Spontaneous Micronucleus Frequencies Research Articles

Evidence of chromosomal damage in scleroderma

Evidence of chromosomal damage in scleroderma

Influence of Polymorphisms of DNA Repair and GST Genes on Genotoxic Damage and Mutagen Sensitivity in Workers Occupationally Exposed to Very Low Doses of Ionizing Radiation

The study investigated the influence of genetic polymorphisms of the enzymes for DNA repair and detoxification of reactive intermediates on spontaneous and bleomycin-induced (BLM) genotoxic damage in 43 workers exposed to very low doses of ionizing radiation (IR) (mean cumulative dose 5.31 mSv) and 43 subjects with no occupational exposure to IR (controls). In all the subjects examined, the frequency of chromosome aberrations (CAs) and micronuclei (MN), both spontaneous and BLM-induced, the Comet assay parameters (tail intensity), the genotypic variants of the DNA repair enzymes XRCC1 (Arg194Trp, Arg280His, Arg399Gln), XRCC3 (Thr241Met), XPD (Lys751Gln), and of the detoxification enzymes GSTM1 and GSTT1 (null genotype) and BLMH (A1450G) were determined. Among the biomarkers considered, only the frequency of total CAs (p < 0.05), and in particular of chromosome breaks (p < 0.01), was found to be significantly higher in the exposed workers than the controls. The frequency of spontaneous MN was higher in subjects with at least one allelic variant in XRCC1 than in carriers of the wild-type, but again only in exposed workers (p = 0.046). Linear regression analysis showed a positive dependency of the frequency of spontaneous chromosome breaks on occupational exposure, and a dependency of the frequency of BLM-induced MN negative on occupational exposure and positive on alcohol consumption and the null GSTM1 genotype. In conclusion, the frequency of chromosome breaks seems to be a useful cytogenetic biomarker for exposure to very low doses of IR, while only the combined effect of different gene variants or genetic, occupational, and lifestyle habits factors seems to be able to modulate the genotoxic effect of very low doses of IR.

Cytokinesis Block Micronucleus Cytome (CBMN Cyt) Assay Biomarkers and Their Association With Radiation Sensitivity Phenotype in Prostate Cancer Cases and DNA Repair Gene hOGG1 (C1245G) Polymorphism.

Prostate cancer (PC) is commonly diagnosed cancer in men but only a few risk factors, such as family history, ethnicity, and age have been established. Chromosomal instability is another possible risk factor but this has not been adequately explained previously. In this study, we tested the hypotheses that peripheral blood lymphocytes (PBL) of PC patients have (1) an abnormally high level of chromosomal instability; (2) that they are hypersensitive to ionizing radiation-induced DNA damage; and (3) that these phenotypes are affected by hOGG1 (C1245G) polymorphism. These experiments were performed using the cytokinesis-block micronucleus Cytome (CBMN cyt) assay in PC cases and controls. We found that spontaneous or radiation-induced (3G) micronucleus (MN) frequency is not significantly different between both groups. However, spontaneous frequency of nucleoplasmic bridges (NPBs) and radiation-induced nuclear buds (NBuds) were significantly higher in patients vs. controls (P < 0.0001; P = 0.0005, respectively). In addition, apoptosis and nuclear division index (NDI) was significantly higher in patients compared to controls after radiation treatment (P = 0.006; P = 0.0002, respectively). Furthermore carriage of at least one G allele of hOGG1 (C1245G) polymorphism was associated with a significantly increased odds ratio (OR) to have a base-line MN, NPB, or NBud frequency greater than medium level compared to homozygotes for C allele (OR:1.94, 1.77, 2.36, respectively, P = 0.02; 0.04, and 0.004, respectively). Our results support the hypotheses that those who develop PC have significantly higher level of genomic instability which is further increased in those who carry G allele of the hOGG1 (C1245G) polymorphism. Environ. Mol. Mutagen. 59:813-821, 2018. © 2018 Wiley Periodicals, Inc.

Decreased Chromosomal Damage in Lymphocytes of Obese Patients After Bariatric Surgery

The number of bariatric surgeries being performed worldwide has markedly risen. While the improvement in obesity-associated comorbidities after bariatric surgery is well-established, very little is known about its impact on cancer risk. The peripheral lymphocyte micronucleus test is a widely used method for the monitoring of chromosomal damage levels in vivo, and micronucleus frequency positively correlates with cancer risk. Therefore, the aim of this study was to compare the micronucleus frequency before and after bariatric surgery in obese subjects. Peripheral blood mononuclear cells were collected from 45 obese subjects before and at two time-points after bariatric surgery (6 and 12 months) to assess spontaneous micronucleus frequency. Consistent with the increased cancer risk previously shown, bariatric surgery-induced weight loss led to a significant reduction in lymphocyte micronucleus frequency after 12 months. Interestingly, comorbidities such as type 2 diabetes mellitus and metabolic syndrome further seemed to have an impact on the lymphocyte micronucleus frequency. Our findings may indicate a successful reduction of cancer risk in patients following weight loss caused by bariatric surgery.

Markers of human extraembryonal cells individual radiosensitivity in vitro

Background: Genotoxic effects of ionizing radiation in early stages of human embryonic development can lead to fatal consequences. At the same time, the radiosensitivity of human embryonic and extraembryonic cells is still poorly studied. In this study, the analysis of DNA double-strand break repair effectiveness in human extraembryonal fibroblasts was carried out. Materials and methods. Extraembryonic human fibroblasts was irradiated by 1 Gy gamma-rays using Theratron Equinox (Cancer Research Institute, Tomsk). The level of DNA double strand breaks was assessed using γH2AX foci. Frequency of cytogenetic damage was assessed using micronucleus test conducted with FISH as a frequency of centromere-negative micronuclei. Results. No significant correlation was observed between both endogenous and residual levels of radiation-induced γH2AX foci and frequency of micronuclei after irradiation. It is suggested to be a result of the specificity of extraembryonal fibroblast radiation-induced response. The spontaneous frequency of centromere-negative micronuclei correlated with radiation-induced frequency of centromere-negative micronuclei. Conclusion. It was shown that human extraembryonal fibroblasts ability to repair radiation-induced DNA damage is likely to be reflected by the repair of spontaneous DNA double strand breaks.

Smad7 foci are present in micronuclei induced by heavy particle radiation

DNA damage and reactive oxygen species (ROS) generated by ionizing radiation (IR) activate DNA damage response (DDR) and cytokine signaling pathways, including double strand break (DSB) repair and TGFβ/Smad signaling pathway. Proteins assembled at IR-induced DSB sites can be visualized as foci, including γH2AX, 53BP1, ATM and ATF2. Unrepaired DSBs are thought to be one origin of micronuclei (MN), an indicator of genotoxic stress and chromosomal instability. Studies have detected γH2AX in IR-induced MN, indicating the presence of DSB in MN. Previously we reported that TGFβ downstream proteins Smad7 and phospho-Smad2 (pSmad2) co-localized with DDR proteins following radiation. Here we studied the status of Smad7 and pSmad2 in MN post high linear energy transfer (LET) radiation in human normal and cancerous cells. We observed γH2AX foci in IR-induced MN, whereas 53BP1 and ATF2 were absent. Interestingly, Smad7 foci, but not pSmad2, were detectable in both spontaneous and IR-induced MN. We compared the effect of particle track structures on the yield of MN using 5.6MeV/u boron (B) and 600MeV/u iron (Fe) particles with similar LET (200 and 180keV/μm, respectively) in human fibroblasts. The frequency of MN induced by B was lower than that by Fe particles, albeit the proportion of Smad7-positive to Smad7-negative MN remained constant. An increased frequency of spontaneous MN, with slightly higher ratio of Smad7 or γH2AX positive, was found in human prostate cancer cells (PC3) compared to normal cells. 24h after 1Gy of Fe particles exposure, the yield of MN increased, and the majority (∼70%) carried γH2AX and Smad7. Phospho-ATM (Ser1981) foci were found in both spontaneous and IR-induced MN in PC3 cells, displaying a much lower frequency compared to γH2AX and Smad7. Our data suggest a unique role of Smad7 in IR-induced MN formation, which may associate with DNA repair, apoptosis and genomic instability.

Influence of DNA-repair gene variants on the micronucleus frequency in thyroid cancer patients

The role of different DNA-repair genes (OGG1, XRCC1, XRCC2 and XRCC3) on both the spontaneous and the induced frequency of micronuclei (MN) has been studied in the lymphocytes of a group of 114 patients with differentiated thyroid cancer (DTC). Induction of MN was achieved by treatment of the lymphocytes with 0.5Gy of gamma-radiation.The selected genes are involved in base-excision repair (BER) (OGG1, Ser326Cys; XRCC1, Arg280His and Arg399Gln), and in homologous recombination repair (HRR) (XRCC2, Arg188His and XRCC3, IVS5-14G). Genotyping was carried out by use of the iPLEX (Sequenom) technique.Results indicate that only the OGG1-Ser326Cys polymorphism was able to modulate the MN frequency. This effect was only observed in the spontaneous MN frequency (P=0.016), but not in the MN frequency induced after irradiation. In addition, a strong correlation was observed between spontaneous and induced MN frequency, which would suggest an underlying genetic background.

An investigation of the relationshıp between coronary atherosclerosis and chromosome damage in human peripheral blood lymphocytes

Background: Coronary atherosclerosis is a disease of the coronary arteries and is a result of cumulative processes that are mostly seen in old-age population. Progressive accumulation of oxidative damage to cellular macromolecules, especially DNA, over a lifetime is one of the important causes of aging. This accumulation of damage may result in chromosome instability and this can be measured by micronucleus (MN) technique. Materials and Methods: In this study, MN technique was applied to observe the relationship between chromosome instability and proliferative effectiveness of lymphocytes and coronary atherosclerosis. Non-toxic levels of hydrogen peroxide were also administered to observe any existing differential effect due to this stress inducing oxidizing agent. Spontaneous and H 2 O 2 stress induced MN frequencies and proliferation state of peripheral lymphocytes were measured from both 30 patients diagnosed as coronary atherosclerosis and 30 healthy control individuals of the same age range. Results: There was no significant difference between spontaneous MN frequencies of patient and control groups (P = 0.08). Also, probability index (PI) values between these groups reflected no difference (P = 0.13). Hydrogen peroxide addition did not change spontaneous MN frequencies either in control (P = 0.23) or in patient group (P = 0.81). This non-toxic H 2 O 2 level failed to show any different effect on patients compared to controls (P = 0.41). Although increasing PI values were observed as a result of H 2 O 2 administration only in patient group, the comparisons between the two groups showed no significant difference (P = 0.34). Conclusion: Chromosome instability and proliferation index that were measured by MN technique in this study have no role in discriminating coronary atherosclerosis patients from healthy individuals.

The effect of aging on micronuclei frequency and proliferation in human peripheral blood lymphocytes

INTRODUCTION:Increase in the instability of cellular genome with an increasing age is the result of an accumulation of cellular damage and mutations. This instability which might be observed as chromosome damage or chromosome losses can be measured by the micronucleus technique.AIM:The aim of this study was to investigate the effect of aging and oxidative stress induced by non-toxic levels of H2O2 on micronuclei induction and their relationship to cell proliferation in human peripheral blood lymphocytes.MATERIALS AND METHODS:Healthy volunteers with different ages were choosen. Spontaneous and H2O2 induced micronuclei frequencies were measured in peripheral blood lymphocytes of 30 volunteers by the micronucleus method.RESULTS:Spontaneous micronuclei frequencies increased first then started to decrease after 50 years of age. This biphasic response was significantly higher than micronucleus (MN) frequencies induced by H2O2 (P < 0.05), which followed the similar shape of response to increasing ages with lower frequencies. Proliferative capacity of cells either treated with H2O2 or not did not differ with an increasing age giving similar responses.CONCLUSION:These results indicate biphasic character of chromosome damage; first increase and decrease after 50 years with an increasing age. But this change pattern was not correlated with the steady state of proliferation capacity of cells through an increasing age. Decreases in H2O2-induced MN frequencies compared to spontaneous MN frequencies may be inducing an apoptosis by H2O2 treatment leading to underscoring damaged cells.

The high frequency of spontaneous micronuclei observed in lymphocytes of systemic sclerosis patients: preliminary results

Aim of the study is to assess the presence of spontaneous chromosome damage in patients affected by limited (lSSc) or diffuse (dSSc) Systemic Slerosis, using the micronucleus (MN) assay. We evaluated MN frequency in cultured peripheral lymphocytes of 18 SSc and in a group of 20 healthy controls. Patients were also classified as ACA+, Scl70+, FAN+ according to the presence of the specific anti-nuclear antibodies. We also explored the hypothesis that the extent of cytogenetic alteration might be related to the severity of the pathological condition and/or to the immunological profile. Compared to controls, the patient group as a whole showed significantly higher MN frequencies (10.8+/-4.5 vs. 27.8+/-13.7, p<0.001). No correlation was found between spontaneous chromosome damage and severity of the disease, being MN frequency 33.1+/-17.0 and 19.8+/-2.7 in lSSc and dSSc, respectively. Interestingly, ACA+ subjects displayed the highest MN frequency (36.9+/-15.0), as compared to patients with different antibody pattern (Scl70+, FAN+; 19.7+/-8.2). Our results confirm the presence of chromosomal damage in circulating lymphocytes of SSc patients and would suggest a key role of antibodies to the centromere in determining the observed cytogenetic anomalies.

A semi-automated micronucleus-centromere assay to assess low-dose radiation exposure in human lymphocytes

Purpose: The in vitro micronucleus (MN) assay is a reliable method to assess radiation-induced chromosomal damage in human peripheral blood lymphocytes. It is used to evaluate in vivo radiation over-exposure and to assess in vitro chromosomal radiosensitivity. A limitation of the MN assay is the relatively high and variable spontaneous MN frequency that restricts low-dose estimation to doses of about 0.3 gray (Gy). As radiation-induced MN mainly contain acentric fragments and spontaneous MN originate from lagging chromosomes, both MN types can be distinguished from each other by using fluorescence in situ hybridisation (FISH) with a pan-centromeric probe. The aim of this study was to investigate if the sensitivity, reliability and processing time of the MN assay can be enhanced by combining the automated MN assay with pan-centromere scoring.Materials and methods: Blood samples from 10 healthy donors were irradiated in vitro with low doses of gamma-rays. Dose response curves were determined for fully-automated and semi-automated MN scoring and semi-automated scoring of centromere negative MN (MNCM−).Results: A good correlation was obtained between fully-automated and semi-automated MN scoring (r2 = 0.9973) and between fully automated MN scoring and semi-automated scoring of MNCM− (r2 = 0.998). With the Wilcoxon test, a significant p value was obtained between 0 and 0.2 Gy for the fully-automated MN analysis, between 0 and 0.1 Gy for semi-automated MN analysis and between 0 and 0.05 Gy for semi-automated scoring of MNCM−.Conclusion: The semi-automated micronucleus-centromere assay combines high-speed MN analysis with a more accurate assessment in the low-dose range which makes it of special interest for large-scale radiation applications.

Micronucleus evaluation in mitogen-stimulated lymphocytes of patients with acromegaly

Acromegaly is a syndrome characterized by a sustained elevation of circulating growth hormone and insulin-like growth factor-1 (IGF-1). Insulin-like growth factor-1 is a potent mitogen and has a role in the transformation of normal cells to malignant cells. This study aims to evaluate the spontaneous micronucleus (MN) frequency by using the cytokinesis-block MN assay to determine genetic damage in the lymphocytes of patients with acromegaly. The study was carried out in 20 patients who had active acromegaly and in 20 age- and sex-matched healthy controls. The MN values were measured in binucleated cells obtained from mitogen-stimulated lymphocytes of patients and control subjects. The distribution of binucleated cells with 1, 2, 3, or more MNs was also measured. We found significantly higher MN frequency values in the lymphocytes of acromegalic patients than in those of the control subjects (2.23 ± 0.68 vs 1.03 ± 0.54, P = .001). The MN frequency increased with increasing IGF-1 levels of acromegalic patients (P = .036, R = 0.47). We observed that the number of binucleated cells with 2 MNs was higher for the majority of patients with acromegaly than for control subjects. Furthermore, the receiver operating characteristic curve (area under the curve = 0.914, P < .0001) was calculated to assess the discriminative power of the MN frequency. Our results indicate that increased MN frequency in the lymphocytes of patients with acromegaly may reflect genomic instability and this increased MN frequency may be associated with elevated levels of circulating growth hormone and IGF-1.

CYTOKINESIS-BLOCK MICRONUCLEUS ASSAY IN HUMAN GLIOMA CELLS EXPOSED TO RADIATION

Biological tests are efficient in reflecting the biological influences of several types of generally harmful exposures. The micronucleus assay is widely used in genotoxicity studies or studies on genomic damage in general. We present methodological aspects of cytokinesis-block micronucleus assay performed in human gliomas irradiated in vitro. Eight human glioblastoma cell lines obtained from DSMZ (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Germany) were gamma-irradiated (60Co) over a dose range of 0-10 Gy. Cytokinesis-block micronucleus assay was performed to quantitate cytogenetic damage. The cells were fixed directly on dishes, stained with fluorochrome DAPI and evaluated under fluorescent and phase contrast microscope. The micronucleus frequency was expressed as a micronuclei (MN) per binucleated cell (BNC) ratio, calculated after scoring at least 100 BNC per dish. The frequency of spontaneous MN ranged from 0.17 to 0.613 (mean: 0.29 ± 0.14). After irradiation increase of MN frequency in the range of 0.312 - 2.241 (mean: 0.98 ± 0.68) was found at 10 Gy. Gliomas are extremely heterogenous in regard to cytogenetic effects of irradiation, as shown in this study by cytokinesis-block micronucleus assay. This test is easily performed on irradiated glioma cell lines and can assist in determining their radiosensitivity. However, in order to obtain reliable and reproducible results, precise criteria for MN scoring must be strictly followed. Simultaneous use of fluorescent and phase contrast equipment improves imaging of morphological details and can further optimize MN scoring.

Flow cytometric scoring of micronucleated erythrocytes: an efficient platform for assessing in vivo cytogenetic damage

The relative simplicity of the micronucleated erythrocyte endpoint has made it amenable to automated scoring approaches. Flow cytometry is one such scoring platform that has been employed successfully. This review describes the evolution and properties of flow cytometry-based scoring of micronucleated erythrocytes. The methodology has become widely applied to rodent blood specimens and the high throughput nature of the technology provides a number of advantages over manual microscopic scoring. For instance, the ability to efficiently survey many dose levels and many more cells per specimen relative to microscopy benefits studies that are designed to identify no observable effect levels or lowest observable effect levels. Furthermore, flow cytometry makes it practical to study species with low spontaneous reticulocyte (RET) counts and micronucleus (MN) frequencies, thereby facilitating integration of blood-based micronucleated reticulocyte (MN-RET) frequency measurements into experiments conducted across species of toxicological interest. This capability enhances genotoxicity assessments that have historically been made in dedicated MN tests performed in one species. Importantly, the feasibility of using MN-RET frequencies in blood from humans as an index of genetic damage in bone marrow opens a critical area of application that had not been practical previously. We conclude with recommendations for additional work that is needed to more fully realise the potential of flow cytometric in vivo MN scoring.

Nuclear damage in peripheral lymphocytes of obese and overweight Italian children as evaluated by the γ‐H2AX focus assay and micronucleus test

Childhood obesity, often characterized by a chronic low-grade inflammation, has been associated with an increased risk of developing some types of cancer later in life. Nuclear γ-H2AX foci represent the first detectable response of cells to DNA tumorigenesis lesions, such as the double-strand breaks (DSBs). An excess of micronucleated peripheral lymphocytes was found in subjects with cancer or inflammation-based diseases. We set out to investigate the expression of genome damage, from DNA lesions to chromosome mutations (micronuclei), in overweight and obese children. Using the γ-H2AX focus assay and micronucleus (MN) test, we analyzed peripheral lymphocytes from 119 Italian children classified as normal weight (n=38), overweight (n=20), or obese (n=61). Cultures treated with bleomycin (BLM) were also set up for each child in both assays to check functioning of the apparatus that ensures DNA integrity. We measured serum TNF-α, IL-6, and C-reactive protein (CRP) as markers of inflammation. Overweight and obese children had significantly higher levels of H2AX phosphorylation (0.0191±0.0039 and 0.0274±0.0029 γ-H2AXF/n) and increased MN frequencies (2.30±0.25 and 2.45±0.22‰) than normal-weight children (0.0034±0.0006 γ-H2AXF/n, and 0.92±0.12‰ MN), while all subjects responded to BLM induction, irrespective of their weight status. The fold increase of spontaneous MN frequencies in overweight and obese subjects was 2.5 and 2.7, respectively, well below the corresponding increase in the γ-H2AX foci (5.6- and 8.0-fold, respectively). IL-6 and CRP mean values were significantly higher in obese and overweight children than in controls. Here, we demonstrated that peripheral cells of overweight and obese children showed increased levels of DSBs, which were not completely repaired as part of them has been converted into micronuclei. Characterization of childhood obesity inflammation could be implemented using molecular markers of genome damage.

Comparison of four different treatment conditions of extended exposure in the in vitro micronucleus assay using TK6 lymphoblastoid cells

In the OECD Guideline 487, a total of four extended exposure treatment conditions are proposed for the in vitro micronucleus (MNvit) assay in the presence and absence of a cytokinesis block and with or without a recovery period. This guideline also states that rodent cell lines and human lymphocytes can be used as shown by many validated studies but that human cell lines such as TK6 and HepG2 are not yet validated. In this present study each extended exposure condition was characterized by investigation using TK6 cells and nine chemicals known to be able to induce micronucleus (MN) in rodent cell lines. The results revealed two concerns: six chemicals did not show significant MN induction in the ‘cytokinesis block without recovery period’; two aneugens showed no dose-dependent cytotoxicity in the ‘cytokinesis block with recovery period’. Further investigation revealed that 3–4 times higher spontaneous MN frequency than that in the other conditions is a possible reason for the low sensitivity, and this high spontaneous MN frequency was not observed in Chinese hamster lung cells under the identical treatment condition. With regard to the two conditions without cytokinesis block, two negative substances were evaluated and found to be negative, suggesting the validity of the TK6 test system for these conditions. Although our findings showed a few concerns for the treatment with cytokinesis block, the TK6 cells were considered to be a reliable cell line to be used for detecting potential inducers of MN in the in vitro micronucleus assay based on the overall results.

Increased chromosome damage in systemic sclerosis skin fibroblasts

Objective: To evaluate chromosome damage, by means of micronucleus frequency, in dermal fibroblasts from affected and non-affected skin from systemic sclerosis (SSc) patients and from controls.Methods: Primary fibroblast cultures were obtained by biopsy from affected and non-affected skin from SSc patients. Control fibroblasts were derived from skin remnants from plastic surgery in healthy adults. The number of micronuclei-bearing cells per 1000 binucleated cells (MN+ cells/1000 BN) was determined in cultures with and without clastogenic stimulus (bleomycin 3 μg/mL).Results: Primary cultures from 10 SSc patients (affected and non-affected skin) and nine controls were analysed by two blinded examiners. In the absence of bleomycin, the frequency of MN+ cells was higher in cultures from affected (14.01 ± 11.96 MN+ cells/1000 BN; p = 0.004) and non-affected (15.41 ± 13.58 MN+ cells/1000 BN; p = 0.005) skin from SSc patients as compared to fibroblasts from healthy controls (4.74 ± 3.30 MN+ cells/1000 BN). In bleomycin-treated cultures, the frequency of MN+ cells was higher in SSc affected (38.03 ± 26.14 MN+ cells/1000 BN; p = 0.041) and non-affected skin (38.47 ± 17.88 MN+ cells/1000 BN; p = 0.034) as compared to healthy control fibroblasts (20.54 ± 13.09 MN+ cells/1000 BN). There was no difference in the frequency of MN+ cells in cultures from affected and non-affected skin of SSc patients.Conclusions: This is the first demonstration that dermal fibroblasts from SSc patients present an increased frequency of spontaneous and clastogen-induced micronuclei. Increased clastogenesis seems to be a widespread phenomenon in SSc because fibroblasts from clinically affected and non-affected skin presented the equivalent increased micronuclei counts.

Spontaneous and bleomycin‐induced chromosome damage in non cancer thyroid patients

Presence of chromosome damage in lymphocytes of patients affected by several diseases, including cancer, was detected by the micronucleus (MN) assay. Individual susceptibility to DNA damage, considered as a risk factor for cancer, can be also evaluated using the bleomycin (BLM) sensitivity test. We aimed to evaluate spontaneous or BLM-induced MN frequencies in autoimmune (AI, n = 19) and non autoimmune (NAI, n = 11) thyroid patients, not receiving (131)I radiometabolic therapy with respect to a control group of 18 healthy subjects. According to thyroid function, patients were also divided into hypothyroid (n = 10), euthyroid (n = 13) or hyperthyroid (n = 7) subjects. Spontaneous MN frequencies of AI and NAI patients did not differ from those of controls. Hypothyroid patients had more elevated MN basal levels (9.00 + or - 1.71 per thousand) than hyperthyroid (3.75 + or - 1.17 per thousand, P < 0.05) and euthyroid (5.38 + or - 0.97 per thousand, P < 0.01) patients or healthy subjects (4.17 + or - 0.63 per thousand, P < 0.01). In particular, the hypothyroid AI group showed the highest value (9.79 + or - 2.26 per thousand, P < 0.01). All thyroid patients responded differently to BLM than controls (39.90 + or - 2.48 per thousand vs. 31.08 + or - 2.51 per thousand, P = 0.0377). The NAI group had BLM-induced MN levels (45.00 + or - 2.56 per thousand) significantly higher (P = 0.0215) than AI patients (36.95 + or - 3.49 per thousand) or healthy subjects (31.08 + or - 2.51 per thousand). No significant difference was seen when patients were stratified according to autoimmunity. We report that hypothyroid patients exhibit a moderate increase in the level of spontaneous genome damage, and that AI thyroid patients resulted to be less sensitive than NAI patients to the mutagen sensitivity test. In prospective, it may be of interest to reinvestigate hypothyroid patients when correction of their dysfunction is achieved.

Spontaneous frequency of micronuclei among the newborns from high level natural radiation areas of Kerala in the southwest coast of India

Purpose: The present study was an attempt to determine the spontaneous frequency of micronuclei (MN) in newborns from High Level Natural Radiation Areas (HLNRA) and the adjoining Normal Level Radiation Areas (NLNRA) of the monazite-bearing Kerala Coast in Southern West India using Cytochalasin Blocked Micronuclei (CBMN) assay.Materials and methods: Human umbilical cord blood samples were collected from a total number of 271 newborns (61 from NLNRA and 210 from HLNRA), born to mothers aged between 17 and 37 years (mean maternal age: 24.08 ± 4.23 years). Lymphocyte cultures were set up following microculture techniques and cultures were terminated at 72 hours. Cytochalasin B at a concentration of 4.0 μg/ml was added to the lymphocyte cultures at 44 h. Enumeration of micronuclei was restricted to Cytochalasin Blocked binucleated (BN) cells only.Results: The frequency of MN among the newborns from NLNRA (1.40 ± 0.12) per 1000 BN cells was not statistically significant as compared to HLNRA newborns (1.33 ± 0.04) per 1000 BN cells. Our data did not show any radiation dose response. Odds ratios (OR) and confidence intervals (CI) have been calculated to see statistical significance in the mean MN frequency among the newborns from various dose groups with respect to control and it did not reveal any significant difference (p > 0.05). A marginal increase in the frequency of micronuclei was observed among the female newborns as compared to males with increasing mothers' age groups except for the mothers aged > 30 years, though not statistically significant.Conclusion: The baseline frequency of micronuclei in HLNRA newborns is not statistically different from NLNRA newborns suggesting that elevated level of naturally occurring radiation has no significant effect on the induction of micronuclei frequency among the newborns.

Radiosensitivity of Peripheral Blood Lymphocytes in Patients with Genital Cancer

Purpose: To evaluate the in vitro radiation sensitivity of the peripheral blood lymphocytes obtained from genital cancer patients (n = 20) in comparison to healthy individuals (n=15). Materials and Methods: Chromosomal radiosensitivity was measured by the cytokinesis-blocked micronucleus (MN) assay. Unstimulated whole blood cultures were exposed in vitro to 2,5 Gy 60Co gamma rays delivered at a high dose-rate (HDR: 25 cGy / min) or low dose-rate (LDR: 0,25 cGy / min). Results: Spontaneous micronucleus frequencies (MNSP) in blood lymphocytes from patients and controls are not statistically different. Cancer patients group had a significantly higher mean baseline micronucleus frequency (p < 0,05) than normal healthy controls at LDR irradiation. After HDR irradiation, we observed no difference in radioinduced micronucleus frequency between the two studied groups of individuals. Conclusion: The results indicate that cancer patients were more radiosensitive than normal subjects at LDR irradiation. Enhanced radiosensitivity could be due to defects in DNA repair genes involved in genital cancer formation.