

PURPOSE: To study, characterize and compare chromosome aberrations and karyotype anomalies among newborns from high (> 1.5 mGy/y) and normal (≤ 1.5 mGy/y) level natural radiation areas of monazite-sand bearing southwest coast of Kerala in India. MATERIALS AND METHODS: Cord blood samples from newborns were collected from selected Government hospitals in heparinized vials and cultures were set up employing standard microculture techniques, slides were prepared, coded and stained with giemsa. Well spread metaphases were analyzed for chromosome aberrations and karyotype anomalies. RESULTS: A total of 1,267,788 metaphases from 27,295 newborns of mothers aged 17-45 years (17,298 from high and 9,997 from normal level radiation areas) were analyzed during 1986-2007. Frequencies of dicentrics in high and normal level radiation areas were 1.90 ± 0.14 and 2.01 ± 0.26 per 10,000 cells, respectively (Relative frequency [RF] = 0.94; 95% CI: 0.71-1.26). Karyotype anomalies had a frequency of 5.49‰ and 6.7‰, respectively (RF = 0.82; 95% CI: 0.60-1.12). No dose-related trend was observed in chromosome aberrations or karyotype anomalies. CONCLUSION: Frequencies of chromosomal aberration and karyotype anomalies between the newborns from the high level natural radiation area (HLNRA) and normal level natural radiation areas (NLNRA) were very similar.

Conventional case-cohort design and analysis for studies of interaction.

BACKGROUND: The case-cohort study design has received significant methodological attention in the statistical and epidemiological literature but has not been used as widely as other cohort-based sampling designs, such as the nested case-control design. Despite its efficiency and practicality for a wide range of epidemiological study purposes, researchers may not yet be aware of the fact that the design can be analysed using standard software with only minor adjustments. Furthermore, although the large number of options for design and analysis of case-cohort studies may be daunting, they can be reduced to a few simple recommendations. METHODS: We review conventional methods for the design and analysis of case-cohort studies and describe empirical comparisons based on a study of radiation, gene polymorphisms and cancer in the Japanese atomic bomb survivor cohort. RESULTS: Stratified, as opposed to simple, random subcohort selection is recommended, especially for studies of gene-environment interaction, which are notorious for lacking statistical power. Methods based on the score-unbiased exact pseudo-likelihood (or its analogue with stratified case-cohort data) are recommended for use in conjunction with the asymptotic variance estimator. CONCLUSIONS: We present an example of how to implement case-cohort analysis methods using SPSS, a popular statistical package that lacks some of the features necessary to directly adapt and implement published methods based on other software platforms. We also illustrate case-control analysis using Epicure, which provides greater risk-modelling flexibility than other software. Our conclusions and recommendations should help investigators to better understand and apply the case-cohort design in epidemiological research.

Correlations between DNA polymorphism and frequencies of gamma-radiation induced and spontaneous cytogenetic damage.

With the aim of developing genetic tests for elevated and reduced radiation sensitivity, the authors studied the correlations between various genotypes and frequencies of spontaneous and radiation-induced chromosome aberrations in human lymphocytes. Cytogenetic analysis and genotyping (19 sites of detoxification and DNA repair genes) were carried out for a group of cleanup workers of the Chernobyl nuclear power plant accident (83 people) and for a uniform control group of volunteers (97 people). In both cases, the frequencies of chromosome type aberrations were higher in carriers of minor alleles of gene XPD [sites T2251G (Lys751Gln) and G862A (Asp312Asn)] and the "positive" genotypes GSTM1/GSTT1. The polymorphism of these genes did not affect the frequency of aberrations induced by gamma radiation in the control group (1 Gy in vitro), which was associated with genotypes by loci OGG1, XRCC1, and CYP1A1. Thus, in the control group, spontaneous and in vitro induced cytogenetic effects are associated with different groups of polymorphic genes. In the cleanup workers group (irradiated in vivo), the elevated frequency of aberrations was observed in the carriers of those genotypes that typically have a higher level of spontaneous (but not in vitro induced) cytogenetic damage in the control. The genotype "minor XPD + insertion GST," having an estimated incidence of 64% in central Russia, was characterized and found to be strongly associated with an elevated frequency of chromosome type aberrations following irradiation in vivo (OR = 6.9; p = 0.008).

Infección por Trypanosoma cruzi y polimorfismo del Citocromo B del ADN mitocondrial en Triatoma maculata de Anzoategui y Portuguesa, Venezuela / Trypanosoma cruzi infection and cytochrome b polimorphism of mitochondrial dna in Triatoma maculata of Anzoátegui and Portuguesa states, Venezuela

Con el fin de entender la dinámica poblacional de Triatoma maculata, se analizó el polimorfismo genético y los índices de infección con Trypanosoma cruzi, utilizando triatominos provenientes de ecotopos y regiones geográficas diferentes. El índice de infección parasitaria para T. maculata, fue de 29.8% a través de la observación directa al microscopio y 40.3% utilizando el método de reacción en cadena de la polimerasa. Los niveles de infección encontrados incrementan la importancia de T. maculata como vector involucrado en el ciclo de transmisión de T. cruzi. El análisis del polimorfismo de longitud de fragmentos de restricción de una región del gen Cyt B, permitió establecer en forma preliminar, diferencias en los patrones de bandas de este gen, según el origen geográfico de cada población. Esto puede asociarse a cambios relacionados con procesos adaptativos involucrados en la colonización de nuevos hábitats. No se observó variación genética para vectores capturados en diferentes ecotopos de una misma localidad. Sin embargo es evidente la participación del vector en el ciclo de transmisión, mostrando que la presencia de T. maculata en las casas no puede ser ignorada

In order to understand more about the populational dynamics of Triatoma maculata, the genetic polimorphism and the infection indexes of Trypanosoma cruzi were analysed, using triatomine obtained from different ecotopes and geographical regions. The parasitic infection index of T. maculata was 29.8% using the microscope direct observation, and 40.3% by the polymerase chain reaction method. Both methods were important for epidemiological screening of the vectors with low potential of infection. The amplification of one region the Cyt B gene of these organisms, followed by a restriction fragments length polymorphism analysis, allowed us to establish different patterns of bands according to the geographic origin of each population, which indicates the lack of migration between individuals of Portuguesa and Anzoátegui states. These genetic differences may be associated with changes in adaptative events involved in the colonization of new habitats. The lack of polymorphism among vectors collected in different habitats of the same region showed an important genetic flow which has epidemiological implications in the risk of transmission of the disease, showing that the presence of T. maculata in houses cannot be ignored

Influence of the TP53 codon 72 polymorphism on the cellular responses to X-irradiation in fibroblasts from nonagenarians.

In mice, genetic modification of the gene encoding p53 affects both cancer incidence and longevity. In humans, we recently found that a TP53 codon 72 Arginine (Arg) to Proline (Pro) polymorphism affected both cancer incidence and longevity as well. The TP53 codon 72 polymorphism has previously been shown to influence the apoptotic potential of human cells in response to oxidative stress. Here, we studied the influence of this polymorphism on the cellular responses to X-irradiation of fibroblasts obtained from nonagenarians. We found that the average clonogenic survival after X-irradiation was similar for the three TP53 codon 72 genotype groups. As described before, X-irradiation did not induce an appreciable degree of apoptosis in human fibroblasts. However, percentages of senescence-associated (SA)-beta-galactosidase positive cells (p < 0.001), micronucleated cells (p < 0.001) and cells displaying abnormal nuclear morphologies (p < 0.001) significantly increased with the radiation dose. Compared to Arg/Arg fibroblasts, Pro/Pro fibroblasts exhibited higher irradiation dose-dependent increases in SA-beta-galactosidase positive cells (p(interaction) = 0.018), micronucleated cells (p(interaction) = 0.005) and cells displaying abnormal nuclear morphologies (p(interaction) = 0.029) at 3 days after irradiation. Possibly, these differences in cellular responses to stress between the TP53 codon 72 genotypes contribute to the differences in cancer incidence and longevity observed earlier for these genotypes.

The genomics revolution and radiotherapy.

The expansion of our knowledge through the Human Genome Project has been accompanied by the development of new high-throughput techniques, which provide extensive capabilities for the analysis of a large number of genes or the whole genome. These assays can be carried out in various clinical samples at the DNA (genome), RNA (transcriptome) or protein (proteome) level. There is a belief that this genomic revolution, i.e. sequencing of the human genome and developments in high-throughput technology, heralds a future of personalised medicine. For clinical oncology, this progress should increase the possibility of predicting individual patient responses to radiotherapy. This review highlights some of the work involving sparsely ionising radiation and the new technologies.

TP53 codon 72 polymorphism in radiation-associated human papillary thyroid cancer.

The study investigated an association between the germline polymorphism at TP53 codon 72 and the development of papillary thyroid cancer (PTC) following exposure to radiation from the Chernobyl accident. TP53 genotype was examined in 48 pediatric/adolescent (age at diagnosis <18 years) and 68 adult post-Chernobyl patient with PTC, 53 adult patients with sporadic PTC and 313 healthy individuals from Russian-Ukrainian population. In addition, we evaluated loss of heterozygosity for TP53 and the allele expression ratio. The genotype of the patients was correlated with clinicopathological data. Arg TP53 homozygotes were found to be significantly underrepresented among adults with post-Chernobyl PTC, but not in children and adolescents when compared with sporadic PTC cases and the general population. In the tumors, cell transformation did not lead to allelic loss or biased TP53 allele expression in heterozygous individuals. None of TP53 genotypes specifically associated with tumor stage and morphology, however there were particular correlations with lymph node status in certain age groups of radiation-associated cases not seen in sporadic PTCs. The findings suggest TP53 allele combinations other than Arg/Arg may contribute to the risk of development of PTC in individuals exposed to radiation during their late childhood, adolescence or in young adulthood.

Tandem duplication and copy number polymorphism of the SRY gene in patients with sex chromosome anomalies and males exposed to natural background radiation.

Mutations in the SRY gene encompassing the HMG box have been well characterized in gonadal dysgenesis, male infertility and other types of sex chromosome related anomalies (SCRA). However, no information is available on copy number status of this gene under such abnormal conditions. Employing 'Taqman Probe Assay' specific to the SRY gene, we screened 16 DNA samples from patients with SCRA and 36 samples from males exposed to high levels of natural background radiation (HNBR). Patients with SCRA showed 2-16 copies of the SRY gene of which, one, Oxen (49, XYYYY) had eight copies with sequences different from one another. Of the 36 HNBR samples, 12 had one copy whereas 24 harboured 2-8 copies of the SRY gene. A HNBR male 33F had one normal and one mutated copy of this gene. Analysis of 25 DNA samples from blood and semen of normal males showed only one copy of this gene. Despite multiple copies in affected males, fluorescence in-situ hybridization (FISH) with SRY probe detected a single signal on the Y chromosome in HNBR males suggesting its possible localized tandem duplication. Copy number status of the other Y-linked loci is envisaged to augment DNA diagnostics facilitating genetic counselling to affected patients.

Diagnostic x rays, DNA repair genes and childhood acute lymphoblastic leukemia.

A model for childhood leukemia proposes that characteristic chromosomal translocations can arise in utero and that for most cases a second hit occurring postnatally will be necessary. Possible causal mechanisms for leukemias are environmental factors such as ionizing radiation from x rays and inherited susceptibility from polymorphisms in DNA repair genes. We performed a case-control study of childhood acute lymphoblastic leukemia measuring reported postnatal x rays in 701 cases aged 0-14 y and in as many population-based controls matched on age and sex. In addition we performed a case-only study in 207 cases to evaluate the interaction between x ray exposure and polymorphisms in DNA repair genes. There was an increase in risk of leukemia with number of x rays: the adjusted odds ratio for two or more x rays vs. none was 1.48 (95% confidence interval: 1.11-1.97). That risk was slightly higher among girls (odds ratio = 1.67). A polymorphism in the APE gene (ex 5) involved in the base excision repair system was suggestive of an increased risk among boys and a reduced risk among girls. HMLH1 (ex 8), a mismatch repair gene, was associated with reduction of risk among girls. Results from the genetic data are still preliminary and must be interpreted with caution especially because of the relatively small number of genotyped cases. However, ionizing radiation from x rays as well as polymorphisms in DNA repair genes are plausible risk factors for childhood leukemia and should be studied more.

[Genomic disorders in the mononuclear blood cells of those who worked in the cleanup of the accident at the Chernobyl Atomic Electric Power Station]. / Issledovanie genomnykh narushenii v mononuklearakh krovi likvidatorov avarii na ChAES.

The results of molecular investigations of blood mononuclears from 120 clean-up workers after 7-9 years of Chernobyl accident with the total exposure radiation doses ranging from 5 to 76 cGr are presented. Structural polymorphism of the leukemia associated bcr and ribosomal RNA (rRNA) genes were studied using Southern blot hybridization. Allelic polymorphism of bcr gene with characteristic for leukemia allele distribution was detected in 16.6%. Rearrangements of rRNA genes were observed in 13% of Chernobyl accident clean-up workers.

Susceptibility to effects of UVB radiation on induction of contact hypersensitivity as a risk factor for skin cancer in humans.

Normal, healthy human volunteers and patients with proved history of non-melanoma skin cancer have been tested for their capacity to develop contact hypersensitivity to dinitrochlorobenzene (DNCB) following exposure of buttock skin to acute, low-dose ultraviolet B (UVB) radiation. Using a radiation protocol that achieves virtually complete depletion of normal-appearing Langerhans cells from irradiated skin, it was learned that approximately 60% of healthy volunteers developed vigorous contact hypersensitivity (CH) when 2000 micrograms DNCB was painted on the irradiated site. These individuals were designated UVB-resistant, and were distinguished from other individuals, designated UVB-susceptible, who failed to develop contact hypersensitivity following an identical treatment protocol. It was then discovered that virtually all (92%) skin cancer patients exposed to UVB and DNCB failed to develop CH, i.e., were UVB-susceptible. In subsequent experiments, epicutaneous application of 2000 micrograms DNCB to unirradiated skin of UVB-susceptible individuals revealed a further distinction between normal persons and skin cancer patients. Approximately 45% of the latter (and none of the former) remained unresponsive (failed to develop contact hypersensitivity following this second attempt at sensitization), implying that they had been rendered immunologically tolerant. These tolerant individuals responded normally to the unrelated hapten, diphencyprone. We conclude that human beings resemble inbred strains of laboratory mice in that some individuals are UVB-susceptible, whereas others are UVB-resistant. Because the incidence of UVB-susceptibility was significantly higher in skin cancer patients, and as specific unresponsiveness could be demonstrated only in these patients, we propose that UVB-susceptibility, as we define it in this hapten system, may be a risk factor for the development of skin cancer.