Average Micronuclei Frequency Research Articles

Genome instability in peripheral blood lymphocytes of patients with heart failure and reduced ejection fraction.

Heart failure (HF) is a complex clinical condition characterized by functional and structural defects of the myocardium, but genetic and environmental factors are considered to play an important role in the development of the disease. In the present study, we investigated the genome instability (DNA and chromosomal damage) in patients with heart failure with reduced ejection fraction (HFrEF) ≤40% and its association with risk factors. The studied population included 48 individuals, of which 29 HFrEF patients (mean age 57.41 ± 5.74 years) and 19 healthy controls (mean age 57.63 ± 6.09 years). The genetic damage index in peripheral blood lymphocytes was analyzed using the comet assay, while micronuclei frequency and nuclear division index were analyzed using the cytokinesis-block micronucleus assay. Our results showed that HFrEF patients had a significantly higher genetic damage index compared with the healthy controls (P < .001). Cytokinesis-block micronucleus assay showed that the average micronucleus frequency in peripheral blood lymphocytes of patients was significantly higher, while the nuclear division index values were significantly lower than in controls (P < .01). Using multiple linear regression analysis, pathological state, ejection fraction, creatinine, glucose, associated disease, residence, proBNP, troponin, urea, ACE-inhibitors, and length of the drug therapy were identified as predictors of DNA and/or chromosomal damage in HF patients. We can conclude that DNA and chromosomal damage was increased in patients with HF, which may be a consequence of disease and/or drug therapy.

Effect of urbanization on the micronucleus frequency in birds from forest fragments

The urbanization contributes to environment contamination, mainly by the increased vehicle traffic. This intense traffic releases in the air chemical compounds with mutagenic properties that can affect the entire ecosystem. The birds for the flight accumulate air, absorbing gases or particles. The absorption of this polluted air may be causative agent responsible for micronucleus (MN) induction in bird erythrocytes. The quantitative analysis of MN in birds can assists in the monitoring environmental quality of various places. The study aimed to quantify the MN frequency in birds and compare their variation: I) among forest fragments distant from and near to urbanization, II) among species and populations of birds and III) to find if there is a relation between vehicle traffic and MN induction. The capture of the birds took place in four forest fragments in central Brazil (two distant and two near the city) using mist nets. Blood smears were prepared with blood collected from the tarso-metatarsal vein. A total of 354 individuals were analyzed and MN were found in 52% of the individuals analyzed. All sampled birds were identified and resulted to belong to 50 different species, only 4 of which were captured in all the 4 forest fragments. The average MN frequency, regardless of bird species and sampling area, was 1.04/10,000 erythrocytes. From an overall analysis carried out on all birds, the MN frequencies were found to vary among forest fragments distant from and near to urbanization (x2 =15.513; p < 0.001) and demonstrated positive correlation between vehicle traffic intensity and the MN frequency (r = 0.988; df=2; p = 0.011). The species of birds presented variation in the amount of MN (x2 = 84.64; df=49; p = 0.001). Therefore, a restricted analysis carried out on the 4 species of birds present in all the forest fragments showed that the populations of Antilophia galeata (x2 =6.029; p = 0.014), Basileuterus culicivorus (x2 =9.53; p = 0.002), Eucometis penicillata (x2 =8.067; p = 0.005) and Myiothlypis flaveola (x2 =4.771; p = 0.029) showed difference in the MN frequency, when compared between forest fragments distant from and near to cities. The analysis demonstrated that birds living in forest areas near to the city presented higher MN frequency, probably because pollutants generated by urbanization affect birds and that the larger the vehicle traffic the greater the induced MN in the birds.

Cytological grading: An alternative to histological grading in oral squamous cell carcinoma.

Background:Micronuclei (MN) in oral exfoliative cells have been shown to indicate a disparaging change in genetic information of the cell. Recent studies showed correlation between the frequency of MN and severity of this damage. Grading of lesions can be used to determine the austerity of this damage. Aims: The aim of this study is to compare the MN frequency in oral exfoliated cells of normal and oral squamous cell carcinoma (OSCC) individuals and to cytologically grade the frequency of MN in cytological smears and to correlate it with histological grading. The objective is to ascertain whether MN frequency in oral exfoliated cells can be a parameter for grading of OSCC.Settings and Design:The study group comprises of 40 subjects (20 controls and 20 OSCC patients) in the age group of 45-85 years.Materials and Methods:The cytosmear was obtained from each group and stained with Papanicolaou (PAP) stain. Twenty cells from each slide were counted for MN and cytological grade of OSCC was assigned based on the average frequency of MN. Cytological grade was correlated with histological grading and the data were recorded. Student's t-test and Spearman's correlation were used for the analysis of the data.Results:Average frequency of MN was 2.5 times higher in OSCC patients when compared to that in controls and the difference was found to be highly significant. Sixty percent correlation was found between cytological grade and histological grade of OSCC and the difference between them was not significant.Conclusions:Cytological grading can be used in grading OSCC, and MN insinuates genotoxic damage occurring in the epithelial cells.

Sublethal exposure of heavy metals induces micronuclei in fish, Channa punctata

Present studies were designed to evaluate toxic potential of three common heavy metals, adequately present in agro-industrial effluents, viz. mercury, arsenic and copper using in vivo micronucleus assay in an actinopterygiian fish, Channa punctata (2 n = 32). Ten days laboratory acclimatized fishes were divided into five groups. Groups I and II served as negative and positive controls, respectively and fishes of group III–V were subjected uninterrupted to sublethal concentrations (10% of 96 h LC 50) of heavy metal compounds, HgCl 2 (0.081 mg L −1), As 2O 3 (6.936 mg L −1) and CuSO 4·5H 2O (0.407 mg L −1) for 24, 48, 72, 96 and 168 h of exposure periods. Significant increase over and above negative control in the frequency of micronuclei was observed in fishes exposed to metal compounds. The average frequency of micronuclei in fishes exposed to Hg(II), As(III) and Cu(II) observed was 9.79, 12.03 and 8.86, respectively. It reveals that the order of induction of micronuclei frequency and toxicity was As > Hg > Cu. Findings depict genotoxic potential of these metal compounds even in sublethal concentrations.

The antibiotic erythromycin did not affect micronucleus frequency in human PHA-stimulated lymphocytes

The aim of the present study was to investigate the ability of the antibiotic erythromycin to induce micro?nucleus (MN) frequency and the cytokinesis block proliferation index (CBPI) in human peripheral blood lymphocytes in vitro using the cytokinesis-block micronucleus test (CBMN). Cell cultures were treated with five different concentrations of erythromycin ranging from 0.68 x 10-4 to 5.45 x 10-4 M. The positive control cells were treated with a known mutagen, mitomycin C (MMC), in a concentration of 1.6 x 10-6 M. None of the tested concentrations of erythromycin significantly changed the average MN frequency or CBPI in PHA-stimulated lymphocytes (p &gt; 0.05).

Treatment of Pregnant Women with a Betamimetic and Verapamil Increases the Micronuclei Frequency in Umbilical Cord Blood Lymphocytes

In prevention of preterm labor, betamimetics are used in gynecological practice mostly combined with antiarrhythmic verapamil because of their therapeutic cardiovascular side effects. The aim of this study was to investigate the influence of a betamimetic (ritodrine hydrochloride, fenoterol or hexoprenaline) and verapamil (administered to mothers) on the frequency of micronuclei (MN) in umbilical cord blood lymphocytes of neonates, using cytokinesis-block micronucleus test. The analyzed sample included 23 babies whose mothers received the therapy and 30 control babies whose mothers received no therapy. The average MN frequency was significantly higher in the neonates whose mothers received the therapy (8.13 +/- 2.69 MN/1000 BN cells), in comparison with the baseline frequency in untreated controls (3.30 +/- 2.63 MN/1000 BN cells), with probability p < 0.05. The highest MN frequency was found in neonates whose mothers received fenoterol and verapamil (2.8-fold i.e. 9.10 +/- 3.00 MN/1000 BN cells), while ritodrine hydrochloride and hexoprenaline combined with verapamil induced 2.3-fold and 2.2-fold higher MN values than in controls (7.50 +/- 3.33 MN/1000 BN cells and 7.29 +/- 0.95 MN/1000 BN cells). Multiple linear regression analysis showed that MN frequency was affected only by the maternal therapeutic treatment, while the neonates' sex, maternal age, cigarette smoking, and therapeutic doses did not affect the MN frequency in umbilical lymphocytes of neonates. We conclude that the treatment of pregnant women with a betamimetic and verapamil significantly increases the MN frequency in umbilical cord blood lymphocytes of neonates, regardless to therapeutic doses.

Micronuclei Frequency in Lymphocytes and Antioxidants in the Blood of Traditional Limited-Resource Farm Workers Exposed to Pesticides

Chronic low-level exposure to synthetic pesticides is implicated in many health conditions that result from the induction of oxidative stress, including cytogenetic damage. The objective of this study was to assess the risk of genotoxicity using micronuclei (MN) formation in lymphocytes and to determine changes in blood antioxidants superoxide dismutase (SOD) in erythrocytes (E) and glutathione (GSH) in E and plasma (PL) in farm workers for six months during a growing season. Blood and urine samples were collected once a month for six months (June to November 2003) from farm workers (n = 15) and urban unexposed controls (n = 10). Lymphocytes from blood were separated by density gradient centrifugation using Histopaque and cultured using the standard technique. There was no significant difference in the cytokinesis blocked proliferation index (CBPI) of lymphocytes between the farm workers and the control group, but there was a 76% increase in average MN frequency in lymphocytes of the farm worker group (P ≤ 0.05). In addition, MN frequency peaked during August as compared to the other months and the controls (P ≤ 0.05). An 18% decline was observed in the activity of E-SOD in the farm worker group (P ≤ 0.05). GSH in E and PL were similar in both groups. These data suggest that the farm workers may be at a greater risk of developing genotoxicity due to continued exposure to pesticides, especially during the intensive growing season.

The Frequency of Micronuclei among Newborns from Kragujevac, Central Serbia, after NATO Bombing in the Spring of 1999

Kragujevac, as an important industrial and economic center of the region, is now placed on the top of the list of environmentally endangered cities in our country. The aim of this biomonitoring study was to evaluate cytogenetic damage in a sample of newborns from Kragujevac after contamination of the environment entailed by the intensive bombing of the industrial zone of this city in the spring of 1999. The frequency of micronuclei (MN) in peripheral blood lymphocytes in a total of 61 phenotypically healthy newborns was analyzed before and after the NATO bombing. Analysis of micronuclei has been performed using the cytokinesis-block technique (CB MN test). Average MN frequency in lymphocytes of newborns before the bombing (N = 25) was 5.77 ± 0.85/1000 analyzed cells. After the bombing (N = 36), the average frequency of MN increased by a factor of 1.4 (8.11 ± 0.85), compared to the control frequency before the bombing. Statistical difference (p < 0.05) was established by Student's t-test. Our data suggest that such changes in genetic material were a direct consequence of contamination of the living environment.

Effect of a low-dose ethinylestradiol and gestodene in combination on the frequency of micronuclei in human peripheral blood lymphocytes of healthy women in vivo

Background Since most oral hormonal contraceptives contain a fixed combination of ethinylestradiol and gestodene as an estrogenic/progestogenic component, we decided to evaluate the possible mutagenic effect of a low-dose contraceptive pills containing 20 μg ethinylestradiol and 75 μg gestodene. Methods A total of 30 healthy women received hormonal contraception during six consecutive menstrual cycles. A single daily dose was 20 μg ethinylestradiol and 75 μg gestodene. The pills were taken orally in a monthly cycle of 3 weeks on and 1 week off. In the investigation of the mutagenic effect of these contraceptive pills in vivo, the cytokinesis block micronucleus (CBMN) test was used, and the frequency of micronuclei (MN) in peripheral blood lymphocytes was determined. Results Average MN frequency in women before therapy was 7.40 ± 0.75 MN/1000 analyzed cells, and after therapy was 7.37 ± 0.59 MN/1000 analyzed cells (p > 0.05). Conclusion Our data suggest that oral contraception with 20 μg ethinylestradiol and 75 μg gestodene in combination during six consecutive menstrual cycles does not induce micronuclei in peripheral blood lymphocytes of women.

Effects of ALDH2 gene polymorphisms and alcohol-drinking behavior on micronuclei frequency in non-smokers

Alcohol abuse is a serious health problem, leading to life-threatening damage to most of the important organ systems. Genotoxic damage is used as an early effect indicator in the surveillance of human exposure to genotoxic substances. Intra- and inter-individual variations of baseline frequencies of micronuclei (MN) in peripheral blood lymphocytes of human populations have been reported previously. Polymorphisms in a few metabolic enzyme genes seem to account for a proportion of this variability, but the impact of specific genetic variants on MN frequencies has not yet been clarified. In 42 healthy Japanese non-smoking men, we investigated the relationship between the MN frequency levels and genetic polymorphisms in three different genes: aldehyde dehydrogenase 2 (ALDH2), X-ray repair cross-complementing group 1 (XRCC1) and excision repair cross-complementing group 2 (ERCC2). Genotyping was performed by PCR–RFLP analysis. The ALDH2 variant (deficient-type) was significantly associated with increased MN frequency levels in subjects with drinking more than three times per week, whereas the XRCC1 and ERCC2 variants seemed to be unrelated to the MN frequency. The ALDH2-deficient habitual drinkers had an average MN frequency of 5.88±0.58 (±S.E.) compared with 3.20±0.80 in the ALDH2-proficient habitual drinkers ( P<0.05). The ALDH2-proficient non-habitual drinkers had the lowest MN frequency (1.56±0.41). Furthermore, subjects with highest levels of mean MN frequency, who consumed more than 100 g of alcohol per week and more than three times per week, had A2 genotype of ALDH2. A significant odds ratio (12.25, P<0.05) for the MN frequency levels above the 50th percentile value was observed for the ALDH2-deficient individuals versus the ALDH2-proficient individuals after adjustment for several confounders. These results strongly suggest that human early genotoxic effect studies based on the cytogenetic markers of MN should take into account both the individual ALDH2 polymorphism and the potential confounding effect of the drinking behavior.

Lack of significant genotoxicity of purified soy isoflavones (genistein, daidzein, and glycitein) in 20 patients with prostate cancer

Genistein may be useful in the prevention or treatment of prostate cancer; however, it causes genetic damage in cultured human cells. The objective was to assess the potential genotoxicity of a purified soy unconjugated isoflavone mixture in men with prostate cancer. Twenty patients with prostate cancer were treated with 300 mg genistein/d for 28 d and then with 600 mg/d for another 56 d. In peripheral lymphocytes, DNA strand breaks were assessed as nuclear tail moment, chromosomal damage was assessed as micronucleus frequency (MF), and translocations of the MLL gene (11q23) were assessed by using fluorescence in situ hybridization. Values are also reported for 6 healthy men. The studies were performed under Investigational New Drug application no. 54 137 at a tertiary referral academic medical center. No changes in group average or individual nuclear tail moment and MF were observed. We observed a single elevated MF value in one subject that exceeded a clinical threshold set before we initiated the study. A significant decrease in average COMET tail moment was observed on day 28 relative to day 0. We detected no genistein-induced rearrangements of the MLL gene in the 3 subjects we studied with this technique. MF increased significantly in lymphocytes exposed in vitro to unconjugated genistein at concentrations > or = 100 micromol/L. Total genistein never exceeded a peak concentration of 27.1 micro mol/L, and unconjugated genistein never exceeded a peak concentration of 0.32 micromol/L. Although isoflavones are capable of inducing genetic damage in vitro, a similar effect was not observed in subjects treated with a purified soy unconjugated isoflavone mixture.

Effect of various doses of gestogens on micronuclei frequency in human peripheral blood lymphocytes of pregnant women.

Since gestogens, in the form of hormonal substitution therapy, have been proposed to have a role in the prevention of threatened spontaneous abortions during the first three months of pregnancy, we decided to evaluate possible genotoxic effects of these preparations. A total of 30 pregnant women, with a diagnosis of threatened spontaneous abortions, received the gestogen therapy in the first 3 months, and a sample of 30 pregnant women without indication for hormonal therapy were included as the control group. For investigation of mutagenic effects of gestogens in vivo the cytokinesis block (CB) micronucleus (MN) test was applied. Average MN frequency in the control group was 6.79 +/- 0.69 MN/1000 cells. The second analysed group included 12 patients with threatened spontaneous abortions, who received gestogen therapy in doses of 100-750 mg. Average MN frequency in these patients before therapy was 11.83 +/- 1.33 MN/1000 cells, and after therapy it was 16.50 +/- 1.32 MN/1000 cells (P < 0.001). The third analysed group comprised nine patients, who received gestogen therapy in doses of 750-2000 mg. Average MN frequency in these patients before therapy was 15.67 +/- 3.00 MN/1000 cells, and after therapy was 23.89 +/- 2.49 MN/1000 cells (P < 0.001). The fourth analysed sample comprised nine patients, treated with gestogen doses of 2000-8400 mg. The average MN frequency in these patients before therapy was 11.89 +/- 1.63 MN/1000 cells, and after therapy was 21.22 +/- 2.80 MN/1000 cells (P < 0.001). The increase of therapeutic gestogen doses was followed by an increase of average MN frequency. The greatest rise of MN frequency (1.8-fold) was observed in the group of patients who were treated with the highest gestogen doses (2000-8400 mg). The smallest increase (1.4-fold) of MN frequency was found in the group of patients whose therapeutic doses were the lowest (100-750 mg).

Effect of intratumoral application of methotrexate in vivo on frequency of micronuclei in peripheral blood lymphocytes

BACKGROUND: Methotrexate is a cytostatic drug belonging to the group of antimetabolites; it is an antagonist of folic acid with an intensive application in gynecology practice. METHODS: The frequency of micronuclei (MN) in peripheral blood lymphocytes of 30 patients with myoma uteri was analyzed before and after intratumoral application of methotrexate (MTX) in total dose from 50 to 115 mg. Analysis of micronuclei was performed by the application of cytokinesis-block technique (CB). RESULTS: Average frequency of MN in lymphocytes of patients before the therapy was 4.6?0.4 MN/1000 analyzed cells. After the completion of therapy with six separate doses during six consecutive menstrual average frequency of MN increased 1.5 times (7.0?0.6) in comparison to control frequency before the therapy. Statistically significant difference (p&lt;0.001) was established by the Student t test. CONCLUSION: Methotrexate intratumoral treatment of the myoma uteri significantly increased micronuclei in peripheral blood lymphocytes.

The micronucleus assay in human lymphocytes after high radiation doses (5–15 Gy)

Individuals can be exposed to high doses (more than 5 Gy) during radiation accidents. It is, of course, helpful to the physician to have biological indicators also for such high doses. The problem with most cytogenetic indicators is, that the response levels off at doses starting around 5–7 Gy of low LET radiation and that the dose-response curve even declines after doses exceeding about 10 Gy. Thus, it may be difficult to decide, whether the dose was, for example, 8 or 14 Gy. We studied how the micronucleus assay can be used to give information also in the high dose range. It turned out that micronucleus frequency itself cannot be used for the estimation of doses exceeding about 5–7 Gy. There are, however, at least three other endpoints that can be determined in the cytochalasin B assay that can assist the decision in the high dose range: (1) the number of mononucleated cells; (2) the ratio of tri- to tetranucleated cells; (3) the average micronucleus frequency in micronucleus positive binucleated cells.

Micronuclei and ageing in a sample of Yugoslavian population

Instability in the organization and expression of the genetic material has been hypothesized as the basic mechanism of ageing. Aim of this study was to quantify the effect of ageing on spontaneous micronuclei (MN) frequency in peripheral blood lymphocytes. Analysis of Yugoslavian population sample (164 tested individuals, age 0-62 years) has performed by application of cytokinesis-block technique (CB). There was an increase of MN frequency with age, from newborns to 40-year-old persons. The total average of MN frequency per 1000 analyzed binuclear cells amounts to 8.03 +/- 0.42, with variation from 0 to 26 MNs. In a sample of 29 newborns the recorded average MN frequency was 6.91 +/- 0.81, while among 69 persons 25-39 years old, the MN frequency was 9.16 +/- 1.00. The lowest average MN frequency, however, was noted in the sample of 34 tested individuals 40 to 62 years of age. An increase with age in MN frequency has been observed in samples of studied individuals from newborns to 40-year-old persons. A decrease of MN frequency in older groups could be explained by a gradual decrease of proliferative cell capacities.

Induction of micronuclei and sister chromatid exchanges by polycyclic and N-heterocyclic aromatic hydrocarbons in cultured human lymphocytes.

Many natural environments are contaminated with carcinogenic polycyclic aromatic hydrocarbons (PAHs) and N-heterocyclic aromatic hydrocarbons (NHAs) as complex mixtures of coal tar, petroleum, and shale oil. These potentially hazardous substances are prevalent at many former tar production and coal gasification sites. Three polycyclic [benzo(a)pyrene (BaP), benz(a)anthracene (BAA), and 7,12-dimethylbenz(a)anthracene (DMBA)] and two N-heterocyclic [7H-dibenzo(c,g)carbazole (DBC), and dibenz(a,j)acridine (DBA)] aromatic hydrocarbons were analyzed for cytotoxic and genotoxic effects on human lymphocytes. All of these polyaromatic compounds are normally present in the environment, except for DMBA. Lymphocytes from healthy donors were isolated from whole blood. The 5-ring polycyclic aromatic BaP consistently induced micronuclei in a linear dose-dependent manner with doses from 0.1-10.0 micrograms/ml, whereas the 4-ring compounds (BAA and DMBA) had no effect on the induction of micronuclei above controls except at 5 and 10 micrograms/ml. Of the two N-heterocyclic compounds, DBC produced a significant increase in micronuclei in lymphocytes, but the dose response tended to plateau above 0.1 microgram/ml. DBA showed an effect on the frequency of micronuclei above controls only at high doses of 5 and 10 micrograms/ml. The average background frequency of micronuclei for 7 lymphocyte donors averaged 3.1 per 1,000 stimulated cells, whereas the average frequency of micronuclei at 10 micrograms/ml BaP was 36.8 per 1,000 stimulated cells. The lowest effective dose in 2 donors for BaP occurred at 0.1 microgram/ml. At a challenge dose of 1 microgram/ml (4 microM) of BaP, considerable variation in micronuclei induction between 7 individuals was observed, ranging from 2-6-fold increases above spontaneous frequency. Over a dose range of 1-10.0 micrograms/ml (4-40 microM), BaP also induced sister chromatid exchanges (SCEs) in lymphocytes, whereas BAA had no effect above controls. Parallel studies of both cytogenetic endpoints showed that the micronucleus assay is a more sensitive indicator of BaP exposure at equivalent doses. Mitotic and replication indices of BaP-exposed lymphocytes showed that cell proliferation is only moderately inhibited even at the highest dose; this shows that bulky DNA-adducts are generally compatible with cell survival. The cytogenetic data are consistent, first-off, with reports that individuals in the population vary widely with respect to the inducibility of the CYP1A1 gene, which is known to be involved in polycyclic aromatic hydrocarbon metabolism, in particular, in BaP. Secondly, the data support the fact that polyaromatic compounds differ with regard to micronucleus induction within the same sample(s) of human lymphocytes, indicating selective metabolism of polyaromatic compounds that may reflect carcinogen sensitivity of the individual.(ABSTRACT TRUNCATED AT 400 WORDS)