Quantification of nitrotyrosine in nitrated proteins

For kinetic studies of protein nitration reactions, we have developed a method for the quantification of nitrotyrosine residues in protein molecules by liquid chromatography coupled to a diode array detector of ultraviolet-visible absorption. Nitrated bovine serum albumin (BSA) and nitrated ovalbumin (OVA) were synthesized and used as standards for the determination of the protein nitration degree (ND), which is defined as the average number of nitrotyrosine residues divided by the total number of tyrosine residues in a protein molecule. The obtained calibration curves of the ratio of chromatographic peak areas of absorbance at 357 and at 280 nm vs. nitration degree are nearly the same for BSA and OVA (relative deviations <5%). They are near-linear at low ND (< 0.1) and can be described by a second-order polynomial fit up to \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$${\hbox{ND}} = 0.5\left( {{R^2} > 0.99} \right)$$\end{document}. A change of chromatographic column led to changes in absolute peak areas but not in the peak area ratios and related calibration functions, which confirms the robustness of the analytical method. First results of laboratory experiments confirm that the method is applicable for the investigation of the reaction kinetics of protein nitration. The main advantage over alternative methods is that nitration degrees can be efficiently determined without hydrolysis or digestion of the investigated protein molecules

A benzene-degrading nitrate-reducing microbial consortium displays aerobic and anaerobic benzene degradation pathways

All sequence data from this study were deposited at the European Bioinformatics Institute under the accession numbers ERS1670018 to ERS1670023. Further, all assigned genes, taxonomy, function, sequences of contigs, genes and proteins can be found in Table S3.In this study, we report transcription of genes involved in aerobic and anaerobic benzene degradation pathways in a benzene-degrading denitrifying continuous culture. Transcripts associated with the family Peptococcaceae dominated all samples (2136% relative abundance) indicating their key role in the community. We found a highly transcribed gene cluster encoding a presumed anaerobic benzene carboxylase (AbcA and AbcD) and a benzoate-coenzyme A ligase (BzlA). Predicted gene products showed >96% amino acid identity and similar gene order to the corresponding benzene degradation gene cluster described previously, providing further evidence for anaerobic benzene activation via carboxylation. For subsequent benzoyl-CoA dearomatization, bam-like genes analogous to the ones found in other strict anaerobes were transcribed, whereas gene transcripts involved in downstream benzoyl-CoA degradation were mostly analogous to the ones described in facultative anaerobes. The concurrent transcription of genes encoding enzymes involved in oxygenase-mediated aerobic benzene degradation suggested oxygen presence in the culture, possibly formed via a recently identified nitric oxide dismutase (Nod). Although we were unable to detect transcription of Nod-encoding genes, addition of nitrite and formate to the continuous culture showed indication for oxygen production. Such an oxygen production would enable aerobic microbes to thrive in oxygen-depleted and nitrate-containing subsurface environments contaminated with hydrocarbons.This study was supported by a grant of BE-Basic-FES funds from the Dutch Ministry of Economic Affairs. The research of A.J.M. Stams is supported by an ERC grant (project 323009) and the gravitation grant “Microbes for Health and Environment” (project 024.002.002) of the Netherlands Ministry of Education, Culture and Science. F. Hugenholtz was supported by the same gravitation grant (project 024.002.002). B. Hornung is supported by Wageningen University and the Wageningen Institute for Environment and Climate Research (WIMEK) through the IP/OP program Systems Biology (project KB-17-003.02-023).info:eu-repo/semantics/publishedVersio

Outcomes of early switching from intravenous to oral antibiotics on medical wards

OBJECTIVES: To evaluate outcomes following implementation of a checklist with criteria for switching from intravenous (iv) to oral antibiotics on unselected patients on two general medical wards. METHODS: During a 12 month intervention study, a printed checklist of criteria for switching on the third day of iv treatment was placed in the medical charts. The decision to switch was left to the discretion of the attending physician. Outcome parameters of a 4 month control phase before intervention were compared with the equivalent 4 month period during the intervention phase to control for seasonal confounding (before-after study; April to July of 2006 and 2007, respectively): 250 episodes (215 patients) during the intervention period were compared with the control group of 176 episodes (162 patients). The main outcome measure was the duration of iv therapy. Additionally, safety, adherence to the checklist, reasons against switching patients and antibiotic cost were analysed during the whole year of the intervention (n = 698 episodes). RESULTS: In 38% (246/646) of episodes of continued iv antibiotic therapy, patients met all criteria for switching to oral antibiotics on the third day, and 151/246 (61.4%) were switched. The number of days of iv antibiotic treatment were reduced by 19% (95% confidence interval 9%-29%, P = 0.001; 6.0-5.0 days in median) with no increase in complications. The main reasons against switching were persisting fever (41%, n = 187) and absence of clinical improvement (41%, n = 185). CONCLUSIONS: On general medical wards, a checklist with bedside criteria for switching to oral antibiotics can shorten the duration of iv therapy without any negative effect on treatment outcome. The criteria were successfully applied to all patients on the wards, independently of the indication (empirical or directed treatment), the type of (presumed) infection, the underlying disease or the group of antibiotics being used

Regulatory network modelling of iron acquisition by a fungal pathogen in contact with epithelial cells

Peer reviewedPublisher PD

(Per)chlorate reduction by an acetogenic bacterium, Sporomusa sp., isolated from an underground gas storage

A mesophilic bacterium, strain An4, was isolated from an underground gas storage reservoir with methanol as substrate and perchlorate as electron acceptor. Cells were Gram-negative, spore-forming, straight to curved rods, 0.5–0.8 μm in diameter, and 2–8 μm in length, growing as single cells or in pairs. The cells grew optimally at 37°C, and the pH optimum was around 7. Strain An4 converted various alcohols, organic acids, fructose, acetoin, and H2/CO2 to acetate, usually as the only product. Succinate was decarboxylated to propionate. The isolate was able to respire with (per)chlorate, nitrate, and CO2. The G+C content of the DNA was 42.6 mol%. Based on the 16S rRNA gene sequence analysis, strain An4 was most closely related to Sporomusa ovata (98% similarity). The bacterium reduced perchlorate and chlorate completely to chloride. Key enzymes, perchlorate reductase and chlorite dismutase, were detected in cell-free extracts

Temporal changes in the epidemiology, management, and outcome from acute respiratory distress syndrome in European intensive care units: a comparison of two large cohorts

Background: Mortality rates for patients with ARDS remain high. We assessed temporal changes in the epidemiology and management of ARDS patients requiring invasive mechanical ventilation in European ICUs. We also investigated the association between ventilatory settings and outcome in these patients. Methods: This was a post hoc analysis of two cohorts of adult ICU patients admitted between May 1–15, 2002 (SOAP study, n = 3147), and May 8–18, 2012 (ICON audit, n = 4601 admitted to ICUs in the same 24 countries as the SOAP study). ARDS was defined retrospectively using the Berlin definitions. Values of tidal volume, PEEP, plateau pressure, and FiO2 corresponding to the most abnormal value of arterial PO2 were recorded prospectively every 24&nbsp;h. In both studies, patients were followed for outcome until death, hospital discharge or for 60&nbsp;days. Results: The frequency of ARDS requiring mechanical ventilation during the ICU stay was similar in SOAP and ICON (327[10.4%] vs. 494[10.7%], p = 0.793). The diagnosis of ARDS was established at a median of 3 (IQ: 1–7) days after admission in SOAP and 2 (1–6) days in ICON. Within 24&nbsp;h of diagnosis, ARDS was mild in 244 (29.7%), moderate in 388 (47.3%), and severe in 189 (23.0%) patients. In patients with ARDS, tidal volumes were lower in the later (ICON) than in the earlier (SOAP) cohort. Plateau and driving pressures were also lower in ICON than in SOAP. ICU (134[41.1%] vs 179[36.9%]) and hospital (151[46.2%] vs 212[44.4%]) mortality rates in patients with ARDS were similar in SOAP and ICON. High plateau pressure (&gt; 29 cmH2O) and driving pressure (&gt; 14 cmH2O) on the first day of mechanical ventilation but not tidal volume (&gt; 8&nbsp;ml/kg predicted body weight [PBW]) were independently associated with a higher risk of in-hospital death. Conclusion: The frequency of and outcome from ARDS remained relatively stable between 2002 and 2012. Plateau pressure &gt; 29 cmH2O and driving pressure &gt; 14 cmH2O on the first day of mechanical ventilation but not tidal volume &gt; 8&nbsp;ml/kg PBW were independently associated with a higher risk of death. These data highlight the continued burden of ARDS and provide hypothesis-generating data for the design of future studies

Computational Homogenization of Architectured Materials

Architectured materials involve geometrically engineered distributions of microstructural phases at a scale comparable to the scale of the component, thus calling for new models in order to determine the effective properties of materials. The present chapter aims at providing such models, in the case of mechanical properties. As a matter of fact, one engineering challenge is to predict the effective properties of such materials; computational homogenization using finite element analysis is a powerful tool to do so. Homogenized behavior of architectured materials can thus be used in large structural computations, hence enabling the dissemination of architectured materials in the industry. Furthermore, computational homogenization is the basis for computational topology optimization which will give rise to the next generation of architectured materials. This chapter covers the computational homogenization of periodic architectured materials in elasticity and plasticity, as well as the homogenization and representativity of random architectured materials

Effects of Once-Weekly Exenatide on Cardiovascular Outcomes in Type 2 Diabetes.

Abstract BACKGROUND: The cardiovascular effects of adding once-weekly treatment with exenatide to usual care in patients with type 2 diabetes are unknown. METHODS: We randomly assigned patients with type 2 diabetes, with or without previous cardiovascular disease, to receive subcutaneous injections of extended-release exenatide at a dose of 2 mg or matching placebo once weekly. The primary composite outcome was the first occurrence of death from cardiovascular causes, nonfatal myocardial infarction, or nonfatal stroke. The coprimary hypotheses were that exenatide, administered once weekly, would be noninferior to placebo with respect to safety and superior to placebo with respect to efficacy. RESULTS: In all, 14,752 patients (of whom 10,782 [73.1%] had previous cardiovascular disease) were followed for a median of 3.2 years (interquartile range, 2.2 to 4.4). A primary composite outcome event occurred in 839 of 7356 patients (11.4%; 3.7 events per 100 person-years) in the exenatide group and in 905 of 7396 patients (12.2%; 4.0 events per 100 person-years) in the placebo group (hazard ratio, 0.91; 95% confidence interval [CI], 0.83 to 1.00), with the intention-to-treat analysis indicating that exenatide, administered once weekly, was noninferior to placebo with respect to safety (P<0.001 for noninferiority) but was not superior to placebo with respect to efficacy (P=0.06 for superiority). The rates of death from cardiovascular causes, fatal or nonfatal myocardial infarction, fatal or nonfatal stroke, hospitalization for heart failure, and hospitalization for acute coronary syndrome, and the incidence of acute pancreatitis, pancreatic cancer, medullary thyroid carcinoma, and serious adverse events did not differ significantly between the two groups. CONCLUSIONS: Among patients with type 2 diabetes with or without previous cardiovascular disease, the incidence of major adverse cardiovascular events did not differ significantly between patients who received exenatide and those who received placebo. (Funded by Amylin Pharmaceuticals; EXSCEL ClinicalTrials.gov number, NCT01144338 .)