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The PAINESD risk score was developed in 2015 as a tool to stratify the risk of acute hemodynamic decompensation during ventricular tachycardia (VT) ablation in structural heart disease patients and further then used for post procedure 30-day mortality prediction. The original cohort however did not include Chagas disease (ChD) patients. We aim to evaluate the relevance of the score in a ChD population. The PAINESD risk score gives weighted values for specific characteristics (chronic obstructive pulmonary disease, age > 60 years, ischemic cardiomyopathy, New York Heart Association [NYHA] functional class 3 or 4, ejection fraction less than 25%, VT storm, and diabetes). The score was applied in a retrospective cohort of ChD VT ablations in a single tertiary center in Brazil. Data were collected by VT study reports and patient record analysis at baseline and on follow-up. Between January 2013 and December 2018, 157 VT catheter ablation procedures in 121 ChD patients were analyzed. Overall, 30-day mortality was 9.0%. Multivariate analysis correlated NYHA functional class (HR 1.78, 95% CI 1.03–3.08, P 0.038) and the need for urgent surgery (HR 31.5, 95% CI 5.38–184.98, P < 0.001), as well as a tendency for VT storm at presentation (HR 2.72, 95% CI 0.87–8.50, P 0.084) as risk factors for the primary endpoint. The median PAINESD risk score in this population was 3 (3–8). The area under the receiver operating characteristic (ROC) curve was 0.64 (95% CI 0.479–0.814). The PAINESD risk score did not perform well in predicting 30-day mortality in ChD patients. Pre-procedure NYHA functional class and the need for urgent surgery due to refractory pericardial bleeding were independently associated with increased 30-day mortality. Prospective studies are needed to take final conclusions in Chagas disease when using PAINESD score.

Coronavirus disease 2019 (COVID-19) is a global pandemic that has severely affected health care systems around the world. During the emergency state declared in Portugal in the months of March and April 2020, there was a severe reduction in medical activity in order to reduce the pressure on health systems. This study aimed to assess the impact of COVID-19 in gastroenterology departments across Portugal and the strategies developed to overcome this challenge.This was a cross-sectional study based on an online survey. A detailed questionnaire concerning different aspects of gastroenterology department activity was sent via e-mail to the heads of gastroenterology departments of Portuguese District Hospitals (Núcleo de Gastroenterologia dos Hospitais Distritais). Two periods were assessed, i.e., the emergency state and the recovery period between May and September. The responses were collected between September and October 2020.A total of 21 hospitals were enrolled (80.8% response rate). Twenty-eight percent of the responders reported healthcare professionals from their unit infected with COVID-19. At least 1 member (mostly fellows) of the department was deployed to another workplace in 66.7% of the hospitals. During the emergency state, 47.6% of the hospitals only performed urgent/emergent endoscopic procedures. In 38.5% of the hospitals the need to ration personal protective equipment led to the suspension of endoscopic training. Regarding the recovery period, nonurgent procedures were restarted in almost all of the centers. The same was reported for the colorectal cancer screening program. Remarkably, 81% of the responders confirmed that they had postponed procedures at patients' request for "fear of getting infected." Remote consultation was maintained in 81% of the hospitals. Globally, the fellows had resumed their training.This study provides a snapshot of the impact and consequences of the first wave of the COVID-19 pandemic across Portuguese hospitals. It is important to understand how the gastroenterology world dealt with the first impact of COVID-19 and what strategies were implemented in order to better prepare for what might follow.A doença por coronavirus 19 (COVID-19) é uma pandemia global que afetou gravemente os serviços de saúde em todo o mundo. Durante o estado de emergência declarado em Portugal nos meses de Março e Abril de 2020, verificou-se uma redução importante na atividade médica. Este estudo teve como objetivo avaliar o impacto da COVID-19 nos Serviços de Gastroenterologia portugueses e as estratégias implementadas para fazer face a este desafio.Tratou-se de um estudo transversal baseado num inquéritoResponderam 21 Hospitais (taxa de resposta: 80,8%). Vinte e oito porcento dos inquiridos tiveram profissionais da sua unidade infectados com COVID-19. Pelo menos um elemento foi mobilizado para outro local de trabalho em 66,7% dos hospitais, maioritariamente internos de formação específica de Gastroenterologia. Durante o estado de emergência, 47,6% dos Hospitais só realizaram endoscopias urgentes/emergentes. Em 38,5%, a necessidade de racionamento de equipamento de proteção individual levou à suspensão do treino em endoscopia. Relativamente ao período de recuperação, os procedimientos não urgentes foram retomados na maioria dos centros. O mesmo se verificou para o programa de rastreio do cancro colorectal. Salienta-se que 81% dos inquiridos confirmaram que adiaram exames a pedido dos doentes por “medo de ficarem infetados”. Mantiveram-se consultas nãopresenciais em 81% dos Hospitais. De modo geral, as atividades de formação dos internos foram progressivamente retomadas.Este estudo oferece o retrato do impacto e consequências da primeira vaga da pandemia nos Hospitais portugueses. É importante compreender como os Serviços de Gastrenterologia nacionais lidaram com o primeiro impacto da COVID-19 e que estratégias foram implementadas de forma a melhor preparar o que se segue.

Background The prevalence of coronary artery disease (CAD) is high among patients with severe aortic stenosis who undergo transcatheter aortic valve replacement (TAVR). Indications for TAVR are now expanding to younger and lower risk patients. During their lifetime, these patients will be at risk of developing CAD and it is expected an increase in coronary angiography and percutaneous coronary intervention (PCI). Aortic prosthesis, particularly if in supra-annular position, may pose important technical difficulties in coronary re-engagement after TAVR. Purpose To evaluate the feasibility to reengage the coronary ostia after TAVR, describe complications and compare technical differences between coronary procedures performed before and after TAVR. Methods Retrospective analysis of 714 patients submitted to TAVR from August 2007 to December 2019. Patients who needed coronary angiography after TAVR were selected. The primary endpoint was the rate of successful coronary ostia cannulation after TAVR, defined by the possibility to selectively cannulate and inject both coronary ostia. Secondary endpoint was complications associated with coronary catheterization after TAVR. Results Among 714 patients, 25 (3.5%) patients were submitted to a total of 28 coronary angiography after TAVR. 14 patients were male (56%), mean age 78.2±6.2 years and 9 (36%) had history of previous coronary revascularization. From the 28 coronary angiographies (balloon-expandable Edwards-Sapien n=11, 44%; self-expandable CoreValve n=10, 40%; Portico n=2, 8%; Symetis n=2, 8%), 25 (89%) met the primary endpoint. Only three was semiselective (Symetis, CoreValve Evolut R and CoreValve TAVR in TAVR), with impossibility to cannulate both coronary arteries, right coronary artery and left coronary artery, respectively. 13 (46%) patients had also indication for PCI and all were successfully performed (Edwards-Sapien n=4, 31%; CoreValve n=6, 46%; Portico n=2, 15%; Symetis n=1, 8%). The main indications for coronary angiography was chronic coronary syndrome (n=12, 43%) and acute coronary syndrome without ST segment elevation (n=7, 25%). Circumflex artery was the most frequently treated vessel (n=6), followed by left anterior descending artery (n=4), right coronary artery (n=3) and left main (n=2). There were no complications reported during or post-procedure. Comparing coronary angiographies before and after TAVR, there were no significant differences regarding arterial access site, catheter diameter, fluoroscopy time and quantity of contrast used in coronary angiography. Conclusion Although the need for coronary angiography was rare in patients after TAVR, selective diagnostic coronary angiographies were possible in 89% (25/28) and PCI was feasible in all patients in whom it was indicated, without any reported complications. Further prospective studies are needed to confirm the great feasibility of performing coronary angiography after TAVR. Funding Acknowledgement Type of funding sources: Public hospital(s). Main funding source(s): Centro Hospitalar Vila Nova de Gaia / Espinho

Background Nowadays, in patients with aortic regurgitation (AR), aortic valve surgery is indicated when severe and symptomatic or those with depressed LVEF. However, clinical outcomes of patients with significant aortic regurgitation are not influenced by these factors only. Recently, a new staging system for severe aortic stenosis has been proposed by Généreux on the basis of the extent of anatomic and functional cardiac damage. If this model could be applicable to an unselected significant AR population has not been tested. Purpose The aim of our study was to evaluate the prevalence of the different stages of extra-aortic valvular cardiac damage by the application of Généreux staging and its impact on prognosis in a large, real world cohort of significant AR patients. Methods This study retrospectively analysed the clinical, Doppler echocardiographic and outcome data in patients with grade III or greater AR between January 2014 and September 2019. According to the extent of cardiac damage on echocardiography, patients were classified as Stage 0 (no cardiac damage), Stage 1 (left ventricular damage), Stage 2 (mitral valve or left atrial damage), Stage 3 (tricuspid valve or pulmonary artery vasculature damage) or Stage 4 (right ventricular damage). Exclusion criteria were severe aortic stenosis and previous valve repair or replacement. The primary end-point was all-cause mortality. Results A total of 572 patients, aged 70.1±13.9 years, 294 (51.3%) men were enrolled. One third of patients were in NYHA I. Based on the proposed classification, 82 patients (14.3%) were classified in stage 0, 130 (22.7%) in stage 1, 276 (48.2%) in stage 2, 68 (11.8%) in stage 3 and 17 (3.0%) in stage 4. Median follow-up time was 3.3±1.9 years. There was a progressive increase in mortality rates according to staging: 8.5% in stage 0, 10.8% in stage 1, 24.9% in stage 2, 42.6% in stage 3 and 52.9% in stage 4 (p Conclusion Our study demonstrated that this new staging system studied for aortic stenosis also provides increased prognostic value to patients with significant aortic regurgitation. This staging system can be helpful to identify the degree of extra-aortic valvular cardiac damage and to optimize the time of valvular intervention. Further prospective studies are needed to confirm the benefit of the applicability of this model in clinical practice. Funding Acknowledgement Type of funding sources: Public hospital(s). Main funding source(s): Centro Hospitalar Vila Nova de Gaia / Espinho Distribution of stages of cardiac damageSurvival analysis according to stage

Background and aim Long-term data on the durability of transcatheter heart valves is scarce. This is of particular interest as indications expand to younger and lower surgical risk patients. We sought to assess the incidence of long-term structural valve dysfunction (SVD) and bioprosthetic valve failure (BVF) in a cohort of patients with TAVR who reached at least 5-year follow-up, as compared to surgical aortic valve replacement (SAVR), performed within the same time-frame at the same institution. Methods and results Consecutive patients with at least 5-year available follow-up, who underwent TAVR between November 2008 to December 2015 in a tertiary single center, were included. From a group of 246 patients undergoing TAVR, 126 had available follow-up data (age at implantation: 83.0 [77.8–87.0] years; EuroScore II: 4.54 [2.60–6.29]%; follow-up: 5.94 [5.06–7.67] years). First generation Corevalve® and Sapien® prosthesis were implanted in 56% and 38% patients, respectively. SVD and BVF were defined according to the new consensus statement from the EAPCI endorsed by the ESC and the EACTS. Mean transaortic pressure gradients decreased from 53.2±1.3 mmHg (pre-TAVR) to 10.4±0.4 mmHg (at discharge or up to one-year after TAVR, p As comparator, from a cohort of 587 patients submitted to biological SAVR, 247 (age 75.0 [70.0–79.0] years; EuroScore II 1.43 [1.06–2.17]%) had available long-term follow-up (6.89 [6.08–8.19] years). Moderate and severe SVD were reported in 42 and 3 patients, respectively (8-year cumulative incidence function to SVD: 3.13%; 95% CI, 2.45–4.21). These events were clinically relevant (BVF) in 19 of them: 8 performed TAVR valve-in-valve procedures and 3 redo SAVR. At the fifth-year of follow-up the incidence of SVD was not statistically different between TAVR (8%) and SAVR (15%), with a p for comparison of 0.137. Conclusions In our population of patients with symptomatic severe aortic stenosis treated with first-generation percutaneous bioprostheses, TAVR was associated with a low incidence of BVF and SVD at the long-term follow-up. These outcomes seem indistinct from those occurring in patients submitted to conventional SAVR Funding Acknowledgement Type of funding sources: None. KM curve reporting probability of SVD

Funding Acknowledgements Type of funding sources: None. Background Randomized controlled trials comparing stress cardiac magnetic resonance (CMR) and single-photon emission computed tomography (SPECT) suggest similar diagnostic accuracy for detecting obstructive coronary artery disease (CAD). However, there are few data on whether or not this remains true in routine clinical practice. The aim of this study was to assess the clinical and angiographic characteristics of patients undergoing invasive coronary angiography (ICA) after a positive stress CMR or positive SPECT, and to compare their positive predictive value with published results from the CE-MARC trial. Methods In this retrospective tertiary-center analysis, we included 429 patients (mean age 67 ± 10 years, 28% women, 42% diabetic) undergoing ICA between January 2016 and December 2020, after a positive stress CMR or positive SPECT. Regarding stress test, an adenosine protocol was performed in all stress CMR and in 76.4% (n = 272) of stress SPECT. Stress test results, including ischemia location and severity, were classified as reported by their primary readers. Patients with missing data on key variables, and those in whom microvascular disease was considered likely in the original stress test report were excluded. Obstructive CAD was defined as any coronary artery stenosis ≥ 50% in a vessel compatible with the ischemic territory on stress testing. Results Out of the total 429 patients, 356 (83%) were referred after a positive SPECT, and 73 (17%) after a positive stress CMR. Patients did not differ regarding age, cardiovascular risk factors, previous revascularization or left ventricular dysfunction, but patients with SPECT were more frequently male (p = 0.046). Overall, 320 patients (75%) had obstructive CAD on ICA. The prevalence of obstructive CAD was similar in patients with positive SPECT vs. positive stress CMR (76.1% vs. 80.8%, respectively, p = 0.385). There were also no significant differences in the prevalence of left main or 3-vessel disease (9.0% vs. 9.6%, p = 0.871, and 19.7% vs. 23.3% p = 0.483, respectively). Revascularization was performed or planned in 59.3% of patients in the SPECT group, and 52.1% of those in the stress CMR group (p = 0.255). The positive predictive values of both techniques were similar to those reported in the CE-MARC trial (Figure), and would increase to 88.1% and 89.4% for SPECT and stress CMR, respectively, if patients reported as having only mild ischemia were excluded. Conclusion In this tertiary center analysis, stress CMR and SPECT showed similar positive predictive values, comparable to those reported in the CE-MARC trial.

Background The preoperative differentiation between salivary gland tumor entities using computed tomography, magnetic resonance imaging (MRI) and ultrasound (US) is still limited. Biopsies are often regarded as indispensable for properly characterizing these various lesions. The aim of this study was to analyze the value of acoustic radiation force impulse (ARFI) sonoelastography as an US differentiation tool when examining parotid gland (PG) lesions. Methods We included 104 patients with PG masses in this study, employing two different US devices using quantitative ARFI-sonoelastography (Siemens Acuson-S3000, n=59; Siemens Acuson-Sequoia, n=45). The ability of sonoelastographic measurements to differentiate between different neoplasms was compared and analyzed for both US machines. Results Quantitative shear wave sonoelastography is limited in its ability to reliably differentiate between tumor entities of the PG as a stand-alone parameter. Measurement results were unsystematically distributed and not transferable between the two US devices. A significant differentiation of benign and malignant lesions was not possible with either US machine (S3000: P=0.770, Sequoia: P=0.382). A differentiation between pleomorphic adenomas (PA) and Warthin tumors was only possible with the Acuson S3000 system (P=0.001, Spearman-Rho =0.492, sensitivity 73.9%, specificity 65.0%). Conclusions A reliable identification and differentiation of PG tumors as well as clinical treatment decisions cannot be made with the sole use of ARFI-sonoelastography. The results emphasize the device-dependence and high error-proneness of this US technique when examining lesions of the PG.

Background Anti-malarial drug resistance remains a key concern for the global fight against malaria. In Ghana sulfadoxine-pyrimethamine (SP) is used for intermittent preventive treatment of malaria in pregnancy and combined with amodiaquine for Seasonal Malaria Chemoprevention (SMC) during the high malaria season. Thus, surveillance of molecular markers of SP resistance is important to guide decision-making for these interventions in Ghana. Methods A total of 4469 samples from uncomplicated malaria patients collected from 2009 to 2018 was submitted to the Wellcome Trust Sanger Institute, UK for DNA sequencing using MiSeq. Genotypes were successfully translated into haplotypes in 2694 and 846 mono infections respectively for pfdhfr and pfdhps genes and the combined pfhdfr/pfdhps genes across all years. Results At the pfdhfr locus, a consistently high (> 60%) prevalence of parasites carrying triple mutants (IRNI) were detected from 2009 to 2018. Two double mutant haplotypes (NRNI and ICNI) were found, with haplotype NRNI having a much higher prevalence (average 13.8%) than ICNI (average 3.2%) across all years. Six pfdhps haplotypes were detected. Of these, prevalence of five fluctuated in a downward trend over time from 2009 to 2018, except a pfdhps double mutant (AGKAA), which increased consistently from 2.5% in 2009 to 78.2% in 2018. Across both genes, pfdhfr/pfdhps combined triple (NRNI + AAKAA) mutants were only detected in 2009, 2014, 2015 and 2018, prevalence of which fluctuated between 3.5 and 5.5%. The combined quadruple (IRNI + AAKAA) genotype increased in prevalence from 19.3% in 2009 to 87.5% in 2011 before fluctuating downwards to 19.6% in 2018 with an average prevalence of 37.4% within the nine years. Prevalence of parasites carrying the quintuple (IRNI + AGKAA or SGEAA) mutant haplotypes, which are highly refractory to SP increased over time from 14.0% in 2009 to 89.0% in 2016 before decreasing to 78.9 and 76.6% in 2017 and 2018 respectively. Though quintuple mutants are rising in prevalence in both malaria seasons, together these combined genotypes vary significantly within season but not between seasons. Conclusions Despite high prevalence of pfdhfr triple mutants and combined pfdhfr/pfdhps quadruple and quintuple mutants in this setting SP may still be efficacious. These findings are significant as they highlight the need to continuously monitor SP resistance, particularly using deep targeted sequencing to ascertain changing resistance patterns.

Confocal laser endomicroscopy is an optical imaging technique that allows in vivo, real-time, microscope-like images of the upper aerodigestive tract's mucosa. The assessment of morphological tissue characteristics for the correct differentiation between healthy and malignant suspected mucosa requires strict evaluation criteria.This study aims to validate an eight-point score for the correct assessment of malignancy.We performed confocal laser endomicroscopy between March and October 2020 in 13 patients. 197 sequences (11.820 images) originated from the marginal area of pharyngeal and laryngeal carcinomas. Specimens were taken at corresponding locations and analyzed in HE staining as a standard of reference. A total of six examiners evaluated the sequences based on a scoring system; they were blinded to the histopathological examination. The primary endpoints are sensitivity, specificity, and accuracy. Secondary endpoints are interrater reliability and receiver operator characteristics.Healthy mucosa showed epithelium with uniform size and shape with distinct cytoplasmic membranes and regular vessel architecture. Confocal laser endomicroscopy of malignant cells demonstrated a disorganized arrangement of variable cellular morphology. We calculated an accuracy, sensitivity, specificity, positive predictive value, and negative predictive value of 83.2%, 81.3%, 85.5%, 86.7%, and 79.7%, respectively, with a κ-value of 0.64, and an area under the curve of 0.86.The results confirm that this scoring system is applicable in the laryngeal and pharyngeal mucosa to classify benign and malignant tissue. A scoring system based on defined and reproducible characteristics can help translate this experimental method to broad clinical practice in head and neck diagnosis.

Introduction New evidence on the role of myocardial revascularization in stable ischemic heart disease (SIHD), recently presented, showed that revascularization guided by the presence of moderate-to severe ischemia relieves angina more effectively than optimal medical therapy (OMT), without a significant benefit in hard clinical endpoints. Aim To assess the representativeness of the ISCHEMIA trial in a real-world population and compare management strategies between patients who fulfill the eligibility criteria of the trial (Group 1, G1) and those who do not (Group 2, G2). Methods and population Single centre retrospective analysis including all consecutive patients referred to coronary angiography (CA) for SIHD from January 2018 to December 2019. Patients were stratified in two groups (G1 and G2) according to the ISCHEMIA trial inclusion and exclusion criteria. G1 was compared with G2 and with a subset of G2 with obstructive coronary artery disease (CAD), defined as ≥70% luminal stenosis in at least one coronary artery or >50% for the left main. Results A total of 1020 patients underwent CA, of whom only 124 (12.2%) would have been eligible for the ISCHEMIA trial (G1). Overall, there were no significant differences in baseline characteristics between the two groups. G1 patients had more extensive and severe disease, presenting more frequently with proximal left anterior descending (LAD) involvement (26.6% vs 10.4%; p Conclusions Patients included in the ISCHEMIA trial are underrepresented in a real-world population of SIHD patients referred to coronary angiography. PCI rates were similar among patients with at least one significant coronary artery stenosis, regardless of previous evidence or severity of ischemia. Our findings underline the need for further refinement in criteria for revascularization in SIHD. Funding Acknowledgement Type of funding source: None

Background Chagas disease (ChD) is a major cause of non-ischemic cardiomyopathy (NICM) in Latin America and is becoming more frequent in other parts of the world, especially due to immigration movements. Ventricular Tachycardia (VT) is more common in this type of NICM than others, and finding an effective treatment strategy still is a challenge. Catheter ablation is an option, but there is poor data regarding its efficacy and safety. Purpose Evaluate the outcomes after VT catheter ablation in ChD patients. Methods Data were collected by VT studies reports and patient record analysis, including comorbidities and clinical status at baseline and on follow-up. We analyzed all-cause mortality, one-year VT recurrence rate and procedure related major complications. Results Between January 2013 and December 2018, 157 catheter ablation procedures in 121 ChD patients were performed in our institution. The mean follow-up time was 22.6±22.1 (mean ± SD) months. Overall post procedure mortality was 33.1%, and mean survival time was 51.2 months (95% CI: 44.8–58. NYHA functional class (p=0.022), ejection fraction (p=0.020) and immediate ablation result (p 0.002) were predictors of all-cause mortality in the follow-up. Clinical VT inducibility after ablation was a predictor of VT recurrence at one year (p=0.04). An epicardial approach was performed in 125 (79%) procedures, and accidental right ventricle (RV) puncture occurred in 23 (18.4%), in which open-chest surgery for bleeding hemostasis was necessary in 4 procedures (3.2%). Conclusion Mortality and recurrence rates in ChD patients after VT ablation were high, and correlated with heart failure severity. Epicardial approach is often necessary in this subset of patients. There was a correlation between immediate ablation results and recurrence. Kaplan-Meier of cumulative survival Funding Acknowledgement Type of funding source: None

Introduction Computed tomography (CT) is often performed before atrial fibrillation (AF) ablation to assess the anatomy of the pulmonary veins and exclude left atrial (LA) and left atrial appendage (LAA) thrombus. With the growing use of new oral anticoagulants (NOACs), a reassessment of the need for systematic thrombus exclusion in this context seems warranted. Objective To evaluate the prevalence of thrombus in LA/LAA in pre-ablation CT in a contemporary cohort of patients predominantly anticoagulated with NOACs. Methods We evaluated 789 consecutive patients (mean age 61±12 years; 38% female; 84% with paroxysmal AF) who underwent pre-ablation CT between Oct/2015 and Oct/2019. ECG-gated CT-angiography was performed using a dual-source 64-slice CT after iodinated contrast injection. Whenever necessary, a second dedicated acquisition was made 60 seconds after the first set of images. Presence of thrombus was defined as a persistent opacification defect. For each patient, thromboembolic risk was assessed with the CHA2DS2-VASc score. Results The median interval between CT and AF ablation was 1 day (IQR 1 – 2 days). The median CHA2DS2-VASc was 2 points (IQR 0 – 3 points), with 590 patients (75%) having CHA2DS2-VASc ≥1. Among the 199 patients (25%) with CHA2DS2-VASc = 0, 118 (59,3%) were anticoagulated with a NOAC and 14 (7%) with a vitamin K antagonist; 67 (34%) were not anticoagulated. Conversely, amongst the 590 patients with CHA2DS2-VASc ≥1, 84% were anticoagulated with a NOAC (n=494), 11% used vitamin K antagonists (n=62), and 34 patients were not anticoagulated (23 with CHA2DS2-VASc = 1). On cardiac CT, 521 (66%) patients were in sinus rhythm. Overall, only one LAA thrombus was found (0.12% [1/789]; 95% CI: 0.0–0.7%) – in a patient with CHA2DS2-VASc = 0, anticoagulated with a NOAC. The median effective radiation dose was 3.2 mSv (IQR 2.1–4.8 mSv). There were 5 minor allergic reactions to iodinated contrast. No strokes were documented within the first 24 hours after ablation. Conclusion In this contemporary cohort of patients with predominantly paroxysmal AF and anticoagulated with NOAC, the prevalence of intracavitary thrombus was extremely low (0.12%). While these findings do not compromise the multipurpose role of pre-ablation CT, they should nevertheless inform future discussions on the risk/benefit and cost/benefit of performing systematic exclusion of LA/LAA thrombi prior to AF ablation. Funding Acknowledgement Type of funding source: None

Background Vascular complications increase morbidity and mortality in transcatheter aortic valve implantation (TAVI). A collagen plug-based closure device - MANTA® was recently introduced as an alternative to the suture-mediated ProGlide® vascular closure device (VCD). Data regarding the efficacy and safety comparing both VCD is scarce. The present study sought to compare the effectiveness of both devices. Methods Single center retrospective analysis on prospectively collected data of 300 consecutive patients who underwent TAVI using MANTA® or ProGlide® since 2018. A 1:1 propensity-score matched population derived by a multivariate logistic regression model based on age, sex, body mass index, pre-procedural haemoglobin, EuroSCORE II, main access calcification and the sheath-to-artery ratio. The primary endpoint was the composite of major or life-threatening bleeding (VARC-2 definition), femoral artery stenosis/dissection, pseudoaneurysm and need for endovascular/surgical bailout intervention. Results The propensity score matching resulted in 129 matched pairs. The median age was 84 years old [IQR 80–87], 42% males with a median EuroSCOREII of 4.29% [IQR 3.05–6.24]. There were no differences in the primary endpoint between MANTA ® and ProGlide® cohorts (3.9% vs 7.8%, p=0.287, respectively). The rates of the primary endpoint with the MANTA® device decreased with center experience, with relatively steep learning curve effect concerning device success. Major or life-threatening bleeding (3.1% vs 5.4%, p=0.540) and pseudoaneurysm (0.8% vs 2.3%, p=0.622) occurred less frequently in MANTA® cohort, but the differences did not reach statistical significance. Endovascular (stent or balloon) or surgical rescue intervention (9.3% vs 5.4%, p=0.341) and femoral artery stenosis/dissection (6.2% vs 3.1%, p=0.376), were also similar rates. In ProGlide® cohort, to achieve VCD success (without primary endpoint events), 15.5% needed more than 2 devices, significantly different from MANTA ® (p Conclusions In patients undergoing transfemoral TAVI, the MANTA® VCD showed a similar efficacy and safety compared to the ProGlide® device and it reduced significantly the need of additional VCDs for completion of hemostasis. These results were obtained despite a clear learning curve associated with MANTA. Funding Acknowledgement Type of funding source: None

Introduction The evolution of percutaneous coronary intervention (PCI) and coronary artery bypass grafting (CABG) techniques made the choice of the optimal revascularization strategy of unprotected left main coronary disease (ULMD) challenging. Scoring systems are useful tools for the decision-making process and for risk stratification. Purpose To evaluate 1) the performance of the SYNTAX score I (SSI) and II (SSII) and Euroscore II (EII) in risk stratification and 2) the outcome predictors of patients (pts) with ULMD, according to the treatment strategy chosen (PCI or CABG). Methods Retrospective single centre cohort study of 440 consecutive pts (age 68±11 years; 76.6% male) with significant ULMD (defined as left main coronary artery stenosis >50%, with no patent arterial or venous bypass graft to left anterior descending artery), who were submitted to PCI (n=135) or CABG (n=307), between January 2006 and December 2018. Median follow-up (FU) was 4.0±1.8 years. The primary outcome was a composite of cardiovascular (CV) death, non-fatal myocardial infarction (MI) and target lesion revascularization (TLR). Results During the FU period, there were 112 (25.5%) CV deaths, 26 (5,9%) non-fatal MI and 53 (12.0%) TLR. Multivariate analysis of pts submitted to PCI showed that SSII and anatomical complete revascularization were independent predictors of the primary outcome (HR 1.045, CI 1.015–1.075, p=0.003 and HR 3,014, CI 1.655–5.489, p In the CABG cohort, only SSII was an independent predictor of the outcome by multivariate analysis (HR 1.061, CI 1.035–1.086, p The ROC curve analysis for all cohort presented no discriminative capacity for SSI (AUC 0.538, CI 0.482–0.593, p=0.186) and a weak discrimination for SSII (AUC 0.659, CI 0.605–0.713, p Conclusion In a real-world ULMD population, the most common risk scores, mainly those integrating anatomical and clinical features, presented a very modest role in the risk stratification, both in chronic and acute coronary syndromes. However, in pts with ULMD submitted to PCI, those risk scores had a more significant role in the risk stratification of these pts. Figure 1. SSI, SSII, EII ROC curves for all cohort Funding Acknowledgement Type of funding source: None

Background & Aim HepaStem is composed of human autologous liver-derived progenitor cells (HALPC) manufactured under GMP conditions and developed by Promethera Biosciences for the treatment of fibro-inflammatory disorders such as acute-on-chronic liver failure (ACLF). ACLF is a life-threatening complication of cirrhosis characterized by an acute liver decompensation associated with multiple organ failure and a very low survival rate. ACLF is linked to multiple cell type dysfunction, immunological imbalance and increased local (liver) and systemic inflammation. Preclinical data demonstrated that HepaStem has immunomodulatory properties. It is expected that HepaStem exerts immunomodulatory effects by interactions with immune cells of the patient and by paracrine effects through the various factors they secrete. It is expected that HepaStem will play a favorable role in treating the immunological disturbances observed in patients with ACLF and ultimately will be able to restore liver homeostasis. Methods, Results & Conclusion After a phase IIa clinical trial showing that IV administration of HepaStem was safe in ACLF patients, Promethera Biosciences initiated beginning of 2020 an efficacy study across multiple centres in Europe. In this context, the strategy followed by Promethera to develop the potency assay for release testing of batches that will be used in clinical trial is based on the immunomodulatory properties as main mechanism of action targeted for this pathology with (1) the development of an in vitro cell-based-assay that directly measures a biological activity of HepaStem representative of the biological effect expected in clinics and (2) then the development of non-biological analytical assay(s) used as surrogate markers to allow robust and easy routine evaluation of the potency. After identification of several well described soluble mediators with immunomodulatory properties, loss- and gain- of function experiments were performed to identify PGE2 as the main mediator associated with the suppression of APC-induced CD4+ T cell proliferation as well as inhibition of pro-inflammatory cytokine secretion such as TNFa. A very good correlation between the concentration of PGE2 in HepaStem conditioned medium after pro-inflammatory stimulation and the ability to inhibit APC-induced CD4+ T cell proliferation in vitro allowed together with the method validation to determine a first specification for the potency assay that will be later refined based on clinical data.

Background Coronary computed tomography angiography (CCTA) provides incremental prognostic information over traditional risk factors in patients with suspected coronary artery disease. However, little is known about the long-term predictive performance of CCTA-derived coronary volumes and mid-diastolic left ventricular (LV) mass. Purpose To assess long-term prognostic value of coronary volumes and mid-diastolic LV mass as novel potential imaging predictors derived from low-dose prospectively ECG-triggered CCTA. Methods Consecutive patients with suspected or known coronary artery disease, referred for low-dose CCTA, were included. Patients with previous revascularization were excluded. The following parameters were evaluated: calcium score, segment involvement score (SIS: 1 point for each coronary segment with presence of plaque), coronary volume, mid-diastolic LV mass and coronary volume indexed to LV mass. Major adverse cardiovascular events (MACE) were defined as all-cause death, non-fatal myocardial infarction and revascularization (PCI or CABG). The association between CCTA measures and the occurrence of events was quantified using cox regression hazard and Kaplan Meier analysis. Results A total of 147 consecutive patients were included in the study. Of them, 93 (63.3%) were male and 79 (53.7%) hat one or more traditional cardiovascular risk factors. There was a weak but statistical significant inverse correlation between indexed coronary volume and both calcium score (R=-0.3, p=0.01) and SIS (R=-0.24, p=0.005). After a median follow-up of 5.8 years 30 MACE occurred in 25 patients, including 3 deaths, 26 revascularizations and 1 non-fatal myocardial infarction. In univariate cox regression hazard analysis calcium score (HR=12.69, 95% CI 2.99–53.83, p Conclusions Indexed coronary volume, derived from low-dose CCTA, independently predicts cardiovascular events. Larger studies are mandated to confirm the predictive value of this potential new biomarker.

Introduction The use of venoarterial extracorporeal membrane oxygenation (VA-ECMO) to support patients in cardiogenic shock has been increasing in Portugal over the past few years. Nonetheless, epidemiologic, prognostic and clinical outcome data are scarce. Purpose We aim to identify clinical variables with prognostic significance in this challenging population, as well as the performance of various risk scores in mortality prediction. Methods All patients that underwent VA-ECMO support at our Cardiac ICU between 2011 and 2018 were included in the analysis. Logistic regression analysis was used to assess the relationship between clinical variables and outcomes. Results Short-term mechanical support with VA-ECMO was given to 40 patients, with a mean age of 52±11 years. At the time of the implant, the mean SOFA score was 11.2±4.0, and mean SAVE score was −4.75±4.6. Mean ECMO support duration was 116±96 hours. In 70% (N=28) of patients, VA-ECMO was successfully weaned. In-hospital mortality was observed in 52.5% of patients, which was in accordance with the predicted mortality by SOFA score (22.5% to 82% in our population risk range) and by SAVE score (60 to 70%). Those who placed the VA-ECMO as a bridge to transplant or to long-term mechanical LV assist device had greater in-hospital mortality rates (91.6 vs 41.9%, p=0.013), as well as those under ≥2 inotropic/vasopressors (69.2 vs 21.4%, p=0.012) or when adrenaline use was needed (100% vs 44.1%, p=0.01). No other between-group differences were observed in what concerns short-term mortality. After logistic regression analysis, independent predictors of in-hospital mortality included AMI setting, number of vasoactive amines used, and necessity of a LV venting device. SAVE score had the greater predictive ability in these patients (AUC = 0.638) among the most utilized clinical risk scores (SOFA score AUC = 0.37; APACHE II score AUC = 0.59; SAPS II score AUC = 0.54). Conclusion In our analysis, patients in profound cardiogenic shock on VA-ECMO support had slightly better survival rates than predicted by classical Risk Scores. The SAVE score may be the most accurate tool to predict in-hospital mortality in this specific, and yet heterogeneous, clinical subset. Other well recognized clinical markers of severity may also help refine short-term prognosis, and potentially improve organ transplant or other destination therapy prioritization.

Background The prognostic impact of the timing where new-onset Atrial Fibrillation (AF) occurs in Acute Coronary Syndrome (ACS) patients isn't well studied. Objective Evaluate the predictors and prognostic impact of early-onset AF (EAF) and late-onset AF (LAF) in ACS patients. Methods We analysed 17016 patients with ACS enrolled in a national multicenter registry from October 2010 to January 2019. Patients with history of valvular disease, valve replacement, AF present at hospital admission or those who died in the first 48 hours were excluded. EAF was defined as AF in the first 48 hours of hospitalization and LAF after 48 hours. The primary endpoint was a composite of death and readmission from cardiovascular causes at 1 year. Results The mean age was 65±13 years; 74% were males and 42% had STEMI). 324 (1.9%) had EAF and 344 (2.0%) had LAF. The predictors of EAF were age ≥75 years (OR 2.04, 95% CI 1.53–2.70, p The predictors of LAF were age ≥75 years (OR 3.15, 95% CI 2.36–4.19, p In STEMI patients only those with LAF had higher symptoms time (245 (IQR 165–400) minutes vs 284 (IQR 200–425) min; p=0.02). During hospitalization, LAF patients had a worse prognosis with a higher rate of heart failure, re-infarction, stroke, major bleeding and death (Table 1). In the follow-up, in multivarite analysis, only LAF was a predictor of the primary endpoint (EAF: HR 0.79, 95% CI 0.50–1.25, p=0.314; LAF: HR 1.45, 95% CI 1.05–2.00, p Complications during hospitalization No AF EAF LAF p Death 1.3 6.5 12.5 Kaplan Meier curve Conclusion Patients with EAF and LAF have different characteristics and outcomes with LAF patients having a worse in-hospital and long term prognosis.

Background Stress hyperglycaemia (SH) is a transient elevation of blood glucose (Gc) associated with acute, severe illness. However, the presence of hyperglycemia does not necessarily indicate the presence of SH, thus, the relative acute Gc rise provided by the ratio between acute and chronic Gc levels (A/C ratio) could more accurately represent so. Objective We investigated the association between the A/C ratio with in-hospital and long term prognosis in Acute Coronary Syndrome (ACS) patients. Methods Retrospective study of patients with ACS included in a single center between Jan/2012 and Dec/2017. Gc and HbA1c levels were measured at hospital admission. To calculate the A/C ratio the published formula 28.7×HbA1c-46.7 to estimate chronic Gc was used. The primary endpoints were a composite of in-hospital death and Killip class (KK) ≥III and a composite of infarction, stroke, heart failure and cardiovascular death (MACCE) in the follow-up. Results We included 404 patients (68±13 years; 72.8% males; 43.6% STEMI). The median A/C ratio was 1.07 (IQR 0.92–1.32). Patients in the highest tertile of the A/C ratio had a higher GRACE risk score (134±32, 139±38, 158±42; p During hospitalization 9 (2.2%) patientss died and 48 (11.9%) had the primary endpoint. The incidence of the in-hospital primary endpoint increased with A/C ratio tertiles (4.4%, 8.2%, 23.0%; p for trend After a median follow-up of 34 months (IQR 19–51), 50 (13%) patients died and 84 (21.9%) had MACCE. After adjusting for admission diagnosis, diabetes mellitus, GRACE and EF, an A/C ratio>1.31 was an independent predictor for the risk of death (HR 2.79, 95% CI: 1.17–6.65; p=0.021) and MACCE (HR 1.79, 95% CI: 1.09–2.93; p=0.020) in the follow-up. Conclusion In ACS patients, the A/C glycemic ratio increased with the severity of the index event and was a predictor of death and MACCE during the follow-up. It is readily available and provides valuable risk stratification and prognostic information.

We investigate the hypothesis that, in Escherichia coli, while the concentration of RNA polymerases differs in different growth conditions, the fraction of RNA polymerases free for transcription remains approximately constant within a certain range of these conditions. After establishing this, we apply a standard model-fitting procedure to fully characterize the in vivo kinetics of the rate-limiting steps in transcription initiation of the Plac/ara-1 promoter from distributions of intervals between transcription events in cells with different RNA polymerase concentrations. We find that, under full induction, the closed complex lasts ∼788 s while subsequent steps last ∼193 s, on average. We then establish that the closed complex formation usually occurs multiple times prior to each successful initiation event. Furthermore, the promoter intermittently switches to an inactive state that, on average, lasts ∼87 s. This is shown to arise from the intermittent repression of the promoter by LacI. The methods employed here should be of use to resolve the rate-limiting steps governing the in vivo dynamics of initiation of prokaryotic promoters, similar to established steady-state assays to resolve the in vitro dynamics.

The psychostimulant amphetamine can be prescribed to ameliorate the symptoms of narcolepsy, attention-deficit hyperactivity disorder and to facilitate weight loss. This stimulant can also have negative effects including toxicity and addiction risk. The impact of amphetamine on gene networks is partially understood and this study addresses this gap in consideration of the physical activity. The striata of mice exposed to either amphetamine or saline treatment were compared in a mouse line selected for home cage physical overactivity, a phenotype that can be mitigated with amphetamine, and in a contemporary control line using RNA-seq. Genes presenting opposite expression patterns between treatments across lines included a pseudogene of coiled-coil-helix-coiled-coil-helix domain containing 2 gene (Chchd2), ribonuclease P RNA component H1 (Rpph1), short stature homeobox 2 (Shox2), transient receptor potential melastatin 6 (Trpm6), and tumor necrosis factor receptor superfamily, member 9 (Tnfrsf9). Genes presenting consistent treatment patterns across lines, albeit at different levels of significance included cholecystokinin (Cck), vasoactive intestinal polypeptide (Vip), arginine vasopressin (Avp), oxytocin/neurophysin (Oxt), thyrotropin releasing hormone (Trh), neurotensin (Nts), angiotensinogen (Agt), galanin (Gal), prolactin receptor (Prlr), and calcitonin receptor (Calcr). Potassium inwardly rectifying channel, subfamily J, member 6 (Kcnj6), and retinoic acid-related (RAR)-related orphan receptor alpha (Rora) were similarly differentially expressed between treatments across lines. Functional categories enriched among the genes presenting line-dependent amphetamine effect included genes coding for neuropeptides and associated with memory and neuroplasticity and synaptic signaling, energy, and redox processes. A line-dependent association between amphetamine exposure and the synaptic signaling genes neurogranin (Nrgn) and synaptic membrane exocytosis 1(Rims1) was highlighted in the gene networks. Our findings advance the understanding of molecular players and networks affected by amphetamine in support of the development of activity-targeted therapies that may capitalize on the benefits of this psychostimulant.

Magnet-enforced targeting of drug-loaded superparamagnetic iron oxide nanoparticles (SPIO-NP) towards malignant tumors appears to be a desirable tool to reduce adverse side effects of systemically applied drugs including biologicals such as specific monoclonal antibodies. In order to make magnetic targeting feasible for tumors in the depth of the body we propose to load red blood cells (RBC) with SPIO-NP and use them as blood-born carriers to target them to endothelial cells of the tumor neo-vasculature. We employ original optoelectronical techniques to monitor the uptake of SPIO-NP. Targeting will be achieved by labeling of RBC with antibodies recognizing receptors specifically expressed by tumor endothelial cells together with strong external magnets placed over tumors. Here, we report magnetic and optoelectronical in vitro experiments showing the feasibility of endothelial cell loading with SPIO-NP, encapsulation of derivatized SPIO-NP in RBC, surface conjugation of RBC with antibodies, their binding to cultured endothelial cells and subsequent internalization.

In Escherichia coli, under optimal conditions, protein aggregates associated with cellular aging are excluded from midcell by the nucleoid. We study the functionality of this process under sub-optimal temperatures from population and time lapse images of individual cells and aggregates and nucleoids within. We show that, as temperature decreases, aggregates become homogeneously distributed and uncorrelated with nucleoid size and location. We present evidence that this is due to increased cytoplasm viscosity, which weakens the anisotropy in aggregate displacements at the nucleoid borders that is responsible for their preference for polar localisation. Next, we show that in plasmolysed cells, which have increased cytoplasm viscosity, aggregates are also not preferentially located at the poles. Finally, we show that the inability of cells with increased viscosity to exclude aggregates from midcell results in enhanced aggregate concentration in between the nucleoids in cells close to dividing. This weakens the asymmetries in aggregate numbers between sister cells of subsequent generations required for rejuvenating cell lineages. We conclude that the process of exclusion of protein aggregates from midcell is not immune to stress conditions affecting the cytoplasm viscosity. The findings contribute to our understanding of E. coli's internal organisation and functioning, and its fragility to stressful conditions.

To understand the role of bile acids (BAs) in cell function, many authors have investigated their effect on biomembrane models which are less complex systems, but there are still many open questions. The present study aims to contribute for the deepening of the knowledge of the interaction between BAs and model membranes, in particular, focusing on the effect of BA mixtures. The cytotoxic deoxycholic acid (DCA), the cytoprotective ursodeoxycholic acid (UDCA), and the equimolar mixture (DCA + UDCA) were investigated. Monolayers and liposomes were taken as model membranes with two lipid compositions: an equimolar mixture of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), sphingomyelin (SM), and cholesterol (Chol)) traditionally associated with the formation of lipid rafts and an equimolar POPC/SM binary mixture. The obtained results showed that DCA causes the fluidization of monolayers and bilayers, leading to the eventual rupture of POPC/SM liposomes at high concentration. UDCA may provide a stabilization of POPC/SM membranes but has a negligible effect on the Chol-containing liposomes. In the case of equimolar mixture DCA/UDCA, the interactions depend not only on the lipid composition but also on the design of the experiment. The BA mixture has a greater impact on the monolayers than do pure BAs, suggesting a cooperative DCA-UDCA interaction that enhances the penetration of UDCA in both POPC/SM and POPC/SM/Chol monolayers. For the bilayers, the presence of UDCA in the mixture decreases the disturbing effect of DCA.

From in vivo single-cell, single-RNA measurements of the activation times and subsequent steady-state active transcription kinetics of a single-copy Lac-ara-1 promoter in Escherichia coli, we characterize the intake kinetics of the inducer (IPTG) from the media, following temperature shifts. For this, for temperature shifts of various degrees, we obtain the distributions of transcription activation times as well as the distributions of intervals between consecutive RNA productions following activation in individual cells. We then propose a novel methodology that makes use of deconvolution techniques to extract the mean and the variability of the distribution of intake times. We find that cells, following shifts to low temperatures have higher intake times, although, counter-intuitively, the cell-to-cell variability of these times is lower. We validate the results using a new methodology for direct estimation of mean intake times from measurements of activation times at various inducer concentrations. The results confirm that E. coli's inducer intake times from the environment are significantly higher, following a shift to a sub-optimal temperature. Finally, we provide evidence that this is likely due to the emergence of additional rate-limiting steps in the intake process at low temperatures, explaining the reduced cell-to-cell variability in intake times. acceptedVersion

Promoters in Escherichia coli include an ‘OFF’ state, during which transcription is halted. Here, we propose a novel empirical method for assessing the time-length spent by promoters in this state. It relies on direct measurements of RNA production kinetics at the single molecule level at different induction levels, followed by an estimation of the RNA production rate under infinite induction, which is then compared to this rate under real, maximum induction. We apply it to the LacO3O1 promoter and infer that, under full induction, on average, 15% of the time between successful transcription events is spent in the OFF state. We verify this result by comparing the kinetics of a mutant strain lacking repressor molecules with that of the inferred rate under infinite induction. We expect this strategy of dissecting the kinetics of transcription repression to be applicable to a wide number of promoters in E. coli. acceptedVersion

Made available in DSpace on 2015-03-18T15:55:24Z (GMT). No. of bitstreams: 0 Previous issue date: 2014-10-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Instituto Nacional de Ciencia e Tecnologia: Obesidade e Diabetes The disruption to glucose homeostasis upon glucocorticoid (GC) treatment in adult male rats has not been fully characterized in older rats or in females. Thus, we evaluated the age-and gender-related changes in glucose homeostasis in GC-treated rats. We injected male and female rats at 3 months and 12 months of age with either dexamethasone (1.0 mg/kg body mass, intraperitoneally) or saline, daily for 5 days. All of the GC-treated rats had decreased body mass and food intake, and adrenal hypotrophy. Increased glycemia was observed in all of the GC-treated groups and only the 3-month-old female rats were not glucose intolerant. Dexamethasone treatment resulted in hyperinsulinemia and hypertriacylglyceridemia in all of the GC-treated rats. The glucose-stimulated insulin secretion (GSIS) was higher in all of the dexamethasone-treated animals, but it was less pronounced in the older animals. The beta-cell mass was increased in the younger male rats treated with dexamethasone. We conclude that dexamethasone treatment induces glucose intolerance in both the 3- and 12-month-old male rats as well as hyperinsulinemia and augmented GSIS. Three-month-old female rats are protected from glucose intolerance caused by GC, whereas 12-month-old female rats developed the same complications that were present in 3- and 12-month-old male rats. Fed Univ Santa Catarina UFSC, Dept Physiol Sci, Ctr Biol Sci, BR-88040900 Florianopolis, SC, Brazil Sao Paulo State Univ UNESP, Dept Biol, Inst Biosci Humanities & Exact Sci, Sao Jose Do Rio Preto, Brazil State Univ Campinas UNICAMP, Dept Struct & Funct Biol, Inst Biol, Campinas, SP, Brazil State Univ Campinas UNICAMP, OCRC, Campinas, SP, Brazil Sao Paulo State Univ UNESP, Dept Biol, Inst Biosci Humanities & Exact Sci, Sao Jose Do Rio Preto, Brazil CNPq: 471397/2011-3

Glucocorticoid (GC) excess alters glucose homeostasis and promotes modifications in murinometric and anthropometric parameters in rodents and humans, respectively. β-hydroxy-β-methylbutyrate (HMB), a leucine metabolite, has been proposed as a nutritional strategy for preventing muscle wasting, but few data regarding its effects on glucose homeostasis are available. Here, we analyzed whether the effects of GC excess on glucose homeostasis may be attenuated or exacerbated by the concomitant ingestion of HMB. Adult Wistar rats (90-days-old) were assigned to four groups: (1) vehicle treated (Ctl), (2) dexamethasone (DEX) treated (Dex), (3) HMB treated (Hmb), and (4) DEX plus HMB treated (DexHmb). Dex groups received DEX (1 mg·kg body weight (BW)−1, intraperitoneal) for 5 consecutive days. HMB groups ingested HMB (320 mg·kg BW−1, oral gavage) for the same 5 days. HMB ingestion did not attenuate the effects of DEX on food intake and body weight loss, changes in masses of several organs, insulin resistance, and glucose intolerance (p > 0.05). In fact, in DexHmb rats, there was increased fasting glycemia and exacerbated glucose intolerance with the main effect attributed to DEX treatment (p < 0.05). HMB exerted no attenuating effect on plasma triacylglycerol levels from DexHmb rats, but it seems to attenuate the lipolysis induced by β-adrenergic stimulation (20 μmol·L−1isoproterenol) in fragments of retroperitoneal adipose tissue from DexHmb rats. Therefore, HMB does not attenuate the diabetogenic characteristics of GC excess. In fact, the data suggest that HMB may exacerbate GC-induced glucose intolerance.

Knowledge and lessons from past accidental exposures in radiotherapy are very helpful in finding safety provisions to prevent recurrence. Disseminating lessons is necessary but not sufficient. There may be additional latent risks for other accidental exposures, which have not been reported or have not occurred, but are possible and may occur in the future if not identified, analyzed, and prevented by safety provisions. Proactive methods are available for anticipating and quantifying risk from potential event sequences. In this work, proactive methods, successfully used in industry, have been adapted and used in radiotherapy. Risk matrix is a tool that can be used in individual hospitals to classify event sequences in levels of risk. As with any anticipative method, the risk matrix involves a systematic search for potential risks; that is, any situation that can cause an accidental exposure. The method contributes new insights: The application of the risk matrix approach has identified that another group of less catastrophic but still severe single-patient events may have a higher probability, resulting in higher risk. The use of the risk matrix approach for safety assessment in individual hospitals would provide an opportunity for self-evaluation and managing the safety measures that are most suitable to the hospital's own conditions.

OBJECTIVE: Declines in working memory are a ubiquitous finding within the cognitive-aging literature. A unitary inhibitory selection mechanism that serves to guide attention toward task-relevant information and resolve interference from task-irrelevant information has been proposed to underlie such deficits. However, inhibition can occur at multiple time points in the memory-processing stream. Here, we tested whether the time point at which inhibition occurs in the memory-processing stream affects age-related memory decline. METHOD: Clinically healthy younger (n = 23) and older (n = 22) adults performed two similar item-recognition working memory tasks. In one task, participants received an instruction cue telling them which words to attend to followed by a memory set, promoting perceptual inhibition at the time of encoding. In the other task, participants received the instruction cue after they received the memory set, fostering inhibition of items already in memory. RESULTS: We found that older and younger adults differed in their ability to inhibit items both during encoding and when items had to be inhibited in memory but that these age differences were exaggerated when irrelevant information had to be inhibited from memory. These results provide insights into the mechanisms that support cognitive changes to memory processes in healthy aging.

Escherichia coli segregates protein aggregates to the poles by nucleoid exclusion. Combined with cell divisions, this generates heterogeneous aggregate distributions in subsequent cell generations. We studied the robustness of this process with differing medium richness and antibiotics stress, which affect nucleoid size, using multimodal, time-lapse microscopy of live cells expressing both a fluorescently tagged chaperone (IbpA), which identifies in vivo the location of aggregates, and HupA-mCherry, a fluorescent variant of a nucleoid-associated protein. We find that the relative sizes of the nucleoid's major and minor axes change widely, in a positively correlated fashion, with medium richness and antibiotic stress. The aggregate's distribution along the major cell axis also changes between conditions and in agreement with the nucleoid exclusion phenomenon. Consequently, the fraction of aggregates at the midcell region prior to cell division differs between conditions, which will affect the degree of asymmetries in the partitioning of aggregates between cells of future generations. Finally, from the location of the peak of anisotropy in the aggregate displacement distribution, the nucleoid relative size, and the spatiotemporal aggregate distribution, we find that the exclusion of detectable aggregates from midcell is most pronounced in cells with mid-sized nucleoids, which are most common under optimal conditions. We conclude that the aggregate management mechanisms of E. coli are significantly robust but are not immune to stresses due to the tangible effect that these have on nucleoid size. IMPORTANCE Escherichia coli segregates protein aggregates to the poles by nucleoid exclusion. From live single-cell microscopy studies of the robustness of this process to various stresses known to affect nucleoid size, we find that nucleoid size and aggregate preferential locations change concordantly between conditions. Also, the degree of influence of the nucleoid on aggregate positioning differs between conditions, causing aggregate numbers at midcell to differ in cell division events, which will affect the degree of asymmetries in the partitioning of aggregates between cells of future generations. Finally, we find that aggregate segregation to the cell poles is most pronounced in cells with mid-sized nucleoids. We conclude that the energy-free process of the midcell exclusion of aggregates partially loses effectiveness under stressful conditions.

Placental explant culture, and cellular cytolysis and cellular differentiation have been previously studied. However, oxidative stress and nitric oxide profiles have not been evaluated in these systems. The aim of this study was to determine the release of lipid peroxidation and nitric oxide from placental explants cultured over a seven day period. Placental explants were maintained for seven days in culture and the medium was changed every 24 hours. The response was assessed in terms of syncytiotrophoblast differentiation (human chorionic gonadotropin, hCG), cellular cytolysis (lactate dehydrogenase, LDH), oxidative stress (thiobarbituric acid reactive substances, TBARS), and nitric oxide (NO). Levels of hCG increased progressively from day two to attain its highest level on days four and five after which it decreased gradually. In contrast, the levels of LDH, TBARS, and NO were elevated in the early days of placental culture when new syncytiotrophoblast from cytotrophoblast were forming and also in the last days of culture when tissue was declining. In conclusion, the levels of NO and lipid peroxidation follow a pattern similar to LDH and contrary to hCG. Future placental explant studies to evaluate oxidative stress and NO should consider the physiological changes inherent during the time of culture.

Escherichia coli is an established model for the study of protein aggregation in vivo, a process that is related to its aging. These aggregates have a predisposition for locating at the cell poles, which, following cell division, results in asymmetric damage distribution by the two poles of the daughter cells. Such preference for polar localization is due the occlusion caused by the nucleoid at midcell. To study this process, it is necessary to correlate the spatial location of the protein aggregates and nucleoids during a cell life cycle. For this, and while neither structure has clear borders, it is necessary to perform their segmentation from fluorescence microscopy images. Here, we propose an adaptation of the method used in automatic Drusen detection in retinal images for detecting and segmenting DAPI-stained nucleoids at each stage of the cell cycle. We use the GPL algorithm to detect the number of nucleoids inside each cell. For the segmentation step, according to the number of nucleoids present in the cell, we use either one or two 3-dimensional, modified Gaussian functions as a point spread function model. By setting the amplitude profile parameter with a value equal to 10, the segmentation threshold is directly obtained from the value of z0. The method was applied to images of cells at different temperatures. Table 1 shows the extracted relative nucleoid major axis length for cells with one and with two nucleoids, in each temperature condition. From Table 1, the mean relative nucleoid major axis length decreases with increasing temperature for both cells with one and with two nucleoids, as expected by visual inspection of the images. Also, the positions of the borders of the nucleoid detected by our method, for cells with one nucleoid, are consistent with regions of anisotropies in aggregates motion reported in our previous studies. Additionally, the locations of the nucleoids for both cells with one and with two nucleoids are in agreement with the observed spatial distribution of the protein aggregates, which remain outside the nucleoid region according to our previous studies. These results demonstrate that the proposed method is able to differentiate between one and two nucleoids, perform their segmentation efficiently, and extract with precision the nucleoid(s) size along both the major and minor cell axis.

Angiogenesis is a key event in tumor growth and metastasis, chronic inflammatory disease, and cardiovascular disease. It is controlled by positive and negative regulators, which include vascular endothelial growth factor (VEGF) as the most active of these. VEGF/VEGF receptors are important targets not only for therapy but also for imaging. Based on the structural study of VEGF, we developed a novel cyclopeptide (cyclo-VEGI) that exhibits powerful antitumor properties. We herein report the design of novel molecules derived from cyclo-VEGI as potential targeting agents in cancer and other angiogenesis-related diseases. We performed selective chemical modification of the most active VEGF-derived cyclopeptide (cyclo-VEGI). Original hydrophilic linkers were synthesized and coupled to cyclo-VEGI. These reactions provide nanocarriers for delivery. The inhibitory effect of the different compounds on VEGF binding was evaluated in competition assays with 125I-VEGF. A fluorescent cyclo-VEGI peptide was synthezised to assess direct binding and internalization of cyclo-VEGI. Chemical modifications of cyclo-VEGI do not diminish the biological activity of cyclo-VEGI as measured in competition assays; in fact, it is even increased. Moreover there is a strong cellular accumulation of the fluorescent-labeled cyclo-VEGI. Conjugates synthesized in this study may be useful leads to design delivery systems for targeting approaches in cancer and other angiogenesis-related diseases. The modified cyclo-VEGIs may have a wide range of applications and represent a useful tool to develop delivery/carrier systems for therapeutic targeting or imaging.