Effect of Pesticide Inoculation,Duration of Composting,and Degradation Time on the Content of Compost Fatty Acids,Quantified Using Two Methods

Compost biobeds can promote biodegradation of pesticides. The microbial community structure changes during the composting process, and simple methods can potentially be used to follow these changes. In this study the microbial identification (MIDI) and ester-linked (EL) procedures were used to determine the composition of fatty acid methyl esters (FAMEs) in composts aged 3 and 12 months, inoculated with 3 recalcitrant pesticides (azoxystrobin, chlorotoluron, and epoxyconazole and a coapplication of all three) after 0, 56, and 125 days of degradation. Pesticide persistence was high, and after 125 days the residue was 22 to 70% of the applied amount depending mostly on the composting age. Seventy-one FAMEs belonging to nine groups were detected. The EL method provided three times as many detections as did the MIDI method and was more sensitive for all FAME groups except alcohol. Thirty-six and five FAMEs were unique to the EL and MIDI methods, respectively. The extraction method was of importance. The EL method provided a higher number of detections for 57 FAMEs, and the MIDI method provided a higher number for 9 FAMEs, while the two methods were equal for 5 FAMEs; thus, the EL method provided a more uniform overall FAME profile. Effects of the other factors were not always clear. Inoculation with pesticide did not influence the FAME profile with the MIDI method, while it influenced cyclopropane and monounsaturated content with the EL method. Composting age and degradation time had an effect on some groups of FAMEs, and this effect was greater with the EL method. The use of some FAMEs as biomarkers to follow microbial community succession was likely influenced by the type of compost and other factors.Plant protection has become a key factor in intensive agriculture, but the widespread use of pesticides can be toxic to nontarget organisms and can lead to ecosystem alterations. Formulated chlorotoluron (a herbicide), azoxystrobin (a fungicide), and epoxyconazole (a fungicide) are commonly used at the postemergence stage mainly for cereals, to control broad-leaved weeds and grasses and to control some foliar and soilborne diseases, respectively.Most information on pesticide persistence is derived from registration documents, few studies appear in the scientific literature, and independent research is always suggested. A recent pesticide database (Footprint, 2007; http://www.eu-footprint.org/ppdb.html, accessed April 2010) reports half-life values (days) in soil, derived from laboratory studies at 20°C, of 73.5 for azoxystrobin, 59 for chlorotoluron, and 226 for epoxyconazole. However, a certain variability exists, and in other research half-lives are longer, i.e., more than 2 years (at 10°C) for epoxyconazole (4), 62 to 107 days for azoxystrobin (10), and 30 to 200 days for chlorotoluron (36), showing that significant degradation should be measured on a time scale of months or years (8, 9) according to the results of a recent review on pesticide persistence (3).The use of such persistent pesticides on agricultural fields can have a high impact on the environment through leaching into aquifers and altering the microbial community and processes in the soil. Investigations of structural and functional changes in soil microbial communities have been done using several methods, including the fatty acid profiling method (14, 27, 29). This procedure has also been used as an early and sensitive method to investigate microorganism succession during the composting process (32), together with the composition of long-chain fatty acids (26, 28).Recent studies show a short-term effect on fatty acids during composting, with the main decrease being between days 3 and 24 of composting (32), and also a fatty acid profile shift in months-long experiments (2).Simple standardized methods to monitor changes in the microbial community structure may have the potential to follow microbial succession and determine compost maturity and quality. Methods to determine the fatty acid composition of the microorganisms have thus been developed, such as the microbial identification (MIDI) system and the ester-linked (EL) procedure (25, 29). The application of these methods to compost samples is recent, and little is known about the extent to which they can give comparable information (32).Compost is an interesting medium for biodepuration systems such as biobeds (20) because the high content and availability of organic matter could have a significant effect on pesticide adsorption and dissipation (16, 34). The effectiveness of the degradative capacity of those systems can be studied using a worst-case scenario, i.e., using pesticides recalcitrant to degradation.Some studies show that less mature compost is better able to absorb and degrade pesticides, due to greater carbon sources for microorganisms to cometabolize and remove pesticides (37). For example, in samples taken from composting facilities (33) it was found that a 6-month-old compost had a greater capacity to remove atrazine from solution, because of its higher organic carbon content, and this suggested that less mature compost may be better suited for environmental applications. However, since most compost is produced in big facilities, much of the compost can remain in the facility and mature, changing physical-chemical and microbiological properties (13) and consequently pesticide removal capacity.The main goal of this laboratory study is to assess the evolution of microorganisms in compost aged 3 and 12 months after inoculation with three recalcitrant pesticides (azoxystrobin, chlorotoluron, and epoxyconazole, the persistence of which is also measured) and a coapplication of all three, through the changes in the fatty acid methyl ester (FAME) profile using two methods, the microbial identification (MIDI) and ester-linked (EL) methods.     

Altered Expression of Fibrosis Genes in Capsules of Failed Ahmed Glaucoma Valve Implants

Purpose

Ahmed glaucoma valve (AGV) implant is an aqueous shunt device used to control intraocular pressure in glaucoma. Implant failure results from impervious encapsulation of the shunt plate causing increased hydraulic resistance and raised intraocular pressure. We hypothesized that deregulation of fibrosis pathway contributes to capsular resistance. We tested this by studying fibrosis related gene expression in failed AGV implants.

Methods

Differential gene expression was examined in failed AGV capsules and compared to normal control tenon. Following total RNA extraction, 84 key genes in fibrosis pathway were examined by real-time PCR using RT² Profiler PCR Array. Relative gene expression was calculated using ΔΔC_t method. Gene specific TaqMan assays were used to validate select genes with ≥2 fold differential expression in the array expression profile.

Results

We observed differential expression in several genes in the fibrosis pathway. Almost half (39/84) of examined genes showed ≥2 fold differential expression in majority of capsules examined on the array. TaqMan assays for select genes including CCN2 (CTGF), THBS1, SERPINE1, THBS2, COL3A1, MMP3, and IL1A in an increased validation sample set showed significant changes in expression (p value from <0.001 to 0.022) at a high frequency in concurrence with our array results.

Conclusions

Pathway-focused analyses identified candidate genes with altered expression providing molecular evidence for deregulation of the fibrosis pathway in AGV failure.     

First Administration of the Fc-Attenuated Anti-β Amyloid Antibody GSK933776 to Patients with Mild Alzheimer’s Disease: A Randomized,Placebo-Controlled Study

Objective

To assess the safety, tolerability, pharmacokinetics, and pharmacodynamics of the Fc-inactivated anti-β amyloid (Aβ) monoclonal antibody (mAb) GSK933776 in patients with mild Alzheimer’s disease (AD) or mild cognitive impairment (MCI).

Methods

This was a two-part, single blind, placebo-controlled, first-time-in-human (FTIH) study of single (n = 18) and repeat dose (n = 32) intravenous GSK933776 0.001–6 mg/kg (ClinicalTrials.gov: NCT00459550). Additional safety data from an open-label, uncontrolled, single dose study of intravenous GSK933776 1–6 mg/kg (n = 18) are included (ClinicalTrials.gov: NCT01424436).

Results

There were no cases of amyloid-related imaging abnormalities-edema (ARIA-E) or –hemorrhage (ARIA-H) after GSK933776 administration in both studies. Three patients across the two studies developed anti-GSK933776 antibodies. Plasma GSK933776 half-life (t_1/2) was 10–15 days after repeat dosing. After each of three administrations of GSK933776, plasma levels of total Aβ42 and Aβ increased whereas plasma levels of free Aβ decreased dose dependently; no changes were observed for placebo. For total Aβ42 the peak:trough ratio was ≤2 at doses ≥3 mg/kg; for total Aβ the ratio was ≤2 at 6 mg/kg. CSF concentrations of Aβ showed increases from baseline to week 12 for Aβ X–38 (week 12:baseline ratio: 1.65; 95%CI: 1.38, 1.93) and Aβ X–42 (week 12:baseline ratio: 1.18; 95%CI: 1.06, 1.30) for values pooled across doses.

Conclusion

In this FTIH study the Fc-inactivated anti-Aβ mAb GSK933776 engaged its target in plasma and CSF without causing brain ARIA-E/H in patients with mild AD or MCI.

Trial Registration

ClinicalTrials.gov NCT00459550     

Serum TGF-β1 as a Biomarker for Type 2 Diabetic Nephropathy: A Meta-Analysis of Randomized Controlled Trials

Background

Abnormal expression of serum TGF-β1 was found in patients with diabetic nephropathy. However, the association of TGF-β1 with the risk of diabetic nephropathy remains unknown. The present study was undertaken to investigate whether such an association exists.

Methods

We searched the Chinese VIP, Wangfang, China National Knowledge Infrastructure, PubMed, Embase, and Google Scholar databases for relevant studies and extracted all eligible data. Stata12 software was used for statistical analysis.

Results

Nine reports met our criteria and were used for data extraction. There were 264 patients and 227 healthy controls from qualified reports in this meta-analysis. The results suggested that serum TGF-β1 levels were significantly up-regulated in patients with diabetic nephropathy; the instrumental variable was 3.94 (95% confidence interval 3.20–4.68, p<0.01).

Conclusions

Meta-analysis suggested that elevated serum TGF-β level in patients with diabetes is associated with a high risk of nephropathy. Further studies are required to validate these observations.     

Nasal Levels of Antimicrobial Peptides in Allergic Asthma Patients and Healthy Controls: Differences and Effect of a Short 1,25(OH)2 Vitamin D3 Treatment

Background

Allergy is often accompanied by infections and lower levels of antimicrobial peptides (AMPs). Vitamin D has been shown to increase expression of selected AMPs. In this study we investigated whether antimicrobial peptide levels in nasal secretions of allergic asthma patients are lower than in healthy controls, and whether administration of the active form of vitamin D (1,25(OH)₂D3) affects these antimicrobial peptide levels.

Methods

The levels of antimicrobial peptides in nasal secretions were compared between 19 allergic asthma patients and 23 healthy controls. The effect of seven days daily oral treatment with 2 μg 1,25(OH)₂D3 on antimicrobial peptides in nasal secretions was assessed in a placebo-controlled cross-over clinical study.

Results

Levels of neutrophil α-defensins (human neutrophil peptides 1–3; HNP1-3) and lipocalin 2 (LCN2; also known as NGAL) were significantly lower in asthmatics, but no differences in LL-37 and SLPI were detected. Treatment with a short-term 1,25(OH)₂D3 caused a small increase in HNP1-3, but not when the asthma and control groups were analyzed separately. LL-37, LCN2 and SLPI did not change after treatment with 1,25(OH)₂D3.

Conclusion

Levels of the antimicrobial peptides HNP1-3 and LCN2 are lower in nasal secretions in asthmatics and are not substantially affected by a short-term treatment with active vitamin D.     

Multiparametric MRI of Epiphyseal Cartilage Necrosis (Osteochondrosis) with Histological Validation in a Goat Model

Purpose

To evaluate multiple MRI parameters in a surgical model of osteochondrosis (OC) in goats.

Methods

Focal ischemic lesions of two different sizes were induced in the epiphyseal cartilage of the medial femoral condyles of goats at 4 days of age by surgical transection of cartilage canal blood vessels. Goats were euthanized and specimens harvested 3, 4, 5, 6, 9 and 10 weeks post-op. Ex vivo MRI scans were conducted at 9.4 Tesla for mapping the T₁, T₂, T_1ρ, adiabatic T_1ρ and T_RAFF relaxation times of articular cartilage, unaffected epiphyseal cartilage, and epiphyseal cartilage within the area of the induced lesion. After MRI scans, safranin O staining was conducted to validate areas of ischemic necrosis induced in the medial femoral condyles of six goats, and to allow comparison of MRI findings with the semi-quantitative proteoglycan assessment in corresponding safranin O-stained histological sections.

Results

All relaxation time constants differentiated normal epiphyseal cartilage from lesions of ischemic cartilage necrosis, and the histological staining results confirmed the proteoglycan (PG) loss in the areas of ischemia. In the scanned specimens, all of the measured relaxation time constants were higher in the articular than in the normal epiphyseal cartilage, consistently allowing differentiation between these two tissues.

Conclusions

Multiparametric MRI provided a sensitive approach to discriminate between necrotic and viable epiphyseal cartilage and between articular and epiphyseal cartilage, which may be useful for diagnosing and monitoring OC lesions and, potentially, for assessing effectiveness of treatment interventions.     

A 21-Day of Adjunctive Corticosteroid Use May Not Be Necessary for HIV-1-Infected Pneumocystis Pneumonia with Moderate and Severe Disease

Background

The current guidelines recommend 21-day adjunctive corticosteroid therapy for HIV-1-infected pneumocystis pneumonia patients (HIV-PCP) with moderate-to-severe disease. Whether shorter adjunctive corticosteroid therapy is feasible in such patients is unknown.

Methods

We conducted a retrospective study to elucidate the proportion of patients with moderate and severe HIV-PCP who required adjunctive corticosteroid therapy for 21 days. The enrollment criteria included HIV-PCP that fulfilled the current criteria for 21-day corticosteroid therapy; PaO₂ on room air of <70mmHg or A-aDO₂ ≥35 mmHg.

Results

The median duration of corticosteroid therapy in the 73 study patients was 13 days (IQR 9–21). Adjunctive corticosteroid therapy was effective and discontinued within 10 and 14 days in 30% and 60% of the patients, respectively. Only 9% of the patients with moderate HIV-PCP (n = 22, A-aDO₂ 35–45 mmHg) received steroids for >14 days, whereas 35% of the patients with severe HIV-PCP (n = 51, A-aDO₂ ≥45 mmHg) required corticosteroid therapy for ≥21 days. Four (13%) of the severe cases died, whereas no patient with moderate disease died. Among patients with severe HIV-PCP, discontinuation of corticosteroid therapy within 14 days correlated significantly with higher baseline CD4 (p = 0.049).

Conclusion

Shorter adjunctive corticosteroid therapy was clinically effective and adjunctive corticosteroid could be discontinued within 14 days in 60% of moderate-to-severe HIV-PCP and 90% of moderate cases.     

Polyunsaturated Fatty Acids Attenuate Diet Induced Obesity and Insulin Resistance,Modulating Mitochondrial Respiratory Uncoupling in Rat Skeletal Muscle

Objectives

Omega (ω)-3 polyunsaturated fatty acids (PUFA) are dietary compounds able to attenuate insulin resistance. Anyway, the precise actions of ω-3PUFAs in skeletal muscle are overlooked. We hypothesized that PUFAs, modulating mitochondrial function and efficiency, would ameliorate pro-inflammatory and pro-oxidant signs of nutritionally induced obesity.

Study Design

To this aim, rats were fed a control diet (CD) or isocaloric high fat diets containing either ω-3 PUFA (FD) or lard (LD) for 6 weeks.

Results

FD rats showed lower weight, lipid gain and energy efficiency compared to LD-fed animals, showing higher energy expenditure and O₂ consumption/CO₂ production. Serum lipid profile and pro-inflammatory parameters in FD-fed animals were reduced compared to LD. Accordingly, FD rats exhibited a higher glucose tolerance revealed by an improved glucose and insulin tolerance tests compared to LD, accompanied by a restoration of insulin signalling in skeletal muscle. PUFAs increased lipid oxidation and reduced energy efficiency in subsarcolemmal mitochondria, and increase AMPK activation, reducing both endoplasmic reticulum and oxidative stress. Increased mitochondrial respiration was related to an increased mitochondriogenesis in FD skeletal muscle, as shown by the increase in PGC1-α and -β.

Conclusions

our data strengthened the association of high dietary ω3-PUFA intake with reduced mitochondrial energy efficiency in the skeletal muscle.     

Impact of Endoscopic Lung Volume Reduction on Right Ventricular Myocardial Function

Introduction

Endoscopic lung volume reduction (ELVR) provides a minimally invasive therapy for patients with severe lung emphysema. As its impact on right ventricular (R_tV) function is undefined, we examined the extent of R_tV functional changes following ELVR, as assessed by use of speckle tracking-based R_tV deformation analysis.

Methods

We enrolled 32 patients with severe emphysematous COPD scheduled for bronchoscopic LVR using endobronchial valves (Zephyr, PulmonX, Inc.), comprising 16 matched clinical responders and 16 non-responders. Echocardiography was conducted one day prior to ELVR and at an eight-week postprocedural interval.

Results

Patients were predominantly of late middle-age (65.8±8.7yrs), male (62.5%) and presented advanced COPD emphysema (means FEV1 and RV: 32.6% and 239.1% of predicted, respectively). After ELVR, R_tV apical longitudinal strain improved significantly in the total study cohort (-7.96±7.02% vs. -13.35±11.48%, p=0.04), whereas there were no significant changes in other parameters of R_tV function such as R_tV global longitudinal strain, TAPSE or pulmonary arterial systolic pressure. In responding patients, 6MWT-improvement correlated with a decrease in NT-proBNP (Pearson´s r: -0.53, p=0.03). However, clinical non-responders did not exhibit any R_tV functional improvement.

Discussion

ELVR beneficially impacts R_tV functional parameters. Speckle tracking-based R_tV apical longitudinal strain analysis allows early determination of R_tV contractile gain and identification of clinical responsiveness.     

Can Spectral CT Imaging Improve the Differentiation between Malignant and Benign Solitary Pulmonary Nodules?

Purpose

To quantitatively assess the value of dual-energy CT (DECT) in differentiating malignancy and benignity of solitary pulmonary nodules.

Materials and Methods

Sixty-three patients with solitary pulmonary nodules detected by CT plain scan underwent contrast enhanced CT scans in arterial phase (AP) and venous phase (VP) with spectral imaging mode for tumor type differentiation. The Gemstone Spectral Imaging (GSI) viewer was used for image display and data analysis. Region of interest was placed on the relatively homogeneous area of the nodule to measure iodine concentration (IC) on iodine-based material decomposition images and CT numbers on monochromatic image sets to generate spectral HU curve. Normalized IC (NIC), slope of the spectral HU curve (λ_HU) and net CT number enhancement on 70keV images were calculated. The two-sample t-test was used to compare quantitative parameters. Receiver operating characteristic curves were generated to calculate sensitivity and specificity.

Results

There were 63 nodules, with 37 malignant nodules (59%) and 26 benign nodules (41%). NIC, λ_HU and net CT number enhancement on 70keV images for malignant nodules were all greater than those of benign nodules. NIC and λ_HU had intermediate to high performances to differentiate malignant nodules from benign ones with the areas under curve of 0.89 and 0.86 respectively in AP, 0.96 and 0.89 respectively in VP. Using 0.30 as a threshold value for NIC in VP, one could obtain sensitivity of 93.8% and specificity of 85.7% for differentiating malignant from benign solitary pulmonary nodules. These values were statistically higher than the corresponding values of 74.2% and 53.8% obtained with the conventional CT number enhancement.

Conclusions

DECT imaging with GSI mode provides more promising value in quantitative way for distinguishing malignant nodules from benign ones than CT enhancement numbers.     

Tumor T1 Relaxation Time for Assessing Response to Bevacizumab Anti-Angiogenic Therapy in a Mouse Ovarian Cancer Model

Purpose

To assess whether T₁ relaxation time of tumors may be used to assess response to bevacizumab anti-angiogenic therapy. Procedures: 12 female nude mice bearing subcutaneous SKOV3ip1-LC ovarian tumors were administered bevacizumab (6.25ug/g, n=6) or PBS (control, n=6) therapy twice a week for two weeks. T₁ maps of tumors were generated before, two days, and 2 weeks after initiating therapy. Tumor weight was assessed by MR and at necropsy. Histology for microvessel density, proliferation, and apoptosis was performed.

Results

Bevacizumab treatment resulted in tumor growth inhibition (p<0.04, n=6), confirming therapeutic efficacy. Tumor T₁ relaxation times increased in bevacizumab treated mice 2 days and 2 weeks after initiating therapy (p<.05, n=6). Microvessel density decreased 59% and cell proliferation (Ki67+) decreased 50% in the bevacizumab treatment group (p<.001, n=6), but not apoptosis.

Conclusions

Findings suggest that increased tumor T₁ relaxation time is associated with response to bevacizumab therapy in ovarian cancer model and might serve as an early indicator of response.     

The Effects of Freezing on Faecal Microbiota as Determined Using MiSeq Sequencing and Culture-Based Investigations

Background

High-throughput sequencing has enabled detailed insights into complex microbial environments, including the human gut microbiota. The accuracy of the sequencing data however, is reliant upon appropriate storage of the samples prior to DNA extraction. The aim of this study was to conduct the first MiSeq sequencing investigation into the effects of faecal storage on the microbiota, compared to fresh samples. Culture-based analysis was also completed.

Methods

Seven faecal samples were collected from healthy adults. Samples were separated into fresh (DNA extracted immediately), snap frozen on dry ice and frozen for 7 days at -80°C prior to DNA extraction or samples frozen at -80°C for 7 days before DNA extraction. Sequencing was completed on the Illumina MiSeq platform. Culturing of total aerobes, anaerobes and bifidobacteria was also completed.

Results

No significant differences at phylum or family levels between the treatment groups occurred. At genus level only Faecalibacterium and Leuconostoc were significantly different in the fresh samples compared to the snap frozen group (p = 0.0298; p = 0.0330 respectively). Diversity analysis indicated that samples clustered based on the individual donor, rather than by storage group. No significant differences occurred in the culture-based analysis between the fresh, snap or -80°C frozen samples.

Conclusions

Using the MiSeq platform coupled with culture-based analysis, this study highlighted that limited significant changes in microbiota occur following rapid freezing of faecal samples prior to DNA extraction. Thus, rapid freezing of samples prior to DNA extraction and culturing, preserves the integrity of the microbiota.     

Care Needs and Clinical Outcomes of Older People with Dementia: A Population-Based Propensity Score-Matched Cohort Study

Objective

To explore the healthcare resource utilization, psychotropic drug use and mortality of older people with dementia.

Design

A nationwide propensity score-matched cohort study.

Setting

National Health Insurance Research database.

Participants

A total of 32,649 elderly people with dementia and their propensity-score matched controls (n=32,649).

Measurements

Outpatient visits, inpatient care, psychotropic drug use, in-hospital mortality and all-cause mortality at 90 and 365 days.

Results

Compared to the non-dementia group, a higher proportion of patients with dementia used inpatient services (1 year after index date: 20.91% vs. 9.55%), and the dementia group had more outpatient visits (median [standard deviation]: 7.00 [8.87] vs. 3.00 [8.30]). Furthermore, dementia cases with acute admission had the highest psychotropic drug utilization both at baseline and at the post-index dates (difference-in-differences: all <0.001). Dementia was associated with an increased risk of all-cause mortality (90 days, Odds ratio (OR)=1.85 [95%CI 1.67-2.05], p<0.001; 365 days, OR=1.59 [1.50-1.69], p<0.001) and in-hospital mortality (90 days, OR=1.97 [1.71-2.27], p<0.001; 365 days, OR=1.82 [1.61-2.05], p<0.001) compared to matched controls.

Conclusions

When older people with dementia are admitted for acute illnesses, they may increase their use of psychotropic agents and their risk of death, particularly in-hospital mortality.     

Dietary Inflammatory Index and Incidence of Cardiovascular Disease in the SUN Cohort

Background

Diet is known to play a key role in atherogenesis and in the development of cardiovascular events. Dietary factors may mediate these processes acting as potential modulators of inflammation. Potential Links between inflammatory properties of diet and the occurrence of cardiovascular events have not been tested previously.

Objective

We aimed to assess the association between the dietary inflammatory index (DII), a method to assess the inflammatory potential of the diet, and incident cardiovascular disease.

Methods

In the prospective, dynamic SUN cohort, 18,794 middle-aged, Spanish university graduates were followed up for 8.9 years (median). A validated 136-item food-frequency questionnaire was used to calculate the DII. The DII is based on scientific evidence about the relationship between diet and inflammatory biomarkers (C-reactive protein, IL-1β, IL-4, IL-6, IL-10 and TNF-α). Cox proportional hazard models were used to estimate hazard ratios (HR) and 95% confidence intervals (CI) for the association between the DII and incident cardiovascular disease (myocardial infarction, stroke or cardiovascular death).

Results

The risk for cardiovascular events progressively increased with each increasing quartile of DII (p_trend = 0.017). The multivariable-adjusted HR for participants in the highest (most pro-inflammatory) vs. the lowest quartile of the DII was 2.03 (95% CI 1.06–3.88).

Conclusions

A pro-inflammatory diet was associated with a significantly higher risk for developing cardiovascular events.     

Impaired Expression of Prostaglandin E2 (PGE2) Synthesis and Degradation Enzymes during Differentiation of Immortalized Urothelial Cells from Patients with Interstitial Cystitis/Painful Bladder Syndrome

Purpose

The differentiated superficial cells of the urothelium restrict urine flow into the bladder wall. We have demonstrated that urothelial cells isolated from bladders of patients with interstitial cystitis/painful bladder syndrome (IC/PBS) fail to release PGE₂ in response to tryptase. This study examines the expression of PGE₂ synthesis and degradation enzymes in urothelial cells during differentiation.

Materials and Methods

We measured immunoprotein expression of cyclooxygenase-2 (COX-2), prostaglandin E₂ synthase (PGES) and 15-hydroxyprostaglandin dehydrogenase (PGDH) in human urothelial cells and in immortalized urothelial cells isolated from the bladders of IC/PBS patients or normal subjects during stratification and differentiation produced by increased calcium and fetal bovine serum (Ca/FBS) in the culture medium for 1, 3 and 7 days.

Results

PGES immunoprotein expression increased during differentiation in normal and IC/PBS urothelial cells. COX-2 expression also increased in cells from normal patients following differentiation. Remarkably, no COX-2 expression was detectable in urothelial cells isolated from 3 out of 4 IC/PBS patients. PGDH immunoprotein expression decreased in normal cells after 1 and 3 days of Ca/FBS addition, but returned to normal after 7 days. PGDH expression was unchanged during differentiation at 1 and 3 days, but was more than 2-fold higher at 7 days compared to day 0 in the IC/PBS cells. Urothelial cells isolated from IC/PBS patients demonstrated no PGE₂ release in response to tryptase under any of the experimental conditions studied.

Conclusions

Taken together, our results indicate that PGE₂ release is compromised during stratification and differentiation in IC/PBS urothelium and may contribute to impaired barrier function.     

Single and Multiple Dose Pharmacokinetics,Pharmacodynamics and Safety of the Novel Lipoprotein-Associated Phospholipase A2 Enzyme Inhibitor Darapladib in Healthy Chinese Subjects: An Open Label Phase-1 Clinical Trial

Background and Objectives

Darapladib is a lipoprotein-associated phospholipase A₂ (Lp-PLA₂) inhibitor. This study evaluated the pharmacokinetics, pharmacodynamics and safety of darapladib in healthy Chinese subjects.

Methods

Twenty-four subjects received darapladib 160 mg orally, approximately 1 hour after a standard breakfast, as a single dose and once daily for 28 days. Non-compartmental methods were used to determine the single and multiple dose pharmacokinetics of darapladib and its metabolite SB-553253. Repeat dose Lp-PLA₂ activity and safety were evaluated.

Results

Systemic exposure (AUC_(0-T), Cmax geometric mean (CVb%)) of darapladib was higher after multiple-dosing (519 ng.h/mL (33.3%), 34.4 ng/mL (49.9%)) compared to single-dose administration (153 ng.h/mL (69.0%), 17.9 ng/mL (55.2%). The steady-state accumulation ratio was less than unity (Rs = 0.80), indicating time-dependent pharmacokinetics of darapladib. Darapladib steady-state was reached by Day 14 of once daily dosing. Systemic exposure to SB-553253 was lower than darapladib with median (SB-553253: darapladib) ratios for AUC_(0-τ) of 0.0786 for single dose and 0.0532 for multiple dose administration. On Day 28, pre-dose and maximum inhibition of Lp-PLA₂ activity was approximately 70% and 75% relative to the baseline value, respectively and was dependent of darapladib concentration. The most common adverse events (≥ 21% subjects) were abnormal faeces, abnormal urine odour, diarrhoea and nasopharyngitis.

Conclusion

Darapladib 160 mg single and repeat doses were profiled in healthy Chinese subjects. Single dose systemic exposure to darapladib in healthy Chinese subjects was consistent with that observed previously in Western subjects whereas steady-state systemic exposure was approximately 65% higher in Chinese than Western subjects. The Lp-PLA₂ activity and adverse event profile were similar in healthy Chinese and previous reports in Western subjects. Ethnic-specific dose adjustment of darapladib is not considered necessary for the Chinese population.

Trial Registration

ClinicalTrials.gov NCT02000804     

Endothelial-Derived Oxidative Stress Drives Myofibroblastic Activation and Calcification of the Aortic Valve

Aims

Oxidative stress is present in and contributes to calcification of the aortic valve, but the driving factors behind the initiation of valve oxidative stress are not well understood. We tested whether the valve endothelium acts as an initiator and propagator of oxidative stress in aortic valve disease.

Methods and Results

Calcified human aortic valves showed side-specific elevation of superoxide in the endothelium, co-localized with high VCAM1 expression, linking oxidative stress, inflammation, and valve degeneration. Treatment with inflammatory cytokine TNFα increased superoxide and oxidative stress and decreased eNOS and VE-cadherin acutely over 48 hours in aortic valve endothelial cells (VEC) and chronically over 21 days in ex vivo AV leaflets. Co-treatment of VEC with tetrahydrobiopterin (BH₄) but not apocynin mitigated TNFα-driven VEC oxidative stress. Co-treatment of ex vivo AV leaflets with TNFα+BH₄ or TNFα+peg-SOD rescued endothelial function and mitigated inflammatory responses. Both BH₄ and peg-SOD rescued valve leaflets from the pro-osteogenic effects of TNFα treatment, but only peg-SOD was able to mitigate the fibrogenic effects, including increased collagen and αSMA expression.

Conclusions

Aortic valve endothelial cells are a novel source of oxidative stress in aortic valve disease. TNFα-driven VEC oxidative stress causes loss of endothelial protective function, chronic inflammation, and fibrogenic and osteogenic activation, mitigated differentially by BH₄ and peg-SOD. These mechanisms identify new targets for tailored antioxidant therapy focused on mitigation of oxidative stress and restoration of endothelial protection.     

Reliability and Validity of Dual-Task Mobility Assessments in People with Chronic Stroke

Background

The ability to perform a cognitive task while walking simultaneously (dual-tasking) is important in real life. However, the psychometric properties of dual-task walking tests have not been well established in stroke.

Objective

To assess the test-retest reliability, concurrent and known-groups validity of various dual-task walking tests in people with chronic stroke.

Design

Observational measurement study with a test-retest design.

Methods

Eighty-eight individuals with chronic stroke participated. The testing protocol involved four walking tasks (walking forward at self-selected and maximal speed, walking backward at self-selected speed, and crossing over obstacles) performed simultaneously with each of the three attention-demanding tasks (verbal fluency, serial 3 subtractions or carrying a cup of water). For each dual-task condition, the time taken to complete the walking task, the correct response rate (CRR) of the cognitive task, and the dual-task effect (DTE) for the walking time and CRR were calculated. Forty-six of the participants were tested twice within 3–4 days to establish test-retest reliability.

Results

The walking time in various dual-task assessments demonstrated good to excellent reliability [Intraclass correlation coefficient (ICC_2,1) = 0.70–0.93; relative minimal detectable change at 95% confidence level (MDC₉₅%) = 29%-45%]. The reliability of the CRR (ICC_2,1 = 0.58–0.81) and the DTE in walking time (ICC_2,1 = 0.11–0.80) was more varied. The reliability of the DTE in CRR (ICC_2,1 = -0.31–0.40) was poor to fair. The walking time and CRR obtained in various dual-task walking tests were moderately to strongly correlated with those of the dual-task Timed-up-and-Go test, thus demonstrating good concurrent validity. None of the tests could discriminate fallers (those who had sustained at least one fall in the past year) from non-fallers.

Limitation

The results are generalizable to community-dwelling individuals with chronic stroke only.

Conclusions

The walking time derived from the various dual-task assessments generally demonstrated good to excellent reliability, making them potentially useful in clinical practice and future research endeavors. However, the usefulness of these measurements in predicting falls needs to be further explored. Relatively low reliability was shown in the cognitive outcomes and DTE, which may not be preferred measurements for assessing dual-task performance.     

Use of Fibrates Monotherapy in People with Diabetes and High Cardiovascular Risk in Primary Care: A French Nationwide Cohort Study Based on National Administrative Databases

Background and Aim

According to guidelines, diabetic patients with high cardiovascular risk should receive a statin. Despite this consensus, fibrate monotherapy is commonly used in this population. We assessed the frequency and clinical consequences of the use of fibrates for primary prevention in patients with diabetes and high cardiovascular risk.

Design

Retrospective cohort study based on nationwide data from the medical and administrative databases of French national health insurance systems (07/01/08-12/31/09) with a follow-up of up to 30 months.

Methods

Lipid-lowering drug-naive diabetic patients initiating fibrate or statin monotherapy were identified. Patients at high cardiovascular risk were then selected: patients with a diagnosis of diabetes and hypertension, and >50 (men) or 60 (women), but with no history of cardiovascular events. The composite endpoint comprised myocardial infarction, stroke, amputation, or death.

Results

Of the 31,652 patients enrolled, 4,058 (12.8%) received a fibrate. Age- and gender-adjusted annual event rates were 2.42% (fibrates) and 2.21% (statins). The proportionality assumption required for the Cox model was not met for the fibrate/statin variable. A multivariate model including all predictors was therefore calculated by dividing data into two time periods, allowing Hazard Ratios to be calculated before (HR_<540) and after 540 days (HR_>540) of follow-up. Multivariate analyses showed that fibrates were associated with an increased risk for the endpoint after 540 days: HR_<540 = 0.95 (95% CI: 0.78–1.16) and HR_>540 = 1.73 (1.28–2.32).

Conclusion

Fibrate monotherapy is commonly prescribed in diabetic patients with high cardiovascular risk and is associated with poorer outcomes compared to statin therapy.     

Center-Related Determinants of VKA Anticoagulation Quality: A Prospective,Multicenter Evaluation

Background

Center-specific TTR (c-TTR) is a measure reporting the mean patient TTR within an anticoagulation clinic describing the quality of anticoagulant monitoring offered by that clinic. c-TTR has a considerable between-center variation, but its determinants are poorly understood.

Objectives

We aimed at evaluating which clinical, procedural or laboratory factors could be associated with c-TTR variability in a multicenter, observational cross-sectional study over a five-year period.

Patients/Methods

Data from 832,204 individual patients followed for VKA therapy in 292 Centers affiliated with the Italian Federation of Anticoagulation Clinics (FCSA) were analyzed. c-TTR was computed based on the TTR of patients followed at each Center, and a mixed linear regression model was used for a predefined set of explanatory variables.

Results

The Center next-visit interval ratio (the mean number of days after a visit with an INR outside the therapeutic range, divided by the days after a visit with an INR within the therapeutic range), the Center mean patient INR and the Center laboratory performance at EQA proficiency testing were the only variables that were independently associated with c-TTR (β-coefficients -17.32, 9.67, and -0.11, respectively; r ² = 0.635).

Conclusions

These findings suggest that c-TTR associates with proactive strategies aimed at keeping patients very close to their target INR with a prompt re-evaluation of those patients with under- or over-therapeutic INR.