Exposure to nanoparticles of magnetite Fe3O4 in three different doses and their influence on selected respiratory parameters of bronchoalveolar lavage after intravenous instillation.

OBJECTIVES: Due to nano-dimensions (less than 100 nm), can nanoparticles probably penetrate through various membranes and travel from the bloodstream to other organs in the body. The aim of our study was to find out whether NPs Fe3O4 (coated with sodium oleate) injected into the tail vein of laboratory Wistar rats pass through the bloodstream to the respiratory tract (in comparison with a control group); and if so whether increasing doses of NPs Fe3O4 have an escalating harmful effect on selected bronchoalveolar lavage (BAL) parameters. METHODS: Wistar rats were intravenously given 3 doses of the suspension of NPs Fe3O4 (0.1% LD50 = 0.0364, 1.0% = 0.364 and 10.0% = 3.64 mg/kg animal body weight). Seven days later, we sacrificed the animals under anaesthesia, performed bronchoalveolar lavage (BAL), and isolated the collected cells. Many inflammatory and cytotoxic BAL parameters were examined. RESULTS: Both inflammatory and cytotoxic BAL parameters affected by Fe3O4 suspension were changed compared to control results, but not all were statistically significant. Thus, the NPs Fe3O4 passed through the bloodstream to the respiratory tract and affected it. The highest concentration of NPs Fe3O4 (10%) had the most influence on BAL parameters (7 of 12 parameters). Only 3 parameters showed a pure dose dependence. CONCLUSION: We assume that the adverse effect of Fe3O4 NPs in our study is probably not correlated with the dose, but rather with the size of the particles or with their surface area.

A considerable proportion of metabolic syndrome-free adults from Bratislava Region, Slovakia, display an increased cardiometabolic burden.

Although the dichotomous classification of metabolic syndrome (MS) enables the classification of individuals as MS-free or presenting MS, it is inconvenient for assessing cardiometabolic risk in MS-free individuals. Continuous MS score allows for estimation of cardiometabolic burden even in MS-free subjects. We used the scores to estimate the proportion of MS-free subjects on high cardiometabolic risk. A total of 876 subjects (62% females) of Central European descent, aged 20-81 years, were included. International Diabetes Federation (IDF) criteria were employed to classify MS. Continuous scores were calculated. We used the receiver operating characteristics (ROC) analysis to estimate the cutoff value to determine the proportion of MS-free subjects on increased risk. Using the waist circumference, 38% of males and 23% of females presented MS. ROC area under the curves (90%-98%) showed an acceptable performance of both scores to classify the presence of MS. Up to 18% of MS-free males and up to 10% of females displayed continuous score ≥ the relevant cutoff point. The waist-to-height ratio performed similar results. Both continuous scores were proven credible for assessing cardiometabolic risk in MS-free subjects. Clinically, this is important for earlier intervention. Despite minor differences between waist circumference and waist-to-height ratio, it would be appropriate to objectify it using reference population. Novelty: The first study using Z-MSS/siMSS (population-specific Z-score/continuous score of MS) to estimate cardiometabolic risk in Slovak adults. A proportion of MS-free Slovak adults is on high cardiometabolic risk. Difference between using waist circumference and the waist-to-height ratio does not seem to be major in our cohort.

Comparison of respiratory toxicity of TiO2 and Fe3O4 nanoparticles after intravenous instillation: an experimental study.

OBJECTIVE: Nanomaterials consist of particles smaller than 100 nm - nanoparticles (NPs). Their nano dimensions allow them to penetrate through various membranes and enter into the bloodstream and disseminate into different body organs. Massive expansion of nanotechnologies together with production of new nanoparticles which have not yet been in contact with living organisms may pose a potential health problem. It is therefore necessary to investigate the health impact of NPs after experimental exposure. Comparison of the effect of TiO2 and NPs Fe3O4 in Wistar rats at time intervals 1, 7, 14 and 28 days was performed by studying the cytotoxic effect in the isolated inflammatory cells from bronchoalveolar lavage (BAL). METHODS: Wistar rats were intravenously (i.v.) given a suspension of NPs TiO2 or Fe3O4 (coated by sodium oleate) via the tail vein. After time intervals of 1, 7, 14 and 28 days, we sacrificed the animals under anaesthesia, performed BAL and isolated the cells. The number of animals in the individual groups was 7-8. We examined the differential count of BAL cells (alveolar macrophages - AM, polymorphonuclear leukocytes - PMN, lymphocytes - Ly); viability and phagocytic activity of AM; the proportion of immature and polynuclear cells and enzymes - cathepsin D - CAT D, lactate dehydrogenase - LDH and acid phosphatase - ACP. RESULTS: We found that TiO2 NPs are relatively inert - without induction of inflammatory and cytotoxic response. Exposure to nanoparticles Fe3O4 induced - under the same experimental conditions - in comparison with the control and TiO2 a more extensive inflammatory and cytotoxic response, albeit only at 1, 7 and 14 days after injection. CONCLUSIONS: The results suggest that TiO2 and Fe3O4 nanoparticles used in our study were transferred from the bloodstream to the respiratory tract, but this effect was not observed at 28 days after i.v. injection, probably due to their removal from the respiratory tract.

Distinct pathways associated with chromosomal aberration frequency in a cohort exposed to genotoxic compounds compared to general population.

Non-specific structural chromosomal aberrations (CAs) observed in peripheral blood lymphocytes of healthy individuals can be either chromosome-type aberrations (CSAs) or chromatid-type aberrations (CTAs) depending on the stage of cell division they are induced in and mechanism of formation. It is important to study the genetic basis of chromosomal instability as it is a marker of genotoxic exposure and a predictor of cancer risk. For that purpose, we conducted two genome-wide association studies (GWASs) on healthy individuals in the presence and absence of apparent genotoxic exposure from the Czech Republic and Slovakia. The pre-GWAS cytogenetic analysis reported the frequencies of CSA, CTA and total CA (CAtot). We performed both linear and binary logistic regression analysis with an arbitrary cut-off point of 2% for CAtot and 1% for CSA and CTA. Using the statistical threshold of 1.0 × 10-5, we identified five loci with in silico predicted functionality in the reference group and four loci in the exposed group, with no overlap between the associated regions. A meta-analysis on the two GWASs identified further four loci with moderate associations in each of the studies. From the reference group mainly loci within genes related to DNA damage response/repair were identified. Other loci identified from both the reference and exposed groups were found to be involved in the segregation of chromosomes and chromatin modification. Some of the discovered regions in each group were implicated in tumourigenesis and autism.

Consumption of a dark roast coffee blend reduces DNA damage in humans: results from a 4-week randomised controlled study.

PURPOSE: To determine the DNA protective effects of a standard coffee beverage in comparison to water consumption. METHODS: The single-blind, randomised controlled study with parallel design included healthy women (n = 50) and men (n = 50) recruited from the general Central European population. The subjects were randomised in a coffee and a control group, with stratification for sex and body mass index. The study comprised two periods of 4 weeks: a preconditioning period, with daily consumption of at least 500 ml water but no coffee, nor tea, nor any other caffeine-containing product. During the subsequent intervention period the coffee group consumed 500 ml of freshly brewed dark roast coffee blend per day, the control group consumed water instead. On the last day of each period, blood was drawn and analysed by comet assay (single-cell gel electrophoresis) to assess the level of DNA damage (strand breakage). RESULTS: At the end of the intervention period the mean level of DNA strand breaks in the coffee group has decreased in comparison to the control group [difference in means 0.23% TI (tail intensity), p = 0.028]. The mean change from baseline (delta value) was - 23% in the coffee group (p = 0.0012). Effects of coffee intake were similar for men and women. During intervention, neither group showed any significant change in body weight or calorie intake. CONCLUSIONS: Our results indicate that regular consumption of a dark roast coffee blend has a beneficial protective effect on human DNA integrity in both, men and women.

Respiratory toxicity of TiO2 nanoparticles after intravenous instillation: an experimental study.

OBJECTIVE: Nanomaterials are materials consisting of particles having one or more dimensions smaller than 100 nm. Nanoparticles (NP) have different properties and effects in comparison with the same particle materials of larger size. They can penetrate through various membranes and get from the bloodstream to other organs in the body. Therefore, in our experiment we have dealt with the impact of nanoparticles TiO2 instilled intravenously (i.v.) (to a tail vein of an animal) on the selected parameters of bronchoalveolar lavage (BAL). The aim of our study was to determine whether TiO2 nanoparticles do pass through the vascular system to the respiratory tract, and if so, how they affect the selected inflammatory and cytotoxic parameters of bronchoalveolar lavage. METHODS: Wistar rats were intravenously given a suspension of TiO2 nanoparticles in saline solution. This suspension contained 10% volume of rat serum in dose: 1.0% from LD50 = 0.592 mg/kg of animal body weight. After the time intervals 1, 7, 14 and 28 days, the animals were sacrificed under anaesthesia; bronchoalveolar lavage was performed and the BAL cells were isolated. We have examined these markers: differential count of BAL cells - alveolar macrophages (AM), polymorphonuclear leukocytes (PMNL), lymphocytes (Ly); viability and phagocytic activity of AM; proportion of immature cells and cathepsin D enzyme levels. RESULTS: Regarding the respiratory toxicity of TiO2 nanoparticles we have found that TiO2 nanoparticles are relatively inert. BAL examined parameters (except the immature form of AM) were not significantly changed after 28 days of instillation compared to the control group. We found that the TiO2 nanoparticles used in our study were transferred from the bloodstream to the respiratory tract, but in a 28-day phase after i.v. instillation have been largely eliminated by the defence mechanism from the respiratory tract. CONCLUSIONS: We suggest low biopersistence and relatively rapid elimination of TiO2 nanoparticles from the lung under used experimental conditions.

Gender-associated differences in the prevalence of central obesity using waist circumference and waist-to-height ratio, and that of general obesity, in Slovak adults.

OBJECTIVES: Central obesity represents an increased risk to develop cardiovascular diseases. Guidelines of international societies suggest estimating central obesity by measuring waist circumference (WC). Robust statistical data in literature provide evidence on the superiority of waist-to-height ratio (WHtR) over WC and body mass index (BMI) for detecting cardiometabolic risk in both genders. Based on measurements of weight, height and waist circumference we compared the prevalence of central obesity using both the above mentioned criteria in the apparently healthy Slovak adults, and compared the prevalence of central obesity to that of general obesity (BMI). METHODS: Data collected from 5,184 individuals (45% males) aged ≥18 years in four cross-sectional studies carried out between the years 2009-2012 were subjected to secondary analysis. RESULTS: Waist circumference underestimated central obesity in males and overestimated in females: 37.3% of males and 41.8% of females presented central obesity according to WC, 54.2% males and 34.9% females according to WHtR. 17.3% of males centrally obese according to WC present WHtR < 0.5; while 7.8% of females centrally obese according to their WHtR do not display increased WC. The frequency of central obesity increased with age. According to BMI, the prevalence of overweight was 39% in males and 22% in females; that of obesity was 17% and 15%, respectively. CONCLUSION: The prevalence of central obesity estimated using WC vs. WHtR differs significantly in Slovak adults. WHtR is considered superior for detection of the risk of future development of cardiovascular afflictions. Thus, further studies addressing the gender-associated discordance of central obesity measures are required to determine whether our results are consistent across geographical regions and ethnic groups.

Association between metabolically healthy central obesity in women and levels of soluble receptor for advanced glycation end products, soluble vascular adhesion protein-1, and the activity of semicarbazide-sensitive amine oxidase.

AIM: To determine the levels of circulating soluble receptor for advanced glycation end products (sRAGE), as a biomarker of risk of metabolic syndrome and cardiovascular disease development in centrally obese (CO) women considered metabolically healthy (COH) in comparison with those metabolically unhealthy (COU). METHODS: 47 lean healthy, 17 COH (presenting waist-to-height ratio ≥0.5 but not elevated blood pressure, atherogenic lipid profile, and insulin resistance), and 50 COU (CO presenting ≥2 risk factors) women aged 40-45 years were included. Anthropometric characteristics, blood chemistry and hematology data, adipokines, markers of inflammation, sRAGE, soluble vascular adhesion protein-1 (sVAP-1), and the activity of semicarbazide sensitive amine oxidase (SSAO) were determined. RESULTS: Central obesity associated with low sRAGE levels (lean healthy: 1503±633 pg/mL; COH: 1103±339 pg/mL, P<0.05; COU: 1106±367 ng/mL, P<0.0.1), hyperleptinemia, and elevated markers of inflammation irrespective of the presence or absence of cardiometabolic risk factors. COU women presented high adiponectin levels. SVAP-1 concentrations and the activity of SSAO were similar in all 3 groups. CONCLUSION: COH women present abnormalities in non-standard markers of cardiometabolic risk (sRAGE, leptin, high sensitive C-reactive protein), supporting the view that there is no healthy pattern of obesity. The clinical impact of our findings for future prognosis of metabolically healthy obese subjects remains to be elucidated in longitudinal studies.

Association of Inflammatory and Oxidative Status Markers with Metabolic Syndrome and Its Components in 40-to-45-Year-Old Females: A Cross-Sectional Study.

Oxidative stress and sterile inflammation play roles in the induction and maintenance of metabolic syndrome (MetS). This study cohort included 170 females aged 40 to 45 years who were categorized according to the presentation of MetS components (e.g., central obesity, insulin resistance, atherogenic dyslipidemia, and elevated systolic blood pressure) as controls not presenting a single component (n = 43), those with pre-MetS displaying one to two components (n = 70), and females manifesting MetS, e.g., ≥3 components (n = 53). We analyzed the trends of seventeen oxidative and nine inflammatory status markers across three clinical categories. A multivariate regression of selected oxidative status and inflammatory markers on the components of MetS was performed. Markers of oxidative damage (malondialdehyde and advanced-glycation-end-products-associated fluorescence of plasma) were similar across the groups. Healthy controls displayed lower uricemia and higher bilirubinemia than females with MetS; and lower leukocyte counts, concentrations of C-reactive protein, interleukine-6, and higher levels of carotenoids/lipids and soluble receptors for advanced glycation end-products than those with pre-MetS and MetS. In multivariate regression models, levels of C-reactive protein, uric acid, and interleukine-6 were consistently associated with MetS components, although the impacts of single markers differed. Our data suggest that a proinflammatory imbalance precedes the manifestation of MetS, while an imbalance of oxidative status accompanies overt MetS. Further studies are needed to elucidate whether determining markers beyond traditional ones could help improve the prognosis of subjects at an early stage of MetS.

Genotoxic effects of occupational exposure to glass fibres - A human biomonitoring study.

As part of a large human biomonitoring study, we conducted occupational monitoring in a glass fibre factory in Slovakia. Shopfloor workers (n = 80), with a matched group of administrators in the same factory (n = 36), were monitored for exposure to glass fibres and to polycyclic aromatic hydrocarbons (PAHs). The impact of occupational exposure on chromosomal aberrations, DNA damage and DNA repair, immunomodulatory markers, and the role of nutritional and lifestyle factors, as well as the effect of polymorphisms in metabolic and DNA repair genes on genetic stability, were investigated. The (enzyme-modified) comet assay was employed to measure DNA strand breaks (SBs) and apurinic sites, oxidised and alkylated bases. Antioxidant status was estimated by resistance to H2O2-induced DNA damage. Base excision repair capacity was measured with an in vitro assay (based on the comet assay). Exposure of workers to fibres was low, but still was associated with higher levels of SBs, and SBs plus oxidised bases, and higher sensitivity to H2O2. Multivariate analysis showed that exposure increased the risk of high levels of SBs by 20%. DNA damage was influenced by antioxidant enzymes catalase and glutathione S-transferase (measured in blood). DNA repair capacity was inversely correlated with DNA damage and positively with antioxidant status. An inverse correlation was found between DNA base oxidation and the percentage of eosinophils (involved in the inflammatory response) in peripheral blood of both exposed and reference groups. Genotypes of XRCC1 variants rs3213245 and rs25487 significantly decreased the risk of high levels of base oxidation, to 0.50 (p = 0.001) and 0.59 (p = 0.001), respectively. Increases in DNA damage owing to glass fibre exposure were significant but modest, and no increases were seen in chromosome aberrations or micronuclei. However, it is of concern that even low levels of exposure to these fibres can cause significant genetic damage.

Genotoxicity testing of PLGA-PEO nanoparticles in TK6 cells by the comet assay and the cytokinesis-block micronucleus assay.

The in vitro genotoxicity of PLGA-PEO (poly-lactic-co-glycolic acid-polyethylene oxide copolymer) nanoparticles was assessed in TK6 cells using the comet assay as well as cytokinesis-block micronucleus (CBMN) assay. The cells were exposed to 0.12-75µg/cm² of PLGA-PEO nanoparticles during 2 and 24h for analysis in the comet assay, and to 3-75µg/cm² of these nanoparticles during 4, 24, 48 and 72h, respectively, for analysis in the CBMN assay. Two different protocols for treatment with cytochalasin B were used. We found that PLGA-PEO was neither cytotoxic (measured by relative cell growth activity and cytokinesis-block proliferation index (CBPI)), nor did it induce DNA strand-breaks (detected by the comet assay) or oxidative DNA lesions (measured by the comet assay modified with lesion-specific enzyme formamidopyrimidine-DNA-glycosylase). There were no statistically significant differences in the frequencies of micronucleated binucleated cells (MNBNCs) between untreated and treated cells in either of the conditions used. This suggests that PLGA-PEO did not have potential genotoxicity. However, using two experimental protocols of the micronucleus assay, PLGA-PEO nanoparticles showed a weak but significant increase in the level of MN in mononucleated cells, in cells treated for 48h with PLGA-PEO nanoparticles when cytochalasin B was added for the last 24h (1st protocol), and in cells treated for 24h with PLGA-PEO nanoparticles followed by washing of NPs and addition of cytochalasin B for another 24h (2nd protocol). It remains unclear whether the increase of MNMNC after treatment with PLGA-PEO nanoparticles is the effect of a possible, weak aneugenic potential or early effect of these particles, or due to another reason. These results suggest that aneugenicity in addition to clastogenicity may be considered as an important biomarker when assessing the genotoxic potential of polymeric nanoparticles.

Are glutathione S transferases involved in DNA damage signalling? Interactions with DNA damage and repair revealed from molecular epidemiology studies.

Glutathione S-transferases (GSTs) are members of a multigene family of isoenzymes that are important in the control of oxidative stress and in phase II metabolism. Acting non-enzymically, GSTs can modulate signalling pathways of cell proliferation, cell differentiation and apoptosis. Using a molecular epidemiology approach, we have investigated a potential involvement of GSTs in DNA damage processing, specifically the modulation of DNA repair in a group of 388 healthy adult volunteers; 239 with at least 5 years of occupational exposure to asbestos, stone wool or glass fibre, and 149 reference subjects. We measured DNA damage in lymphocytes using the comet assay (alkaline single cell gel electrophoresis): strand breaks (SBs) and alkali-labile sites, oxidised pyrimidines with endonuclease III, and oxidised purines with formamidopyrimidine DNA glycosylase. We also measured GST activity in erythrocytes, and the capacity for base excision repair (BER) in a lymphocyte extract. Polymorphisms in genes encoding three GST isoenzymes were determined, namely deletion of GSTM1 and GSTT1 and single nucleotide polymorphism Ile105Val in GSTP1. Consumption of vegetables and wine correlated negatively with DNA damage and modulated BER. GST activity correlated with oxidised bases and with BER capacity, and differed depending on polymorphisms in GSTP1, GSTT1 and GSTM1. A significantly lower BER rate was associated with the homozygous GSTT1 deletion in all asbestos site subjects and in the corresponding reference group. Multifactorial analysis revealed effects of sex and exposure in GSTP1 Ile/Val heterozygotes but not in Ile/Ile homozygotes. These variants affected also SBs levels, mainly by interactions of GSTP1 genotype with exposure, with sex, and with smoking habit; and by an interaction between sex and smoking. Our results show that GST polymorphisms and GST activity can apparently influence DNA stability and repair of oxidised bases, suggesting a potential new role for these proteins in DNA damage processing via DNA damage signalling.

Associations between environmental factors and incidence of cutaneous melanoma. Review.

BACKGROUND: Cutaneous melanoma is one of the most serious skin cancers. It is caused by neural crest-derived melanocytes - pigmented cells normally present in the epidermis and, sometimes, in the dermis. METHODS: We performed a review of current knowledge on the risk factors of cutaneous melanoma. Relevant studies were identified using the PubMed, Science Direct, Medline, Scopus, Scholar Google and ISI Web of Knowledge databases. RESULTS: Melanoma incurs a considerable public health burden owing to the worldwide dramatic rise in incidence since the mid-1960s. Ultraviolet radiation exposure is the predominant environmental risk factor. The role of geographical (latitude) and individual factors such as skin type, life style, vitamin D levels and antioxidant protection, sunburn, and exposure to other environmental factors possibly contributing to melanoma risk (such as cosmetics including sunscreen, photosensitising drugs, and exogenous hormones) are reviewed in this article. Recently, both rare high risk susceptibility genes and common polymorphic genes contributing to melanoma risk have been identified. CONCLUSIONS: Cutaneous melanoma is a complex cancer with heterogeneous aetiology that continues to increase in incidence. Introduction of new biomarkers may help to elucidate the mechanism of pathogenesis and individual susceptibility to the disease, and make both prevention and treatment more effective.

Bladder cancer, a review of the environmental risk factors.

BACKGROUND: Many epidemiological studies and reviews have been performed to identify the causes of bladder cancer. The aim of this review is to investigate the links between various environmental risk factors and cancer of the bladder. METHODS: A systematic literature search was performed using PubMed, Science Direct, Scopus, Scholar Google and Russian Google databases to identify reviews and epidemiological studies on bladder cancer risk factors associated with the environment published between 1998 and 2010. Only literature discussing human studies was considered. RESULTS: Smoking, mainly cigarette smoking, is a well known risk factor for various diseases, including bladder cancer. Another factor strongly associated with bladder cancer is exposure to arsenic in drinking water at concentrations higher than 300 µg/l. The most notable risk factor for development of bladder cancer is occupational exposure to aromatic amines (2-naphthylamine, 4-aminobiphenyl and benzidine) and 4,4'-methylenebis(2-chloroaniline), which can be found in the products of the chemical, dye and rubber industries as well as in hair dyes, paints, fungicides, cigarette smoke, plastics, metals and motor vehicle exhaust. There are also data suggesting an effect from of other types of smoking besides cigarettes (cigar, pipe, Egyptian waterpipe, smokeless tobacco and environmental tobacco smoking), and other sources of arsenic exposure such as air, food, occupational hazards, and tobacco. Other studies show that hairdressers and barbers with occupational exposure to hair dyes experience enhanced risk of bladder cancer. For example, a study related to personal use of hair dyes demonstrates an elevated bladder cancer risk for people who used permanent hair dyes at least once a month, for one year or longer. CONCLUSION: Smoking, in particular from cigarettes, exposure to arsenic in drinking water, and occupational exposure to aromatic amines and 4,4'-methylenebis(2-chloroaniline) are well known risk factors for various diseases including bladder cancer. Although the number of chemicals related to occupational exposure is still growing, it is worth noting that it may take several years or decades between exposure and the subsequent cancer.

Polymorphic Variants of Interleukin-13 R130Q and Interleukin-4 T589C in Children with and without Cow's Milk Allergy.

Cow's milk allergy (CMA) is one of the most frequent types of food allergy. The aim of the study was to investigate whether IL-13 R130Q and IL-4 T589C polymorphisms are associated with the risk of CMA in young Ukrainian children. A total of 120 children (age range: 1−3 years) participated in the study and were divided into two groups: CMA (n = 60) and healthy controls (CNT, n = 60). The CMA group had two subgroups: specific oral tolerance induction (SOTI, n = 30) and milk elimination diet (MED, n = 30). The CNT group had two subgroups: positive family history of allergy (+FHA, n = 24) and negative family history of allergy (−FHA, n = 36). In the CMA group, molecular genetic testing of CC, CT, and TT genotypes of single nucleotide IL-4 T589C gene polymorphisms showed significantly higher rates of the CC genotype compared to healthy controls (92.2% vs. 58.8%; p < 0.01). In the CMA group, molecular genetic testing of GG, GA, and AA genotypes of single nucleotide IL-13 R130Q gene polymorphisms showed significantly higher rates of GA and AA polymorphic locus genotypes compared to healthy controls (43.5% vs. 22.4%, p < 0.05 and 8.7% vs. 0%, p < 0.05, respectively). In future studies, the genotypic and allelic distribution of these polymorphic variants will be determined in children with CMA and healthy children.

A pooled analysis of molecular epidemiological studies on modulation of DNA repair by host factors.

Levels of DNA damage represent the dynamics between damage formation and removal. Therefore, to better interpret human biomonitoring studies with DNA damage endpoints, an individual's ability to recognize and properly remove DNA damage should be characterized. Relatively few studies have included DNA repair as a biomarker and therefore, assembling and analyzing a pooled database of studies with data on base excision repair (BER) was one of the goals of hCOMET (EU-COST CA15132). A group of approximately 1911 individuals, was gathered from 8 laboratories which run population studies with the comet-based in vitro DNA repair assay. BER incision activity data were normalized and subsequently correlated with various host factors. BER was found to be significantly higher in women. Although it is generally accepted that age is inversely related to DNA repair, no overall effect of age was found, but sex differences were most pronounced in the oldest quartile (>61 years). No effect of smoking or occupational exposures was found. A body mass index (BMI) above 25 kg/m2 was related to higher levels of BER. However, when BMI exceeded 35 kg/m2, repair incision activity was significantly lower. Finally, higher BER incision activity was related to lower levels of DNA damage detected by the comet assay in combination with formamidopyrimidine DNA glycosylase (Fpg), which is in line with the fact that oxidatively damaged DNA is repaired by BER. These data indicate that BER plays a role in modulating the steady-state level of DNA damage that is detected in molecular epidemiological studies and should therefore be considered as a parallel endpoint in future studies.

Vitamin C levels in blood are influenced by polymorphisms in glutathione S-transferases.

PURPOSE: Glutathione S-transferases (GSTs) are intimately involved in combating oxidative stress and in detoxifying xenobiotics. Our objective was to examine possible interactions between polymorphisms in GST genes and plasma vitamin C, tocopherols and carotenoids in 149 reference subjects and 239 subjects occupationally exposed to mineral fibres (asbestos, rock wool, glass fibre), agents that induce oxidative stress. METHODS: Deletion of GSTM1 and GSTT1, and substitution 105Ile/Val in GSTP1 genes were determined by PCR, antioxidants in plasma were measured by HPLC. RESULTS: Tocopherols and carotenoids were affected by age, sex, smoking, occupational exposure to fibres, but not by GST polymorphisms. Vitamin C level was influenced by sex, smoking and occupational exposure. Subjects with deletion of GST had lower vitamin C levels compared with subjects carrying the functional gene variant. Vitamin C levels varied according to GSTM1 polymorphism in the whole group (p < 0.05), in all reference subjects (p < 0.05), in the asbestos factory reference group (p < 0.05), and according to GSTT1 polymorphism in reference group of the rock wool plant (p < 0.05). Vitamin C levels were approximately 20% lower in subjects with both functionally deficient genes in the whole group (p < 0.01) and in all non-exposed subjects (p < 0.05). CONCLUSIONS: The correspondence of lower vitamin C levels with non-functional GST isoenzymes may indicate a causal connection between two antioxidant defence pathways, also the underlying mechanism is not yet clear. It seems that supplementation by natural antioxidants is particularly important for subjects with unfavourable genetic makeup and in those exposed to oxidative stress.

DNA repair gene polymorphisms and chromosomal aberrations in healthy, nonsmoking population.

Nonspecific structural chromosomal aberrations (CAs) can be found at around 1% of circulating lymphocytes from healthy individuals but the frequency may be higher after exposure to carcinogenic chemicals or radiation. The frequency of CAs has been measured in occupational monitoring and an increased frequency of CAs has also been associated with cancer risk. Alterations in DNA damage repair and telomere maintenance are thought to contribute to the formation of CAs, which include chromosome type of aberrations and chromatid type of aberrations. In the present study, we used the result of our published genome-wide association studies to extract data on 153 DNA repair genes from 866 nonsmoking persons who had no known occupational exposure to genotoxic substances. Considering an arbitrary cut-off level of P< 5 × 10-3, single nucleotide polymorphisms (SNPs) tagging 22 DNA repair genes were significantly associated with CAs and they remained significant at P < 0.05 when adjustment for multiple comparisons was done by the Binomial Sequential Goodness of Fit test. Nucleotide excision repair pathway genes showed most associations with 6 genes. Among the associated genes were several in which mutations manifest CA phenotype, including Fanconi anemia, WRN, BLM and genes that are important in maintaining genome stability, as well as PARP2 and mismatch repair genes. RPA2 and RPA3 may participate in telomere maintenance through the synthesis of the C strand of telomeres. Errors in NHEJ1 function may lead to translocations. The present results show associations with some genes with known CA phenotype and suggest other pathways with mechanistic rationale for the formation of CAs in healthy nonsmoking population.

DNA Repair Gene Polymorphisms and Chromosomal Aberrations in Exposed Populations.

DNA damage and unrepaired or insufficiently repaired DNA double-strand breaks as well as telomere shortening contribute to the formation of structural chromosomal aberrations (CAs). Non-specific CAs have been used in the monitoring of individuals exposed to potential carcinogenic chemicals and radiation. The frequency of CAs in peripheral blood lymphocytes (PBLs) has been associated with cancer risk and the association has also been found in incident cancer patients. CAs include chromosome-type aberrations (CSAs) and chromatid-type aberrations (CTAs) and their sum CAtot. In the present study, we used data from our published genome-wide association studies (GWASs) and extracted the results for 153 DNA repair genes for 607 persons who had occupational exposure to diverse harmful substances/radiation and/or personal exposure to tobacco smoking. The analyses were conducted using linear and logistic regression models to study the association of DNA repair gene polymorphisms with CAs. Considering an arbitrary cutoff level of 5 × 10-3, 14 loci passed the threshold, and included 7 repair pathways for CTA, 4 for CSA, and 3 for CAtot; 10 SNPs were eQTLs influencing the expression of the target repair gene. For the base excision repair pathway, the implicated genes PARP1 and PARP2 encode poly(ADP-ribosyl) transferases with multiple regulatory functions. PARP1 and PARP2 have an important role in maintaining genome stability through diverse mechanisms. Other candidate genes with known roles for CSAs included GTF2H (general transcription factor IIH subunits 4 and 5), Fanconi anemia pathway genes, and PMS2, a mismatch repair gene. The present results suggest pathways with mechanistic rationale for the formation of CAs and emphasize the need to further develop techniques for measuring individual sensitivity to genotoxic exposure.

DNA damage in circulating leukocytes measured with the comet assay may predict the risk of death.

The comet assay or single cell gel electrophoresis, is the most common method used to measure strand breaks and a variety of other DNA lesions in human populations. To estimate the risk of overall mortality, mortality by cause, and cancer incidence associated to DNA damage, a cohort of 2,403 healthy individuals (25,978 person-years) screened in 16 laboratories using the comet assay between 1996 and 2016 was followed-up. Kaplan-Meier analysis indicated a worse overall survival in the medium and high tertile of DNA damage (p < 0.001). The effect of DNA damage on survival was modelled according to Cox proportional hazard regression model. The adjusted hazard ratio (HR) was 1.42 (1.06-1.90) for overall mortality, and 1.94 (1.04-3.59) for diseases of the circulatory system in subjects with the highest tertile of DNA damage. The findings of this study provide epidemiological evidence encouraging the implementation of the comet assay in preventive strategies for non-communicable diseases.