Soluble metals as well as the insoluble particle fraction are involved in cellular DNA damage induced by particulate matter

Soluble metals as well as the insoluble particle fraction are involved in cellular DNA damage induced by particulate matter. Knaapen AM, Shi T, Borm PJ, Schins RP. Particle Research Core, Research Institute for Environmental Health (IUF), Dusseldorf, Germany. Exposure to ambient particulate matter has been reported to be associated with increased rates of lung cancer. Previously we showed that total suspended particulate matter (PM) induces oxidative DNA damage in epithelial lung cells. The aim of the present study was to further investigate the mechanism of PM-induced DNA damage, in which soluble iron-mediated hydroxyl radical (.OH) formation is thought to play a crucial role. Using electron spin resonance (ESR) we showed that PM suspensions as well as their particle-free, water-soluble fractions can generate .OH in the presence of hydrogen peroxide (H2O2), an effect which was abrogated by both deferoxamine and catalase. In addition, PM was also found to induce the .OH-specific DNA lesion 8-hydroxydeoxyguanosine (8-OHdG) in the presence of H2O2 as assessed by dot-blot analysis of calf thymus DNA using an 8-OHdG antibody. In human alveolar epithelial cells (A549), both PM suspensions and the particle-free soluble fraction elicited formation of DNA strand breaks (comet-assay). Unlike the acellular DNA assay, in epithelial cells the DNA-damaging capacity of the particle suspensions appeared to be stronger than that of their corresponding particle-free filtrates. In conclusion, our findings demonstrate that the water-soluble fraction of PM elicits DNA damage via transition metal-dependent .OH formation, implicating an important role of H2O2. Moreover, our data indicate that direct 'particle' effects contribute to the genotoxic hazard of ambient particulate matter in lung target cell

Mechanisms of neutrophil-induced DNA damage in respiratory tract epithelial cells

Mechanisms of neutrophil-induced DNA damage in respiratory tract epithelial cells. Knaapen AM, Schins RP, Polat D, Becker A, Borm PJ. Particle Research Core, Institute for Environmental Health (IUF), Dusseldorf, Germany. Reactive oxygen species (ROS) released by neutrophils have been suggested to play an important role in cancer development. Since the mechanisms underlying this effect in the respiratory tract are still unclear, we evaluated DNA damage induced by neutrophils in respiratory tract epithelial cells in vitro and in vivo. For in vitro studies, rat lung epithelial cells (RLE) were co-incubated with activated neutrophils, neutrophil-conditioned medium, or hydrogen peroxide. For in vivo studies, we considered the human nose as a target organ, comparing neutrophilic inflammation in the nasal lavage fluid with the oxidative DNA lesion 8-hydroxydeoxyguanosine (8-OHdG) in epithelial cells obtained by nasal brush. Our in vitro data show that human neutrophils are able to induce both 8-OHdG and strand breaks in DNA from RLE cells. Our data also suggest that DNA damage induced by neutrophils is inhibited when neutrophil-derived H2O2 is consumed by myeloperoxidase. In contrast, in the nose no association between neutrophil numbers and 8-OHdG was found. Therefore, it remains unclear whether neutrophils pose a direct genotoxic risk for the respiratory tract epithelium during inflammation, andmore in vivo studies are needed to elucidate the possible association between neutrophils and genotoxicity in the lung

Polymorphisms in the promoter of the tumor necrosis factor: a gene in coal miners.

Department of Health Risk Analysis and Toxicology, Maastricht University, The Netherlands. Tumor necrosis factor-alpha (TNF) is recognized as a central mediator of mineral dust-induced lung fibrosis, and genetic polymorphisms of the TNF promoter have been reported to influence levels of TNF production. To assess whether polymorphisms within the TNF promoter gene are associated with susceptibility to coal workers' pneumoconiosis (CWP), the DNA of 78 coal miners was typed for G-to-A transitions at positions -238 and -308. Our results show that frequency of A-308 genotype (T2) is significantly overpresented in coal miners with CWP (50%), as compared with miners without CWP (25%) and controls (29%). After correction for cumulative dust exposure and smoking, the A-308 transition genotype is still associated with the presence of CWP (OR = 3.0, 95% CI = 1.0-9.0). Both A-238 and A-308 transition genotypes were related to TNF release from endotoxin-stimulated blood monocytes; only the A-238 transition and not the A-308 transition was associated to coal dust-induced TNF release. In summary, this study shows that the A-308 transition is related to CWP, but this relation is not paralleled by a different TNF release in this genotype. A larger number of patients coupled to frequent TNF release are required to evaluate genotype screening to estimate individual health risks for effects of coal mine dust exposure. Publication Types: Clinical Trial Controlled Clinical Tria

Ambient particulate matter induces oxidative DNA damage in lung epithelial cells

Although epidemiological studies have established a correlation between pm10 levels and acute cardiovascular and respiratory complications, hardly any data is available on possible chronic effects such as cancer. The purpose of this study was to investigate the production of free radicals by ambient particulate matter (tsp) and to link these data to oxidative dna damage in lung epithelial cells. In line with previous findings on pm10, supercoiled plasmid dna was depleted by tsp as well as tsp supernatant (p <.001), and this effect was reduced in the presence of mannitol (5 mm). Using electron spin resonance (esr) and the spin trap dimethyl-1-pyrroline n-oxide (dmpo) we were able to show that hydroxyl radicals (oh) are formed from both tsp and tsp supernatant. The dmpo-oh signal was completely abrogated when tsp was preincubated with deferoxamine (5 mm), showing the importance of iron and other soluble metals in this process. Atomic absorption spectroscopy (aas) analysis of the tsp supernatant showed the presence of soluble fe, v,and ni (respectively 253.0, 14.7, and 76.0 mug/g insoluble tsp). To investigate the biological significance of oh formation by tsp,8-hydroxydeoxyguanosine (8-oxodg) was measured in a rat type ii cell line by immunocytochemistry. The formation of this hydroxyl-radical-specific dna adduct was increased twofold (p <.01) after incubation with tsp supernatants, and this effect was inhibited by deferoxamine (p <.01). In summary, our results provide direct evidence that ambient particulate matter generates hydroxyl radicals in acellular systems. Furthermore, we showed that these particulates induce the hydroxyl-radical-specific dna lesion 8-oxodg in lung target cells via an iron-mediated mechanism

Oxidant-induced DNA damage by quartz in alveolar epithelial cells

Oxidant-induced DNA damage by quartz in alveolar epithelial cells. Schins RP, Knaapen AM, Cakmak GD, Shi T, Weishaupt C, Borm PJ. Institut fur Umweltmedizinische Forschung, Heinrich-Heine Universitat Dusseldorf, Auf'm Hennekamp 50, 40225 Dusseldorf, Germany. [email protected] Respirable quartz has recently been classified as a human carcinogen. Although, studies with quartz using naked DNA as a target suggest that formation of oxyradicals by particles may play a role in the DNA-damaging properties of quartz, it is not known whether this pathway is important for DNA damage in the target cells for quartz carcinogenesis, i.e. alveolar epithelial cells. Therefore, we determined in vitro DNA damage by DQ12 quartz particles in rat and human and alveolar epithelial cells (RLE, A549) using the single cell gel electrophoresis/comet assay. The radical generation capacity of quartz was analysed by electron spin resonance (ESR) and by immunocytochemical analysis of the hydroxyl radical-specific DNA lesion 8-hydroxydeoxyguanosine (8-OHdG) in the epithelial cells. Quartz particles as well as the positive control hydrogen peroxide, caused a dose-dependent increase in DNA strand breaks in both cell lines. DNA damage by quartz was significantly reduced in the presence of the hydroxyl-radical scavengers mannitol or DMSO. The involvement of hydroxyl radicals was further established by ESR measurements and was also demonstrated by the ability of the quartz to induce formation of 8-OHdG. In conclusion, our data show that quartz elicits DNA damage in rat and human alveolar epithelial cells and indicate that these effects are driven by hydroxyl radical-generating properties of the particle