Institute for Lung Health

Leicester, United Kingdom

Institute for Lung Health

Leicester, United Kingdom
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Wagner T.,FH Dortmund | Kroll A.,Eawag - Swiss Federal Institute of Aquatic Science and Technology | Haramagatti C.R.,University of Bayreuth | Lipinski H.-G.,FH Dortmund | Wiemann M.,Institute for Lung Health
PLoS ONE | Year: 2017

Darkfield and confocal laser scanning microscopy both allow for a simultaneous observation of live cells and single nanoparticles. Accordingly, a characterization of nanoparticle uptake and intracellular mobility appears possible within living cells. Single particle tracking allows to measure the size of a diffusing particle close to a cell. However, within the more complex system of a cell's cytoplasm normal, confined or anomalous diffusion together with directed motion may occur. In this work we present a method to automatically classify and segment single trajectories into their respective motion types. Single trajectories were found to contain more than one motion type. We have trained a random forest with 9 different features. The average error over all motion types for synthetic trajectories was 7.2%. The software was successfully applied to trajectories of positive controls for normal- and constrained diffusion. Trajectories captured by nanoparticle tracking analysis served as positive control for normal diffusion. Nanoparticles inserted into a diblock copolymer membrane was used to generate constrained diffusion. Finally we segmented trajectories of diffusing (nano-)particles in V79 cells captured with both darkfield- and confocal laser scanning microscopy. The software called "TraJClassifier" is freely available as Ima-geJ/Fiji plugin via https://git.io/v6uz2. © 2017 Wagner et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.


Brand P.L.P.,Isala Hospital | Brand P.L.P.,University of Groningen | Caudri D.,Erasmus Medical Center | Eber E.,Medical University of Graz | And 10 more authors.
European Respiratory Journal | Year: 2014

Since the publication of the European Respiratory Society Task Force report in 2008, significant new evidence has become available on the classification and management of preschool wheezing disorders. In this report, an international consensus group reviews this new evidence and proposes some modifications to the recommendations made in 2008. Specifically, the consensus group acknowledges that wheeze patterns in young children vary over time and with treatment, rendering the distinction between episodic viral wheeze and multiple-trigger wheeze unclear in many patients. Inhaled corticosteroids remain first-line treatment for multiple-trigger wheeze, but may also be considered in patients with episodic viral wheeze with frequent or severe episodes, or when the clinician suspects that interval symptoms are being under reported. Any controller therapy should be viewed as a treatment trial, with scheduled close follow-up to monitor treatment effect. The group recommends discontinuing treatment if there is no benefit and taking favourable natural history into account when making decisions about long-term therapy. Oral corticosteroids are not indicated in mild-to-moderate acute wheeze episodes and should be reserved for severe exacerbations in hospitalised patients. Future research should focus on better clinical and genetic markers, as well as biomarkers, of disease severity. Copyright © ERS 2014.


Murphy A.C.,Institute for Lung Health | Murphy A.C.,University of Leicester | Brightling C.E.,Institute for Lung Health | Brightling C.E.,University of Leicester | And 8 more authors.
Thorax | Year: 2012

Medication non-adherence and the clinical implications in difficult-to-control asthma were audited. Prescription issue data from 115 patients identified sub-optimal adherence (<80%) in 65% of patients on inhaled corticosteroids (ICS) or combined ICS/long-acting β2 agonist (LABA). In those using separate ICS and LABA, adherence to LABA (50%) was significantly better than to ICS (14.3%). Patients with sub-optimal ICS adherence had reduced FEV 1 and higher sputum eosinophil counts. Adherence ratio was an independent predictor of previous ventilation for acute severe asthma (p=0.008). The majority of patients with difficult-to-control asthma are non-adherent with their asthma medication. Non-adherence is correlated with poor clinical outcomes.


PubMed | German Federal Institute for Risk Assessment, Robert Koch Institute RKI, Institute of Energy and Environmental Technology IUTA e.V., University of Munster and Institute for Lung Health
Type: | Journal: Particle and fibre toxicology | Year: 2015

Oxidative stress, a commonly used paradigm to explain nanoparticle (NP)-induced toxicity, results from an imbalance between reactive oxygen species (ROS) generation and detoxification. As one consequence, protein carbonyl levels may become enhanced. Thus, the qualitative and quantitative description of protein carbonylation may be used to characterize how biological systems respond to oxidative stress induced by NPs.We investigated a representative panel of 24 NPs including functionalized amorphous silica (6), zirconium dioxide (4), silver (4), titanium dioxide (3), zinc oxide (2), multiwalled carbon nanotubes (3), barium sulfate and boehmite. Surface reactivities of all NPs were studied in a cell-free system by electron spin resonance (ESR). NRK-52E cells were treated with all NPs, analyzed for viability (WST-1 assay) and intracellular ROS production (DCFDA assay). Carbonylated proteins were assessed by 1D and/or 2D immunoblotting and identified by matrix assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF/TOF). In parallel, tissue homogenates from rat lungs intratracheally instilled with silver NPs were studied.Eleven NPs induced elevated levels of carbonylated proteins. This was in good agreement with the surface reactivity of the NPs as obtained by ESR and the reduction in cell viability as assessed by WST-1 assay. By contrast, results obtained by DCFDA assay were deviating. Each NP induced an individual pattern of protein carbonyls on 2D immunoblots. Affected proteins comprised cytoskeletal components, proteins being involved in stress response, or cytoplasmic enzymes of central metabolic pathways such as glycolysis and gluconeogenesis. Furthermore, induction of carbonyls upon silver NP treatment was also verified in rat lung tissue homogenates.Analysis of protein carbonylation is a versatile and sensitive method to describe NP-induced oxidative stress and, therefore, can be used to identify NPs of concern. Furthermore, detailed information about compromised proteins may aid in classifying NPs according to their mode of action.


PubMed | German Federal Institute for Risk Assessment, BASF, University of Duisburg - Essen, University of Munster and Institute for Lung Health
Type: | Journal: Toxicology and applied pharmacology | Year: 2016

Numbers of engineered nanomaterials (ENMs) are steadily increasing. Therefore, alternative testing approaches with reduced costs and high predictivity suitable for high throughput screening and prioritization are urgently needed to ensure a fast and effective development of safe products. In parallel, extensive research efforts are targeted to understanding modes of action of ENMs, which may also support the development of new predictive assays. Oxidative stress is a widely accepted paradigm associated with different adverse outcomes of ENMs. It has frequently been identified in in vitro and in vivo studies and different assays have been developed for this purpose. Fluorescent dye based read-outs are most frequently used for cell testing in vitro but may be limited due to possible interference of the ENMs. Recently, other assays have been put forward such as acellular determination of ROS production potential using methods like electron spin resonance, antioxidant quantification or the use of specific sensors. In addition, Omics based approaches have gained increasing attention. In particular, redox proteomics can combine the assessment of oxidative stress with the advantage of getting more detailed mechanistic information. Here we propose a comprehensive testing strategy for assessing the oxidative stress potential of ENMs, which combines acellular methods and fast in vitro screening approaches, as well as a more involved detailed redox proteomics approach. This allows for screening and prioritization in a first tier and, if required, also for unraveling mechanistic details down to compromised signaling pathways.


PubMed | German Federal Institute for Risk Assessment, H+ Technology, Institute for Lung Health, Saarland University and 3 more.
Type: Journal Article | Journal: Nanotoxicology | Year: 2016

Lung lining fluid is the first biological barrier nanoparticles (NPs) encounter during inhalation. As previous inhalation studies revealed considerable differences between surface functionalized NPs with respect to deposition and toxicity, our aim was to investigate the influence of lipid and/or protein binding on these processes. Thus, we analyzed a set of surface functionalized NPs including different SiO2 and ZrO2 in pure phospholipids, CuroSurf(TM) and purified native porcine pulmonary surfactant (nS). Lipid binding was surprisingly low for pure phospholipids and only few NPs attracted a minimal lipid corona. Additional presence of hydrophobic surfactant protein (SP) B in CuroSurf(TM) promoted lipid binding to NPs functionalized with Amino or PEG residues. The presence of the hydrophilic SP A in nS facilitated lipid binding to all NPs. In line with this the degree of lipid and protein affinities for different surface functionalized SiO2 NPs in nS followed the same order (SiO2 Phosphateunmodified SiO2


Alrashdan Y.A.,University of Sydney | Alkhouri H.,University of Sydney | Chen E.,University of Sydney | Lalor D.J.,University of Sydney | And 9 more authors.
American Journal of Physiology - Lung Cellular and Molecular Physiology | Year: 2012

CXCL10 (IP10) is involved in mast cell migration to airway smooth muscle (ASM) bundles in asthma. We aimed to investigate the role of cytokineinduced MAPK activation in CXCL10 production by ASM cells from people with and without asthma. Confluent growth-arrested ASM cells were treated with inhibitors of the MAPKs ERK, p38, and JNK and transcription factor NF-κB, or vehicle, and stimulated with IL-1ββ, TNF-α, or IFN-γ, alone or combined (cytomix). CXCL10 mRNA and protein, JNK, NF-κB p65 phosphorylation, and Ik-Bα protein degradation were assessed using real-time PCR, ELISA, and immunoblotting, respectively. Cytomix, IL-1β, and TNF-αinduced CXCL10 mRNA expression more rapidly in asthmatic than nonasthmatic ASM cells. IL-1β and/or TNF-αcombined with IFN-γsynergistically increased asthmatic ASM cell CXCL10 release. Inhibitor effects were similar in asthmatic and nonasthmatic cells, but cytomix-induced release was least affected, with only JNK and NF-κB inhibitors halving it. Notably, JNK phosphorylation was markedly less in asthmatic compared with nonasthmatic cells. However, in both, the JNK inhibitor SP600125 reduced JNK phosphorylation and CXCL10 mRNA levels but did not affect CXCL10 mRNA stability or Ik-Bα degradation. Together, the JNK and NF-κB inhibitors completely inhibited their CXCL10 release. We concluded that, in asthmatic compared with nonasthmatic ASM cells, JNK activation was reduced and CXCL10 gene expression was more rapid following cytomix stimulation. However, in both, JNK activation did not regulate early events leading to NF-κB activation. Thus JNK and NF-κB provide independent therapeutic targets for limiting CXCL10 production and mast cell migration to the ASM in asthma. © 2012 the American Physiological Society.


The two poorly soluble iron containing solid aerosols of siderite (FeCO 3) and magnetite (Fe 3O 4) were compared in a 4-week inhalation study on rats at similar particle mass concentrations of approximately 30 or 100mg/m 3. The particle size distributions were essentially identical (MMAD ≈1.4 μm). The iron-based concentrations were 12 or 38 and 22 or 66mg Fe/m 3 for FeCO 3 and Fe 3O 4, respectively. Modeled and empirically determined iron lung burdens were compared with endpoints suggestive of pulmonary inflammation by determinations in bronchoalveolar lavage (BAL) and oxidative stress in lung tissue during a postexposure period of 3 months. The objective of study was to identify the most germane exposure metrics, that are the concentration of elemental iron (mg Fe/m 3), total particle mass (mg PM/m 3) or particle volume (μl PM/m 3) and their associations with the effects observed. From this analysis it was apparent that the intensity of pulmonary inflammation was clearly dependent on the concentration of particle-mass or -volume and not of iron. Despite its lower iron content, the exposure to FeCO 3 caused a more pronounced and sustained inflammation as compared to Fe 3O 4. Similarly, borderline evidence of increased oxidative stress and inflammation occurred especially following exposure to FeCO 3 at moderate lung overload levels. The in situ analysis of 8-oxoguanine in epithelial cells of alveolar and bronchiolar regions supports the conclusion that both FeCO 3 and Fe 3O 4 particles are effectively endocytosed by macrophages as opposed to epithelial cells. Evidence of intracellular or nuclear sources of redox-active iron did not exist. In summary, this mechanistic study supports previous conclusions, namely that the repeated inhalation exposure of rats to highly respirable pigment-type iron oxides cause nonspecific pulmonary inflammation which shows a clear dependence on the particle volume-dependent lung overload rather than any increased dissolution and/or bioavailability of redox-active iron. © 2011 Informa Healthcare USA, Inc.


Szatkowski L.,University of Nottingham | Murray R.,University of Nottingham | Hubbard R.,University of Nottingham | Agrawal S.,Institute for Lung Health | And 2 more authors.
Thorax | Year: 2015

Using data from The Health Improvement Network and Hospital Episode Statistics, we investigate smoking prevalence, number of smokers treated and opportunities for cessation intervention among patients treated in National Health Service (NHS) hospitals in England from April 2010 to March 2011. Our results show that approximately 1.1 million smokers are treated in English hospitals each year, receiving a total of 2.6 million episodes of care. These findings suggest that delivering smoking cessation as a routine component of hospital care, as recommended by recent National Institute for Health and Care Excellence guidance, could achieve marked reductions in the prevalence of smoking and improve the cost-effectiveness of NHS hospitals.


PubMed | Miltenyi Biotec GmbH and Institute for Lung Health
Type: | Journal: Frontiers in immunology | Year: 2016

Extracellular vesicles (EVs) are specifically loaded with nucleic acids, lipids, and proteins from their parental cell. Therefore, the constitution of EVs reflects the type and status of the originating cell and EVs in melanoma patients plasma could be indicative for the tumor. Likewise, EVs might influence tumor progression by regulating immune responses. We performed a broad protein characterization of EVs from plasma of melanoma patients and healthy donors as well as from T cells, B cells, natural killer (NK) cells, monocytes, monocyte-derived dendritic cells (moDCs), and platelets using a multiplex bead-based platform. Using this method, we succeeded in analyzing 58 proteins that were differentially displayed on EVs. Hierarchical clustering of protein intensity patterns grouped EVs according to their originating cell type. The analysis of EVs from stimulated B cells and moDCs revealed the transfer of surface proteins to vesicles depending on the cell status. The protein profiles of plasma vesicles resembled the protein profiles of EVs from platelets, antigen-presenting cells and NK cells as shown by platelet markers, co-stimulatory proteins, and a NK cell subpopulation marker. In comparison to healthy plasma vesicles, melanoma plasma vesicles showed altered signals for platelet markers, indicating a changed vesicle secretion or protein loading of EVs by platelets and a lower CD8 signal that might be associated with a diminished activity of NK cells or T cells. As we hardly detected melanoma-derived vesicles in patients plasma, we concluded that blood cells induced the observed differences. In summary, our results question a direct effect of melanoma cells on the composition of EVs in melanoma plasma, but rather argue for an indirect influence of melanoma cells on the vesicle secretion or vesicle protein loading by blood cells.

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