The increased chromosomal DNA damage in patients with Familial Mediterranean Fever.

Familial Mediterranean Fever (FMF) is an inherited autoinflammatory disease. In this study, we aimed to assess chromosomal DNA damage and cell proliferation by using cytokinesis-block micronucleus cytome (CBMN-cyt) assay in the peripheral blood lymphocytes of untreated FMF patients carrying M694V and R202Q mutations, which are the most common MEFV gene mutations in Turkish society. The study included 20 untreated FMF patients with M694V and R202Q mutations and 20 healthy individuals of similar age and sex as the control group. Micronucleus (MN), nucleoplasmic bridges (NPBs), and nuclear buds (NBUDs) were scored in the obtained bi-nucleated (BN) cells. Additionally, the nuclear division index (NDI) was calculated using the scores of mononuclear, binuclear, and multinuclear cells. We found that MN and NPBs frequencies in FMF patients were significantly higher than in controls, and number of metaphases was significantly lower (respectively, p < 0.05, p < 0.01, and p < 0.01). However, there was no significant difference in NBUDs frequencies and NDI values between FMF patients and controls (p > 0.05). Our study is the first to evaluate FMF patients' lymphocytes using the CBMN-cyt assay, as no previous research has been found in this respect. Increased MN and NPB frequencies may be useful as biomarkers for chromosomal DNA damage, and may indicate a potential for elevated cancer risk in untreated FMF patients.

Low magnesium in conjunction with high homocysteine increases DNA damage in healthy middle aged Australians.

PURPOSE: Magnesium is one of the most common elements in the human body and plays an important role as a cofactor of enzymes required for DNA replication and repair and many other biochemical mechanisms including sensing and regulating one-carbon metabolism deficiencies. Low intake of magnesium can increase the risk of many diseases, in particular, chronic degenerative disorders. However, its role in prevention of DNA damage has not been studied fully in humans so far. Therefore, we tested the hypothesis that magnesium deficiency either on its own or in conjunction with high homocysteine (Hcy) induces DNA damage in vivo in humans. METHODS: The present study was carried out in 172 healthy middle aged subjects from South Australia. Blood levels of magnesium, Hcy, folate and vitamin B12 were measured. Cytokinesis-Block Micronucleus cytome assay was performed to measure three DNA damage biomarkers: micronuclei (MN), nucleoplasmic bridges (NPBs) and nuclear buds (NBuds) in peripheral blood lymphocytes. RESULTS: Data showed that magnesium and Hcy are significantly inversely correlated with each other (r = - 0.299, p < 0.0001). Furthermore, magnesium is positively correlated both with folate (p = 0.002) and vitamin B12 (p = 0.007). Magnesium is also significantly inversely correlated with MN (p < 0.0001) and NPB (p < 0.0001). Individuals with low magnesium and high Hcy exhibited significantly higher frequency of MN and NPBs compared to those with high magnesium and low Hcy (p < 0.0001). Furthermore, there was an interactive effect between these two factors as well in inducing MN (p = 0.01) and NPB (p = 0.048). CONCLUSIONS: The results obtained in the present study indicate for the first time that low in vivo levels of magnesium either on its own or in the presence of high Hcy increases DNA damage as evident by higher frequencies of MN and NPBs.

The Relationship between Telomere Length and Nucleoplasmic Bridges and Severity of Disease in Prostate Cancer Patients.

Telomeres are repetitive nucleotide (TTAGGG) sequences that stabilize the chromosome ends and play an important role in the prevention of cancer initiation and progression. Nucleoplasmic bridges (NPBs) are formed when chromatids remain joined together during mitotic anaphase either due to mis-repair of DNA breaks or due to chromatid end fusion as a result of telomere loss or telomere dysfunction. We tested the hypotheses that (i) telomere length (TL) is shorter in prostate cancer (PC) patients relative to healthy age-matched individuals, (ii) TL differs in different stages of PC and (iii) shorter TL is significantly correlated with NPBs formation in PC cases. TL was measured in whole blood by well-established quantitative PCR method and the frequency of NPBs was measured in lymphocytes using cytokinesis-block micronucleus cytome (CBMNcyt) assay. Our results indicate that TL is shorter and NPBs are increased in PC patients relative to age-matched healthy controls. Furthermore, TL was significantly shorter (p = 0.03) in patients with a Gleason score more than 7 and there was also a significant trend of decreasing TL across all three stages (p trend = 0.01; Gleason score <7, 7 and >7). Furthermore, TL was significantly inversely correlated with NPB frequency in PC patients (r = -0.316; p = 0.001) but not in controls (r = 0.163; p = 0.06) and their relationships became stronger with higher Gleason scores. More studies are required that can confirm our observations and explore mechanistic differences in the role of telomeres in NPB formation in PC cases relative to non-cancer cases.

Re-evaluation of CBMN test reference values of persons continuously occupationally exposed to low doses of ionizing radiation in Serbia.

When established, cytokinesis-block micronucleus (CBMN) test reference values should be periodically evaluated according to the recommendations of reference documents. The biodosimetry cytogenetic laboratory of the Serbian Institute of Occupational Health established the CBMN test reference range for people occupationally exposed to ionizing radiation in 2016. Since then, new occupationally exposed persons have been subjected to micronucleus testing, resulting in the need for re-evaluation of existing CBMN test values. The examined population comprised 608 occupationally exposed subjects - 201 from the previous laboratory database and 407 newly examined. Comparison of groups based on gender, age and cigarette consumption did not show significant differences, although certain CBMN values differed significantly between the old and new groups. Duration of occupational exposure, gender, age and smoking habit influenced micronuclei frequency in all three analyzed groups, while no relation was found between type of work and micronucleus test parameters. Since the mean values of all tested parameters in the new group of examinees are within previously established reference ranges, existing values can be used in further research.

Genotoxic risk in humans and acute toxicity in rats of a novel oral high-dose coenzyme Q10 oleogel.

Coenzyme Q10 (CoQ10) supplementation has demonstrated to be safe and effective in primary and secondary CoQ10 deficiencies. Previously, we have designed a high-dose CoQ10 oleogel (1 g/disk) with excipients used in quantities that do not represent any toxic risk. However, it was necessary to demonstrate their safety in the final formulation. Following this purpose, an acute toxicity study of the oleogel in rats was performed. Furthermore, the genotoxic risk was evaluated in human volunteers after CoQ10 supplementation with oleogel and compared to the solid form (1 g/three 00-size-capsules). In addition, the general health status and possible biochemical changes of the participants were determined using serum parameters. Results suggested the absence of adverse effects caused by the interaction of the components in the oleogel formulation. Therefore, we conclude that the designed novel high-dose CoQ10 oleogel was safe for oral consumption.

Dose-effect relationships of 12C6+ ions-induced dicentric plus ring chromosomes, micronucleus and nucleoplasmic bridges in human lymphocytes in vitro.

PURPOSE: The objective of this research was to explore the dose-effect relationships of dicentric plus ring (dic + r), micronucleus (MN) and nucleoplasmic bridges (NPB) induced by carbon ions in human lymphocytes. MATERIALS AND METHODS: Venous blood samples were collected from three healthy donors. 12C6+ ions beam was used to irradiate the blood samples at the energy of 330 MeV and linear energy transfer (LET) of 50 keV/µm with a dose rate of 1 Gy/min in the spread-out Bragg peak. The irradiated doses were 0 (sham irradiation), 1, 2, 3, 4, 5 and 6 Gy. Dic + r chromosomes aberrations were scored in metaphases. The cytokinesis-block micronucleus cytome (CBMN) was conducted to analyze MN and NPB. The maximum low-dose relative biological effectiveness (RBEM) values of the induction of dic + r, MN and NPB in human lymphocytes for 12C6+ ions irradiation was calculated relative to 60Co γ-rays. RESULTS: The frequencies of dic + r, MN and NPB showed significantly increases in a dose-depended manner after exposure to 12C6+ ions. The distributions of dic + r and MN exhibited overdispersion, while the distribution of NPB agreed with Poisson distribution at all doses. Linear-quadratic equations were established based on the frequencies of dic + r and MN. The dose-response curves of NPB frequencies followed a linear model. The derived RBEM values for dic + r, MN and NPB in human lymphocytes irradiated with 12C6+ ions were 8.07 ± 2.73, 2.69 ± 0.20 and 4.00 ± 2.69 in comparison with 60Co γ-rays. CONCLUSION: The dose-response curves of carbon ions-induced dic + r, MN and NPB were constructed. These results could be helpful to improve radiation risk assessment and dose estimation after exposed to carbon ions irradiation.

Effects of the DNA repair inhibitors, cytosine arabinoside and 3-aminobenzamide, on the frequency of radiation-induced micronuclei, nuclear buds, and nucleoplasmic bridges.

BACKGROUND: Micronuclei (MN), nuclear bud (NBud), and nucleoplasmic bridge (NPB) are suggested as biomarkers for radiation exposure; however, they have not been extensively studied to understand the underlying mechanisms responsible for their formation. OBJECTIVES: To (1) validate NBud and NPB within the cytokinesis-blocked micronucleus (CBMN) assay as biomarkers for radiation exposure and (2) determine the effects of the DNA repair inhibitors, cytosine arabinoside (Ara C) and 3-aminobenzamide (3-AB) on radiation-induced MN, NBud, and NPB formation. METHODS: Human blood samples were irradiated with 0-3 Gy X-rays and subsequently treated with Ara C and 3-AB. CBMN and chromosome aberration assays were carried out to measure MN, NBud, and NPB and dicentric chromosomes, respectively. RESULTS: The frequency of radiation-induced MN, NBud, and NPB increased in a dose-dependent manner. The frequency of MN, NBud, and NPB was highly and positively correlated with the dicentric chromosome, a standard biomarker for biodosimetry (r > 0.98, p < 0.0001). Furthermore, Ara C increased the frequency of MN, NBud, and NPB, whereas 3-AB increased the frequency of MN and NPB, but not NBud. Further, the potentiating effect of Ara C on the frequency of MN, NBud, and NPB was greater than that of 3-AB. CONCLUSION: Our results validate NBuds and NPBs as independent valuable markers of radiation exposure. Additionally, we suggest that different mechanisms are likely involved in the formation of NBuds and NPBs following X-irradiation; however, additional studies are warranted to better understand the contribution of distinct DNA repair pathways to the formation of radiation-induced damages.

Cytotoxicity and genotoxicity of MWCNT-7 and crocidolite: assessment in alveolar epithelial cells versus their coculture with monocyte-derived macrophages.

In the past years, several in vitro studies have addressed the pulmonary toxicity of multi-walled carbon nanotubes (MWCNT) and compared it with that caused by asbestos fibers, but their conclusions have been somewhat inconsistent and difficult to extrapolate to in vivo. Since cell coculture models were proposed to better represent the in vivo conditions than conventional monocultures, this work intended to compare the cytotoxicity and genotoxicity of MWCNT-7 (Mitsui-7) and crocidolite using A549 cells grown in a conventional monoculture or in coculture with THP-1 macrophages. Although a decrease in A549 viability was noted following exposure to a concentration range of MWCNT-7 and crocidolite, no viability change occurred in similarly exposed cocultures. Early events indicating epithelial to mesenchymal transition (EMT) were observed which could explain apoptosis resistance. The comet assay results were similar between the two models, being positive and negative for crocidolite and MWCNT-7, respectively. An increase in the micronucleus frequency was detected in the cocultured A549-treated cells with both materials, but not in the monoculture. On the other hand, exposure of A549 monocultures to MWCNT-7 induced a highly significant increase in nucleoplasmic bridges in which those were found embedded. Our overall results demonstrate that (i) both materials are cytotoxic and genotoxic, (ii) the presence of THP-1 macrophages upholds the viability of A549 cells and increases the aneugenic/clastogenic effects of both materials probably through EMT, and (iii) MWCNT-7 induces the formation of nucleoplasmic bridges in A549 cells.

Effects of age and gender on the baseline and 2 Gy 60Co γ-ray-induced nucleoplasmic bridges frequencies in the peripheral blood lymphocytes of Chinese population.

Baseline nucleoplasmic bridges (NPB) vary widely in the general population from different regions in the same country or from different countries. The baseline NPB level in the normal Chinese population and the factors affecting the baseline and radiation-induced NPB levels have not been explored yet. The cytokinesis-block micronucleus cytome assay was conducted on the peripheral blood samples of 121 healthy individuals for the baseline NPB and 52 healthy individuals for the 2 Gy γ-ray-induced NPB level. The effects of age and gender on the baseline NPB and 2 Gy γ-ray-induced NPB level were evaluated. The overall baseline NPB in the peripheral blood lymphocytes of 121 healthy adults from the general population in China was 0.46 ±â€¯0.20 per 1000 binucleated (BN) cells. The overall baseline NPB in males (0.56 ±â€¯0.15 per 1000 BN cells) was higher than that in females (0.36 ±â€¯0.22 per 1000 BN cells, P < 0.05). The effect of age on the baseline NPB was not significant, except for females in the 40-year age group. The overall 2 Gy γ-ray-induced NPB frequency for male donors was lower than that for female donors (P < 0.01). No evident trend of the radiation-induced NPB level with increasing age was observed for both genders. For the baseline micronucleus (MN) and radiation-induced MN levels, the effects of gender and age were confirmed. Therefore, the gender of donors affects the baseline and radiation-induced levels of NPB and MN. In addition, the effect of the age of the donors on the baseline and radiation-induced NPB levels showed no clear pattern and needed to be further investigated.

The Frequency of Lymphocytes Containing Dumbbell-Shaped Nuclei Depends on Ionizing Radiation Dose and Correlates with Appearance of Chromosomal Aberrations.

Nuclear anomalies of different types appear in cells in response to the action of ionizing radiation after the passage of the first mitotic division. In this article, we present the results of the study of the frequency of occurrence of three types of nuclear anomalies ("tailed" nuclei, nucleoplasmic bridges, and dumbbell-shaped nuclei) in vitro in human lymphocytes cultured with cytochalasin B when exposed to X-rays at doses of 0.0, 0.1, 0.2, 0.4, 0.5, 0.7, 1.0, 1.5, and 2.0 Gy. To stop the cell cycle of cultured lymphocytes after the first mitotic division, a cytokinesis block was performed using cytochalasin B. Dose-dependent curves of the occurrence of lymphocytes containing "tailed" nuclei, nucleoplasmic bridges, or dumbbell-shaped nuclei after irradiation have been constructed. At the same time, frequencies of occurrence of chromosomal aberrations (dicentric and ring chromosomes) in the culture of lymphocytes exposed to the same radiation doses were studied. Comparison of the frequencies of occurrence of dicentric and ring chromosomes with frequencies of occurrence of nuclear anomalies allows us to conclude that these nuclear anomalies are formed as a result of chromosomal aberrations arising in lymphocytes under the action of ionizing radiation. More than that, most of the chromosomal aberrations are converted into dumbbell-shaped nuclei in vitro in the culture of lymphocytes in the cytochalasin block.

Nucleoplasmic bridges as a biomarker of DNA damage exposed to radon.

Radon is a naturally occurring radionuclide in the environment, during decay it emits high linear energy transfer (LET) alpha particles. When radon exposure is accompanied by smoking it has been reported that lung cancer risk is higher. Blood samples were collected after prior consent, 25 smokers and 25 non smokers (only males) exposed in vitro to radon gas with doses ranging between 0.3-12.6mGy Ionizing radiation is a strong clastogenic agent and a potent inducer of MN. Cytokinesis-Blocked Micro Nucleus (CBMN) assay has proven to be a reliable, thoroughly validated and standardised technique in the field of radiation biology. In view of this, an invitro study has been undertaken using CBMN assay to suggest Nucleoplasmic bridges (NPBs) as a potential bio-dosimeter and to investigate the existence of a possible influence of smoking on genetic damage induced by ionizing radiation. The mean percentages frequencies of micronuclei, nucleoplasmic bridges and nuclear buds among the exposed smoker group were 4.82±0.271, 0.6±0.04 and 0.12±0.02 respectively and these values for exposed non-smoker group were 2.78±0.285, 0.4±0.04 and 0.06±0.01 respectively. Spearman rank correlation for the frequency of nucleoplasmic bridge with respect to dose was considered to be significant (P<0.05) in non-smokers. This study is first of its kind to investigate NPB as a biomarker of early DNA damage induced by radon.

Nucleoplasmic bridges and tailed nuclei are signatures of radiation exposure in Oreochromis mossambicus using erythrocyte micronucleus cytome assay (EMNCA).

Gamma radiation-induced genetic perturbations in aquatic vertebrates is largely unknown at low-dose rate, especially in the wake of a nuclear disaster and/or other environmental outbreaks. Freshwater fish, Oreochromis mossambicus subjected to low-dose rate (2 mGy/min) at 2.5-, 5-, and 10-Gy doses, were analyzed for "exposure signatures" in blood samples drawn on days 3, 6, 12, 18, and 30, respectively. Significant dose-dependent increments in micronuclei frequency and other anomalies such as nucleoplasmic bridges and tailed nuclei were observed and exhibit a strong positive correlation, suggesting that they could be used as prospective signatures of radiation exposure. Similarly increased incidence of apoptosis and DNA repair machinery circuits at high and low doses were noted. This work highlighted "cytogenetic signatures" in fish and the sensitivity of these endpoints toward low-dose rate of radiation exposure.

Effects of low oxygen levels on G2-specific cytogenetic low-dose hyper-radiosensitivity in irradiated human cells.

Low-dose hyper-radiosensitivity (HRS) has been reported in normal human lymphoblastoid cell lines for exposures at ≤ 20 cGy, but the cytogenetic effects of oxygen (O2 ) levels in tissue culture medium on HRS have not been evaluated. We asked whether HRS was lost in G2-irradiated cells grown in atmospheres of 2.5% or 5% O2 , compared to responses by cells cultured in ambient O2 (21%). The results indicate a loss of HRS when cells are cultured and irradiated either in 2.5% or 5% O2 . We then evaluated whether low O2 levels either before or after exposure were responsible for the loss of HRS. For cells irradiated in 5% O2 , subsequent immediate re-oxygenation to ambient O2 levels restored the HRS effect, while cells cultured and irradiated at ambient O2 levels and then transferred to 5% O2 exhibited little or no HRS, indicating that ambient O2 levels after, but not before, radiation substantially affect the amounts of cytogenetic damage. HRS was not observed when cells were irradiated in G1. At doses of 40-400 cGy there was significantly less cytogenetic damage when cells were recovering from radiation at low O2 levels than at ambient O2 levels. Here we provide the first cytogenetic evidence for the loss of HRS at low O2 levels in G2-irradiated cells; these results suggest that at low O2 levels for all doses evaluated there is either less damage to DNA, perhaps because of lower amounts of reactive oxygen species, or that DNA damage repair pathways are activated more efficiently.

Evaluation of the mechanism of nucleoplasmic bridge formation due to premature telomere shortening in agricultural workers exposed to mixed pesticides: indication for further studies.

Agricultural workers are often exposed to high levels of pesticides over prolonged periods of time. We attempted to determine whether exposure to multiple pesticides shortens relative telomere length (RTL) and causes nucleoplasmic bridge (NPB) formation via the mechanism of telomere-end fusion in the lymphocytes of agricultural workers. For measuring RTL, we used quantitative fluorescent in situ hybridization, while NPB frequency was measured as part of the cytome assay. Multivariate analysis of variances taking into account confounding factors (age, gender, years of exposure, smoking, and alcohol intake) did not show a decrease, but rather an increase of RTL in agricultural workers compared to control individuals. In the exposed population, NPB frequency was significantly higher compared to controls (6 times, p<0.05). Multiple regression between NPB, RTL, and confounding factors was not significant. Using Spearman correlation, we did not find proof for our initial hypothesis. Our hypothesis that telomere shortening is a mechanism of NPB origin was not proven, indicating that telomere-end fusion is not a mechanism of NPB formation under our experimental conditions for agricultural workers.

Low concentration of exogenous carbon monoxide protects mammalian cells against proliferation induced by radiation-induced bystander effect.

Radiation-induced bystander effect (RIBE) has been proposed to have tight relationship with the irradiation-caused secondary cancers beyond the irradiation-treated area after radiotherapy. Our previous studies demonstrated a protective effect of low concentration carbon monoxide (CO) on the genotoxicity of RIBE after α-particle irradiation. In the present work, a significant inhibitory effect of low-dose exogenous CO, generated by tricarbonyldichlororuthenium (II) dimer [CO-releasing molecule (CORM-2)], on both RIBE-induced proliferation and chromosome aberration was observed. Further studies on the mechanism revealed that the transforming growth factor ß1/nitric oxide (NO) signaling pathway, which mediated RIBE signaling transduction, could be modulated by CO involved in the protective effects. Considering the potential of exogenous CO in clinical applications and its protective effect on RIBE, the present work aims to provide a foundation for potential application of CO in radiotherapy.

Cytogenetic characterization of low-dose hyper-radiosensitivity in Cobalt-60 irradiated human lymphoblastoid cells.

The dose-effect relationships of cells exposed to ionizing radiation are frequently described by linear quadratic (LQ) models over an extended dose range. However, many mammalian cell lines, when acutely irradiated in G2 at doses ≤0.3Gy, show hyper-radiosensitivity (HRS) as measured by reduced clonogenic cell survival, thereby indicating greater cell lethality than is predicted by extrapolation from high-dose responses. We therefore hypothesized that the cytogenetic response in G2 cells to low doses would also be steeper than predicted by LQ extrapolation from high doses. We tested our hypothesis by exposing four normal human lymphoblastoid cell lines to 0-400cGy of Cobalt-60 gamma radiation. The cytokinesis block micronucleus assay was used to determine the frequencies of micronuclei and nucleoplasmic bridges. To characterize the dependence of the cytogenetic damage on dose, univariate and multivariate regression analyses were used to compare the responses in the low- (HRS) and high-dose response regions. Our data indicate that the slope of the response for all four cell lines at ≤20cGy during G2 is greater than predicted by an LQ extrapolation from the high-dose responses for both micronuclei and bridges. These results suggest that the biological consequences of low-dose exposures could be underestimated and may not provide accurate risk assessments following such exposures.

Investigation of genome instability in patients with non-alcoholic steatohepatitis.

AIM: To evaluate the occurrence of micronucleus (MN), nucleoplasmic bridges (NPBs) and nuclear buds (NBUDs) in the mitogen-stimulated lymphocytes of patients with non-alcoholic steatohepatitis (NASH). METHODS: The study was performed in 25 (9 females, 16 males) patients newly diagnosed with NASH, and 25 healthy subjects of similar ages and genders were used as a control group. None of the controls was known to be receiving any drugs for medical or other reasons or using alcohol. Hepatosteatosis was further excluded by abdominal ultrasound imaging in the control group. The numbers of MN, NPBs and NBUDs scored in binucleated (BN) cells were obtained from the mitogen-stimulated lymphocytes of patients and control subjects. Statistical comparisons of the numbers of BN cells with MN, NPBs and NBUDs and ages between the patients with NASH and control subjects were performed. RESULTS: The mean ages of the patients and the control group were 41.92 ± 13.33 and 41.80 ± 13.09 years (P > 0.05), respectively. The values of the mean body mass index (BMI), HOMA-IR, hemoglobin, creatinin, aspartate aminotransferase, alanine aminotransferase, triglyceride, high density lipoprotein, and low density lipoprotein were 31.19 ± 4.62 kg/m(2) vs 25.07 ± 4.14 kg/m(2), 6.71 ± 4.68 vs 1.40 ± 0.53, 14.73 ± 1.49 g/dL vs 14.64 ± 1.30 g/dL, 0.74 ± 0.15 mg/dL vs 0.80 ± 0.13 mg/dL, 56.08 ± 29.11 U/L vs 16.88 ± 3.33 U/L, 92.2 ± 41.43 U/L vs 15.88 ± 5.88 U/L, 219.21 ± 141.68 mg/dL vs 102.56 ± 57.98 mg/dL, 16.37 ± 9.65 mg/dL vs 48.72 ± 15.31 mg/dL, and 136.75 ± 30.14 mg/dL vs 114.63 ± 34.13 mg/dL in the patients and control groups, respectively. The total numbers and frequencies of BN cells with MN, NPBs and NBUDs, which were scored using the CBMN cytome assay on PHA-stimulated lymphocytes, were evaluated in the patients with NASH and control group. We found significantly higher numbers of MN, NPBs and NBUDs in the BN cells of patients with NASH than in those of the control subjects (21.60 ± 9.32 vs 6.88 ± 3.91; 29.28 ± 13.31 vs 7.84 ± 3.96; 15.60 ± 5.55 vs 4.20 ± 1.63, respectively, P < 0.0001). CONCLUSION: The increased numbers of MN, NPBs and NBUDs observed in the lymphocytes obtained from patients with NASH may reflect genomic instability.

Genetic biomarkers for neoplastic colorectal cancer in peripheral lymphocytes.

BACKGROUND: Loss of genomic stability appears as a key step in colorectal carcinogenesis. Micronucleus (MN) designates a chromosome fragment or an entire chromosme which lags behind mitosis. MN may be noticed as an additional nucleus within the cytoplasm cell during the intermediate mitosis phases. We tested the hypothesis that MN and its related anomalies may be associated with the presence of neoplastic colorectal lesions. METHOD: Peripheral blood lymphocytes were cultured and microscopically examined. The frequency of micronuclei (FMN) and the presence of nucleoplasmic bridges (NPB) in binucleated cells were compared in patients with of without colorectal neoplastic lesions. RESULTS: We included 45 patients undergoing colonoscopy, 23 males and 22 females, with a median age of 59. 17 patients had polyps, 11 colorectal cancer (CRC) and 17 had a normal colonoscopy. The FMN was significantly higher in women than in men (8.14 vs 4.17, p=0.008); NPB were significantly less frequent in patients with advanced adenomas (>10mm or vilous) or CRC (p=0.044) when compared with patients with normal colonoscopy, hiperplastic polyps or non-advanced adenomas. CONCLUSION: Micronuclei are more frequent in women, but its frequency was not significantly different in patients with advanced adenomas or CRC. Null or low frequency values for nucleoplasmic bridges presence in peripheral lymphocyte may be predictive for advanced adenomas and colorectal cancer.

Heavy metals and trace elements in hair and urine of a sample of arab children with autistic spectrum disorder.

UNLABELLED: General information: Autism is a severe developmental disorder which involves social withdrawal, communication deficits, and stereotypic/repetitive behavior. The pathophysiological etiologies which precipitate autism symptoms remain elusive and controversial in many cases, but both genetic and environmental factors (and their interactions) have been implicated. While autism is considered multicausal, environmental factors have received significant attention. International discussion has ocused on neurotoxins such as mercury and lead, suggesting that these and other toxic metals contribute to the development of the disorder. An epidemiological study released in 2006 (Palmer et al.) linking Toxic Release Inventory (TRI) data on mercury to special education data in Texas reported a 61% increase in autism prevalence rates (or 17% adjusted) per 1000 pounds of mercury released into the environment (1). We attempted to further evaluate whether exposure to variable environmental contributes to the genesis of autistic spectrum disorder, and thus is a factor increasing the risk for developing autism symptoms in utero or in early childhood. PURPOSE: The purpose of this study is to examine possible environmental risk factors and sources of exposure to mercury and other heavy metals in children with autism spectrum disorder versus controls. Through laboratory diagnostics we are able to distinguish between present and past exposure (i.e. hair analysis measurements reflect past exposure), urinary excretion levels of unprovoked urine represent immediate exposure. By assessing a spectrum of trace elements and heavy metals in hair and urine of both autistic and control groups, we focused on the participants≈ past and present exposure. METHODOLOGY: The participants were 25 Autistic Spectrum Disorder (ASD) children (22 boys and 3 girls) between the age of 3 and 9 years. They were either diagnosed previously by other psychiatrist, psychologist, and developmental pediatrician or suspected by their parents as being autistic. All children were attendants to the Child Psychiatric Clinic in Erfan Psychiatric Hospital in Jeddah, KSA. Samples were collected during the period of June 2006 to March 2008. A control group of 25 children without any psychiatric or medical disorders was age-matched and sex-matched. All parents signed informed consent forms. All autistic children were subjected to a full clinical child psychiatric sheet for the diagnosis of autism spectrum disorder and exclusion of other psychiatric disorders according to the Diagnostic and Statistical Manual of Mental Disorders, 4th Edition (DSM IV). The severity of autistic symptomatology was measured by the Childhood Autism Rating Scale (CARS) and Autism Behavior Checklist (ABC) using the Arabic versions. Both groups were subjected to the Questionnaire on Exposure to Heavy Metals, Physical Symptoms, and Child Development. Hair and baseline urine samples (i.e. unprovoked urine) were taken from both groups and sent to the German clinical and environmental laboratory Micro Trace Minerals Gmbh, for the detection of heavy metals and trace elements levels where metal testing was performed via ICP-MS spectroscopy utilizing cell technique. RESULTS: By comparing the ASD Group to the Control Group, we found a statistically significant difference in the mean hair levels of arsenic, cadmium, barium, cerium and lead (p=0.01, 0.03, 0.003, 0.003, and 0.03 respectively), and in the mean hair levels of magnesium and zinc (p=0.001 and 0.003 respectively). There were also statistically significant differences in the mean urine levels of aluminum, barium, cerium, mercury, and lead (p=0.004, 002, 0.014, 0.006 and 0.004 respectively), and in the mean urine levels of copper and germanium (p=0.049 and 0.02 respectively). An agreement was found in both specimen (hair and urine) for barium and lead. The statistically significant differences in mean hair levels of arsenic, cadmium, and cerium were not supported by urine baseline levels. Also, the statistically significant magnesium and zinc levels of hair were not supported by urine levels. A disagreement was also found with copper and germanium concentrations.

Efficacy of HUMN criteria for scoring the micronucleus assay in human lymphocytes exposed to a low concentration of p, p'-DDT

The cytokinesis-block micronucleus (CBMN) assay is one of the standard cytogenetic tools employed to assess chromosomal damage subsequent to exposure to genotoxic/cytotoxic agents, and is widely applicable to plant, animal and human cells. In the present study, the CBMN assay was used to assess the baseline damage in binuclear human peripheral blood lymphocytes exposed to 25 µg/L p,p'-DDT for 1, 2, 24, and 48 h by measuring the frequency of micronuclei, nucleoplasmic bridges and nuclear buds. These new scoring criteria facilitated the detection of different types of clastogenic and aneugenic effects induced by this type of pollutant. With these criteria, CBMN can also be used to measure nucleoplasmic bridges which are considered to be consequences of chromosome rearrangements and nuclear buds which are biomarkers of altered gene amplification and gene dosage. The total number of micronuclei observed in binuclear human peripheral blood lymphocytes of the exposed samples (ranging from 32 to 47) was significantly greater (P < 0.05) than that detected in the unexposed (0 time) control sample, where the total number of micronuclei was 7. The number of nucleoplasmic bridges and nuclear buds obtained after 24 and 48 h was also significantly (P < 0.05) greater in the samples treated with p,p'-DDT than in the unexposed control samples. Thus, our results confirmed the usefulness of the new criteria applicable for the CBMN assay employed in measuring the DNA damage and its role of a sensitive cytogenetic biomarker.