Utilisation of a novel ProteaseTag™ activity immunoassay for the specific measurement of neutrophil elastase in patients with Cystic Fibrosis

Neutrophil elastase (NE), a biomarker of infection and inflammation, correlates with the severity of several respiratory diseases including cystic fibrosis (CF) however, its detection and quantification in biological samples is confounded by a lack of robust methodologies. Standard assays using chromogenic or fluorogenic substrates are not specific when added to complex samples containing multiple proteolytic and hydrolytic enzymes, resulting in an over-estimation of the target protease. ELISA systems measure total protein levels which can be a mixture of latent, active and protease-inhibitor complexes. We have therefore developed a novel immunoassay (NE-Tag ELISA), incorporating an activity dependent ProteaseTag™ and a specific antibody step, which is selective and specific for the capture of active NE. The objective of this study was to clinically validate NE-Tag ELISA for the detection of active NE in sputum from CF patients. Sputum (n=45) was recovered from CF patients hospitalised for acute exacerbation. Sol was recovered and analysed for NE activity using the NE-Tag ELISA and two fluorogenic substrate-based assays [1. Suc-AAPV-AMC (Sigma) and 2. InnozymeTM Immunocapture assay (Calbiochem)]. NE activity between assays and with a range of clinical parameters was correlated.A highly significant correlation was shown between assays. NE activity (NE-Tag) further correlated appropriately with clinical parameters: inversely with FEV1 (p = 0.036) and positively with CRP (p = 0.035), neutrophils and total white cell counts (p < 0.001). The InnozymeTM assay showed similar correlations with the clinical parameters (with the exception of CRP). No correlations with any of the clinical parameters were observed when NE was measured using the standard fluorogenic substrate.

Validation of non‑stationary precipitation series for site‑specific impact assessment: comparison of two statistical downscaling techniques

Statistical downscaling (SD) methods have become a popular, low-cost and accessible means of bridging the gap between the coarse spatial resolution at which climate models output climate scenarios and the finer spatial scale at which impact modellers require these scenarios, with various different SD techniques used for a wide range of applications across the world. This paper compares the Generator for Point Climate Change (GPCC) model and the Statistical DownScaling Model (SDSM)—two contrasting SD methods—in terms of their ability to generate precipitation series under non-stationary conditions across ten contrasting global climates. The mean, maximum and a selection of distribution statistics as well as the cumulative frequencies of dry and wet spells for four different temporal resolutions were compared between the models and the observed series for a validation period. Results indicate that both methods can generate daily precipitation series that generally closely mirror observed series for a wide range of non-stationary climates. However, GPCC tends to overestimate higher precipitation amounts, whilst SDSM tends to underestimate these. This infers that GPCC is more likely to overestimate the effects of precipitation on a given impact sector, whilst SDSM is likely to underestimate the effects. GPCC performs better than SDSM in reproducing wet and dry day frequency, which is a key advantage for many impact sectors. Overall, the mixed performance of the two methods illustrates the importance of users performing a thorough validation in order to determine the influence of simulated precipitation on their chosen impact sector.

Age- and Sex-Specific Causal Effects of Adiposity on Cardiovascular Risk Factors

Observational studies have reported different effects of adiposity on cardiovascular risk factors across age and sex. Since cardiovascular risk factors are enriched in obese individuals, it has not been easy to dissect the effects of adiposity from those of other risk factors. We used a Mendelian randomization approach, applying a set of 32 genetic markers to estimate the causal effect of adiposity on blood pressure, glycemic indices, circulating lipid levels, and markers of inflammation and liver disease in up to 67,553 individuals. All analyses were stratified by age (cutoff 55 years of age) and sex. The genetic score was associated with BMI in both nonstratified analysis (P = 2.8 × 10(-107)) and stratified analyses (all P < 3.3 × 10(-30)). We found evidence of a causal effect of adiposity on blood pressure, fasting levels of insulin, C-reactive protein, interleukin-6, HDL cholesterol, and triglycerides in a nonstratified analysis and in the <55-year stratum. Further, we found evidence of a smaller causal effect on total cholesterol (P for difference = 0.015) in the ≥55-year stratum than in the <55-year stratum, a finding that could be explained by biology, survival bias, or differential medication. In conclusion, this study extends previous knowledge of the effects of adiposity by providing sex- and age-specific causal estimates on cardiovascular risk factors.

Genome-wide gene pathway analysis of psychotic illness symptom dimensions based on a new schizophrenia-specific model of the OPCRIT

Empirically derived phenotypic measurements have the potential to enhance gene-finding efforts in schizophrenia. Previous research based on factor analyses of symptoms has typically included schizoaffective cases. Deriving factor loadings from analysis of only narrowly defined schizophrenia cases could yield more sensitive factor scores for gene pathway and gene ontology analyses. Using an Irish family sample, this study 1) factor analyzed clinician-rated Operational Criteria Checklist items in cases with schizophrenia only, 2) scored the full sample based on these factor loadings, and 3) implemented genome-wide association, gene-based, and gene-pathway analysis of these SCZ-based symptom factors (final N= 507). Three factors emerged from the analysis of the schizophrenia cases: a manic, a depressive, and a positive symptom factor. In gene-based analyses of these factors, multiple genes had q<. 0.01. Of particular interest are findings for PTPRG and WBP1L, both of which were previously implicated by the Psychiatric Genomics Consortium study of SCZ; results from this study suggest that variants in these genes might also act as modifiers of SCZ symptoms. Gene pathway analyses of the first factor indicated over-representation of glutamatergic transmission, GABA-A receptor, and cyclic GMP pathways. Results suggest that these pathways may have differential influence on affective symptom presentation in schizophrenia.

Age-specific risk factor profiles of adenocarcinomas of the esophagus: a pooled analysis from the international BEACON consortium

Esophageal (EA) and esophagogastric junction (EGJA) adenocarcinoma have been steadily increasing in frequency in younger people, however the etiology of these cancers is poorly understood. We therefore investigated associations of body- mass index (BMI), cigarette smoking, alcohol consumption, gastroesophageal reflux, and use of non-steroidal anti-inflammatory drugs (NSAIDs) in relation to age-specific risks of EA and EGJA. We pooled individual participant data from eight population-based, case-control studies within the international Barrett’s and Esophageal Adenocarcinoma Consortium (BEACON). The analysis included 1,363 EA patients, 1,472 EGJA patients, and 5,728 control participants. Multivariable logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for age-specific (<50, 50–59, 60–69, ≥70 years) cancer outcomes, as well as interactions by age. BMI, smoking status and pack-years, recurrent gastroesophageal reflux, and frequency of gastroesophageal reflux were positively associated with EA and EGJA in each age group. Early-onset EA (<50 years) had stronger associations with recurrent gastroesophageal reflux (OR=8.06, 95%CI: 4.52, 14.37; Peffect modification=0.01) and BMI (ORBMI ≥30 vs. <25=4.19, 95%CI: 2.23, 7.87; Peffect modification=0.04), relative to older age groups. In contrast, inverse associations of NSAID use were strongest in the oldest age group (≥70 years), although this apparent difference was not statistically significant. Age-specific associations with EGJA showed similar, but slightly weaker patterns and no statistically significant differences by age were observed. Our study provides evidence that associations between obesity and gastroesophageal reflux are stronger among earlier onset EA cancers.

Olefin / Paraffin separation using Task-Specific Materials based on Ionic Liquids: Advances in Gas Processing

In this work, 1-hexene was extracted from its mixtures with n-hexane in varying ratios using a task specific ionic liquid. Herein, the ionic liquid (IL) 1-butyl-3-methylimidazolium nitrate, [BMIM][NO3], was used and examined with and without the addition of a metal salt. The impact of water on both selectivity and distribution coefficient was also tested. Four potential metal salts were investigated, the results of which demonstrate that the dissolution of transition-metal salts in the IL improves the separation of 1-hexene from n-hexane through metal-olefin complexation. Additionally, the presence of water in IL solutions containing metal salt enhances this selectivity. Finally, UNIFAC was used to correlate the experimental LLE data with good accuracy.

, Lexicographic Preferences in Discrete Choice Experiments: Consequences on Individual-Specific Willingness to Pay Estimates

In discrete choice experiments respondents are generally assumed to consider all of the attributes across each of the alternatives, and to choose their most preferred. However, results in this paper indicate that many respondents employ simplified lexicographic decision-making rules, whereby they have a ranking of the attributes, but their choice of an alternative is based solely on the level of their most important attribute(s). Not accounting for these simple decision-making heuristics introduces systemic errors and leads to biased point estimates, as they are a violation of the continuity axiom and a departure from the use of compensatory decision-making. In this paper the implications of lexicographic preferences are examined. In particular, using a mixed logit specification this paper investigates the sensitivity of individual-specific willingness to pay (WTP) estimates conditional on whether lexicographic decision-making rules are accounted for in the modelling of discrete choice responses. Empirical results are obtained from a discrete choice experiment that was carried out to address the value of a number of rural landscape attributes in Ireland

Cadmium and lead in vegetable and fruit produce selected from specific regional areas of the UK

Cadmium and lead were determined in fruit and vegetable produce (~1300 samples) collected from a field and market basket study of locally grown produce from the South-West of Britain (Devon and Cornwall). These were compared with similarly locally grown produce from the North-East of Britain (Aberdeenshire). The concentrations of cadmium and lead in the market basket produce were compared to the maximum levels (ML) set by the European Union (EU). For cadmium 0.2% of the samples exceeded the ML, and 0.6% of the samples exceeded the ML for lead. The location of cadmium and lead in potatoes was performed using laser ablation ICP-MS. All tested samples exhibited higher lead concentrations, and most exhibited increased concentrations of cadmium in the potato skin compared to the flesh. The concentrations of cadmium and lead found in fruits and vegetables sampled during this study do not increase concern about risk to human health.

Monitoring of lineage-specific chimaerism allows early prediction of response following donor lymphocyte infusions for relapsed chronic myeloid leukaemia

Donor lymphocyte infusions (DLI) have been shown to enhance the graft-versus-leukaemia (GVL) effect and induce haematological and molecular remission in patients with relapsed CML following allogeneic bone marrow transplantation (BMT). The potent donor cell-mediated cytolysis following DLI may lead to a short period of aplasia before the re-establishment of donor haematopoiesis. The absence of detectable donor cells in patients prior to DLI infusion may result in permanent aplasia in certain patients. We report on four patients who relapsed 1, 3, 6.5 and 7 years post-BMT for chronic phase CML and were treated with DLI from their original BMT donor. Polymorphic short tandem repeats (STRs) were used to assess haematological chimaerism both prior to and following DLI. At the time of relapse, STR-PCR indicated the presence of donor cells in all four patients, at levels ranging from 1-40%. A clinical and molecular response was seen in 4/4 patients following a short period of cytopenia and all patients remain in clinical remission with a follow-up of 2 months-3 years post-DLI. STR-PCR indicated that a response was occurring during the period of pancytopenia when metaphase analysis was unsuccessful. Lineage-specific analysis of the cellular response to DLI was monitored using STR-PCR of peripheral blood (PB) and bone marrow (BM) lymphocyte-enriched fractions and CD2-positive and -negative T cell fractions. In one patient BM and PB CD34-positive and -negative fractions were also assessed. A change in the ratio of donor:recipient cells in the PB lymphocyte fraction was the earliest molecular indication of an anti-leukaemic response. Subsequent conversion to donor chimaerism occurred in the other lineages and the granulocyte fraction was the last lineage to convert. In conclusion, lineage-specific STR-PCR permits detailed monitoring of subtle changes in donor/recipient cell dynamics in specific lineages following DLI during the crucial pancytopenic phase and may be a useful predictor of haematological response to DLI therapy.

The TGF-β-inducible miR-23a cluster attenuates IFN-γ levels and antigen-specific cytotoxicity in human CD8+ T cells

Cytokine secretion and degranulation represent key components of CD8(+) T-cell cytotoxicity. While transcriptional blockade of IFN-γ and inhibition of degranulation by TGF-β are well established, we wondered whether TGF-β could also induce immune-regulatory miRNAs in human CD8(+) T cells. We used miRNA microarrays and high-throughput sequencing in combination with qRT-PCR and found that TGF-β promotes expression of the miR-23a cluster in human CD8(+) T cells. Likewise, TGF-β up-regulated expression of the cluster in CD8(+) T cells from wild-type mice, but not in cells from mice with tissue-specific expression of a dominant-negative TGF-β type II receptor. Reporter gene assays including site mutations confirmed that miR-23a specifically targets the 3'UTR of CD107a/LAMP1 mRNA, whereas the further miRNAs expressed in this cluster-namely, miR-27a and -24-target the 3'UTR of IFN-γ mRNA. Upon modulation of the miR-23a cluster by the respective miRNA antagomirs and mimics, we observed significant changes in IFN-γ expression, but only slight effects on CD107a/LAMP1 expression. Still, overexpression of the cluster attenuated the cytotoxic activity of antigen-specific CD8(+) T cells. These functional data thus reveal that the miR-23a cluster not only is induced by TGF-β, but also exerts a suppressive effect on CD8(+) T-cell effector functions, even in the absence of TGF-β signaling.