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Background Obstructive sleep apnoea increases risk of respiratory depression with administration of sedatives, narcotics or anxiolytics. To reduce adverse events during hospital admission, we implemented STOP-BANG screening to prompt respiratory monitoring for inpatients receiving these medications. This study reports on protocol development, implementation and an initial analysis over 5 years to evaluate implementation success and outcomes.Intervention The STOP-BANG measure was embedded in the nurse navigator at admission. If the score was ≥3 and sedatives, narcotics and/or anxiolytics were ordered, the provider was prompted to monitor patients with continuous pulse oximetry and/or capnography.Methods We assessed the impact of the intervention using a retrospective pre–post design. Preprotocol data from all adult inpatients over a 2.5-year period, and postprotocol data from all adult inpatients from over a 5-year period, were extracted from the electronic health record. Outcomes included use of monitoring; adverse events during hospitalisation were included to evaluate the effects of the intervention: mortality, rate of rapid response team events, reversal and/or rescue, intensive care unit admission and orders for positive airway pressure equipment.Results The combined preprotocol and postprotocol sample included 254 121 patients. After protocol implementation, overall mortality for patients receiving sedatives, narcotics or anxiolytics decreased slightly from 2.1% to 1.9% (p

This article presents an original analysis of possession in Mojeño Trinitario, an Arawak language, teasing apart the levels of lexical possessive noun classes, morphological derivation between noun classes, and adnominal possessive constructions. The three possessive noun classes (obligatorily prefixed, optionally prefixed and non-prefixable) do not exactly correspond to the three possessive noun classes of the traditional analysis of Arawak languages, as in the analysis proposed here some of the traditionally called “alienable” nouns, i.e. nouns occurring as either possessed or unpossessed, are classified as non-prefixable nouns because they need some additional derivational morphology to be prefixable. The two adnominal possessive constructions (basic, through prefixation and modification; and indirect, via a generic possessive noun) are described, and their distribution is discussed. The article then addresses the question of how the notion of “alienability contrast” could explain the phenomena described, at either the lexical or phrasal level. In the process, it offers frequency counts on how often nouns of the different possessive classes occur as possessed in discourse, and it discusses whether the three noun classes are open or closed classes. It concludes that the alienability contrast is at best only a partial explanation for the expression of possession in Mojeño Trinitario. This article ends by developing how this innovative account of possession in Mojeño Trinitario applies to other Arawak languages.

This introductory article outlines the central topic of the present special issue, viz. the contrast between alienable and inalienable possession, and how this contrast is reflected in the grammars and lexicons of natural languages. It sketches the historical background of the alienability contrast in the linguistic literature and points to a number of biases that need to be overcome in order to (1) advance our understanding of the contrast and (2) face the limits of its explanatory potential. Specifically, the present introduction, as well as the contributions to the present issue, proposes to move beyond prototypical possessive relationships (ownership, part-whole, and kinship relations), prototypical possessor categories (human possessors) as well as prototypical possessee categories (artifacts, body-parts and kin). In addition, the issue contains three contributions dealing with Amazonian languages, thus filling in an important gap in previous crosslinguistic studies on possession. The data presented in the special issue show that many morphosyntactic phenomena that have been explained in terms of the alienability contrast – or are amenable to such explanations – cannot be reduced to it, and are sometimes even better described without recourse to alienability at all. The present article thus concludes that the alienability contrast is at best regarded as a heuristic tool in exploring linguistic data, and cautions that, if used as the only explanatory principle, it could actually hamper an adequate description of the data.

Abstract Cystic fibrosis transmembrane conductance regulator modulators have revolutionized cystic fibrosis (CF) care in the past decade. This study explores the CF-related mortality trends in the US from 1999 to 2020. We extracted CF-related mortality data from the CDC WONDER database. CF age-standardized mortality rates (ASMRs) were identified by ICD-10 code E84 and were stratified by demographic and geographical variables. Temporal trends were analyzed using Joinpoint modeling. CF-related ASMRs decreased from 1.9 to 1.04 per million population (p = 0.013), with a greater reduction in recent years. This trend was replicated in both sexes. The median age of death increased from 24 to 37 years. CF mortality rates decreased across sex, white race, non-Hispanic ethnicity, census regions, and urbanization status. Incongruent trends were reported in non-white races and Hispanic ethnicity. A lower median age of death was observed in women, non-white races, and Hispanic ethnicity. SARS-CoV-2 infection was the primary cause of death in 1.7% of CF decedents in 2020. The national CF-related mortality rates declined and the median age of death among CF decedents increased significantly indicating better survival in the recent years. The changes were relatively slow during the earlier period of the study, followed by a greater decline lately. We observed patterns of sex, ethnic, racial, and geographical disparities associated with the worsening of the gap between ethnicities, narrowing of the gap between races and rural vs. urban counties, and closing of the gap between sexes over the study period.

Conventional agricultural practices severely deplete the soil of essential organic matter and nutrients, increasing its vulnerability to disease, drought, and flooding. Permaculture is a form of agroecology adopting a whole ecosystem approach to create a set of principles and design frameworks for enriching soil fertility, but there is little scientific evidence of its efficiency. This study compares two permaculture managed sites with a conventional arable site to investigate the effect of permaculture management on soil fertility. We used phospholipid fatty acid analysis to estimate microbial abundance and diversity and related these to measured soil nutrients and carbon stocks. The potential of permaculture management to mitigate soil greenhouse gas emissions was assessed during a laboratory soil incubation and measurement of greenhouse gases via gas chromatography. Overall, the permaculture managed allotments had three times higher microbial biomass, one and a half times higher nitrogen, and four times higher carbon content than the arable site. Permaculture soils had larger carbon dioxide and nitrous oxide fluxes compared to arable soil, but all sites had a mean negative flux in methane. Permaculture management by use of organic amendments and no-dig practices provides a constant slow release of nutrients and build-up of organic matter and carbon and consequently promotes greater bacterial and fungal biomass within the soil.

Improper storage conditions or processing of milk leads to potential spoilage and illness, due in part to temperature abuse, allowing bacteria present to grow and spoil the product. However, certain proteins naturally found in raw milk, such as lactoferrin, have reported antibacterial properties. The levels of lactoferrin required to effectively inhibit growth of pathogens have not been investigated thoroughly. This study aimed to examine various concentrations of lactoferrin as a potential biopreservative and as an antimicrobial against the common dairy pathogens Salmonella enterica and Escherichia coli O157:H7. Minimum inhibitory concentration assays were conducted on raw bovine milk in which the bacteria were exposed to varying concentrations of lactoferrin. In the raw milk system, the growth of E. coli O157:H7 was significantly decreased at levels greater than 14.05 mg/mL lactoferrin based on the reduction of tetrazolium salts. For S. enterica, only lactoferrin concentrations at or above 112.5 mg/mL in the milk resulted in reduced growth. Taken together, these results indicate that lactoferrin may have biopreservative potential. To fully examine the practicality and effectiveness of lactoferrin as an antimicrobial additive, a similar study should be conducted using additional (gram-positive) pathogens, such as Bacillus cereus and Listeria monocytogenes. If effective, lactoferrin could prolong the shelf life of dairy products and help reduce the incidence of foodborne illnesses in developing countries with limited refrigeration capability.

BackgroundEmerging data suggests a possible role for cysteamine as an adjunct treatment for pulmonary exacerbations of cystic fibrosis (CF) that continue to be a major clinical challenge. There are no studies investigating the use of cysteamine in pulmonary exacerbations of CF. This exploratory randomized clinical trial was conducted to answer the question: In future pivotal trials of cysteamine as an adjunct treatment in pulmonary exacerbations of CF, which candidate cysteamine dosing regimens should be tested and which are the most appropriate, clinically meaningful outcome measures to employ as endpoints?Methods and findingsMulticentre double-blind randomized clinical trial. Adults experiencing a pulmonary exacerbation of CF being treated with standard care that included aminoglycoside therapy were randomized equally to a concomitant 14-day course of placebo, or one of 5 dosing regimens of cysteamine. Outcomes were recorded on days 0, 7, 14 and 21 and included sputum bacterial load and the patient reported outcome measures (PROMs): Chronic Respiratory Infection Symptom Score (CRISS), the Cystic Fibrosis Questionnaire-Revised (CFQ-R); FEV1, blood leukocyte count, and inflammatory markers. Eighty nine participants in fifteen US and EU centres were randomized, 78 completed the 14-day treatment period. Cysteamine had no significant effect on sputum bacterial load, however technical difficulties limited interpretation. The most consistent findings were for cysteamine 450mg twice daily that had effects additional to that observed with placebo, with improved symptoms, CRISS additional 9.85 points (95% CI 0.02, 19.7) p = 0.05, reduced blood leukocyte count by 2.46x109 /l (95% CI 0.11, 4.80), p = 0.041 and reduced CRP by geometric mean 2.57 nmol/l (95% CI 0.15, 0.99), p = 0.049.ConclusionIn this exploratory study cysteamine appeared to be safe and well-tolerated. Future pivotal trials investigating the utility of cysteamine in pulmonary exacerbations of CF need to include the cysteamine 450mg doses and CRISS and blood leukocyte count as outcome measures.Clinical trial registrationNCT03000348; www.clinicaltrials.gov.

Cenomanian gas deposits account for about 80% of natural gas production in Russia. Today, mature Cenomanian gas fields are depleted by more than 75% on average, the reservoir pressure in the productive formation zone has decreased by almost 90% compared to the initial pressure and in some places reaches 1.0-1.5 MPa. Reduction of pressure in the pay zone leads to active intrusion of water from the aquifer. The liquid is loading the well, and gas production decreases. The technology of gas well operation with foaming agent is widespread in the world, which is characterized by a high level of efficiency, including economic. The use of technology leads to a solution to the liquid loading and requires a preliminary selection of the optimal concentration of surfactants. The optimal concentration should ensure the creation of a relatively stable foam and provide a minimum pressure gradient in the production tubing. The article presents the results of studies of CTAB foaming agent influence on the process of liquid removal and pressure gradient in production tubing. Sequential studies to determine CMC, foamability, foam stability, and pressure gradient under foam flow were conducted on small-scale and large-scale facilities. These results can be reliable when using CTAB foaming agent, foaming agents based on it, or other cationic foaming agents in Cenomanian gas wells under the condition of reservoir water inflow. Besides, the effect of pressure on the optimal concentration of the foaming agent was also studied in this work.

Educators and students, discussing the future of nuclear age education at a symposium, generally agreed that knowledge and values are separated in society. Sources of information and of the proceedings are listed along with the Association for Supervision and Curriculum Development's (ASCD) resolution on nuclear freeze. (MLF)

AbstractMicrotubules are filamentous proteins that act as a substrate for the translocation of motor proteins. As such, they may be envisioned as a scaffold for the self-assembly of functional materials and devices. Physisorption, self-assembly and combing are here investigated as a potential prelude to microtubule-templated self-assembly. Dense films of self-assembled microtubules were successfully produced, as well as patterns of both dendritic and non-dendritic bundles of microtubules. They are presented in the present paper and the mechanism of their formation is discussed.

Abstract Background Malignant gliomas are lethal cancers, highly dependent on angiogenesis and treatment options and prognosis still remain poor for patients with recurrent glioblastoma multiforme (GBM). Ephs and ephrins have many well-defined functions during embryonic development of central nervous system such as axon mapping, neural crest cell migration, hindbrain segmentation and synapse formation as well as physiological and abnormal angiogenesis. Accumulating evidence indicates that Eph and ephrins are frequently overexpressed in different tumor types including GBM. However, their role in tumorigenesis remains controversial, as both tumor growth promoter and suppressor potential have been ascribed to Eph and ephrins while the function of EphA7 in GBM pathogenesis remains largely unknown. Methods In this study, we investigated the immunohistochemical expression of EphA7 in a series of 32 primary and recurrent GBM and correlated it with clinical pathological parameters and patient outcome. In addition, intratumor microvascular density (MVD) was quantified by immunostaining for endothelial cell marker von Willebrand factor (vWF). Results Overexpression of EphA7 protein was predictive of the adverse outcome in GBM patients, independent of MVD expression (p = 0.02). Moreover, high density of MVD as well as higher EphA7 expression predicted the disease outcome more accurately than EphA7 variable alone (p = 0.01). There was no correlation between MVD and overall survival or recurrence-free survival (p > 0.05). However, a statistically significant correlation between lower MVD and tumor recurrence was observed (p = 0.003). Conclusion The immunohistochemical assessment of tissue EphA7 provides important prognostic information in GBM and would justify its use as surrogate marker to screen patients for tyrosine kinase inhibitor therapy.

Abstract Background The rapid desensitization of the human prostacyclin (IP) in response to agonist binding has been shown in cell culture. Phosphorylation of the IP receptor by protein kinase C (PKC) has been suggested to be involved in this process. Methods and results In this study we investigated the vasodilatory effects of iloprost, a stable prostacyclin analogue, in perfused rabbit lungs. Continuous infusion of the thromboxane mimetic U46619 was employed to establish stable pulmonary hypertension. A complete loss of the vasodilatory response to iloprost was observed in experiments with continuous iloprost perfusion, maintaining the intravascular concentration of this prostanoid over a 180 min period. When lungs under chronic iloprost infusion were acutely challenged with inhaled iloprost, a corresponding complete loss of vasoreactivity was observed. This desensitization was not dependent on upregulation of cAMP-specific phosphodiesterases or changes in adenylate cyclase activity, as suggested by unaltered dose-response curves to agents directly affecting these enzymes. Application of a prostaglandin E1 receptor antagonist 6-isopropoxy-9-oxoxanthene-2-carboxylic acid (AH 6809) or the PKC inhibitor bisindolylmaleimide I (BIM) enhanced the vasodilatory response to infused iloprost and partially prevented tachyphylaxis. Conclusion A three-hour infusion of iloprost in pulmonary hypertensive rabbit lungs results in complete loss of the lung vasodilatory response to this prostanoid. This rapid desensitization is apparently not linked to changes in adenylate cyclase and phosphodiesterase activation, but may involve PKC function and co-stimulation of the EP1 receptor in addition to the IP receptor by this prostacyclin analogue.

Artificial intelligence (AI)-based digital phenotyping, including computational speech analysis, increasingly allows for the collection of diagnostically relevant information from an ever-expanding number of sources. Such information usually assesses human behaviour, which is a consequence of the nervous system, and so digital phenotyping may be particularly helpful in diagnosing neurological illnesses such as Alzheimer's disease. As illustrated by the use of computational speech analysis of Alzheimer's disease, however, neurological illness also introduces ethical considerations beyond commonly recognised concerns regarding machine learning and data collection in everyday environments. Individuals' decision-making capacity cannot be assumed. Understanding of analytical results will likely be limited even as the personal significance of those results is both highly sensitive and personal. In a traditional clinical evaluation, there is an opportunity to ensure that information is relayed in a way that is highly customised to the individual's ability to understand results and make decisions, and privacy is closely protected. Can any such assurance be offered as digital phenotyping technology continues to advance? AI-supported digital phenotyping offers great promise in neurocognitive disorders such as Alzheimer's disease, but it also poses ethical challenges. We outline some of these risks as well as strategies for risk mitigation., Competing Interests: Competing interests: FRD has nothing to disclose. PSP has nothing to disclose. KBC has nothing to disclose. WJ has nothing to disclose. MD wishes to disclose an affiliation with the American College of Physicians through a contract with his institution for consultation on ethics policy issues. VD has nothing to disclose. PWF has nothing to disclose. MHM has nothing to disclose. LEH has nothing to disclose., (© Author(s) (or their employer(s)) 2025. No commercial re-use. See rights and permissions. Published by BMJ Group.)

Background/objectives: Kaposi sarcoma (KS) is a common lymphatic endothelial cancer among children with and without HIV in central and eastern Africa. Despite its clinical heterogeneity, its various clinical phenotypes are often grouped together in staging and treatment algorithms. Patients with KS tumor-associated edema, referring to hard, non-pitting lesions which often lead to chronic disability, represent a unique, understudied subgroup of children with KS. To continue our work defining the distinct phenotypes of pediatric KS, this study aimed to assess the clinical progression and outcomes of KS edema in children., Methods: A retrospective cohort study was conducted at Kamuzu Central Hospital in Lilongwe, Malawi, focusing on children diagnosed with KS edema between 2010 and 2023., Results: We identified 52 children with KS edema, representing 27% of all patients with KS. Initial chemotherapy resulted in a clinical response in 92% of patients, but 46% experienced relapse or disease progression with a median time to first relapse of 12 months. Multiple progressions were common, with 31% of patients experiencing two or more events. Event-free survival at two years was 32%, dropping to 24% at five years, while overall survival was 73% at two years and 57% at five years. Relapse was more common among patients with KS edema versus those without it (relative risk = 2.1; 95%CI, 1.4-3.2; p < 0.001). Eight patients (15%) relapsed with visceral disease, five of whom originally presented with KS edema alone., Conclusions: Patients with KS edema have a unique, relapsing-remitting pattern of disease with a high risk of relapse relative to other forms of KS with subsequent long-term mortality, even after initial positive treatment responses. Late relapse and mortality with visceral disease are possible even among children presenting initially with KS edema alone. Children with KS edema require long-term follow-up, and novel treatment approaches tailored towards preventing frequent relapse are needed.

Introduction: There is insufficient evidence to determine if non-invasive transcutaneous bilirubin (TcB) measurement can replace serum bilirubin (SBR) in assessing rebound hyperbilirubinaemia after phototherapy., Objective: To investigate if TcB can safely guide management of neonates after phototherapy., Subjects: 100 well neonates ≥35 weeks' gestation who had received inpatient phototherapy., Method: Measurement of both helix (manufacturer's recommendation) and earlobe TcB coincidentally with routine SBR 12 hours after cessation of phototherapy. All mothers gave written informed consent., Results: Gestation ranged from 35 +0 to 41 +5 (median 37 +6 ) weeks; birth weight 2018-4566 (median 3230) g; age 55-222 (median 109) hours at testing. 86% neonates were Caucasian. Outcomes determined by SBR included restarting phototherapy (n=0), repeat SBR next day (n=29), no further routine follow-up (n=71).TcB and SBR measurements were unpredictably inconsistent. Helix TcB tended to underestimate SBR (mean difference 50.1 (95% CI 113.9 to -13.7) μmols/L); for earlobe TcB mean difference was -13.4 (95% CI 46.3 to -73.2) μmols/L (overestimate), but bias was greater over the range of mean differences. No demographic factor predicted consistency between TcB and SBR. TcB was 25% (helix) and 76% (earlobe) sensitive in predicting repeat phototherapy and/or repeat SBR; specificities were 92% and 58%, respectively. Adding a safety margin of 120 μmols/L to helix TcB value could have safely avoided invasive SBR measurement in 50/98 (51%) babies., Conclusions: Consistency between TcB and rebound SBR is unpredictable in well neonates > 35 weeks' gestation but adopting a wide safety margin has potential to reduce blood sampling. Recommencement of phototherapy is uncommon in this population., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

Nutrition security is challenging in regions where resources are limited and food production is naturally constrained. In low- and middle-income countries (LMICs), undernutrition is high for many reasons, including lack of nutritional diversity and low high-quality protein content. Interest in the role of animal-source food (ASF) in reducing nutrition insecurity is increasing, as evidence from LMICs suggests that consumption of ASF is strongly associated with reduction in stunting, improved diet quality, and overall nutrition, particularly in early stages of life. We review the strengths and limitations of ASF consumption in terms of accessibility, safety, and nutritional benefits compared to non-ASF sources. We present a critical discussion on existing barriers to ASF consumption and its future directions in LMICs. Understanding the role of ASF in improving nutrition security in LMICs is crucial to optimizing public health, designing appropriate interventions, and implementing effective policy in resource-poor settings.

This study aimed to provide further insight into the evolutionary dynamics of SARS-CoV-2 by analyzing the case of a 40-year-old man who had previously undergone autologous hematopoietic stem cell transplantation due to a diffuse large B-cell lymphoma. He developed a persistent SARS-CoV-2 infection lasting at least 218 days and did not manifest a humoral immune response to the virus during this follow-up period. Whole-genome sequencing and viral cultures confirmed a persistent infection with a replication-positive virus that had undergone genetic variation for at least 196 days after symptom onset.

The aim of this study was to screen the chemical components of Solanum elaeagnifolium leaves and assess their therapeutic attributes with regard to their antioxidant, antibacterial, and antidiabetic activities. The antidiabetic effects were explored to determine the α-amylase and α-glucosidase inhibitory potential of the leaf extract. To identify the active antidiabetic drugs from the extracts, the GC-MS-screened molecules were docked with diabetes-related proteins using the glide module in the Schrodinger Tool. In addition, molecular dynamics (MD) simulations were performed for 100 ns to evaluate the binding stability of the docked complex using the Desmond module. The ethyl acetate had a significant total phenolic content (TPC), with a value of 79.04 ± 0.98 mg/g GAE. The ethanol extract was tested for its minimum inhibitory concentration (MIC) for its bacteriostatic properties. It suppressed the growth of B. subtilis, E. coli, P. vulgaris, R. equi and S. epidermis at a dosage of 118.75 µg/mL. Moreover, the IC50 values of the ethanol extract were determined to be 17.78 ± 2.38 in the α-amylase and and 27.90 ± 5.02 µg/mL in α-glucosidase. The in-silico investigation revealed that cyclolaudenol achieved docking scores of -7.94 kcal/mol for α-amylase. Likewise, the α-tocopherol achieved the docking scores of -7.41 kcal/mol for glycogen phosphorylase B and -7.21 kcal/mol for phosphorylase kinase. In the MD simulations, the cyclolaudenol and α-tocopherol complexes exhibited consistently stable affinities with diabetic proteins throughout the trajectory. Based on these findings, we conclude that this plant could be a good source for the development of novel antioxidant, antibacterial, and antidiabetic agents.Communicated by Ramaswamy H. Sarma.

Introduction: Handover is crucial during the transfer of patient care between healthcare professionals.Successful handover maintains patients' safety and high-quality care. The electronic handover of patients' information is achievable, feasible, and potentially improves the quality of patient care. However, the introduction of the electronic handover is relatively recent and challenging for healthcare providers, especially nurses., Objectives: Since Sultan Bin Abdulaziz Humanitarian City (SBAHC) has recently implemented an electronic handover system among nurses, the main aim of the current study was to develop a tool to assess the perception and barriers to electronic handover systems among nurses working at SBAHC and evaluate its psychometric properties., Methods: The tool content and face validity were assessed by the content validity ratio (CVR). Exploratory and confirmatory factor analysis were used as validity methods, while the test-retest and inter-item consistency were used as reliability measures. The sample size was planned as five times the number of questions and a total of 200 nurses participated in the study., Results: The criteria required for factor analysis were met as indicated by the results of the Kaiser-Mayer-Olkin test and Bartlett's test of sphericity. The reliability results showed a Cronbach's alpha coefficient ranging from 0.858 to 0.910 for the perception subscale and from 0.564 to 0.789 for the barrier subscale, and an overall interclass correlation of 0.986, p<0.001., Conclusions: The developed SBAHC electronic handover tool was valid and reliable, and it is advisable to consider it at the initial stages of implanting an electronic handover system to identify obstacles that are faced by the staff to be considered and addressed by the higher management., Competing Interests: The current study is evaluating the psychometric properties of the Electronic Handover Tool that was developed by the authors of Sultan Bin Abdulaziz Humanitarian City Nurses. The tool might be copyrighted in the future., (Copyright © 2023, Al Baker et al.)

Avian endoparasites play important roles in conservation, biodiversity and host evolution. Currently, little is known about the epidemiology of intestinal helminths and protozoans infecting wild birds of Britain and Ireland. This study aimed to determine the rates of parasite prevalence, abundance and infection intensity in wild passerines. Fecal samples ( n = 755) from 18 bird families were collected from 13 sites across England, Wales and Ireland from March 2020 to June 2021. A conventional sodium nitrate flotation method allowed morphological identification and abundance estimation of eggs/oocysts. Associations with host family and age were examined alongside spatiotemporal and ecological factors using Bayesian phylogenetically controlled models. Parasites were detected in 20.0% of samples, with corvids and finches having the highest prevalences and intensities, respectively. Syngamus (33%) and Isospora (32%) were the most prevalent genera observed. Parasite prevalence and abundance differed amongst avian families and seasons, while infection intensity varied between families and regions. Prevalence was affected by diet diversity, while abundance differed by host age and habitat diversity. Infection intensity was higher in birds using a wider range of habitats, and doubled in areas with feeders present. The elucidation of these patterns will increase the understanding of parasite fauna in British and Irish birds.

Therapeutic antibodies have broad indications across diverse disease states, such as oncology, autoimmune diseases, and infectious diseases. New research continues to identify antibodies with therapeutic potential as well as methods to improve upon endogenous antibodies and to design antibodies de novo. On April 27-30, 2022, experts in antibody research across academia and industry met for the Keystone symposium "Antibodies as Drugs" to present the state-of-the-art in antibody therapeutics, repertoires and deep learning, bispecific antibodies, and engineering., (© 2022 New York Academy of Sciences.)

Background: Quantifying grit with the Short Grit Scale (Grit-S) has shown ability to predict success in various academic and professional domains. Grit has yet to be analyzed in patients with cancer., Methods: This study is a longitudinal analysis of prospectively distributed Grit-S surveys to patients undergoing radical or partial nephrectomy. Patients who completed a preoperative Grit-S survey with confirmed renal cell carcinoma (RCC) were included in the analysis. The relationship between preoperative grit scores and overall survival (OS) was determined using Cox proportional-hazard models and Kaplan-Meier analysis., Results: A total of 323 patients with RCC that completed the Grit-S survey prior to nephrectomy were included in the study. Median Grit score was 3.9. Most patients were male (67.5%), White (69.3%), and greater than 60 years old (57.0%) with a median age of 62 at the time of surgery. Patients scoring above or below the median grit score had similar baseline characteristics. As a binary variable, lower preoperative grit was significantly associated with shorter OS [hazard ratio (HR) =2.02, 95% confidence interval (CI): 1.12-3.63, P=0.019] on multivariable analysis. Unit changes in grit were not significantly associated with OS (HR =0.77, 95% CI: 0.53-1.14, P=0.193)., Conclusions: Lower grit scores may predict decreased OS in RCC patients undergoing nephrectomy. The Grit-S survey may have utility in preoperative evaluation. Further research assessing grit in other malignancies and how to psychologically optimize patients prior to surgery are needed., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://tau.amegroups.com/article/view/10.21037/tau-22-408/coif). The authors have no conflicts of interest to declare., (2022 Translational Andrology and Urology. All rights reserved.)

The current research explored the gender stereotype about psychology and its effect on perceptions of people's fit in the field. Across six studies ( N  = 1,516), results showed that participants believed that women represent the majority of people in both the major and profession of psychology. Also, participants associated psychology more strongly with femininity than masculinity and assigned more feminine and less masculine traits to people studying psychology than to people studying a stereotypically masculine career. In terms of fit within the field, participants rated psychology as less likely to meet the needs of men compared to women, especially after learning that the field was majority women. Overall, the studies provide evidence for an association between femininity and psychology and suggest that the stereotype affects perceptions of men's and women's fit within the field.

The present study aimed to establish a population pharmacokinetic (PopPK) modeling of benznidazole (BZD) in Brazilian patients with chronic Chagas disease. This was part of a Brazilian prospective cohort study with eight patients diagnosed with Chagas disease during the beginning of BZD treatment up to the 60th day. On the 15th day of treatment, a blood sampling was collected and analyzed. A one-compartment PK model was developed using Pmetrics. Patients with an average age of 50.3 (SD: 6.2) years old, 6 female patients and 2 males, 70.2 kg (14.2), receiving a 5 mg/Kg/day dose were included. PK parameters estimated for CL, V and Ka were 6.27 L/h, 38.97 L and 1.66 h-1, respectively. This is the first study to establish a population pharmacokinetic modeling of BZD in Brazilian patients with chronic Chagas disease. Therefore, further studies are needed to obtain the complete characterization of BZD pharmacokinetics.

Background: Besides the well-accepted role in lipid metabolism, high-density lipoprotein (HDL) also seems to participate in host immune response against infectious diseases., Objective: We used a quantitative proteomic approach to test the hypothesis that alterations in HDL proteome associate with severity of Coronavirus disease 2019 (COVID-19)., Methods: Based on clinical criteria, subjects (n=41) diagnosed with COVID-19 were divided into two groups: a group of subjects presenting mild symptoms and a second group displaying severe symptoms and requiring hospitalization. Using a proteomic approach, we quantified the levels of 29 proteins in HDL particles derived from these subjects., Results: We showed that the levels of serum amyloid A 1 and 2 (SAA1 and SAA2, respectively), pulmonary surfactant-associated protein B (SFTPB), apolipoprotein F (APOF), and inter-alpha-trypsin inhibitor heavy chain H4 (ITIH4) were increased by more than 50% in hospitalized patients, independently of sex, HDL-C or triglycerides when comparing with subjects presenting only mild symptoms. Altered HDL proteins were able to classify COVID-19 subjects according to the severity of the disease (error rate 4.9%). Moreover, apolipoprotein M (APOM) in HDL was inversely associated with odds of death due to COVID-19 complications (odds ratio [OR] per 1-SD increase in APOM was 0.27, with 95% confidence interval [CI] of 0.07 to 0.72, P=0.007)., Conclusion: Our results point to a profound inflammatory remodeling of HDL proteome tracking with severity of COVID-19 infection. They also raise the possibility that HDL particles could play an important role in infectious diseases., (Copyright © 2021. Published by Elsevier Inc.)

Chagas disease (CD) is still a neglected disease. Infected individuals are diagnosed late, being treated in worse clinical conditions. Thus, this study aimed to analyze the prevalence and the factors associated with new confirmed cases of CD identified by serological screening in an endemic region of Minas Gerais State, Brazil. This is an analytical cross-sectional study with data from a project of the Research Center in Tropical Medicine of Sao Paulo- Minas Gerais (SaMi-Trop) conducted in two municipalities. Data collection included a questionnaire with closed questions, a venous blood collection and an ELISA serological test for CD. A total of 2,038 individuals with no previous diagnosis of CD participated in the study. The result of the serological test for CD was adopted as the dependent variable. The independent variables addressed personal issues, health conditions and lifetime housing. A descriptive analysis of individual variables was performed. Subsequently, a bivariate analysis was performed using the Pearson's chi-square test. Households sheltering individuals positive for CD were georeferenced, and the analysis of spatial distribution was performed using the quartic function to estimate the density of the nucleus. Among the participants, 188 (9.2 %) were positive for CD. The profile of participants with CD was associated with place of residence, age, relative/family member with CD and living conditions. It is noteworthy that there are still patients with CD who are unaware of their diagnosis in both, rural and urban areas.

The ubiquitin conjugating enzyme UBE2W catalyzes non-canonical ubiquitination on the N-termini of proteins, although its substrate repertoire remains unclear. To identify endogenous N-terminally-ubiquitinated substrates, we discover four monoclonal antibodies that selectively recognize tryptic peptides with an N-terminal diglycine remnant, corresponding to sites of N-terminal ubiquitination. Importantly, these antibodies do not recognize isopeptide-linked diglycine (ubiquitin) modifications on lysine. We solve the structure of one such antibody bound to a Gly-Gly-Met peptide to reveal the molecular basis for its selective recognition. We use these antibodies in conjunction with mass spectrometry proteomics to map N-terminal ubiquitination sites on endogenous substrates of UBE2W. These substrates include UCHL1 and UCHL5, where N-terminal ubiquitination distinctly alters deubiquitinase (DUB) activity. This work describes an antibody toolkit for enrichment and global profiling of endogenous N-terminal ubiquitination sites, while revealing functionally relevant substrates of UBE2W., (© 2021. The Author(s).)

SARS-CoV-2 infection poses a global health crisis. In parallel with the ongoing world effort to identify therapeutic solutions, there is a critical need for improvement in the prognosis of COVID-19. Here, we report plasma proteome fingerprinting that predict high (hospitalized) and low-risk (outpatients) cases of COVID-19 identified by a platform that combines machine learning with matrix-assisted laser desorption ionization mass spectrometry analysis. Sample preparation, MS, and data analysis parameters were optimized to achieve an overall accuracy of 92%, sensitivity of 93%, and specificity of 92% in dataset without feature selection. We identified two distinct regions in the MALDI-TOF profile belonging to the same proteoforms. A combination of SDS-PAGE and quantitative bottom-up proteomic analysis allowed the identification of intact and truncated forms of serum amyloid A-1 and A-2 proteins, both already described as biomarkers for viral infections in the acute phase. Unbiased discrimination of high- and low-risk COVID-19 patients using a technology that is currently in clinical use may have a prompt application in the noninvasive prognosis of COVID-19. Further validation will consolidate its clinical utility., (© 2021 Lazari et al.)

Mesenchymal stem cells are the focus of intense research in bone development and regeneration. The potential of microparticles as modulating moieties of osteogenic response by utilizing their architectural features is demonstrated herein. Topographically textured microparticles of varying microscale features are produced by exploiting phase-separation of a readily soluble sacrificial component from polylactic acid. The influence of varying topographical features on primary human mesenchymal stem cell attachment, proliferation and markers of osteogenesis is investigated. In the absence of osteoinductive supplements, cells cultured on textured microparticles exhibit notably increased expression of osteogenic markers relative to conventional smooth microparticles. They also exhibit varying morphological, attachment and proliferation responses. Significantly altered gene expression and metabolic profiles are observed, with varying histological characteristics in vivo. This study highlights how tailoring topographical design offers cell-instructive 3D microenvironments which allow manipulation of stem cell fate by eliciting the desired downstream response without use of exogenous osteoinductive factors., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Nonoperational treatments for Morton's neuroma remain controversial because it is believed that sclerosing injections do not change nerve fibers on a cellular level. Up to 80% success rates with 4% ethanol sclerosing have been documented, and the remainder required operational removal of the painful nerve. We sought to evaluate the histologic characteristics of Morton's neuromas treated with 4% ethanol sclerosing injection versus corticosteroid injection alone in patients who required removal of the nerve for pain relief., Methods: A retrospective histologic review was performed of 23 consecutive patients who were treated with either sclerosing injection or nonsclerosing injection and underwent nerve removal between September 1, 2012, and February 28, 2015., Results: Of 19 patients who met the inclusion criteria, eight received sclerosing injections and 11 received nonsclerosing injections. Intraneural fibrosis was more severe in the nonsclerosing injection group (P = .008)., Conclusion: Histologic changes are seen in Morton's neuroma with the use of 4% ethanol sclerosing injection, contrary to findings from previous studies., (American Podiatric Medical Association.)

Introduction: The use of activity trackers has increased both among private consumers and in healthcare. It is therefore relevant to consider whether a consumer-graded activity tracker is comparable to or may substitute a research-graded activity tracker, which could further increase the use of activity trackers in healthcare and rehabilitation. Such use will require knowledge of their accuracy as the clinical implications may be significant. Studies have indicated that activity trackers are not sufficiently accurate, especially at lower walking speeds. The present study seeks to inform decision makers and healthcare personnel considering implementing physical activity trackers in clinical practice. This study investigates the criterion validity of the consumer-graded Garmin Vivosmart ® HR and the research-graded StepWatch ™ 3 compared with manual step count (gold standard) at different walking speeds under controlled conditions., Methods: Thirty participants, wearing Garmin Vivosmart ® HR at the wrist and StepWatch ™ 3 at the ankle, completed six trials on a treadmill at different walking speeds: 1.6 km/h, 2.4 km/h, 3.2 km/h, 4.0 km/h, 4.8 km/h, and 5.6 km/h. The participants were video recorded, and steps were registered by manual step count. Medians and inter-quartile ranges (IQR) were calculated for steps and differences in steps between manually counted steps and the two devices. In order to assess the clinical relevance of the tested devices, the mean absolute percentage error (MAPE) was determined at each speed. A MAPE ≤3% was considered to be clinically irrelevant. Furthermore, differences between manually counted steps and steps recorded by the two devices were presented in Bland-Altman style plots., Results: The median of differences in steps between Garmin Vivosmart ® HR and manual step count ranged from -49.5 (IQR = 101) at 1.6 km/h to -1 (IQR = 4) at 4.0 km/h. The median of differences in steps between StepWatch ™ 3 and manual step count were 4 (IQR = 14) at 1.6 km/h and 0 (IQR = 1) at all other walking speeds. The results of the MAPE showed that differences in steps counted by Garmin Vivosmart ® HR were clinically irrelevant at walking speeds 3.2-4.8 km/h (MAPE: 0.61-1.27%) as the values were below 3%. Differences in steps counted by StepWatch ™ 3 were clinically irrelevant at walking speeds 2.4-5.6 km/h (MAPE: 0.08-0.35%)., Conclusion: Garmin Vivosmart ® HR tended to undercount steps compared with the manual step count, and StepWatch ™ 3 slightly overcounted steps compared with the manual step count. Both the consumer-graded activity tracker (Garmin Vivosmart ® HR) and the research-graded (StepWatch ™ 3) are valid in detecting steps at selected walking speeds in healthy adults under controlled conditions. However, both activity trackers miscount steps at slow walking speeds, and the consumer graded activity tracker also miscounts steps at fast walking speeds., Competing Interests: The authors declare that they have no competing interests., (© 2020 Svarre et al.)

Background: Depression and anxiety are common after heart transplant, and in a regional heart transplant center servicing northern California, willingness to participate in treatment can be a major barrier. Introduction: Our primary aim is to design a remote cognitive-behavioral therapy (CBT) intervention. This study is the first step in the process. Through a survey to the cohort of heart transplant recipients ( N  = 230), managed within the Kaiser Permanente Northern California's Heart Transplant Service, we aimed to assess symptoms of stress, depression, and anxiety, patient willingness to participate in a CBT intervention, and preference between video and telephone. We proposed to patients a five-visit intervention, with the first and last visits in person and the three middle visits by video. Materials and Methods: One hundred twenty of 230 heart transplant recipients returned the 12-question Likert-like survey. Statistical tests included chi-square, fisher exact test, t -tests, and a logistic regression model. Results: Patients who reported two or more symptoms of stress, depression, and anxiety were 5.67 times more likely to engage in a remote CBT intervention (odds ratio = 5.67; 95% confidence interval 1.63-19.78; p  = 0.006). Patients experiencing stress with depression were the most willing group to participate in a CBT intervention. The 12 patients who met the study criteria of 3 years post-transplant and experiencing at least one mental health symptom were invited to participate in the CBT intervention. All patients (12) who met the CBT intervention criteria were not willing to participate in the two in-person visits. All were willing to participate in a telephone-only CBT intervention. Discussion: Among the heart transplant recipients in this cohort, there is more willingness to participate in a CBT intervention when distress is higher and there is a preference for telephone visits as the modality for treatment delivery. Conclusions: Based on the findings, the CBT intervention will be shorter in duration; instead of five visits, there will be four visits; and it will be conducted by telephone only. The new intervention will be tested with 8 to 10 patients, changed, and then it will need to be empirically tested.

The pre-transplant weight loss required of end-stage renal disease patients is often unachievable. Though robot-assisted procedures among extremely obese have shown minimal complication, long-term outcomes are understudied. Previously, we reported no difference in 6-month patient and graft survival among 28 robot-assisted transplant cases (2009-2013) and 28 open controls (2004-2010). Groups were frequency-matched on age, sex, race, donor compatibility, disease, and dialysis history. Cases had greater median pre-transplant body mass index (BMI; 42.3 (31.1-64.3) vs 36.8 (30.0-51.1)). Here, we compared patient and graft survival through 5 years post-transplant. Infection, wound complications, and significant re-hospitalizations were collected. One-, three-, and five-year graft survival were 100%, 100%, and 89.3% among cases, and 96.4%, 85.7%, and 78.6% among controls. Rejection within 1 year was greater among cases (11 vs 8). Five-year rates were similar (P = 0.54). Post-transplant BMI remained comparable. No cases and eight controls experienced surgical site infection (SSI). Two cases and one control experienced hernias. Post-transplant diabetes was documented among five cases and six controls. Three deaths occurred among cases, two among controls. This is the most extensive known follow-up of such obese recipients of robot-assisted transplant. Our procedure is a promising pathway to transplant and decreased mortality for those deemed too high risk for conventional surgery., (© 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Major histocompatibility complex Class I (MHCI) and Class II (MHCII) presented peptides powerfully modulate T cell immunity and play a vital role in generating effective anti-tumor and anti-viral immune responses in mammals. Characterizing these MHCI or MHCII presented peptides can help generate therapeutic treatments, afford information on T cell mediated biomarkers, provide insight into disease progression, and reduce adverse anti-drug side effects from engineered biotherapeutics. Here, we explore the tools and techniques commonly employed to discover both MHCI- and MHCII-presented peptides. We describe complementary strategies that enhance the characterization of these peptides and the informatics tools employed for both predicting and characterizing MHCI- and MHCII-presented epitopes. The evolution of methodologies for isolating MHC-presented peptides is discussed, as are the mass spectrometric workflows that can be employed for their characterization. We provide a perspective on where this field is headed, and how these tools may be applicable to the discovery and monitoring of epitopes in a variety of scenarios., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Chronic Obstructive Pulmonary Disease (COPD) is a highly prevalent condition characterized by inflammation and progressive obstruction of the airways. At present, there is no treatment that suppresses the chronic inflammation of the disease, and COPD patients often succumb to the condition. Excessive oxidative stress caused by smoke inhalation is a major driving force of the disease. The transcription factor NRF2 is a critical player in the battle against oxidative stress and its function is impaired in COPD. Increasing NRF2 activity may therefore be a viable therapeutic option for COPD treatment. We show that down regulation of KEAP1, a NRF2 inhibitor, protects primary human lung epithelial cells from cigarette-smoke-extract (CSE) induced cell death in an established in vitro model of radical stress. To identify new potential drug targets with a similar effect, we performed a siRNA screen of the 'druggable' genome using a NRF2 transcriptional reporter cell line. This screen identified multiple genes that when down regulated increased NRF2 transcriptional activity and provided a survival benefit in the in vitro model. Our results suggest that inhibiting components of the ubiquitin-proteasome system will have the strongest effects on NRF2 transcriptional activity by increasing NRF2 levels. We also find that down regulation of the small GTPase Rab28 or the Estrogen Receptor ESRRA provide a survival benefit. Rab28 knockdown increased NRF2 protein levels, indicating that Rab28 may regulate NRF2 proteolysis. Conversely ESRRA down regulation increased NRF2 transcriptional activity without affecting NRF2 levels, suggesting a proteasome-independent mechanism., Competing Interests: The authors have read the journal's policy and the authors of this manuscript have the following competing interests: CI, SK and JR are employees of Boehringer Ingelheim Pharma GbH&Co. MH, BS and SS have been employees of Cenix BioScience GmbH. FS and TK have been funded in part by Boehringer Ingelheim Pharma GbH & Co. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Background: Article citations are a well-accepted method of evaluating the influence or impact of a particular article. Other medical specialties have published the top cited articles in their specialty. To date, an analysis of the foot and ankle podiatric medicine-related article citation method has not been published., Methods: Three citation indices were used on October 10, 2014, February 20, 2015, and May 1, 2015, to determine the top 100 cited foot and ankle-related articles., Results: Most of the top 100 cited foot and ankle podiatric medicine-related articles were published in The Journal of Bone and Joint Surgery by medical doctors in the United States in the past two decades, with most of the article topics being the diabetic foot or trauma. The predominant level of evidence for the articles is evenly distributed among levels III, IV, and V., Conclusions: Podiatric medical research and publication has made great strides during the past two decades, particularly in the diabetic foot, but continued research and peer-reviewed journal publication in additional areas regarding the foot and ankle must become a priority in the podiatric medical community.

Cases of pulmonary embolism and pulmonary artery hypertension caused by choriocarcinoma represent a rare clinical emergency. We report a case of a 25-year-old woman who presented with pulmonary embolism and hypertension and died soon after complete pulmonary embolectomy. A related literature review revealed that almost all of these patients had previously experienced a spontaneous abortion (average, 6 months) and were not pregnant., (© 2016 the Journal of Biomedical Research. All rights reserved.)

As the number of clinical trials exploring cell therapy rises, a thorough understanding of the limits of cell delivery is essential. We used an extensive toolset comprising various standard and multiplex assays for the assessment of cell delivery postejection. Primary human mesenchymal stem cell (hMSC) suspensions were drawn up into 100-µl Hamilton syringes with 30- and 34-gauge needles attached, before being ejected at rates ranging from 10 to 300 µl/minute. Effects of ejection rate, including changes in viability, apoptosis, senescence, and other key aspects of cellular health, were evaluated. Ejections at slower flow rates resulted in a lower percentage of the cell dose being delivered, and apoptosis measurements of samples ejected at 10 µl/minute were significantly higher than control samples. Immunophenotyping also revealed significant downregulation of CD105 expression in samples ejected at 10 µl/minute (p < .05). Differentiation of ejected hMSCs was investigated using qualitative markers of adipogenesis, osteogenesis, and chondrogenesis, which revealed that slower ejection rates exerted a considerable effect upon the differentiation capacity of ejected cells, thereby possibly influencing the success of cell-based therapies. The findings of this study demonstrate that ejection rate has substantial impact on the percentage of cell dose delivered and cellular health postejection., (©AlphaMed Press.)

The current study has investigated the use of decellularised, demineralised bone extracellular matrix (ECM) hydrogel constructs for in vivo tissue mineralisation and bone formation. Stro-1-enriched human bone marrow stromal cells were incorporated together with select growth factors including VEGF, TGF-β3, BMP-2, PTHrP and VitD3, to augment bone formation, and mixed with alginate for structural support. Growth factors were delivered through fast (non-osteogenic factors) and slow (osteogenic factors) release PLGA microparticles. Constructs of 5 mm length were implanted in vivo for 28 days within mice. Dense tissue assessed by micro-CT correlated with histologically assessed mineralised bone formation in all constructs. Exogenous growth factor addition did not enhance bone formation further compared to alginate/bone ECM (ALG/ECM) hydrogels alone. UV irradiation reduced bone formation through degradation of intrinsic growth factors within the bone ECM component and possibly also ECM cross-linking. BMP-2 and VitD3 rescued osteogenic induction. ALG/ECM hydrogels appeared highly osteoinductive and delivery of angiogenic or chondrogenic growth factors led to altered bone formation. All constructs demonstrated extensive host tissue invasion and vascularisation aiding integration and implant longevity. The proposed hydrogel system functioned without the need for growth factor incorporation or an exogenous inducible cell source. Optimal growth factor concentrations and spatiotemporal release profiles require further assessment, as the bone ECM component may suffer batch variability between donor materials. In summary, ALG/ECM hydrogels provide a versatile biomaterial scaffold for utilisation within regenerative medicine which may be tailored, ultimately, to form the tissue of choice through incorporation of select growth factors.

Deletion of exon 9 from Cullin-3 (CUL3, residues 403-459: CUL3(Δ403-459)) causes pseudohypoaldosteronism type IIE (PHA2E), a severe form of familial hyperkalaemia and hypertension (FHHt). CUL3 binds the RING protein RBX1 and various substrate adaptors to form Cullin-RING-ubiquitin-ligase complexes. Bound to KLHL3, CUL3-RBX1 ubiquitylates WNK kinases, promoting their ubiquitin-mediated proteasomal degradation. Since WNK kinases activate Na/Cl co-transporters to promote salt retention, CUL3 regulates blood pressure. Mutations in both KLHL3 and WNK kinases cause PHA2 by disrupting Cullin-RING-ligase formation. We report here that the PHA2E mutant, CUL3(Δ403-459), is severely compromised in its ability to ubiquitylate WNKs, possibly due to altered structural flexibility. Instead, CUL3(Δ403-459) auto-ubiquitylates and loses interaction with two important Cullin regulators: the COP9-signalosome and CAND1. A novel knock-in mouse model of CUL3(WT) (/Δ403-459) closely recapitulates the human PHA2E phenotype. These mice also show changes in the arterial pulse waveform, suggesting a vascular contribution to their hypertension not reported in previous FHHt models. These findings may explain the severity of the FHHt phenotype caused by CUL3 mutations compared to those reported in KLHL3 or WNK kinases., (© 2015 The Authors. Published under the terms of the CC BY 4.0 license.)

Electrospinning is a highly adaptable method producing porous 3D fibrous scaffolds that can be exploited in in vitro cell culture. Alterations to intrinsic parameters within the process allow a high degree of control over scaffold characteristics including fiber diameter, alignment and porosity. By developing scaffolds with similar dimensions and topographies to organ- or tissue-specific extracellular matrices (ECM), micro-environments representative to those that cells are exposed to in situ can be created. The airway bronchiole wall, comprised of three main micro-environments, was selected as a model tissue. Using decellularized airway ECM as a guide, we electrospun the non-degradable polymer, polyethylene terephthalate (PET), by three different protocols to produce three individual electrospun scaffolds optimized for epithelial, fibroblast or smooth muscle cell-culture. Using a commercially available bioreactor system, we stably co-cultured the three cell-types to provide an in vitro model of the airway wall over an extended time period. This model highlights the potential for such methods being employed in in vitro diagnostic studies investigating important inter-cellular cross-talk mechanisms or assessing novel pharmaceutical targets, by providing a relevant platform to allow the culture of fully differentiated adult cells within 3D, tissue-specific environments.

Background: Mast cell localization within the airway smooth muscle (ASM)-bundle plays an important role in the development of airway hyper-responsiveness (AHR). Genomewide association studies implicate the 'alarmin' IL-33 in asthma, but its role in mast cell-ASM interactions is unknown., Objectives: We examined the expression and functional role of IL-33 in bronchial biopsies of patients with and without asthma, ex vivo ASM, mast cells, cocultured cells and in a mouse model system., Methods: IL-33 protein expression was assessed in human bronchial tissue from 9 healthy controls, and 18 mild-to-moderate and 12 severe asthmatic patients by immunohistochemistry. IL-33 and ST2 mRNA and protein expression in human-derived ASM, epithelial and mast cells were assessed by qPCR, immunofluorescence and/or flow cytometry and ELISA. Functional assays were used to assess calcium signalling, wound repair, proliferation, apoptosis and contraction. AHR and inflammation were assessed in a mouse model., Results: Bronchial epithelium and ASM expressed IL-33 with the latter in asthma correlating with AHR. ASM and mast cells expressed intracellular IL-33 and ST2. IL-33 stimulated mast cell IL-13 and histamine secretion independent of FcεR1 cross-linking and directly promoted ASM wound repair. Coculture of mast cells with ASM activated by IL-33 increased agonist-induced ASM contraction, and in vivo IL-33 induced AHR in a mouse cytokine installation model; both effects were IL-13 dependent., Conclusion: IL-33 directly promotes mast cell activation and ASM wound repair but indirectly promotes ASM contraction via upregulation of mast cell-derived IL-13. This suggests that IL-33 may present an important target to modulate mast cell-ASM crosstalk in asthma., (© 2015 The Authors. © Journal compilation 2015 Blackwell Munksgaard.)

Tissue engineering strategies can be applied to enhancing osseous integration of soft tissue grafts during ligament reconstruction. Ligament rupture results in a hemarthrosis, an acute intra-articular bleed rich in osteogenic human mesenchymal stem cells (hMSCs). With the aim of identifying an appropriate biomaterial with which to combine hemarthrosis fluid-derived hMSCs (HF-hMSCs) for therapeutic application, this work has investigated the biocompatibility of microparticles manufactured from two forms of poly(D,L-lactic-co-glycolic acid) (PLGA), one synthesized with equal monomeric ratios of lactic acid to glycolic acid (PLGA 50:50) and the other with a higher proportion of lactic acid (PLGA 85:15) which confers a longer biodegradation time. The surfaces of both types of microparticles were functionalized by plasma polymerization with allylamine to increase hydrophilicity and promote cell attachment. HF-hMSCs attached to and spread along the surface of both forms of PLGA microparticle. The osteogenic response of HF-hMSCs was enhanced when cultured with PLGA compared with control cultures differentiated on tissue culture plastic and this was independent of the type of polymer used. We have demonstrated that surface engineered PLGA microparticles are an appropriate biomaterial for combining with HF-hMSCs and the selection of PLGA is relevant only when considering the biodegradation time for each biomedical application., (© 2013 Wiley Periodicals, Inc.)

Current clinical treatments for skeletal conditions resulting in large-scale bone loss include autograft or allograft, both of which have limited effectiveness. In seeking to address bone regeneration, several tissue engineering strategies have come to the fore, including the development of growth factor releasing technologies and appropriate animal models to evaluate repair. Ex vivo models represent a promising alternative to simple in vitro systems or complex, ethically challenging in vivo models. We have developed an ex vivo culture system of whole embryonic chick femora, adapted in this study as a critical size defect model to investigate the effects of novel bone extracellular matrix (bECM) hydrogel scaffolds containing spatio-temporal growth factor-releasing microparticles and skeletal stem cells on bone regeneration, to develop a viable alternative treatment for skeletal degeneration. Alginate/bECM hydrogels combined with poly (d,l-lactic-co-glycolic acid) (PDLLGA)/triblock copolymer (10-30% PDLLGA-PEG-PDLLGA) microparticles releasing VEGF, TGF-β3 or BMP-2 were placed, with human adult Stro-1+ bone marrow stromal cells, into 2mm central segmental defects in embryonic chick femurs. Alginate/bECM hydrogels loaded with HSA/VEGF or HSA/TGF-β3 demonstrated a cartilage-like phenotype, with minimal collagen I deposition, comparable to HSA-only control hydrogels. The addition of BMP-2 releasing microparticles resulted in enhanced structured bone matrix formation, evidenced by increased Sirius red-stained matrix and collagen expression within hydrogels. This study demonstrates delivery of bioactive growth factors from a novel alginate/bECM hydrogel to augment skeletal tissue formation and the use of an organotypic chick femur defect culture system as a high-throughput test model for scaffold/cell/growth factor therapies for regenerative medicine., (Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

There is an unmet need for improved, effective tissue engineering strategies to replace or repair bone damaged through disease or injury. Recent research has focused on developing biomaterial scaffolds capable of spatially and temporally releasing combinations of bioactive growth factors, rather than individual molecules, to recapitulate repair pathways present in vivo. We have developed an ex vivo embryonic chick femur critical size defect model and applied the model in the study of novel extracellular matrix (ECM) hydrogel scaffolds containing spatio-temporal combinatorial growth factor-releasing microparticles and skeletal stem cells for bone regeneration. Alginate/bovine bone ECM (bECM) hydrogels combined with poly(d,l-lactic-co-glycolic acid) (PDLLGA)/triblock copolymer (10-30% PDLLGA-PEG-PLDLGA) microparticles releasing dual combinations of vascular endothelial growth factor (VEGF), chondrogenic transforming growth factor beta 3 (TGF-β3) and the bone morphogenetic protein BMP2, with human adult Stro-1+bone marrow stromal cells (HBMSCs), were placed into 2mm central segmental defects in embryonic day 11 chick femurs and organotypically cultured. Hydrogels loaded with VEGF combinations induced host cell migration and type I collagen deposition. Combinations of TGF-β3/BMP2, particularly with Stro-1+HBMSCs, induced significant formation of structured bone matrix, evidenced by increased Sirius red-stained matrix together with collagen expression demonstrating birefringent alignment within hydrogels. This study demonstrates the successful use of the chick femur organotypic culture system as a high-throughput test model for scaffold/cell/growth factor therapies in regenerative medicine. Temporal release of dual growth factors, combined with enriched Stro-1+HBMSCs, improved the formation of a highly structured bone matrix compared to single release modalities. These studies highlight the potential of a unique alginate/bECM hydrogel dual growth factor release platform for bone repair., (Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Electrospinning is a common technique used to fabricate fibrous scaffolds for tissue engineering applications. There is now growing interest in assessing the ability of collector plate design to influence the patterning of the fibres during the electrospinning process. In this study, we investigate a novel method to generate hybrid electrospun scaffolds consisting of both random fibres and a defined three-dimensional (3D) micro-topography at the surface, using patterned resin formers produced by rapid prototyping (RP). Poly(D,L-lactide-co-glycolide) was electrospun onto the engineered RP surfaces and the ability of these formers to influence microfibre patterning in the resulting scaffolds visualized by scanning electron microscopy. Electrospun scaffolds with patterns mirroring the microstructures of the formers were successfully fabricated. The effect of the resulting fibre patterns and 3D geometries on mammalian cell adhesion and proliferation was investigated by seeding enhanced green fluorescent protein labelled 3T3 fibroblasts onto the scaffolds. Following 24 h and four days of culture, the seeded scaffolds were visually assessed by confocal macro- and microscopy. The patterning of the fibres guided initial cell adhesion to the scaffold with subsequent proliferation over the geometry resulting in the cells being held in a 3D micro-topography. Such patterning could be designed to replicate a specific in vivo structure; we use the dermal papillae as an exemplar here. In conclusion, a novel, versatile and scalable method to produce hybrid electrospun scaffolds has been developed. The 3D directional cues of the patterned fibres have been shown to influence cell behaviour and could be used to culture cells within a similar 3D micro-topography as experienced in vivo.