RESUMEN
Tobacco smoking continues to be the largest preventable cause of premature morbidity and mortality throughout the world, including chronic respiratory diseases such as asthma and chronic obstructive pulmonary disease. Although most smokers are highly motivated to quit and many smoking cessation therapies are available, cessation rates remain very low. Recent research strongly suggests that variation in genetic background is an important determinant of smoking behaviour and addiction. Since these genetic variants might also influence the response to smoking cessation pharmacotherapies, it is likely that assessment of genetic background could be a promising tool to guide selection of the most effective cessation treatment for an individual smoker. Recently, it has been shown that genetic variants in the dopaminergic system, opioid receptors, the bupropion-metabolising enzyme CYP2B6 and the nicotine-metabolising enzyme CYP2A6 may play an important role in predicting smoking cessation responses to nicotine replacement therapy and bupropion treatment. Despite the progress that has been made, several challenges will still have to be overcome before genetically tailored smoking cessation therapy can be implemented in standard clinical practice.
Asunto(s)
Enfermedades Pulmonares Obstructivas/genética , Enfermedades Pulmonares Obstructivas/terapia , Cese del Hábito de Fumar , Hidrocarburo de Aril Hidroxilasas/genética , Citocromo P-450 CYP2A6 , Citocromo P-450 CYP2B6 , Dopamina/metabolismo , Femenino , Predisposición Genética a la Enfermedad , Variación Genética , Humanos , Masculino , Modelos Genéticos , Oxidorreductasas N-Desmetilantes/genética , Serotonina/metabolismo , Fumar/efectos adversos , Resultado del TratamientoRESUMEN
AIMS: To determine the induction of 8-hydroxy-2'-deoxyguanosine (8-OHdG) by fine (<2.5 microm) and coarse (10-2.5 microm) particulate matter (PM) sampled over time at one sampling location, and to relate the observed effects to the hydroxyl radical (*OH) generating activities and transition metal content of these samples, and to meteorological parameters. METHODS: Weekly samples of coarse and fine PM were analysed for H(2)O(2) dependent *OH formation using electron spin resonance (ESR) and formation of 8-OHdG in calf thymus DNA using an immuno-dotblot assay. Immunocytochemistry was used to determine 8-OHdG formation in A549 human epithelial lung cells. To determine temporal effects, samples from six weeks in summer and six weeks in autumn/winter were compared using ESR and the dotblot assay. Concentrations of leachable V, Cr, Fe, Ni, and Cu were determined by inductively coupled plasma mass spectrometry. RESULTS: Both PM fractions elicited *OH generation as well as 8-OHdG formation in calf thymus DNA and in A549 cells. 8-OHdG formation in the naked DNA was significantly related to *OH generation, but not to metal concentrations except for copper. A significantly higher *OH generation was observed for coarse PM, but not fine PM collected during the autumn/winter season; this was not due to differences in sampled mass or metal content. Specific weather conditions under which increased *OH formation in the coarse mode was observed suggest that other, as yet unknown, anthropogenic components might affect the radical generating capacity of PM. CONCLUSIONS: Both coarse and fine PM are able to generate *OH, and induce formation of 8-OHdG. When considered at equal mass, *OH formation shows considerable variability with regard to the fraction of PM, as well as the sampling season. The toxicological implications of this heterogeneity in *OH formation by PM, as can be easily determined by ESR, need further investigation.
Asunto(s)
Desoxiadenosinas/metabolismo , Radical Hidroxilo/química , Animales , Bovinos , ADN/metabolismo , Relación Dosis-Respuesta Inmunológica , Espectroscopía de Resonancia por Spin del Electrón , Radical Hidroxilo/metabolismo , Conceptos Meteorológicos , Tamaño de la Partícula , Timo/metabolismoRESUMEN
Epidemiological studies have shown an association between ambient levels of particulate matter (PM) and increased mortality from cardiovascular diseases. However, the underlying mechanisms are still not clear. We hypothesised that PM, when translocated after inhalation, could affect vascular smooth muscle function. Therefore, total suspended particulate matter (TSP) was sampled and investigated for its ability to affect aortic muscle contraction. Both TSP and TSP supernatant (TSP-sup) induced a concentration-dependent relaxation of phenylephrine (PE)-precontracted aortic rings. Relaxation induced by 100 microg/ml TSP was 51.5 +/- 3.1% of total contraction. At 60 and 100 microg/ml, relaxation induced by TSP was significantly higher compared to TSP-sup. Ultrafine TiO2, used as a model to investigate the role of ultrafine particles, did not show an effect. Soluble iron, present in TSP suspensions, seems not to be involved, as chelating with deferoxamine did not affect TSP-induced relaxation. However, TSP effects were inhibited by Trolox, suggesting a role of oxidants. Nudation of aortic rings showed that effects of TSP were only partly endothelium-dependent, while preincubation with L-NAME increased TSP-induced relaxation. From these data, we conclude that both the particle core and soluble components of TSP can affect the smooth muscle function, leading to changes in the vascular contractile response.
Asunto(s)
Contaminantes Atmosféricos/efectos adversos , Aorta/fisiología , Contracción Muscular/efectos de los fármacos , Músculo Liso/efectos de los fármacos , Administración por Inhalación , Animales , Antioxidantes/administración & dosificación , Antioxidantes/farmacología , Aorta/efectos de los fármacos , Cromanos/administración & dosificación , Cromanos/farmacología , Técnicas In Vitro , Masculino , Músculo Liso/fisiología , Oxidantes , Tamaño de la Partícula , Ratas , Ratas Endogámicas Lew , SolubilidadRESUMEN
Although epidemiological studies have established a correlation between PMIO 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 PMIO, supercoiled plasmid DNA was depleted by JSP as well as JSP 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 hydroxy/radicals ('OH) are formed from both JSP and JSP 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 µ/g insoluble TSP). To investigate the biological significance of OH formation by TSP, 8-hydroxydeoxyguanosine (8-oxodC) 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 hydroxyI radicals in acellular systems. Furthermore, we showed that these particulates induce the hydroxyl-radical-specific DNA lesion 8-oxodC in lung target cells via an iron-mediated mechanism.
RESUMEN
Inflammation has been recognized as a contributing factor in the pathogenesis of some cancers. In the lung, inflammation is characterized by an influx of polymorphonuclear leukocytes (PMN) that release a variety of reactive oxygen species (ROS). The aim of the present study was to investigate the direct effect of PMN on oxidative DNA damage in lung target cells. Therefore, rat alveolar epithelial cells (RLE) were coincubated with PMN or hydrogen peroxide. Known to be correlated with the incidence of cancer, 7-hydro-8-oxo-2'deoxyguanosine (8-oxodG) was used as an effect marker for oxidative damage. Viability of the RLE, when coincubated with PMN, decreased to 43%, dependent on the ratio between PMN and RLE. After washing off PMN, 8-oxodG levels were significantly increased in RLE, but the highest levels were observed in the washed off PMN fraction. In addition, to avoid washing off procedures, immunohistochemical analysis was used to measure the 8-oxodG levels specifically in the RLE and similar results were obtained. In addition, inhibitor experiments showed that antioxidants ameliorated oxidative DNA damage. Our data provide evidence that ROS released by PMN as well as H2O2, cause oxidative DNA damage in epithelial cells.
Asunto(s)
Daño del ADN , Neutrófilos/fisiología , Alveolos Pulmonares/patología , Animales , Células Cultivadas , Células Epiteliales/efectos de los fármacos , Células Epiteliales/patología , Humanos , Peróxido de Hidrógeno/farmacología , Masculino , Oxidación-Reducción , Alveolos Pulmonares/citología , RatasRESUMEN
Inflammatory cells and their reactive oxygen metabolites can cause mutagenic effects in lung cells. The purpose of this study was to investigate the ability of activated neutrophils to modulate DNA binding of benzo[a]pyrene (B[a]P), a known carcinogen, in lung target cells. Equivalent numbers of rat lung epithelial cells (RLE-6TN cell line) and freshly isolated human blood neutrophils (PMN) were coincubated in vitro for 2 hr after addition of benzo[a]pyrene (0.5 microM) or two of its trans-diol metabolites, with or without stimulation with phorbol myristate acetate (PMA). DNA adducts of B[a]P-metabolites were determined in target cells using 32P-postlabeling; oxidative DNA damage (7-hydro-8-oxo-2'-deoxyguanosine [8-oxodG]) was evaluated by high performance liquid chromatography with electrochemical detection. Increased DNA adducts were observed in lung cells coincubated with polymorphonuclear leukocytes (PMN). Activation of PMN with PMA, or addition of more activated PMN in relation to the number of lung cells, further increased the number of adducts, the latter in a dose-response manner. Incubation with B[a]P-4,5-diol did not result in any adduct formation, while B[a]P-7,8-diol led to a significant number of adducts. Moreover, PMA-activated PMN strongly enhanced adduct formation by B[a]P-7,8-diol, but not 8-oxodG, in lung cells. The addition of antioxidants to the coincubations significantly reduced the number of adducts. Results suggest that an inflammatory response in the lung may increase the biologically effective dose of polycyclic aromatic hydrocarbons (PAHs), and may be relevant to data interpretation and risk assessment of PAH-containing particulates.