The SEURAT-1 Approach towards Animal Free Human Safety Assessment

SEURAT-1 is a European public-private research consortium that is working towards animal-free testing of chemical compounds and the highest level of consumer protection. A research strategy was formulated based on the guiding principle to adopt a toxicological mode-of-action framework to describe how any substance may adversely affect human health. The proof of the initiative will be in demonstrating the applicability of the concepts on which SEURAT-1 is built on three levels: (i) Theoretical prototypes for adverse outcome pathways are formulated based on knowledge already available in the scientific literature on investigating the toxicological modes-of-action leading to adverse outcomes (addressing mainly liver toxicity); (ii) adverse outcome pathway descriptions are used as a guide for the formulation of case studies to further elucidate the theoretical model and to develop integrated testing strategies for the prediction of certain toxicological effects (i.e., those related to the adverse outcome pathway descriptions); (iii) further case studies target the application of knowledge gained within SEURAT-1 in the context of safety assessment. The ultimate goal would be to perform ab initio predictions based on a complete understanding of toxicological mechanisms. In the near-term, it is more realistic that data from innovative testing methods will support read-across arguments. Both scenarios are addressed with case studies for improved safety assessment. A conceptual framework for a rational integrated assessment strategy emerged from designing the case studies and is discussed in the context of international developments focusing on alternative approaches for evaluating chemicals using the new 21st century tools for toxicity testing

Modulation of glutaredoxin in the lung and sputum of cigarette smokers and chronic obstructive pulmonary disease

BACKGROUND: One typical feature in chronic obstructive pulmonary disease (COPD) is the disturbance of the oxidant/antioxidant balance. Glutaredoxins (Grx) are thiol disulfide oxido-reductases with antioxidant capacity and catalytic functions closely associated with glutathione, the major small molecular weight antioxidant of human lung. However, the role of Grxs in smoking related diseases is unclear. METHODS: Immunohistochemical and Western blot analyses were conducted with lung specimens (n = 45 and n = 32, respectively) and induced sputum (n = 50) of healthy non-smokers and smokers without COPD and at different stages of COPD. RESULTS: Grx1 was expressed mainly in alveolar macrophages. The percentage of Grx1 positive macrophages was significantly lower in GOLD stage IV COPD than in healthy smokers (p = 0.021) and the level of Grx1 in total lung homogenate decreased both in stage I–II (p = 0.045) and stage IV COPD (p = 0.022). The percentage of Grx1 positive macrophages correlated with the lung function parameters (FEV1, r = 0.45, p = 0.008; FEV1%, r = 0.46, p = 0.007, FEV/FVC%, r = 0.55, p = 0.001). Grx1 could also be detected in sputum supernatants, the levels being increased in the supernatants from acute exacerbations of COPD compared to non-smokers (p = 0.013) and smokers (p = 0.051). CONCLUSION: The present cross-sectional study showed that Grx1 was expressed mainly in alveolar macrophages, the levels being decreased in COPD patients. In addition, the results also demonstrated the presence of Grx1 in extracellular fluids including sputum supernatants. Overall, the present study suggests that Grx1 is a potential redox modulatory protein regulating the intracellular as well as extracellular homeostasis of glutathionylated proteins and GSH in human lung

Role of N-acetylcysteine in the management of COPD

The importance of the underlying local and systemic oxidative stress and inflammation in chronic obstructive pulmonary disease (COPD) has long been established. In view of the lack of therapy that might inhibit the progress of the disease, there is an urgent need for a successful therapeutic approach that, through affecting the pathological processes, will influence the subsequent issues in COPD management such as lung function, airway clearance, dyspnoea, exacerbation, and quality of life. N-acetylcysteine (NAC) is a mucolytic and antioxidant drug that may also influence several inflammatory pathways. It provides the sulfhydryl groups and acts both as a precursor of reduced glutathione and as a direct reactive oxygen species (ROS) scavenger, hence regulating the redox status in the cells. The changed redox status may, in turn, influence the inflammation-controlling pathways. Moreover, as a mucolytic drug, it may, by means of decreasing viscosity of the sputum, clean the bronchi leading to a decrease in dyspnoea and improved lung function. Nevertheless, as successful as it is in the in vitro studies and in vivo studies with high dosage, its actions at the dosages used in COPD management are debatable. It seems to influence exacerbation rate and limit the number of hospitalization days, however, with little or no influence on the lung function parameters. Despite these considerations and in view of the present lack of effective therapies to inhibit disease progression in COPD, NAC and its derivatives with their multiple molecular modes of action remain promising medication once doses and route of administration are optimized

Can Preening Contribute to Influenza A Virus Infection in Wild Waterbirds?

Wild aquatic birds in the Orders Anseriformes and Charadriiformes are the main reservoir hosts perpetuating the genetic pool of all influenza A viruses, including pandemic viruses. High viral loads in feces of infected birds permit a fecal-oral route of transmission. Numerous studies have reported the isolation of avian influenza viruses (AIVs) from surface water at aquatic bird habitats. These isolations indicate aquatic environments have an important role in the transmission of AIV among wild aquatic birds. However, the progressive dilution of infectious feces in water could decrease the likelihood of virus/host interactions. To evaluate whether alternate mechanisms facilitate AIV transmission in aquatic bird populations, we investigated whether the preen oil gland secretions by which all aquatic birds make their feathers waterproof could support a natural mechanism that concentrates AIVs from water onto birds' bodies, thus, representing a possible source of infection by preening activity. We consistently detected both viral RNA and infectious AIVs on swabs of preened feathers of 345 wild mallards by using reverse transcription–polymerase chain reaction (RT-PCR) and virus-isolation (VI) assays. Additionally, in two laboratory experiments using a quantitative real-time (qR) RT-PCR assay, we demonstrated that feather samples (n = 5) and cotton swabs (n = 24) experimentally impregnated with preen oil, when soaked in AIV-contaminated waters, attracted and concentrated AIVs on their surfaces. The data presented herein provide information that expands our understanding of AIV ecology in the wild bird reservoir system

It Costs to Be Clean and Fit: Energetics of Comfort Behavior in Breeding-Fasting Penguins

), seabirds known to fast for up to one month during incubation shifts ashore.A time budget was estimated from focal and scan sampling field observations and the energy cost of comfort activities was calculated from the associated increase in heart rate (HR) during comfort episodes, using previously determined equations relating HR to energy expenditure. We show that incubating birds spent 22% of their daily time budget in comfort behavior (with no differences between day and night) mainly devoted to preening (73%) and head/body shaking (16%). During comfort behavior, energy expenditure averaged 1.24 times resting metabolic rate (RMR) and the corresponding energy cost (i.e., energy expended in excess to RMR) was 58 kJ/hr. Energy expenditure varied greatly among various types of comfort behavior, ranging from 1.03 (yawning) to 1.78 (stretching) times RMR. Comfort behavior contributed 8.8–9.3% to total daily energy expenditure and 69.4–73.5% to energy expended daily for activity. About half of this energy was expended caring for plumage.This study is the first to estimate the contribution of comfort behavior to overall energy budget in a free-living animal. It shows that although breeding on a tight energy budget, king penguins devote a substantial amount of time and energy to comfort behavior. Such findings underline the importance of comfort behavior for the fitness of colonial seabirds

The Depletion of Nuclear Glutathione Impairs Cell Proliferation in 3t3 Fibroblasts

BACKGROUND:Glutathione is considered essential for survival in mammalian cells and yeast but not in prokaryotic cells. The presence of a nuclear pool of glutathione has been demonstrated but its role in cellular proliferation and differentiation is still a matter of debate. PRINCIPAL FINDINGS:We have studied proliferation of 3T3 fibroblasts for a period of 5 days. Cells were treated with two well known depleting agents, diethyl maleate (DEM) and buthionine sulfoximine (BSO), and the cellular and nuclear glutathione levels were assessed by analytical and confocal microscopic techniques, respectively. Both agents decreased total cellular glutathione although depletion by BSO was more sustained. However, the nuclear glutathione pool resisted depletion by BSO but not with DEM. Interestingly, cell proliferation was impaired by DEM, but not by BSO. Treating the cells simultaneously with DEM and with glutathione ethyl ester to restore intracellular GSH levels completely prevented the effects of DEM on cell proliferation. CONCLUSIONS:Our results demonstrate the importance of nuclear glutathione in the control of cell proliferation in 3T3 fibroblasts and suggest that a reduced nuclear environment is necessary for cells to progress in the cell cycle

Antioxidant therapies in COPD

Oxidative stress is an important feature in the pathogenesis of COPD. Targeting oxidative stress with antioxidants or boosting the endogenous levels of antioxidants is likely to be beneficial in the treatment of COPD. Antioxidant agents such as thiol molecules (glutathione and mucolytic drugs, such as N-acetyl-L-cysteine and N-acystelyn), dietary polyphenols (curcumin, resveratrol, green tea, catechins/quercetin), erdosteine, and carbocysteine lysine salt, all have been reported to control nuclear factor-kappaB (NF-κ B) activation, regulation of glutathione biosynthesis genes, chromatin remodeling, and hence inflammatory gene expression. Specific spin traps such as α-phenyl-N-tert-butyl nitrone, a catalytic antioxidant (ECSOD mimetic), porphyrins (AEOL 10150 and AEOL 10113), and a superoxide dismutase mimetic M40419 have also been reported to inhibit cigarette smoke-induced inflammatory responses in vivo. Since a variety of oxidants, free radicals, and aldehydes are implicated in the pathogenesis of COPD, it is possible that therapeutic administration of multiple antioxidants will be effective in the treatment of COPD. Various approaches to enhance lung antioxidant capacity and clinical trials of antioxidant compounds in COPD are discussed