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Aims: Using a model of isolated and Langendorff-perfused rat heart we analysed whether activation of β3-adrenergic receptors (β3-ARs) influences ventricular lusitropic performance. We also focused on the NOS/NO/cGMP/PKG cascade as the signal transduction mechanism. Methods: Hearts were treated with increasing concentrations (from 10−12 to 10−6 m) of BRL37344, a selective β3-AR agonist, and cardiac performance was evaluated by analysing both lusitropic parameters and coronary motility. Cardiac preparations were also perfused with BRL37344 in the presence of either isoproterenol (ISO) or nadolol, or pertussis toxin (PTx), or selective inhibitors of the NOS/NO/cGMP/PKG pathway. Results: BRL37344 caused a significant concentration-dependent reduction in (LVd P/d t)min, a decrease in half time relaxation significant starting from 10−12 m, and an increase in (LVd P/d t)max/(LVd P/d t)min ratio ( T/− t). BRL37344 abolished the ISO-mediated positive lusitropism. β3-AR-dependent effects on relaxation were insensitive to β1/β2-AR inhibition by nadolol (100 nm), and were abolished by Gi/o protein inhibition by PTx (0.01 nm). NO scavenging by haemoglobin (10 μm), and nitric oxide synthase (NOS) inhibition by NG-monomethyl-l-arginine (10 μm) revealed the involvement of NO signalling in BRL37344 response. Pre-treatment with inhibitors of either soluble guanylate cyclase (ODQ; 10 μm) or PKG (KT5823; 100 nm) abolished β3-AR-dependent negative lusitropism. In contrast, anantin (10 nm), an inhibitor of particulate guanylate cyclase, did not modify the effect of BRL37344 on relaxation. Conclusion: Taken together, our findings provide functional evidence for β3-AR modulation of ventricular relaxation in the rat heart which involves PTx-sensitive inhibitory Gi protein and occurs via an NO-cGMP-PKG cascade. Whether the effects of β3-AR stimulation on lusitropism are beneficial or detrimental remains to be established. [ABSTRACT FROM AUTHOR]

Leachate, an effluent produced during solid waste decomposition, interacts directly with soil, mainly in dumpsite areas. Studies on terrestrial animal exposure to leachate are, however, lacking. Plants are the most frequently studied organisms, while animal studies, especially earthworms, are limited. Nevertheless, ecotoxicological assessments involving earthworms are crucial due to their role in soil health and ecosystem maintenance, which are paramount in understanding potential terrestrial ecosystem leachate effects. In this context, this study aimed to evaluate behavioral effects, sublethal cytotoxicity and antioxidant system alterations in Eisenia andrei earthworms chronically exposed to leachate from a closed dumpsite. Cytotoxicity was determined by coelomocyte density, viability and cell typing, while antioxidant system alterations were assessed through superoxide dismutase (SOD), catalase (CAT), glutathione-S-transferase (GST), reduced glutathione (GSH) and metallothionein (MT) determinations. Malondialdehyde (MDA) and protein carbonylation (PTC) levels were also determined as oxidative effect markers. Finally, the Biomarker Response Index (BRI) was assessed, aiming to quantitatively integrate the results of the investigated endpoints and establish a biological health state (BHS) for each leachate concentration. Leachate exposure led to leak responses at concentrations of up to 50%, but attraction at higher concentrations. Decreased cell density (28%) was observed after 48 days and reduced viability (50%), after 14 days of leachate exposure. The observed cell typing changes indicate anti-inflammatory immune system effects. Leachate exposure led to several antioxidant system alterations, increasing SOD (2-6 %), CAT (5-35 %) and GST (5-70 %) activities and GSH (7-37%) and MT (3-67%) levels. Earthworm antioxidant defenses were, however, able to prevent lipid peroxidation, which decreased (11-37%) following leachate exposure to concentrations above 12.5%, and PTC, which increased at 42 days (26%) and reduced at 56 days (12 %). This is the first PTC assessment in leachate-exposed earthworms. The increased carbonylation levels observed after 42 days alongside MDA decreases highlight the need for further research employing oxidative effect biomarkers other than MDA. Finally, an integrated approach employing the BRI was carried out, revealing mild initial changes evolving to moderate to major effects at the highest leachate exposure concentration, with an effect attenuation detected at the end of the experiment. In this sense, this study brings forth a significant novelty, employing a biomarker previously not assessed in earthworms, demonstrating an oxidative effect, alongside the use of the BRI as an integrative tool for the endpoints applied in this assessment., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Simulation in health-care professions has grown in the last few decades. We provide an overview of the history of simulation in other fields, the trajectory of simulation in health professions education, and research in medical education, including the learning theories and tools to assess and evaluate simulation programs. We also propose future directions for simulation and research in health professions education., Competing Interests: Disclosure M.E. McBride is a paid consultant for the American Heart Association., (Copyright © 2022 Elsevier Inc. All rights reserved.)

The number of adults with congenital heart disease is rapidly increasing, resulting in more emergency care needs of this unique population. Concomitantly, the number of physicians trained in adult congenital heart disease (ACHD) care is insufficient, leading to physicians with limited experience assuming primary responsibility for the management of acute illness. We developed a simulation-based curriculum consisting of three cases and linked assessment instruments for fellows in multiple training programs to enhance their experience in this growing field. A 40-min asynchronous didactic presentation on ACHD emergencies was provided between pretests and posttests. Each participant was given checklist, global, and timeliness scores, and a second rater scored a subset to assess inter-rater reliability. Twenty-two participants across multiple disciplines completed the study. Our results demonstrate a significant and meaningful improvement in checklist scores, as well as a significant improvement in the secondary measures of global and efficiency performance from the first simulation to the second. Comfort levels for trainees improved significantly on post-test surveys. Inter-rater reliability was greater than 0.6 for all assessments. In conclusion, our novel simulation-based educational curriculum improved trainee performance in managing emergencies in adults with congenital heart disease, and we provide validity evidence for use of our checklist in training fellows for formative feedback., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Leachate is a variable effluent from waste management systems generated during waste collection and on landfills. Twenty-two leachate samples from waste collection trucks and a landfill were collected from March to December 2019 in the municipality of Rio de Janeiro (Brazil) and were analyzed for Human Adenovirus (HAdV), bacterial indicators and physico-chemical parameters. For viral analysis, samples were concentrated by ultracentrifugation and processed for molecular analysis using QIAamp Fast DNA Stool mini kit® for DNA extraction followed by nested-PCR and qPCR/PMA-qPCR TaqMan® system. HAdV was detected by nested-PCR in 100% (9/9) and 83.33% (12/13) of the truck and landfill leachate samples, respectively. Viral concentrations ranged from 8.31 × 10 1 to 6.68 × 10 7 genomic copies per 100 ml by qPCR and PMA-qPCR. HAdV species A, B, C, and F were characterized using nucleotide sequencing. HAdV were isolated in A549 culture cells in 100% (9/9) and 46.2% (6/13) from truck and landfill leachate samples, respectively. Regardless of the detection methods, HAdV concentration was predicted by the quantity of total suspended solids. A quantitative microbial risk assessment was performed to measure the probability of gastrointestinal (GI) illness attributable to inadvertent oral ingestion of truck leachate, revealing the higher probability of disease for the direct splashing into the oral cavity (58%) than for the gloved hand-to-mouth (33%). In a scenario where waste collectors do not wear gloves as protective personal equipment, the risk increases to 67%. This is the first study revealing infectious HAdV in solid waste leachate and indicates a potential health risk for waste collectors., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Leachate from solid waste landfill is a dark liquid of variable composition and possible source of contamination of groundwater and surface waters. This study aims to assess skimmed milk flocculation and ultracentrifugation as viral concentration methods associated to different nucleic acid extraction protocols in order to establish a methodology for virus recovery from sanitary landfill leachate. Spiking experiments using human adenovirus (HAdV) and bacteriophage PP7 revealed the association of QIAamp Fast DNA Stool mini kit® nucleic acid extraction and ultracentrifugation as an effective method for recovering HAdV (346.18%) and PP7 (523.97%) when compared to organic flocculation method (162.64% for HAdV and 0.61% for PP7) that presented PCR inhibition in all undiluted samples. Ultracentrifugation applied in three landfill samples confirm efficiency of the methodology detecting HAdV in all samples with a mean of 3.44E + 06 ± 1.56E + 06 genomic copies/mL. Nucleotide sequencing characterized HAdV as belonging to group B and F. JC polyomavirus (JCPyV) was also investigated in those samples; however, detection was not observed. Methodologies for detection of viruses in leachate can be useful to generate data for future health risk analysis of workers who have contact with solid urban waste, as well as populations exposed to different environmental matrices contaminated by these effluents.

Introduction: During the course of fellowship training, pediatric critical care fellows are expected to develop a broad and in-depth understanding of the pathophysiology of multiple disease processes. The simulation-based pediatric critical care pathophysiology curriculum we present uses scenarios created by pediatric critical care fellows to teach complex pathophysiology., Methods: Each of the three representative cases presented covered a specific pathophysiologic process and required participants to acutely manage (1) an 18-year-old patient with altered mental status in the setting of hepatic encephalopathy; (2) an 8-year-old patient with sepsis, coagulopathy, and acute kidney injury; or (3) a 12-year-old patient with status epilepticus. Each case could be conducted in a simulation suite or an acute care unit bed. We assessed learners' knowledge and attitudes at the end of these simulations with a structured debriefing session and via completion of an evaluation form. The simulations were then followed by a 30-minute interactive didactic session on the topic., Results: Each scenario had six fellow participants who completed evaluations. After completing each of the three case scenarios presented, the majority of participating pediatric critical care fellows indicated that the content was relevant and sufficiently challenging. They also indicated that these simulation scenarios would improve their clinical practice., Discussion: This fellow-developed simulation curriculum is novel, highlighting the relevance for critical care fellows' understanding of realistic clinical scenarios while promoting advanced management skills with a pathophysiology focus., (© 2020 Mannarino et al.)

Introduction: The World Health Organization (WHO) identifies 8.7 million new cases of tuberculosis (TB) annually around the world. The unfavorable outcomes of TB treatment prevent the achievement of the WHO's cure target., Goal: To evaluate existing intersections in the conceptions relative to the knowledge of TB, the experience of the illness and the treatment., Methods: Doctors, medical students and patients were selected from a public university in Rio de Janeiro, Brazil, from 2011 to 2013. The data were obtained by semi-structured individual and focus group interviews, participant observation and a field journal. The inclusion of patients was interrupted due to saturation, and the inclusion of doctors and medical students stopped due to exhaustion. The theoretical background included symbolic Interactionism, and the analysis used rounded Theory. The analysis prioritized the actions/interactions axis., Results: Twenty-three patients with pulmonary TB, seven doctors and 15 medical students were included. In the interviews, themes such as stigma, self-segregation, and difficulties in assistance emerged, in addition to defense mechanisms such as denial, rationalization, isolation and other mental mechanisms, including guilt, accountability and concealment of the disease. Aspects related to the assistance strategy, the social support network, bonding with the healthcare staff and the doctor-patient relationship were highlighted as adherence enablers. Doctors and students recommended an expansion of the theoretical and practical instruction on TB during medical students' education. The existence of health programs and policies was mentioned as a potential enabler of adherence., Conclusion: The main concepts identified were the stigma, self-segregation, guilt, responsibility, concealment and emotional repercussions. In relation to the facilitation of therapeutic adherence, the concepts identified were the bonds with healthcare staff, the doctor-patient relationship, assistance and educational health strategies.

Objective: To compare the efficacy of a three-dimensional (3D) haptic interface to a two-dimensional (2D) mouse interface for a screen-based simulation (SBS) neonatal intubation (NI) training intervention. Primary hypothesis: a haptic interface is more effective than a mouse interface for SBS training intervention for NI. Secondary hypothesis: SBS training, regardless of interface, will result in improved NI performance on a neonatal airway simulator., Methods: 45 participants were randomised to either a haptics or a mouse interface to complete an identical SBS training intervention for NI over a five-month period. Participants completed pre- and post-training surveys to assess demographics, experience, knowledge and attitudes. The primary outcome of participants' NI skills performance was assessed on a neonatal manikin simulator. Skills were measured pre- and post- training by number of attempts and time to successfully intubate, and airway visualization., Results: The demographics, training and experience were similar between groups. There was no difference in the improvement in skills, knowledge, attitudes or satisfaction ratings pre- and post-training between the groups. There was a significant decrease in number of attempts to intubate a neonatal airway simulator (2.89 vs 1.96, p<0.05) and improvement in the percent of subjects intubating in <30 seconds (22% vs 27%, p=0.02) from pre- to post-training in the study population overall., Conclusion: Using a haptic interface did not have an advantage over a mouse interface in improving NI skills, knowledge, attitudes, or satisfaction. Overall, a SBS training intervention for NI improved skills measured on a neonatal airway simulator., Competing Interests: Competing interests: None declared., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

Context: We previously reported that differentiated thyroid cancer (DTC) has higher aggressiveness and poorer prognosis in patients with Graves' disease (GD) than DTC in euthyroid control patients. Subsequent studies on this issue reached controversial conclusions. Genetic and environmental factors, as well as the lack of appropriate control subjects and/or inadequate patient follow-up, may account for these discrepancies., Objective: The aim of this study was to investigate the long-term disease-specific mortality of nonoccult DTCs occurring in patients with GD compared with DTCs in matched euthyroid control patients., Patients and Design: The previously described cohorts of nonoccult DTCs occurring in either patients with GD (DTC-GD, n = 21) or matched euthyroid DTC control patients (n = 70) were compared again after a longer follow-up (50-363.6 months; median, 165.6 months) to compare the major clinical endpoints of persistent/recurrent disease and overall survival. Both cohorts were recruited in 1982-1994 at a single institution. All patients had undergone total thyroidectomy and were followed up according to a standardized protocol., Results: Persistent/recurrent disease was more frequent in DTC-GD patients than in control patients (P = .0119). Disease-specific mortality was also significantly higher in DTC-GD patients (6 of 21, 28.6%) than in euthyroid control patients (2 of 70, 2.9%) (P = .0001). At the last visit, the percentage of disease-free patients was 57.1% (12 of 21) in the DTC-GD group vs 87.1% (61 of 70) in the control group (P = .0025)., Conclusions: Nonoccult DTCs occurring in patients with GD cause increased disease-specific mortality compared with DTCs in matched euthyroid control patients. These findings emphasize the need for early diagnosis and aggressive treatment of nonoccult DTCs in patients with GD.

The aim of this work was to investigate in the avascular heart of the frog Rana esculenta the influence of nitric oxide (NO) on ventricular systolic and diastolic functions by using a novel image analysis technique. The external volume variations of the whole ventricle were monitored during the heart cycle by video acquisition(visible light) and analysed by an appropriately developed software with a specific formula for irregular convex solids. The system, which measures the rate of volume changes and the ejection fraction, directly determined the volumetric behaviour of the working frog heart after stimulation or inhibition of NOS-NOcGMP pathway. End-diastolic volume (EDVext), end-systolic volume (ESVext), contraction and relaxation velocities (dV/dtsys and dV/dtdia, respectively), stroke volume (SV) and ejection fraction (EF), were measured before and after perfusion with NOS substrate (L-arginine), NO donor (SIN-1), cGMP analogue (8-Br-cGMP),NOS inhibitors (NG-monomethyl-L-arginine, L-NMMA; L-N(5)-(1-iminoethyl)-ornithine, L-NIO; 7-Nitroindazole,7-NI) and guanylyl cyclase inhibitor (ODQ). The results showed that NO reduces ventricular systolicfunction improving diastolic filling, while NOS inhibition increases contractility impairing ventricular filling capacity. The presence of activated eNOS (p-eNOS) was morphologically documented, further supporting that the mechanical activity of the ventricular pump in frog is influenced by a tonic release of NOS-generated NO.

Aims: Using a model of isolated and Langendorff-perfused rat heart we analysed whether activation of beta3-adrenergic receptors (beta3-ARs) influences ventricular lusitropic performance. We also focused on the NOS/NO/cGMP/PKG cascade as the signal transduction mechanism., Methods: Hearts were treated with increasing concentrations (from 10(-12) to 10(-6) m) of BRL(37344), a selective beta3-AR agonist, and cardiac performance was evaluated by analysing both lusitropic parameters and coronary motility. Cardiac preparations were also perfused with BRL(37344) in the presence of either isoproterenol (ISO) or nadolol, or pertussis toxin (PTx), or selective inhibitors of the NOS/NO/cGMP/PKG pathway., Results: BRL(37344) caused a significant concentration-dependent reduction in (LVdP/dt)(min), a decrease in half time relaxation significant starting from 10(-12) m, and an increase in (LVdP/dt)(max)/(LVdP/dt)(min) ratio (T/-t). BRL(37344) abolished the ISO-mediated positive lusitropism. beta3-AR-dependent effects on relaxation were insensitive to beta(1)/beta2-AR inhibition by nadolol (100 nm), and were abolished by G(i/o) protein inhibition by PTx (0.01 nm). NO scavenging by haemoglobin (10 microm), and nitric oxide synthase (NOS) inhibition by NG-monomethyl-l-arginine (10 microm) revealed the involvement of NO signalling in BRL(37344) response. Pre-treatment with inhibitors of either soluble guanylate cyclase (ODQ; 10 microm) or PKG (KT(5823); 100 nm) abolished beta3-AR-dependent negative lusitropism. In contrast, anantin (10 nm), an inhibitor of particulate guanylate cyclase, did not modify the effect of BRL(37344) on relaxation., Conclusion: Taken together, our findings provide functional evidence for beta3-AR modulation of ventricular relaxation in the rat heart which involves PTx-sensitive inhibitory Gi protein and occurs via an NO-cGMP-PKG cascade. Whether the effects of beta3-AR stimulation on lusitropism are beneficial or detrimental remains to be established.

The catecholamine release-inhibitory catestatin [Cts; human chromogranin (Cg) A(352-372), bovine CgA(344-364)] is a vasoreactive and anti-hypertensive peptide derived from CgA. Using the isolated avascular frog heart as a bioassay, in which the interactions between the endocardial endothelium and the subjacent myocardium can be studied without the confounding effects of the vascular endothelium, we tested the direct cardiotropic effects of bovine Cts and its interaction with beta-adrenergic (isoproterenol, ISO) and endothelin-1 (ET-1) signaling. Cts dose-dependently decreased stroke volume and stroke work, with a threshold concentration of 11 nM, approaching the in vivo level of the peptide. Cts reduced contractility by inhibiting phosphorylation of phospholamban (PLN). Furthermore, the Cts effect was abolished by pretreatment with either nitric oxide synthase (N(G)-monomethyl-l-arginine) or guanylate cyclase (ODQ) inhibitors, or an ET(B) receptor (ET(BR)) antagonist (BQ-788). Cts also noncompetitively inhibited the positive inotropic action of ISO. In addition, Cts inhibited the positive inotropic effect of ET-1, mediated by ET(A) receptors, and did not alter the negative inotropic ET-1 influence mediated by ET(BR). Cts action through ET(BR) was further suggested when, in the presence of BQ-788, Cts failed to inhibit the positive inotropism of both ISO and ET-1 stimulation and PLN phosphorylation. We concluded that the cardiotropic actions of Cts, including the beta-adrenergic and ET-1 antagonistic effects, support a novel role of this peptide as an autocrine-paracrine modulator of cardiac function, particularly when the stressed heart becomes a preferential target of both adrenergic and ET-1 stimuli.

Vasostatins (VSs), i.e. the main biologically active peptides generated by the proteolytic processing of chromogranin A (CGA) N-terminus, exert negative inotropism in vertebrate hearts. Here, using isolated working eel (Anguilla anguilla) and frog (Rana esculenta) heart preparations, we have studied the role of the cytoskeleton in the VSs-mediated inotropic response. In both eel and frog hearts, VSs-mediated-negative inotropy was abolished by treatment with inhibitors of cytoskeleton reorganization, such as cytochalasin-D (eel: 10 nM; frog: 1 nM), an inhibitor of actin polymerisation, wortmannin (0.01 nM), an inhibitor of PI3-kinase (PI3-K)/protein kinase B (Akt) signal-transduction cascade, butanedione 2-monoxime (BDM) (eel: 100 nM; frog: 10 nM), an antagonist of myosin ATPase, and N-(6-aminohexil)-5-chloro-1-naphthalenesulfonamide (W7) (eel: 100 nM; frog: 1 nM), a calcium-calmodulin antagonist. These results demonstrate that changes in cytoskeletal dynamics play a crucial role in the negative inotropic influence of VSs on eel and frog hearts.

Chromogranin A (CGA) N-terminal fragments corresponding to residues 1-76 and 1-113, named vasostatins for their inhibitory effects on vascular tension, have been postulated as important homeostatic regulators of the cardiovascular system. We have used an in vitro isolated working frog (Rana esculenta) heart as a bioassay to study the effects of exogenous human recombinant CGA 1-76 (VS-1) and human CGA 7-57 synthetic peptide on cardiac performance. Under basal conditions, the concentration-response curves of the two peptides exhibited a significant negative inotropism. This vasostatin response was unaffected by pretreatment with either Triton X-100 or two nitric oxide synthase inhibitors, i.e., N(G)-monomethyl-L-arginine and L-N5 (5)(1-iminoethyl) ornithine or the soluble guanylate cyclase inhibitor 1H-(1,2,4) oxadiazolo-(4,3-a) quinoxalin-1-one, indicating an endocardial endothelium-nitric oxide-cGMP-independent mechanism. The negative inotropism was also unaffected by either adrenergic (i.e., phentolamine and propranolol) or muscarinic (atropine) receptor or G proteins (pertussis toxin) inhibition. On the contrary, it was dependent from both extracellular Ca(2+) and K(+) channels, since it was abolished by pretreatment to either the Ca(2+) channel inhibitors lanthanum and diltiazem or the K(+) channel inhibitors Ba(2+), 4-aminopyridine, tetraethylammonium chloride, and glibenclamide. In conclusion, the findings that vasostatins exert an inhibitory modulation on basal cardiac performance and counteract, as previously reported, the adrenergic-mediated positive inotropism, strongly support a cardio-regulatory role for these peptides.