Micronucleated Cells Research Articles

Butachlor induced dissipation of mitochondrial membrane potential, oxidative DNA damage and necrosis in human peripheral blood mononuclear cells

Butachlor is a systemic herbicide widely applied on rice, tea, wheat, beans and other crops; however, it concurrently exerts toxic effects on beneficial organisms like earthworms, aquatic invertebrates and other non-target animals including humans. Owing to the associated risk to humans, this chloroacetanilide class of herbicide was investigated with the aim to assess its potential for the (i) interaction with DNA, (ii) mitochondria membrane damage and DNA strand breaks and (iii) cell cycle arrest and necrosis in butachlor treated human peripheral blood mononuclear (PBMN) cells. Fluorescence quenching data revealed the binding constant (Ka=1.2×104M−1) and binding capacity (n=1.02) of butachlor with ctDNA. The oxidative potential of butachlor was ascertained based on its capacity of inducing reactive oxygen species (ROS) and substantial amounts of promutagenic 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG) adducts in DNA. Also, the discernible butachlor dose-dependent reduction in fluorescence intensity of a cationic dye rhodamine (Rh-123) and increased fluorescence intensity of 2′,7′-dichlorodihydro fluorescein diacetate (DCFH-DA) in treated cells signifies decreased mitochondrial membrane potential (ΔΨm) due to intracellular ROS generation. The comet data revealed significantly greater Olive tail moment (OTM) values in butachlor treated PBMN cells vs untreated and DMSO controls. Treatment of cultured PBMN cells for 24h resulted in significantly increased number of binucleated micronucleated (BNMN) cells with a dose dependent reduction in the nuclear division index (NDI). The flow cytometry analysis of annexin V−/7-AAD+ stained cells demonstrated substantial reduction in live population due to complete loss of cell membrane integrity. Overall the data suggested the formation of butachlor–DNA complex, as an initiating event in butachlor-induced DNA damage. The results elucidated the oxidative role of butachlor in intracellular ROS production, and consequent mitochondrial dysfunction, oxidative DNA damage, and chromosomal breakage, which eventually triggers necrosis in human PBMN cells.

An ethanolic extract of black cohosh causes hematological changes but not estrogenic effects in female rodents

Black cohosh rhizome (Actaea racemosa) is used as a remedy for pain and gynecological ailments; modern preparations are commonly sold as ethanolic extracts available as dietary supplements. Black cohosh was nominated to the National Toxicology Program (NTP) for toxicity testing due to its widespread use and lack of safety data. Several commercially available black cohosh extracts (BCE) were characterized by the NTP, and one with chemical composition closest to formulations available to consumers was used for all studies. Female B6C3F1/N mice and Wistar Han rats were given 0, 15 (rats only), 62.5 (mice only), 125, 250, 500, or 1000mg/kg/day BCE by gavage for 90days starting at weaning. BCE induced dose-dependent hematological changes consistent with a non-regenerative macrocytic anemia and increased frequencies of peripheral micronucleated red blood cells (RBC) in both species. Effects were more severe in mice, which had decreased RBC counts in all treatment groups and increased micronucleated RBC at doses above 125mg/kg. Dose-dependent thymus and liver toxicity was observed in rats but not mice. No biologically significant effects were observed in other organs. Puberty was delayed 2.9days at the highest treatment dose in rats; a similar magnitude delay in mice occurred in the 125 and 250mg/kg groups but not at the higher doses. An additional uterotrophic assay conducted in mice exposed for 3days to 0.001, 0.01, 0.1, 1, 10, 100 and 500mg/kg found no estrogenic or anti-estrogenic activity. These are the first studies to observe adverse effects of BCE in rodents.

Female parthenogenetic apomixis and androsporogenetic parthenogenesis in embryonal cells of Araucaria angustifolia: interpolation of progenesis and asexual heterospory in an artificial sporangium

Cell fate, development timing and occurrence of reproductive versus apomictic development in gymnosperms are shown to be influenced by culture conditions in vitro. In this study, female parthenogenetic apomixis (fPA), androsporogenetic parthenogenesis (mAP) and progenesis were demonstrated using embryonal initials of Araucaria angustifolia in scaled-up cell suspensions passing through a single-cell bottleneck in darkness and in an artificial sporangium (AS). Expression was based on defined nutrition, hormones and feedforward-adaptive feedback process controls at 23-25°C and in darkness. In fPA, the nucleus of an embryonal initial undergoes endomitosis and amitosis, forming a diploid egg-equivalent and an apoptotic ventral canal nucleus in a transdifferentiated archegonial tube. Discharge of egg-equivalent cells as parthenospores and their dispersal into the aqueous culture medium were followed by free-nuclear conifer-type proembryogenesis. This replaced the plesiomorphic and central features of proembryogenesis in Araucariaceae. Protoplasmic fusions of embryonal initials were used to reconstruct heterokaryotic expressions of fPA in multiwell plates. In mAP, restitutional meiosis (automixis) was responsible for androsporogenesis and the discharge of monads, dyads, tetrads and polyads. In a display of progenesis, reproductive development was brought to an earlier ontogenetic stage and expressed by embryonal initials. Colchicine increased polyploidy, but androspore formation became aberrant and fragmented. Aberrant automixis led to the formation of chromosomal bouquets, which contributed to genomic silencing in embryonal initials, cytomixis and the formation of pycnotic micronucleated cells. Dispersal of female and male parthenospores displayed heteromorphic asexual heterospory in an aqueous environment.

Exposure of thermoelectric power-plant workers to volatile organic compounds from fuel oil: Genotoxic and cytotoxic effects in buccal epithelial cells

Thermoelectric power-plant workers are constantly exposed to high levels of potentially genotoxic gaseous substances, such as volatile organic compounds (VOCs) from the combustion of fuel oil or the processing of naphtha. The aim of the present study was to estimate the association between such occupational exposure and the frequency of micronucleated cells and cells with other nuclear anomalies.Buccal epithelial cells were collected from a total of 44 power-plant workers (exposed group) and 47 administrative workers (non-exposed group), and examined for the frequency of micronucleated cells (MNC) and of cells with other nuclear anomalies (ONA: pyknosis, karyolysis, and karyorrhexis) by means of the micronucleus assay. The frequencies of MNC and ONA per 1000 cells in the exposed group (1.8‰ and 82.4‰, respectively) were significantly higher than in the non-exposed group (0.2‰ and 58.3‰, respectively). The exposed group had a twelve-fold increase in risk for formation of MNC compared with non-exposed individuals (RR=12.1; 95% CI, 5.0–29.2; P<0.001). The confounding factors analyzed (age, smoking status, alcohol consumption, and mouthwash use) did not show any significant association with the frequency of MNC or ONA.The findings of this study show that workers from power plants exposed to VOCs have a significantly elevated risk for DNA damage. Therefore, bio-monitoring of DNA damage is recommended for this group of workers.

Differential Bystander Signaling Between Radioresistant Chondrosarcoma Cells and Fibroblasts After X-Ray, Proton, Iron Ion and Carbon Ion Exposures

Chondrosarcoma is well known as a radioresistant tumor, but the mechanisms underlying that resistance are still unclear. The bystander effect is well documented in the field of radiation biology. We investigated the bystander response induced by X-rays, protons, carbon ions, and iron ions in chondrosarcoma cells using a transwell insert co-culture system that precludes physical contact between targeted and bystander cells. Human chondrosarcoma cells were irradiated with 0.1-, 0.5-, 1-, and 2-Gy X-rays, protons, carbon ions or iron ions using a transwell insert co-culture system. Formation of micronuclei and p53 binding protein 1 staining in bystander and irradiated cells were analyzed and bystander signaling between mixed cultures of chondrosarcoma cells, and normal human skin fibroblasts was investigated. In this study, we show that the fraction of cells with DNA damages in irradiated chondrosarcoma cells showed dose-dependent increases with all beams. However, the fraction of cells with DNA damages in all bystander chondrosarcoma cells did not show any change from the levels in control cells. In the bystander signaling between mixed cultures of chondrosarcoma cells and fibroblasts, the amount of micronucleus formation in all bystander chondrosarcoma cells co-cultured with irradiated fibroblasts were the same as the levels for control cells. However, all bystander fibroblasts co-cultured with irradiated chondrosarcoma cells showed significant increases in the fraction of micronucleated cells compared to the rate of control cells. We conclude that chondrosarcoma cells in the transwell insert co-culture system could release bystander stimulations but could not develop bystander responses.

Cytogenetic status of human lymphocytes after exposure to low concentrations of p,p′-DDT, and its metabolites (p,p′-DDE, and p,p′-DDD) in vitro

Despite that the use of DDT has been restricted for more than 40 years to malaria affected areas, low doses of this pesticide and its metabolites DDE and DDD can be found in the environment around the world. Although it has been shown that these pollutants induce cell and DNA damage, the mechanisms of their cytogenotoxic activity remains largely unknown. This study looks into their possible genotoxic effects, at doses that can be found in body fluids, on human lymphocytes using the cytokinesis-block micronucleus assay and the comet assay. After exposure for 1, 6, and 24h compounds p,p′-DDT (0.1μgmL−1), p,p′-DDE (4.1μgmL−1), and p,p′-DDD (3.9μgmL−1) showed increase in DNA damage. The most significant results were observed at exposure period of 24h where number of micronucleated cells increased from control 2.5±0.71 to 23.5±3.54, 13.5±0.71, and 16.5±6.36 for DDT, DDE, and DDD, respectively. Similar effect was observed using comet test where the percentage of DNA in comets tail increased from control 1.81±0.16 to 17.24±0.55, 11.21±0.56 and 9.28±0.50 for each compound, respectively. At the same time Fpg-comet assay failed to report induction of oxidative DNA damage of these pollutants. Additionally, the type of cell death was determined using diffusion assay and necrosis dominated. Our findings suggest that even at low concentrations, these pesticides could induce cytogenetic damage to human peripheral blood lymphocytes and in that manner have the impact on human health as well.

Genotoxicity assessment in smokeless tobacco users

India has a long history of tobacco, which includes chewing tobacco and smoking tobacco in various forms. Initially, the smokeless tobacco chewing habit was seen among the majority of the farmers who cultivated tobacco; but in recent years, smokeless tobacco is available in many forms and is cheaper as well and hence it is widely being used among literate and illiterate people. The subjects of our study are living in hilly regions of Yerkaud in Salem district, South India. Most of the inhabitants of our study area are illiterate and more particularly they are unaware of the health effects due to tobacco use. Recent epidemiological reports have strongly indicated the association of cancer risk with usage of smokeless tobacco. The prime aim of our study is to evaluate the genotoxic effects of tobacco use by analysing the cytogenetic end points such as chromosome aberrations in peripheral blood and micronucleus in peripheral blood and buccal cells. About 85 smokeless tobacco users were enrolled for the study and same numbers of age- and sex-matched nontobacco users were also enrolled to serve as controls. The result of our study revealed that tobacco users displayed varied levels of elevated chromosomal damage and micronucleated cells than nontobacco users. The variation in the extent of genetic damage was dependent on the duration of the tobacco use. In conclusion, this study might be helpful in creating awareness on the hazards of the smokeless tobacco products among the global population as a whole for those who chose such products as a cheap alternative to tobacco smoke.

Micronucleus assay as a triage tool for borderline cases of cervical dysplasia

Objective: In cases of cervical carcinogenesis the gradient from low to high-grade lesions is characterized by increasing nuclear atypia and the failure of cellular differentiation. These phenotypic changes are presumed to be accompanied by increased genetic instability that can be documented using the micronucleus (MN) assay in exfoliated cervical cells. Material and methods: This blinded prospective study included 169 samples for final analysis; 119 normal cervical cytology, 18 preinvasive lesions and 33 frank carcinoma. A thin smear was prepared from each sample on a glass slide, which was then stained with acridine orange (a DNA specific dye). Around thousand cells per slide were counted under fluorescent microscope, 20-40 X magnification. Number of micro-nucleated cells and total number of micronuclei were recorded to calculate MN cell ratio and total MN ratio. Results: Micro-nucleated cells as well as the total number of micronuclei show increasing trend towards malignization. By ROC curve analysis, for MN cell ratio of 0.1999, the sensitivity is found to be as high as 91.4% with a specificity of 82.8%. Similar results were obtained when total MN ratios were analyzed. Advantage of combining both the ratios together does not confer any additional benefit in terms of sensitivity and specificity. Key words: micro-nucleus (MN); cervical cancer; lowgrade squamous intraepithelial lesion (LSIL); highgrade squamous intraepithelial lesion (HSIL). DOI: http://dx.doi.org/10.4038/sljog.v33i3.4018 Sri Lanka Journal of Obstetrics and Gynaecology 2011; 33 : 104-111

In vitro genotoxic perspective of Tamiflu

The aim of this study was to investigate the genotoxic and/or cytotoxic effects of Tamiflu, commercial form of the oseltamivir antiviral and most frequently prescribed for the treatment of influenza infections, on cultured human peripheral lymphocytes by using sister chromatid exchange (SCE), chromosomal aberration (CA), and cytokinesis-blocked micronucleus (CBMN) assays. Cells were treated with 0.5, 1, 2μg/mL oseltamivir, the Tamiflu capsule ingredient, for 24 or 48h in the absence or presence of an exogenous metabolic activation system (S9 mix). The test chemical did not demonstrate any genotoxic effect dose-dependently but it showed a weak cytotoxicity on cells in this study. On the other hand, some concentrations of Tamiflu (2μg/mL without S9 mix for 48h and 1μg/mL with S9 mix) induced SCE and also decreased significantly the proliferation index (PI) (48 h period) and the nuclear division index (NDI) (24h period) (P<0.05) in the absence of S9 mix. Considering the results, Tamiflu did not induce significant increases of CA or micronucleated cells in vitro in cultured peripheral blood lymphocytes under the treatment conditions used but weak SCE induction was observed. On the other hand, the weak cytotoxic effects observed disappeared in the cultures treated in presence of the S9 mix.

Analysis of micronuclei in buccal epithelial cells in patients subjected to panoramic radiography

Ionizing radiation is a well-known carcinogen in humans. Chromosomal aberrations and formation of micronuclei in cell cytoplasm are early biological evidence of carcinogenesis. This study was undertaken to assess the genotoxic effect of panoramic radiography in the buccal epithelial cells. The study included 60 healthy individuals (median age 23.5 years; age range 12-65 years) who underwent panoramic radiographic examination. Exfoliated buccal epithelial cells were obtained immediately before and 10 days after radiation exposure. The cells were stained with Giemsa and evaluated for micronuclei by scoring 1000 cells per sample. The paired 't ' test was used to find out the significance of difference in the number of micronuclei before and after x-ray exposure. The Karl Pearson correlation coefficient was used to find out the correlation between age and micronucleated cell frequencies and number of micronucleus per 1000 cells. The ANOVA test was used to find out if there were significant differences in micronucleated cell frequencies between different age-groups. Student's unpaired 't' test was used to find out the significance of difference in micronucleated cell frequencies and number of micronucleus per 1000 cells between genders. The paired 't' test showed that micronucleated cell frequencies (P = 0.02) and number of micronucleus per 1000 cells (P = 0.047) were significantly higher after radiographic exposure. The mean number of micronucleated cells before and after radiation exposure were 0.48 ± 0.14 and 0.51 ± 0.15, respectively. There was statistically significant increase in the frequency of micronuclei in buccal epithelial cells after exposure to panoramic radiography. The correlation of micronucleus frequency with age and gender was statistically nonsignificant. The results indicate that panoramic radiography may induce genotoxic effects in buccal epithelial cells. Considering this risk, panoramic radiography should be used cautiously.

Evaluation of micronuclei in tobacco users: A study in Punjabi population

Introduction:The assessment of micronuclei in exfoliated cells is a promising tool for the study of epithelial carcinogens and can be used to detect chromosome breakage or mitotic interference, thought to be relevant to carcinogenesis.Aim:The present study aimed to detect micronuclei in exfoliated oral mucosal cells in individuals using various tobacco forms from the last 5 years.Materials and Methods:A total of 75 healthy male subjects (25 smokeless tobacco users, 25 smokers, and 25 non–tobacco users) were selected for the study. Smears were obtained with moistened wooden spatula from buccal mucosa and fixed with 95% alcohol. All the cytologic smears were stained by Papanicolaou technique. From each slide, ~1000 cells were examined under the 400× magnification and where micronucleated (MN) cells were located, they were examined under the 1000× magnification.Result:MN cells were found to be significantly higher in smokeless tobacco users than in smokers and controls.Conclusion:A positive correlation is found between increased micronucleus frequency and tobacco-using habits. So micronucleus assay can be used as a biomarker of genotoxicity.

Effects of Short-Term Consumption of a Large Amount of Tea Catechins on Chromosomal Damage, Oxidative Stress Markers, Serum Lipid, Folic Acid, and Total Homocysteine Levels: A Randomized, Double-Blind, Controlled Study

Objective: To evaluate the effects of short-term consumption of a large amount of tea catechins on chromosomal damage, oxidative stress markers, serum lipid, folic acid, and total homocysteine levels in middle-aged healthy volunteers.Methods: Forty volunteers (40-63 years) participated in a randomized, double-blind study. After a 1-week washout, the catechin group consumed approximately 1069 mg/day of total catechins for 1 week. The micronucleated binucleate cells (MNi) frequency in the cytokinesis-block micronucleus cytome assay, urinary 8-hydroxydeoxyguanosine (8-OHdG), isoprostane, lymphocyte and plasma vitamin C, serum lipid, folic acid, and total homocysteine levels were measured at the beginning and end of the intervention.Results: No significant differences were observed between the catechin and placebo groups in terms of MNi frequency, urinary 8-OHdG, isoprostane, or lymphocyte and plasma vitamin C levels. The serum LDL-cholesterol level in the catechin group significantly decreased compared with pre-intervention period, and there was a decreased tendency in the catechin group compared with the placebo group, but the difference was not significant (P= 0.105). The serum folic acid level decreased (P= 0.073) and the total homocysteine level significantly increased in the catechin group (P= 0.029). No serious adverse events were observed during the study.Conclusions: A large amount of tea catechins, which corresponds to approximately 10 cups of green tea per day for 1 week, seemed to be well tolerated, and did not influence chromosomal damage and the oxidative stress markers. Further long-term and large-scale studies are required to clarify the long-term effect of the consumption of a large amount of tea catechins on these markers as well as on improving dyslipidemia.Trial Registration: ClinicalTrials. gov ID, NCT00448513

Micronucleus investigation in human buccal epithelial cells of gutkha users.

Background:Gutkha is a cheap and convenient betel quid substitute, which is popular among all age groups. Various studies reveal its carcinogenic nature that leads to oral submucosus fibrosis and increases the chances of oral cancer. The micronucleus (MN) assay in exfoliated mucosal cells is a useful method for observing genetic damage in humans.Aim:To observe the genotoxic effect of gutkha on human buccal epithelial cells.Materials and Methods:The MN assay was performed to assess the frequency of MN in human buccal epithelial cells. The study comprises 60 individuals of which 30 individuals were gutkha chewers and another 30 were nonusers (control). The MN frequency was scored to estimate the genotoxic damage.Results:In gutkha users, the frequency of MN was highly significant (17.4 ± 0.944) as compared with nonusers (control) groups (4.53 ± 0.331) (P < 0.001).Conclusions:The MN assay in human buccal epithelial cells is a useful and minimally invasive method for monitoring genetic damage in humans. Asignificantly higher frequency of micronucleated cells are found among gutkha users.

Genotoxic potential of methyleugenol and selected methyleugenol metabolites in cultured Chinese hamster V79 cells

Methyleugenol is a substituted alkenylbenzene classified by the European Union's Scientific Committee on Food as a genotoxic carcinogen. We addressed cytotoxicity, genotoxicity and mutagenicity caused by methyleugenol and selected oxidative methyleugenol metabolites in Chinese hamster lung fibroblasts V79 cells. Cytotoxicity was measured by two cell proliferation assays, water soluble tetrazolium salt (WST) 1 and sulforhodamine B (SRB) assays. Genotoxicity was determined by using single cell gel electrophoreses (comet assay) and the in vitro micronuclei test, while mutagenicity was investigated with the hypoxanthinephosphoribosyl transferase (hprt) assay. Methyleugenol and 1'-hydroxymethyleugenol showed no or marginal cytotoxic effects, but caused DNA strand breaks at concentrations ≥10 μM. The metabolites methyleugenol-2',3'-epoxide and 3'-oxomethylisoeugenol exhibited growth inhibitory properties with IC(50)-values of 70-90 μM after 48 h or 72 h of incubation. These metabolites significantly enhanced cytotoxicity and DNA damage after 1 h of incubation. Overall, no increase in formamidopyrimidine DNA glycosylase sensitive sites were detected with the comet assay. The metabolites 1'-hydroxymethyleugenol and methyleugenol-2',3'-epoxide exceeded the DNA strand breaking properties of the parent compound methyleugenol. However, only 3'-oxomethylisoeugenol and methyleugenol-2',3'-epoxide induced the formation of micronucleated cells in comparison to the negative control. These compounds were found to be not or rather weakly mutagenic at the hprt locus. In summary, phase I metabolites exceeded the cytotoxic and genotoxic properties of the parent compound methyleugenol.

Micronucleus assay of buccal mucosa cells in smokers with the history of smoking less and more than 10 years

Cigarette smoking causes severe health problems such as cancer. Micronuclei are structures that present after genomic damages to the cells. The present study is aimed at evaluating the micronucleus assay of buccal mucosa cells in smokers who smoked less or more than 10 years. The present study has been a historical cohort study. The smokers were divided into two groups: First group include individuals with a smoking history less than 10 years (14 samples) and second group with the smoking history of more than 10 years (26 samples).The control group consisted of nonsmokers (23 samples). The exfoliated buccal mucosa cells were scrapped using spatula and were spread over the glass slide. Feulgen method was used for micronucleus staining. 500 cells per subjects were counted. The presence of micronucleus in all subjects and the mean percentage of micronucleus in nuclei were determined. Data were subjected to statistical analysis using T-test. The mean number of micronucleus of buccal mucosa cells in nonsmokers, first group (smoking history less than 10 years) and second group (smoking history more than 10 years) was 0.94±0.94, 1.89±0.62 and 2.01±0.93 respectively. The difference was statistically significant (P<0.002). Considering the number of micronuclei of the buccal mucosa cells, the difference between groups 1 and 2 was not significant (P<0.6). The mean percentage of micronucleated cells in nonsmokers, group 1 and group 2 was 2.26±2.17%, 13.9±5.90 and 14.3±7.97, respectively. The difference was statistically significant (P<0.001).The difference between the percentage of the cells with micronucleus in smokers with a smoking history of less or more than 10 years was not significant (P<0.6). The mean number of micronuclei in buccal mucosa cells of the nonsmokers was significantly lower than that of the smokers. However, the mean number of micronucleus of buccal mucosa cells in smokers who smoked more than 10 years was higher than smokers who smoked less than 10 years. Increasing the smoking duration could heighten the frequency of micronucleus; however, the difference was not significant.

Comparing the level of bystander effect in a couple of tumor and normal cell lines

Radiation-induced bystander effect refers to radiation responses which occur in non-irradiated cells. The purpose of this study was to compare the level of bystander effect in a couple of tumor and normal cell lines (QU-DB and MRC5). To induce bystander effect, cells were irradiated with 0.5, 2, and 4 Gy of 60Co gamma rays and their media were transferred to non-irradiated (bystander) cells of the same type. Cells containing micronuclei were counted in bystander subgroups, non-irradiated, and 0.5 Gy irradiated cells. Frequencies of cells containing micronuclei in QU-DB bystander subgroups were higher than in bystander subgroups of MRC5 cells (P < 0.001). The number of micronucleated cells counted in non-irradiated and 0.5 Gy irradiated QU-DB cells was also higher than the corresponding values for MRC5 cells (P < 0.001). Another difference between the two cell lines was that in QU-DB bystander cells, a dose-dependent increase in the number of micronucleated cells was observed as the dose increased, but at all doses the number of micronucleated cells in MRC5 bystander cells was constant. It is concluded that QU-DB cells are more susceptible than MRC5 cells to be affected by bystander effect, and in the two cell lines there is a positive correlation between DNA damages induced directly and those induced due to bystander effect.

Significance of micronucleus in cervical intraepithelial lesions and carcinoma

Background:The micronuclei (MNi) test on exfoliated cells has been successfully used to screen population groups at risk for cancers of oral cavity, urinary bladder, cervix and esophagus. Their frequency appears to increase in carcinogen-exposed tissues long before any clinical symptoms are evident. There are only limited numbers of studies on MN scoring in cervical pre-neoplastic and neoplastic conditions.Aims:To compare the micronucleus (MN) score in the whole spectrum of cervical lesions including normal, inflammatory, abnormal squamous cells of undetermined significance (ASC-US), abnormal squamous cells cannot exclude HSIL (ASC-H), low-grade squamous intraepithelial lesion (LSIL), high-grade squamous intraepithelial lesion (HSIL) and invasive cancer (IC) and to evaluate the role of MN as a biomarker in different pre-neoplastic and neoplastic lesions.Materials and Methods:A total of 221 slides, comprised of normal (32), inflammatory (32), ASC-US (31), ASC-H (31), LSIL (32), HSIL (31) and IC (32), were studied. All the cases were reviewed by two pathologists independently. Histopathological correlation was done in a few cases of ASC-US, ASC-H, HSIL and IC which were available in the department. Two observers separately and independently counted the number of micronucleated cells per 1,000 epithelial cells in oil immersion magnification and were expressed as MN score per 1,000 cells.Results:The mean MN scores ± SD in normal, inflammatory, ASC-US, ASC-H, LSIL, HSIL and IC cases of cervical lesions were 0.84±0.68, 1.06±0.84, 3±0.73, 4.78±1.43, 4.06±1.13, 8.03±1.64 and 10.5±2.01, respectively. MN scores of IC and HSIL were significantly high compared to normal (P<0.000), inflammatory (P<0.000), ASC-US (P<0.000), ASC-H (P<0.000) and LSIL (P<0.000) group (analysis of variance test). LSIL showed significant difference with the normal (P<0.000), inflammatory (P=0.001), ASC-US (P=0.028), HSIL (P<0.000) and IC (<0.000), but not with the ASC-H (P=0.64) group.Conclusions:MN scoring on the epithelial cells of cervix could be used as a biomarker in cancer screening. This is an easy, simple, reliable, reproducible and objective test which can be performed on routinely stained pap smears.

The role of mutations in the genes FLG and GSTM1, and cytogenetic aberrations in the developmentof allergic occupational dermatitis in the oilfield workers of the north of Siberia

Background. The objective of the investigation was to determine frequencies of micronucleated cells in the buccal epithelium in relation to polymorphism of genes FLG and GSTM1 among oilfield workers suffered from allergic occupational dermatitis. Methods. There were 209 male oilfield workers examined with cases of allergic professional dermatitis at the age of 21-39 years old. as a control group there were used 207 male workers without allergic occupational dermatitis. we used genealogical method, micronucleous test and polymerase chain reaction to detect restriction fragments of genes GSTM1 and FLG. Results. it was detected that oilfield workers with genotypes contented homozygous mutated alleles GSTM1 (0/0) gene showed the higher frequency of micronucleated epithelial cells. it was not detected for FLG gene allele variants. among the examined oilfield workers with allergic occupational dermatitis there was nobody that would have both mutated alleles of FLG (282de14/282de14) and homozygous GSTM1 (0/0) genes, while in the control group we found some individuals of this type. Conclusions. The significantly higher levels of cytogenetic aberrations in the oilfield workers suffered from allergic occupational dermatitis are the result of combined effects of various factors on the genome. The investigation can provide the basis for the elaboration of new scientific criteria for hiring staff in oilfields of west siberia.

Cytogenetic biomonitoring in children submitting to a complete set of radiographs for orthodontic planning

To evaluate the DNA damage (micronucleus) and cellular death (pyknosis, karyolysis, and karyorrhexis) in exfoliated buccal mucosa cells from children undergoing orthodontic radiographs. A total of 25 healthy children undergoing orthodontic therapy partook in a complete set of orthodontic radiographs (lateral cephalographic, posteroanterior cephalographic, panoramic, full periapical exam, and bitewing). The micronucleus test in the buccal exfoliated cells was applied. The paired-samples t-test and the Wilcoxon test were used to compare the frequencies of alterations before and after X-ray exposure. We found no statistically significant differences (P > .05) between micronucleated buccal mucosa cells before and after exposure to radiation. However, radiation did cause other nuclear alterations closely related to cytotoxicity (P = .007). According to the micronucleus test, the complete set of radiographs requested in the orthodontic planning may not be a factor that induces chromosomal damage, but it is able to promote cytotoxicity.

Studies on the Genotoxicity ofGutkha

Nowadays, smokeless tobacco consumption in the form of gutkha is receiving more popularity among the youth of India. In the present investigation, cytogenetic studies were carried out on 50 gutkha consumers and compared with controls, 50 men who did not have habit of smoking, alcohol drinking and consuming gutkha, tobacco and areca nut in any form. Frequency of chromosome aberrations (CA) and sister chromatid exchanges (SCE) were evaluated in peripheral blood lymphocytes, whereas frequency of micronucleated cells (MNC) was evaluated in exfoliated buccal mucosa. A significant increase (p 31) compared with low age group individuals viz CA (1.67-2.81), SCE (4.65-5.80), MNC (0.36-0.70). The increased frequency of these end points was found to be significantly correlated ( p<.001) with duration of consumption (b = 0.193, 0.039, 0.032 for CA, SCE and MNC respectively) and number of pouches consumed per day (b being 0.193, 0.441, 0.429 for CA, SCE and MNC respectively). Besides this, significant dif ference was found between the mean values of CA and SCE among gutkha consumers and gutkha consumers + drinkers + smokers and also significant increase was found between gutkha consumers + smokers and gutkha consumers + drinkers + smokers for CA.