Otimização do envelhecimento da poliamida 11 em produtos químicos de injeção submarina

Nos sistemas de produção de petróleo offshore a injeção submarina de produtos químicos é necessária para garantir a integridade dos componentes do sistema e melhorar produtividade da extração. Devido ao alto custo associado a qualquer manutenção no sistema de injeção, fabricantes de produtos químicos necessitam cumprir exigentes testes de qualificação nos seus produtos. Um dos testes, o de compatibilidade com materiais não metálicos, exige análise das alterações das propriedades de materiais poliméricos após envelhecidos nos produtos químicos por 30, 60 e 90 dias. O longo período de tempo necessário neste teste dificulta o desenvolvimento de novos produtos químicos e gera altos custos com equipamentos. O trabalho visou otimizar o tempo de ensaio de imersão necessário para avaliação do envelhecimento da Poliamida 11. Através do processo de injeção produziu-se diversos corpos de prova de poliamida 11, que foram envelhecidos em diferentes produtos químicos por 15 e 30 dias. Para analisar as alterações causados pelo envelhecimento, as amostras foram avaliadas quanto a variação de massa e volume, alterações das propriedades mecânicas e análise térmica de DSC. Para comprovar a veracidade dos resultados obtidos e avaliar possíveis tendências dos efeitos com o tempo de envelhecimento, os valores obtidos foram comparados com os resultados de ensaios de imersão por 30, 60 e 90 dias. Foi possível concluir que realizando ensaio de DSC em amostras de Poliamida 11 após quinze dias de envelhecimento, já é possível identificar se o produto químico ataca agressivamente o material, evitando a necessidade de um longo tempo de imersão para se obter a mesma conclusão

Solid-state NMR spectroscopy of 10% 13C labeled ubiquitin: spectral simplification and stereospecific assignment of isopropyl groups

We describe the simplification of 13C-13C correlation spectra obtained from a microcrystalline protein sample expressed on a growth medium of 10% fully 13C labeled glucose diluted in 90% natural abundance glucose as compared to a fully labeled sample. Such a labeling scheme facilitates the backbone and side-chain resonance assignment of Phe, Tyr, His, Asp, Asn, Ile, Lys and Pro and yields an unambiguous stereospecific assignment of the valine Cγ1, Cγ2 13C resonances and of Leucine Cδ

Efficient production of a functional G protein-coupled receptor in E. coli for structural studies

G protein-coupled receptors (GPCRs) are transmembrane signal transducers which regulate many key physiological process. Since their discovery, their analysis has been limited by difficulties in obtaining sufficient amounts of the receptors in high-quality, functional form from heterologous expression hosts. Albeit highly attractive because of its simplicity and the ease of isotope labeling for NMR studies, heterologous expression of functional GPCRs in E. coli has proven particularly challenging due to the absence of the more evolved protein expression and folding machinery of higher eukaryotic hosts. Here we first give an overview on the previous strategies for GPCR E. coli expression and then describe the development of an optimized robust protocol for the E. coli expression and purification of two mutants of the turkey β 1 -adrenergic receptor (β 1 AR) uniformly or selectively labeled in 15 N or 2 H, 15 N. These mutants had been previously optimized for thermal stability using insect cell expression and used successfully in crystallographic and NMR studies. The same sequences were then used for E. coli expression. Optimization of E. coli expression was achieved by a quantitative analysis of losses of receptor material at each step of the solubilization and purification procedure. Final yields are 0.2-0.3 mg receptor per liter culture. Whereas both expressed mutants are well folded and competent for orthosteric ligand binding, the less stable YY-β 1 AR mutant also comprises the two native tyrosines Y 5.58 and Y 7.53 , which enable G protein binding. High-quality 1 H- 15 N TROSY spectra were obtained for E. coli -expressed YY-β 1 AR in three different functional states (antagonist, agonist, and agonist + G protein-mimicking nanobody-bound), which are identical to spectra obtained of the same forms of the receptor expressed in insect cells. NdeI and AgeI restriction sites introduced into the expression plasmid allow for the easy replacement of the receptor gene by other GPCR genes of interest, and the provided quantitative workflow analysis may guide the respective adaptation of the purification protocol

Unraveling the complexity of protein backbone dynamics with combined 13C and 15N solid-state NMR relaxation measurements

Typically, protein dynamics involve a complex hierarchy of motions occurring on different time scales between conformations separated by a range of different energy barriers. NMR relaxation can in principle provide a site-specific picture of both the time scales and amplitudes of these motions, but independent relaxation rates sensitive to fluctuations in different time scale ranges are required to obtain a faithful representation of the underlying dynamic complexity. This is especially pertinent for relaxation measurements in the solid state, which report on dynamics in a broader window of time scales by more than 3 orders of magnitudes compared to solution NMR relaxation. To aid in unraveling the intricacies of biomolecular dynamics we introduce 13C spin–lattice relaxation in the rotating frame (R1ρ) as a probe of backbone nanosecond-microsecond motions in proteins in the solid state. We present measurements of 13C′ R1ρ rates in fully protonated crystalline protein GB1 at 600 and 850 MHz 1H Larmor frequencies and compare them to 13C′ R1, 15N R1 and R1ρ measured under the same conditions. The addition of carbon relaxation data to the model free analysis of nitrogen relaxation data leads to greatly improved characterization of time scales of protein backbone motions, minimizing the occurrence of fitting artifacts that may be present when 15N data is used alone. We also discuss how internal motions characterized by different time scales contribute to 15N and 13C relaxation rates in the solid state and solution state, leading to fundamental differences between them, as well as phenomena such as underestimation of picosecond-range motions in the solid state and nanosecond-range motions in solution

Metabolic reprogramming by 3-Iodothyronamine (T1AM): a new perspective to reverse obesity through co-regulation of sirtuin 4 and 6 expression

Obesity is a complex disease associated with environmental and genetic factors. 3-Iodothyronamine (T1AM) has revealed great potential as an effective weight loss drug. We used metabolomics and associated transcriptional gene and protein expression analysis to investigate the tissue specific metabolic reprogramming effects of subchronic T1AM treatment at two pharmacological daily doses (10 and 25 mg/kg) on targeted metabolic pathways. Multi-analytical results indicated that T1AM at 25 mg/kg can act as a novel master regulator of both glucose and lipid metabolism in mice through sirtuin-mediated pathways. In liver, we observed an increased gene and protein expression of Sirt6 (a master gene regulator of glucose) and Gck (glucose kinase) and a decreased expression of Sirt4 (a negative regulator of fatty acids oxidation (FAO)), whereas in white adipose tissue only Sirt6 was increased. Metabolomics analysis supported physiological changes at both doses with most increases in FAO, glycolysis indicators and the mitochondrial substrate, at the highest dose of T1AM. Together our results suggest that T1AM acts through sirtuin-mediated pathways to metabolically reprogram fatty acid and glucose metabolism possibly through small molecules signaling. Our novel mechanistic findings indicate that T1AM has a great potential as a drug for the treatment of obesity and possibly diabetes

Which Lynch syndrome screening programs could be implemented in the "real world"? A systematic review of economic evaluations

Purpose: Lynch syndrome (LS) screening can significantly reduce cancer morbidity and mortality in mutation carriers. Our aim was to identify cost-effective LS screening programs that can be implemented in the "real world."Methods: We performed a systematic review of full economic evaluations of genetic screening for LS in different target populations; health outcomes were estimated in life-years gained or quality-adjusted life-years.Results: Overall, 20 studies were included in the systematic review. Based on the study populations, we identified six categories of LS screening program: colorectal cancer (CRC)-based, endometrial cancer-based, general population-based, LS family registry-based, cascade testing-based, and genetics clinic-based screening programs. We performed an in-depth analysis of CRC-based LS programs, classifying them into three additional subcategories: universal, age-targeted, and selective. In five studies, universal programs based on immunohistochemistry, either alone or in combination with the BRAF test, were cost-effective compared with no screening, while in two studies age-targeted programs with a cutoff of 70 years were cost-effective when compared with age-targeted programs with lower age thresholds. Conclusion: Universal or <70 years-age-targeted CRC-based LS screening programs are cost-effective and should be implemented in the "real world

Measuring routine childhood vaccination coverage in 204 countries and territories, 1980-2019 : a systematic analysis for the Global Burden of Disease Study 2020, Release 1

Background Measuring routine childhood vaccination is crucial to inform global vaccine policies and programme implementation, and to track progress towards targets set by the Global Vaccine Action Plan (GVAP) and Immunization Agenda 2030. Robust estimates of routine vaccine coverage are needed to identify past successes and persistent vulnerabilities. Drawing from the Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2020, Release 1, we did a systematic analysis of global, regional, and national vaccine coverage trends using a statistical framework, by vaccine and over time. Methods For this analysis we collated 55 326 country-specific, cohort-specific, year-specific, vaccine-specific, and dosespecific observations of routine childhood vaccination coverage between 1980 and 2019. Using spatiotemporal Gaussian process regression, we produced location-specific and year-specific estimates of 11 routine childhood vaccine coverage indicators for 204 countries and territories from 1980 to 2019, adjusting for biases in countryreported data and reflecting reported stockouts and supply disruptions. We analysed global and regional trends in coverage and numbers of zero-dose children (defined as those who never received a diphtheria-tetanus-pertussis [DTP] vaccine dose), progress towards GVAP targets, and the relationship between vaccine coverage and sociodemographic development. Findings By 2019, global coverage of third-dose DTP (DTP3; 81.6% [95% uncertainty interval 80.4-82 .7]) more than doubled from levels estimated in 1980 (39.9% [37.5-42.1]), as did global coverage of the first-dose measles-containing vaccine (MCV1; from 38.5% [35.4-41.3] in 1980 to 83.6% [82.3-84.8] in 2019). Third- dose polio vaccine (Pol3) coverage also increased, from 42.6% (41.4-44.1) in 1980 to 79.8% (78.4-81.1) in 2019, and global coverage of newer vaccines increased rapidly between 2000 and 2019. The global number of zero-dose children fell by nearly 75% between 1980 and 2019, from 56.8 million (52.6-60. 9) to 14.5 million (13.4-15.9). However, over the past decade, global vaccine coverage broadly plateaued; 94 countries and territories recorded decreasing DTP3 coverage since 2010. Only 11 countries and territories were estimated to have reached the national GVAP target of at least 90% coverage for all assessed vaccines in 2019. Interpretation After achieving large gains in childhood vaccine coverage worldwide, in much of the world this progress was stalled or reversed from 2010 to 2019. These findings underscore the importance of revisiting routine immunisation strategies and programmatic approaches, recentring service delivery around equity and underserved populations. Strengthening vaccine data and monitoring systems is crucial to these pursuits, now and through to 2030, to ensure that all children have access to, and can benefit from, lifesaving vaccines. Copyright (C) 2021 The Author(s). Published by Elsevier Ltd.Peer reviewe

Why Are Outcomes Different for Registry Patients Enrolled Prospectively and Retrospectively? Insights from the Global Anticoagulant Registry in the FIELD-Atrial Fibrillation (GARFIELD-AF).

Background: Retrospective and prospective observational studies are designed to reflect real-world evidence on clinical practice, but can yield conflicting results. The GARFIELD-AF Registry includes both methods of enrolment and allows analysis of differences in patient characteristics and outcomes that may result. Methods and Results: Patients with atrial fibrillation (AF) and ≥1 risk factor for stroke at diagnosis of AF were recruited either retrospectively (n = 5069) or prospectively (n = 5501) from 19 countries and then followed prospectively. The retrospectively enrolled cohort comprised patients with established AF (for a least 6, and up to 24 months before enrolment), who were identified retrospectively (and baseline and partial follow-up data were collected from the emedical records) and then followed prospectively between 0-18 months (such that the total time of follow-up was 24 months; data collection Dec-2009 and Oct-2010). In the prospectively enrolled cohort, patients with newly diagnosed AF (≤6 weeks after diagnosis) were recruited between Mar-2010 and Oct-2011 and were followed for 24 months after enrolment. Differences between the cohorts were observed in clinical characteristics, including type of AF, stroke prevention strategies, and event rates. More patients in the retrospectively identified cohort received vitamin K antagonists (62.1% vs. 53.2%) and fewer received non-vitamin K oral anticoagulants (1.8% vs . 4.2%). All-cause mortality rates per 100 person-years during the prospective follow-up (starting the first study visit up to 1 year) were significantly lower in the retrospective than prospectively identified cohort (3.04 [95% CI 2.51 to 3.67] vs . 4.05 [95% CI 3.53 to 4.63]; p = 0.016). Conclusions: Interpretations of data from registries that aim to evaluate the characteristics and outcomes of patients with AF must take account of differences in registry design and the impact of recall bias and survivorship bias that is incurred with retrospective enrolment. Clinical Trial Registration: - URL: http://www.clinicaltrials.gov . Unique identifier for GARFIELD-AF (NCT01090362)

Risk profiles and one-year outcomes of patients with newly diagnosed atrial fibrillation in India: Insights from the GARFIELD-AF Registry.

BACKGROUND: The Global Anticoagulant Registry in the FIELD-Atrial Fibrillation (GARFIELD-AF) is an ongoing prospective noninterventional registry, which is providing important information on the baseline characteristics, treatment patterns, and 1-year outcomes in patients with newly diagnosed non-valvular atrial fibrillation (NVAF). This report describes data from Indian patients recruited in this registry. METHODS AND RESULTS: A total of 52,014 patients with newly diagnosed AF were enrolled globally; of these, 1388 patients were recruited from 26 sites within India (2012-2016). In India, the mean age was 65.8 years at diagnosis of NVAF. Hypertension was the most prevalent risk factor for AF, present in 68.5% of patients from India and in 76.3% of patients globally (P < 0.001). Diabetes and coronary artery disease (CAD) were prevalent in 36.2% and 28.1% of patients as compared with global prevalence of 22.2% and 21.6%, respectively (P < 0.001 for both). Antiplatelet therapy was the most common antithrombotic treatment in India. With increasing stroke risk, however, patients were more likely to receive oral anticoagulant therapy [mainly vitamin K antagonist (VKA)], but average international normalized ratio (INR) was lower among Indian patients [median INR value 1.6 (interquartile range {IQR}: 1.3-2.3) versus 2.3 (IQR 1.8-2.8) (P < 0.001)]. Compared with other countries, patients from India had markedly higher rates of all-cause mortality [7.68 per 100 person-years (95% confidence interval 6.32-9.35) vs 4.34 (4.16-4.53), P < 0.0001], while rates of stroke/systemic embolism and major bleeding were lower after 1 year of follow-up. CONCLUSION: Compared to previously published registries from India, the GARFIELD-AF registry describes clinical profiles and outcomes in Indian patients with AF of a different etiology. The registry data show that compared to the rest of the world, Indian AF patients are younger in age and have more diabetes and CAD. Patients with a higher stroke risk are more likely to receive anticoagulation therapy with VKA but are underdosed compared with the global average in the GARFIELD-AF. CLINICAL TRIAL REGISTRATION-URL: http://www.clinicaltrials.gov. Unique identifier: NCT01090362

Improved risk stratification of patients with atrial fibrillation: an integrated GARFIELD-AF tool for the prediction of mortality, stroke and bleed in patients with and without anticoagulation.

OBJECTIVES: To provide an accurate, web-based tool for stratifying patients with atrial fibrillation to facilitate decisions on the potential benefits/risks of anticoagulation, based on mortality, stroke and bleeding risks. DESIGN: The new tool was developed, using stepwise regression, for all and then applied to lower risk patients. C-statistics were compared with CHA2DS2-VASc using 30-fold cross-validation to control for overfitting. External validation was undertaken in an independent dataset, Outcome Registry for Better Informed Treatment of Atrial Fibrillation (ORBIT-AF). PARTICIPANTS: Data from 39 898 patients enrolled in the prospective GARFIELD-AF registry provided the basis for deriving and validating an integrated risk tool to predict stroke risk, mortality and bleeding risk. RESULTS: The discriminatory value of the GARFIELD-AF risk model was superior to CHA2DS2-VASc for patients with or without anticoagulation. C-statistics (95% CI) for all-cause mortality, ischaemic stroke/systemic embolism and haemorrhagic stroke/major bleeding (treated patients) were: 0.77 (0.76 to 0.78), 0.69 (0.67 to 0.71) and 0.66 (0.62 to 0.69), respectively, for the GARFIELD-AF risk models, and 0.66 (0.64-0.67), 0.64 (0.61-0.66) and 0.64 (0.61-0.68), respectively, for CHA2DS2-VASc (or HAS-BLED for bleeding). In very low to low risk patients (CHA2DS2-VASc 0 or 1 (men) and 1 or 2 (women)), the CHA2DS2-VASc and HAS-BLED (for bleeding) scores offered weak discriminatory value for mortality, stroke/systemic embolism and major bleeding. C-statistics for the GARFIELD-AF risk tool were 0.69 (0.64 to 0.75), 0.65 (0.56 to 0.73) and 0.60 (0.47 to 0.73) for each end point, respectively, versus 0.50 (0.45 to 0.55), 0.59 (0.50 to 0.67) and 0.55 (0.53 to 0.56) for CHA2DS2-VASc (or HAS-BLED for bleeding). Upon validation in the ORBIT-AF population, C-statistics showed that the GARFIELD-AF risk tool was effective for predicting 1-year all-cause mortality using the full and simplified model for all-cause mortality: C-statistics 0.75 (0.73 to 0.77) and 0.75 (0.73 to 0.77), respectively, and for predicting for any stroke or systemic embolism over 1 year, C-statistics 0.68 (0.62 to 0.74). CONCLUSIONS: Performance of the GARFIELD-AF risk tool was superior to CHA2DS2-VASc in predicting stroke and mortality and superior to HAS-BLED for bleeding, overall and in lower risk patients. The GARFIELD-AF tool has the potential for incorporation in routine electronic systems, and for the first time, permits simultaneous evaluation of ischaemic stroke, mortality and bleeding risks. CLINICAL TRIAL REGISTRATION: URL: http://www.clinicaltrials.gov. Unique identifier for GARFIELD-AF (NCT01090362) and for ORBIT-AF (NCT01165710)