Nasal Lipopolysaccharide Challenge and Cytokine Measurement Reflects Innate Mucosal Immune Responsiveness

<div><p>Background</p><p><b>P</b>ractical methods of monitoring innate immune mucosal responsiveness are lacking. Lipopolysaccharide (LPS) is a component of the cell wall of Gram negative bacteria and a potent activator of Toll-like receptor (TLR)-4. To measure LPS responsiveness of the nasal mucosa, we administered LPS as a nasal spray and quantified chemokine and cytokine levels in mucosal lining fluid (MLF).</p><p>Methods</p><p>We performed a 5-way cross-over, single blind, placebo-controlled study in 15 healthy non-atopic subjects (n = 14 <i>per protocol</i>). Doses of ultrapure LPS (1, 10, 30 or 100μg/100μl) or placebo were administered by a single nasal spray to each nostril. Using the recently developed method of nasosorption with synthetic adsorptive matrices (SAM), a series of samples were taken. A panel of seven cytokines/chemokines were measured by multiplex immunoassay in MLF. mRNA for intercellular cell adhesion molecule-1 (ICAM-1) was quantified from nasal epithelial curettage samples taken before and after challenge.</p><p>Results</p><p>Topical nasal LPS was well tolerated, causing no symptoms and no visible changes to the nasal mucosa. LPS induced dose-related increases in MLF levels of IL-1β, IL-6, CXCL8 (IL-8) and CCL3 (MIP-1α) (AUC at 0.5 to 10h, compared to placebo, p<0.05 at 30 and 100μg LPS). At 100μg LPS, IL-10, IFN-α and TNF-α were also increased (p<0.05). Dose-related changes in mucosal ICAM-1 mRNA were also seen after challenge, and neutrophils appeared to peak in MLF at 8h. However, 2 subjects with high baseline cytokine levels showed prominent cytokine and chemokine responses to relatively low LPS doses (10μg and 30μg LPS).</p><p>Conclusions</p><p>Topical nasal LPS causes dose-dependent increases in cytokines, chemokines, mRNA and cells. However, responsiveness can show unpredictable variations, possibly because baseline innate tone is affected by environmental factors. We believe that this new technique will have wide application in the study of the innate immune responses of the respiratory mucosa.</p><p>Key Messages</p><p>Ultrapure LPS was used as innate immune stimulus in a human nasal challenge model, with serial sampling of nasal mucosal lining fluid (MLF) by nasosorption using a synthetic absorptive matrix (SAM), and nasal curettage of mucosal cells. A dose response could be demonstrated in terms of levels of IL-1β, IL-6, CXCL8 and CCL3 in MLF, as well as ICAM-1 mRNA in nasal curettage specimens, and levels of neutrophils in nasal lavage. Depending on higher baseline levels of inflammation, there were occasional magnified innate inflammatory responses to LPS.</p><p>Trial Registration</p><p>Clinical Trials.gov <a href="https://clinicaltrials.gov/ct2/show/NCT02284074?term=nasal+lipopolysaccharide&rank=1" target="_blank">NCT02284074</a></p></div

Measurement of the inclusive and dijet cross-sections of b-jets in pp collisions at sqrt(s) = 7 TeV with the ATLAS detector

The inclusive and dijet production cross-sections have been measured for jets containing b-hadrons (b-jets) in proton-proton collisions at a centre-of-mass energy of sqrt(s) = 7 TeV, using the ATLAS detector at the LHC. The measurements use data corresponding to an integrated luminosity of 34 pb^-1. The b-jets are identified using either a lifetime-based method, where secondary decay vertices of b-hadrons in jets are reconstructed using information from the tracking detectors, or a muon-based method where the presence of a muon is used to identify semileptonic decays of b-hadrons inside jets. The inclusive b-jet cross-section is measured as a function of transverse momentum in the range 20 < pT < 400 GeV and rapidity in the range |y| < 2.1. The bbbar-dijet cross-section is measured as a function of the dijet invariant mass in the range 110 < m_jj < 760 GeV, the azimuthal angle difference between the two jets and the angular variable chi in two dijet mass regions. The results are compared with next-to-leading-order QCD predictions. Good agreement is observed between the measured cross-sections and the predictions obtained using POWHEG + Pythia. MC@NLO + Herwig shows good agreement with the measured bbbar-dijet cross-section. However, it does not reproduce the measured inclusive cross-section well, particularly for central b-jets with large transverse momenta.Comment: 10 pages plus author list (21 pages total), 8 figures, 1 table, final version published in European Physical Journal

Understanding the interplay between social and spatial behaviour

According to personality psychology, personality traits determine many aspects of human behaviour. However, validating this insight in large groups has been challenging so far, due to the scarcity of multi-channel data. Here, we focus on the relationship between mobility and social behaviour by analysing trajectories and mobile phone interactions of ∼1000 individuals from two high-resolution longitudinal datasets. We identify a connection between the way in which individuals explore new resources and exploit known assets in the social and spatial spheres. We show that different individuals balance the exploration-exploitation trade-off in different ways and we explain part of the variability in the data by the big five personality traits. We point out that, in both realms, extraversion correlates with the attitude towards exploration and routine diversity, while neuroticism and openness account for the tendency to evolve routine over long time-scales. We find no evidence for the existence of classes of individuals across the spatio-social domains. Our results bridge the fields of human geography, sociology and personality psychology and can help improve current models of mobility and tie formation

The nasal mucosal late allergic reaction to grass pollen involves type 2 inflammation (IL-5 and IL-13), the inflammasome (IL-1β), and complement

Non-invasive mucosal sampling (nasosorption and nasal curettage) was used following nasal allergen challenge with grass pollen in subjects with allergic rhinitis, in order to define the molecular basis of the late allergic reaction (LAR). It was found that the nasal LAR to grass pollen involves parallel changes in pathways of type 2 inflammation (IL-4, IL-5 and IL-13), inflammasome-related (IL-1β), and complement and circadian-associated genes. A grass pollen nasal spray was given to subjects with hay fever followed by serial sampling, in which cytokines and chemokines were measured in absorbed nasal mucosal lining fluid, and global gene expression (transcriptomics) assessed in nasal mucosal curettage samples. Twelve of 19 subjects responded with elevations in interleukin (IL)-5, IL-13, IL-1β and MIP-1β/CCL4 protein levels in the late phase. In addition, in these individuals whole-genome expression profiling showed upregulation of type 2 inflammation involving eosinophils and IL-4, IL-5 and IL-13; neutrophil recruitment with IL-1α and IL-1β; the alternative pathway of complement (factor P and C5aR); and prominent effects on circadian-associated transcription regulators. Baseline IL-33 mRNA strongly correlated with these late-phase responses, whereas a single oral dose of prednisone dose-dependently reversed most nasal allergen challenge-induced cytokine and transcript responses. This study shows that the LAR to grass pollen involves a range of inflammatory pathways and suggests potential new biomarkers and therapeutic targets. Furthermore, the marked variation in mucosal inflammatory events between different patients suggests that in the future precision mucosal sampling may enable rational specific therapy

Prenatal exposures and exposomics of asthma

This review examines the causal investigation of preclinical development of childhood asthma using exposomic tools. We examine the current state of knowledge regarding early-life exposure to non-biogenic indoor air pollution and the developmental modulation of the immune system. We examine how metabolomics technologies could aid not only in the biomarker identification of a particular asthma phenotype, but also the mechanisms underlying the immunopathologic process. Within such a framework, we propose alternate components of exposomic investigation of asthma in which, the exposome represents a reiterative investigative process of targeted biomarker identification, validation through computational systems biology and physical sampling of environmental medi

Fulvestrant-induced expression of ErbB3 and ErbB4 receptors sensitizes oestrogen receptor-positive breast cancer cells to heregulin β1

Introduction We have previously reported that induction of epidermal growth factor receptor and ErbB2 in response to antihormonal agents may provide an early mechanism to allow breast cancer cells to evade the growth-inhibitory action of such therapies and ultimately drive resistant cell growth. More recently, the other two members of the ErbB receptor family, ErbB3 and ErbB4, have been implicated in antihormone resistance in breast cancer. In the present study, we have investigated whether induction of ErbB3 and/or ErbB4 may provide an alternative resistance mechanism to antihormonal action in a panel of four oestrogen receptor (ER)-positive breast cancer cell lines. Methods MCF-7, T47D, BT474 and MDAMB361 cell lines were exposed to fulvestrant (100 nM) for seven days, and effects on ErbB3/4 expression and signalling, as well as on cell growth, were assessed. Effects of heregulin β1 (HRGβ1) were also examined in the absence and presence of fulvestrant to determine the impact of ER blockade on the capacity of this ErbB3/4 ligand to promote signalling and cell proliferation. Results Fulvestrant potently reduced ER expression and transcriptional activity and significantly inhibited growth in MCF-7, T47D, BT474 and MDAMB361 cells. However, alongside this inhibitory activity, fulvestrant also consistently induced protein expression and activity of ErbB3 in MCF-7 and T47D cells and ErbB4 in BT474 and MDAMB361 cell lines. Consequently, fulvestrant treatment sensitised all cell lines to the actions of the ErbB3/4 ligand HRGβ1 with enhanced ErbB3/4-driven signalling activity, reexpression of cyclin D1 and significant increases in cell proliferation being observed when compared to untreated cells. Indeed, in T47D and MDAMB361 HRGβ1 was converted from a ligand having negligible or suppressive growth activity into one that potently promoted cell proliferation. Consequently, fulvestrant-mediated growth inhibition was completely overridden by HRGβ1 in all four cell lines. Conclusions These findings suggest that although antihormones such as fulvestrant may have potent acute growth-inhibitory activity in ER-positive breast cancer cells, their ability to induce and sensitise cells to growth factors may serve to reduce and ultimately limit their inhibitory activity

Specific detection of H5N1 avian influenza A virus in field specimens by a one-step RT-PCR assay

BACKGROUND: Continuous outbreaks of the highly pathogenic H5N1 avian influenza A in Asia has resulted in an urgent effort to improve current diagnostics to aid containment of the virus and lower the threat of a influenza pandemic. We report here the development of a PCR-based assay that is highly specific for the H5N1 avian influenza A virus. METHODS: A one-step reverse-transcription PCR assay was developed to detect the H5N1 avian influenza A virus. The specificity of the assay was shown by testing sub-types of influenza A virus and other viral and bacterial pathogens; and on field samples. RESULTS: Validation on 145 field specimens from Vietnam and Malaysia showed that the assay was specific without cross reactivity to a number of other infuenza strains as well as human respiratory related pathogens. Detection was 100% from allantoic fluid in H5N1 positive samples, suggesting it to be a reliable sampling source for accurate detection. CONCLUSION: The assay developed from this study indicates that the primers are specific for the H5N1 influenza virus. As shown by the field tested results, this assay would be highly useful as a diagnostic tool to help identify and control influenza epidemics

Rapid semi-automated quantitative multiplex tandem PCR (MT-PCR) assays for the differential diagnosis of influenza-like illness

<p>Abstract</p> <p>Background</p> <p>Influenza A, including avian influenza, is a major public health threat in developed and developing countries. Rapid and accurate detection is a key component of strategies to contain spread of infection, and the efficient diagnosis of influenza-like-illness is essential to protect health infrastructure in the event of a major influenza outbreak.</p> <p>Methods</p> <p>We developed a multiplexed PCR (MT-PCR) assay for the simultaneous diagnosis of respiratory viruses causing influenza-like illness, including the specific recognition of influenza A haemagglutinin subtypes H1, H3, and H5. We tested several hundred clinical specimens in two diagnostic reference laboratories and compared the results with standard techniques.</p> <p>Results</p> <p>The sensitivity and specificity of these assays was higher than individual assays based on direct antigen detection and standard PCR against a range of control templates and in several hundred clinical specimens. The MT-PCR assays provided differential diagnoses as well as potentially useful quantitation of virus in clinical samples.</p> <p>Conclusions</p> <p>MT-PCR is a potentially powerful tool for the differential diagnosis of influenza-like illness in the clinical diagnostic laboratory.</p

The Cooperation between hMena Overexpression and HER2 Signalling in Breast Cancer

hMena and the epithelial specific isoform hMena11a are actin cytoskeleton regulatory proteins belonging to the Ena/VASP family. EGF treatment of breast cancer cell lines upregulates hMena/hMena11a expression and phosphorylates hMena11a, suggesting cross-talk between the ErbB receptor family and hMena/hMena11a in breast cancer. The aim of this study was to determine whether the hMena/hMena11a overexpression cooperates with HER-2 signalling, thereby affecting the HER2 mitogenic activity in breast cancer. In a cohort of breast cancer tissue samples a significant correlation among hMena, HER2 overexpression, the proliferation index (high Ki67), and phosphorylated MAPK and AKT was found and among the molecular subtypes the highest frequency of hMena overexpressing tumors was found in the HER2 subtype. From a clinical viewpoint, concomitant overexpression of HER2 and hMena identifies a subgroup of breast cancer patients showing the worst prognosis, indicating that hMena overexpression adds prognostic information to HER2 overexpressing tumors. To identify a functional link between HER2 and hMena, we show here that HER2 transfection in MCF7 cells increased hMena/hMena11a expression and hMena11a phosphorylation. On the other hand, hMena/hMena11a knock-down reduced HER3, AKT and p44/42 MAPK phosphorylation and inhibited the EGF and NRG1-dependent HER2 phosphorylation and cell proliferation. Of functional significance, hMena/hMena11a knock-down reduced the mitogenic activity of EGF and NRG1. Collectively these data provide new insights into the relevance of hMena and hMena11a as downstream effectors of the ErbB receptor family which may represent a novel prognostic indicator in breast cancer progression, helping to stratify patients