Physical and textural properties of functional edible protein films from soybean using an innovative 3D printing technology.

Increasing market demand for sustainable, environmentally friendly edible film materials has called for the development of new customizable production methods utilizing emerging technologies such as 3D printing. We hereby report a new method to generate functional edible soy protein isolate films prepared from three types of soybeans (AR-R11-7999, MO-S17-17168, and MO-S17-19874R) using an innovative 3D printing technology. The protein contents in AR-R11-7999, MO-S17-17168, and MO-S17-19874R soybean meals and their corresponding protein isolates were 40.0, 39.1, and 39.9; and 84.5, 84.7, and 87.3 % (w/w, dry basis), respectively. Response surface methodology was used to maximize the tensile and puncture strength and minimize the thickness of the 3D-printed edible films using protein concentration, plasticizer concentration (glycerol), and drying time as the independent variables. The optimized film production conditions were determined as soy protein concentration: 8.91%, plasticizer concentration: 3.00%, and drying time: 3.98 h with a desirability value of 0.7428. The optimized conditions were then successfully verified with the original soybean lot with a nonsignificant difference in physical properties. At the optimized conditions, the 3D-printed edible films using three soybean lots revealed: 0.108-0.114 mm thickness; 14.79-16.07 MPa tensile strength; 6.97-8.20 N puncture strength; 90.81-91.53, -1.89 to -1.31, and 14.85-17.25 were color parameters L*, a*, and b*, respectively; 1.22-1.36 g/cm3 density; and 104.4-105.7% elongation at break ratio (%). PRACTICAL APPLICATION: Edible soy protein films produced by an extrusion-based 3D printing approach are highly customizable and precise, and could be produced at an industrial scale. This newly produced environment-friendly soy protein-based edible film can serve as an alternate packaging to synthetic plastics and reduce the environmental landfill problem while adding value to soybean produced in the mid-south United States.

The quantitative impact of COVID-19 on surgical training in the United Kingdom.

BACKGROUND: COVID-19 has had a global impact on all aspects of healthcare including surgical training. This study aimed to quantify the impact of COVID-19 on operative case numbers recorded by surgeons in training, and annual review of competency progression (ARCP) outcomes in the UK. METHODS: Anonymized operative logbook numbers were collated from electronic logbook and ARCP outcome data from the Intercollegiate Surgical Curriculum Programme database for trainees in the 10 surgical specialty training specialties.Operative logbook numbers and awarded ARCP outcomes were compared between predefined dates. Effect sizes are reported as incident rate ratios (IRR) with 95 per cent confidence intervals. RESULTS: Some 5599 surgical trainees in 2019, and 5310 in surgical specialty training in 2020 were included. The IRR was reduced across all specialties as a result of the COVID-19 pandemic (0.62; 95 per cent c.i. 0.60 to 0.64). Elective surgery (0.53; 95 per cent c.i. 0.50 to 0.56) was affected more than emergency surgery (0.85; 95 per cent c.i. 0.84 to 0.87). Regional variation indicating reduced operative activity was demonstrated across all specialties. More than 1 in 8 trainees in the final year of training have had their training extended and more than a quarter of trainees entering their final year of training are behind their expected training trajectory. CONCLUSION: The COVID-19 pandemic has had a major effect on surgical training in the UK. Urgent, coordinated action is required to minimize the impacts from the reduction in training in 2020.

The impact of COVID-19 on surgical training: a systematic review.

BACKGROUND: Coronavirus disease (COVID-19) has caused global disruption to health care. Non-urgent elective surgical cases have been cancelled, outpatient clinics have reduced and there has been a reduction in the number of patients presenting as an emergency. These factors will drastically affect the training opportunities of surgical trainees. The aim of this systematic review is to describe the impact of COVID-19 on surgical training globally. METHODS: The review was performed in line with Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines and registered with the Open Science Framework (OSF). Medline, EMBASE, PubMed and the Cochrane Central Register of Controlled Trials were searched. RESULTS: The searches identified 499 articles, 29 of which were included in the review. This contained data from more than 20 countries with 5260 trainees and 339 programme directors. Redeployment to non-surgical roles varied across studies from 6% to 35.1%. According to all of the studies, operative experience has been reduced. Knowledge learning had been switched to online platforms across 17 of the studies and 7 reported trainees had increased time to devote to educational/academic activities. All of the studies reporting on mental health report negative associations with increased stress, ranging from 54.9% to 91.6% of trainees. CONCLUSIONS: The impact of COVID-19 on surgical trainees has been experienced globally and across all specialities. Negative effects are not limited to operative and clinical experience, but also the mental health and wellbeing of trainees. Delivery of surgical training will need to move away from traditional models of learning to ensure trainees are competent and well supported.

Effect of publishing surgeon-specific outcomes on surgical training.

BACKGROUND: Over the past 10 years, the National Health Service in England has started to publish surgeon-specific outcomes publicly. The aim of this study was to investigate how this has affected training case exposure for surgeons in training. METHODS: Anonymized data were collected from the Intercollegiate Surgical Curriculum Programme database for operations in each specialty with published surgeon outcomes, involving surgical trainees on an approved training programme between 1 January 2011 and 31 December 2016. Trainee and supervisor involvement in operations before and after the start of publication of surgeon-specific outcomes were compared using mixed-effects models. RESULTS: A total of 163 076 recorded operative procedures were included. A statistically significant improvement in exposure to training procedures was observed for anterior resection of rectum, carotid endarterectomy, gastrectomy, meningioma excision, prostatectomy and thyroidectomy following the introduction of publication of surgeon outcomes. In coronary artery bypass grafting (CABG) and total hip replacement (THR), however, there was a reduction in involvement in training procedures. This was apparent for both trainee and supervisor involvement in CABG, and for trainee involvement in THR. CONCLUSION: Exposure to training procedures has improved rather than declined in the UK in the majority of surgical specialties, since the publication of surgeon-specific outcomes.

The effects of human factor related issues on assessors during the recruitment process for general and vascular surgery in the UK.

INTRODUCTION: Healthcare professionals increasingly recognise how human factors (HF) can contribute to medical error. An understanding of HF is also important during other high stakes activities such as summative assessments or examinations. National organisations hosting such events need to be aware of potential fatigue, boredom and stress that can occur in interviewers during these often repetitive activities. METHODS: A previously validated questionnaire based around the well known HF analysis and classification system (HFACS) was used to evaluate four factors (care and support, asking questions about the role as an interviewer, working within the rules and boundaries, and stress and pressure) at the 2018 UK general and vascular surgery trainee national selection process. RESULTS: A total of 92 questionnaires were completed and analysed (48% response rate). After recoding for negative phrased questions, no significant differences were found between years of experience in the selection process and mean scores obtained for all four factor items. Interviewers had a positive experience during national selection with mean factor scores ranging from 3.84 to 3.98 (out of a maximum satisfaction score of 5). CONCLUSIONS: Organisations need to plan carefully and recognise the human element to ensure that their interviewers are cared for during any high stakes assessment such as national selection. Our work suggests that a positive assessor experience will further help contribute to a reliable and fair recruitment process.

Expression of growth mediators in the gingival crevicular fluid of patients with aggressive periodontitis undergoing periodontal surgery.

OBJECTIVES: To describe changes in growth factor mediators in the gingival crevicular fluid (GCF) of patients with aggressive periodontitis (AgP) undergoing regenerative (GTR) and access flap (AF) surgery. MATERIALS AND METHODS: This was a 12-month, single-blind, split-mouth RCT involving 18 AgP patients with a bilateral intrabony defect which was treated with GTR or AF. GCF was collected prior to surgery and at subsequent follow-up visits from 3 days to 12 months post-operatively, and the levels of angiopoietin-1 (Ang-1), vascular-endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), bone morphogenetic protein-2 (BMP-2), osteoprotegerin (OPG), tissue inhibitor of metalloproteinase 1 (TIMP-1), keratinocyte growth factor (KGF) and platelet-derived growth factor-AB (PDGF-AB) were measured. At baseline, 6 and 12 months post-surgery, periodontal clinical parameters were evaluated. ANOVA was applied to test for differences in the amount of mediators (p < 0.05). RESULTS: Higher amounts of BMP-2 and OPG and a higher area under the curve (AUC) of KGF at the GTR versus AF sites were observed. The maximum change in the amount of KGF correlated significantly with periodontal clinical parameters at the GTR sites at 6 and 12 months. The AUC over 30 days of the amount of Ang-1, VEGF and KGF significantly correlated with periodontal clinical parameters at the AF sites at 6 months. CONCLUSIONS: AF and GTR differentially affected the profile of the growth mediators in GCF, and significant correlations between certain GCF mediators and periodontal clinical outcomes were identified. CLINICAL RELEVANCE: GCF components represent attractive prognostic markers for periodontal tissues undergoing repair or regeneration. However, the available evidence is not robust enough to suggest the use of a specific marker, and future adequately powered studies are warranted to identify the most relevant mediators that could be applied in clinical practice.

Absorption and Metabolism of Peptide WDHHAPQLR Derived from Rapeseed Protein and Inhibition of HUVEC Apoptosis under Oxidative Stress.

WDHHAPQLR (RAP) is an antioxidative peptide derived from rapeseed protein. Although the health benefits from RAP, due to its antioxidant activities, have been determined by chemical methods, a systematic assessment regarding the absorption, metabolism, and antioxidation processes of RAP is still lacking attention. Hence, Caco-2 cell monolayer models and animal experiments were used to evaluate the absorption and bioavailability of RAP. As expected, RAP could be absorbed by intestinal epithelial cells, and the Papp was 0.82 ± 0.19 × 10-6 cm/s. Three main fragments, RAP, DHHAPQLR, and WDHHAP were transported by the paracellular pathway, and QLR was transported by PepT1. An important modified product of RAP (EGDHHAPQLR) was found to contribute to the elimination of intracellular reactive oxygen species. The absolute bioavailability of RAP was 3.56%, and three degradation products of RAP were also detected in rat serum. More importantly, RAP exerts its antioxidant activity by inhibiting the apoptosis of oxidative stress cells. RAP could downregulate the expression of Bax and caspase-3 and upregulate the expression of Bcl-2 in H2O2-induced HUVECs (human umbilical vein endothelial cells). In general, using in vitro and in vivo experimental models, the in vivo absorption and transformation processes of RAP and its antioxidative molecular mechanisms by inhibiting apoptosis of cells were revealed.

Long-term results and recurrence patterns from SCALOP: a phase II randomised trial of gemcitabine- or capecitabine-based chemoradiation for locally advanced pancreatic cancer.

BACKGROUND: SCALOP, a randomised, phase II trial, tested the activity and safety of gemcitabine (GEM)-based and capecitabine (CAP)-based chemoradiation (CRT) for locally advanced pancreatic cancer (LAPC). Here we present the long-term outcomes. METHODS: Eligibility: histologically proven LAPC ⩽7 cm. Following 12 weeks of induction GEMCAP chemotherapy (three cycles: GEM 1000 mg m-2 days 1, 8, 15; CAP 830 mg m-2 days 1-21 q28 days) patients with stable/responding disease, tumour ⩽6 cm, and WHO Performance Status 0-1 were randomised to receive one cycle GEMCAP followed by CAP (830 mg m-2 b.d. on weekdays only) or GEM (300 mg m-2 weekly) with radiation (50.4 Gy per 28 fractions). RESULTS: One-hundred fourteen patients (28 UK centres) were registered between 24 December 2009 and 25 October 2011, and 74 were randomised (CAP-RT=36; GEM-RT=38). At the time of this analysis, 105 of the 114 patients had died and the surviving 9 patients had been followed up for a median of 10.9 months (IQR: 2.9-18.7). Updated median OS was 17.6 months (95% CI: 14.6-22.7) in the CAP-CRT arm and 14.6 months (95% CI: 11.1-16.0) in the GEM-CRT arm (intention-to-treat adjusted hazard ratio (HR): 0.68 (95% CI: 0.38-1.21, P=0.185)); median progression-free survival (PFS) was 12.0 months (95% CI: 10.0-15.2) in the CAP-CRT arm and 10.4 months (95% CI: 8.8-12.7) in the GEM-CRT arm (intention-to-treat adjusted HR: 0.60 (95% CI: 0.32-1.14, P=0.120)). In baseline multivariable model, age ⩾65 years, better performance status, CA19.9<613 IU l-1, and shorter tumour diameter predicted improved OS. CAP-CRT, age ⩾65 years, better performance status, CA19.9 <46 IU ml-1 predicted improved OS and PFS in the pre-radiotherapy model. Nine-month PFS was highly predictive of OS. CONCLUSIONS: CAP-CRT remains the superior regimen. SCALOP showed that patients with CA19.9 <46 IU ml-1 after induction chemotherapy are more likely to benefit from CRT.

Long-term results and recurrence patterns from SCOPE-1: a phase II/III randomised trial of definitive chemoradiotherapy +/- cetuximab in oesophageal cancer.

BACKGROUND: The SCOPE-1 study tested the role of adding cetuximab to conventional definitive chemoradiotherapy (dCRT), and demonstrated greater toxicity and worse survival outcomes. We present the long-term outcomes and patterns of recurrence. METHODS: SCOPE-1 was a phase II/III trial in which patients were randomised to cisplatin 60 mg m-2 (day 1) and capecitabine 625 mg m-2 bd (days 1-21) for four cycles +/- cetuximab 400 mg m-2 day 1 then by 250 mg m-2 weekly. Radiotherapy consisted of 50 Gy/25# given concurrently with cycles 3 and 4. Recruitment was between February 2008 and February 2012, when the IDMC recommended closure on the basis of futility. RESULTS: About 258 patients (dCRT=129; dCRT+cetuximab (dCRT+C)=129) were recruited from 36 centres. About 72.9% (n=188) had squamous cell histology. The median follow-up (IQR) was 46.2 (35.9-48.3) months for surviving patients. The median overall survival (OS; months; 95% CI) was 34.5 (24.7-42.3) in dCRT and 24.7 (18.6-31.3) in dCRT+C (hazard ratio (HR)=1.25, 95% CIs: 0.93-1.69, P=0.137). Median progression-free survival (PFS; months; 95% CI) was 24.1 (15.3-29.9) and 15.9 (10.7-20.8) months, respectively (HR=1.28, 95% CIs: 0.94-1.75; P=0.114). On multivariable analysis only earlier stage, full-dose RT, and higher cisplatin dose intensity were associated with improved OS. CONCLUSIONS: The mature analysis demonstrates that the dCRT regimen used in the study provided useful survival outcomes despite its use in patients who were largely unfit for surgery or who had inoperable disease. Given the competing risk of systemic and local failure, future studies should continue to focus on enhancing local control as well as optimising systemic therapy.

Impact of performance in a mandatory postgraduate surgical examination on selection into specialty training.

BACKGROUND: The Intercollegiate Membership of the Royal College of Surgeons (MRCS) examination is undertaken by large numbers of trainees in the UK and internationally as a mandatory step within surgical training. Unlike some high-stakes medical examinations, the MRCS is yet to be validated. A quantitative study was undertaken to assess its predictive validity by investigating the relationship between MRCS (Parts A and B) and national selection interview scores for general and vascular surgery in the UK. METHODS: Pearson correlation coefficients were used to examine the linear relationship between each assessment, and linear regression analyses were employed to identify potential independent predictors of the national selection score. All UK medical graduates who attempted the interview in 2011-2015 were included. RESULTS: Some 84·4 per cent of the candidates (1231 of 1458) were matched with MRCS data. There was a significant positive correlation between the first attempt score at Part B of the MRCS examination and the national selection score (r = 0·38, P < 0·001). In multivariable analysis, 17 per cent of variance in the national selection first attempt score was explained by the Part B MRCS score and number of attempts (change in R 2 value of 0·10 and 0·07 respectively; P < 0·001). Candidates who required more than two attempts at Part B were predicted to score 8·1 per cent less than equally matched candidates who passed at their first attempt. CONCLUSION: This study supports validity of the MRCS examination, and indicates its predictive value regarding entry into specialist training.

Meta-analysis of operative experiences of general surgery trainees during training.

BACKGROUND: General surgical training curricula around the world set defined operative numbers to be achieved before completion of training. However, there are few studies reporting total operative experience in training. This systematic review aimed to quantify the published global operative experience at completion of training in general surgery. METHODS: Electronic databases were searched systematically for articles in any language relating to operative experience in trainees completing postgraduate general surgical training. Two reviewers independently assessed citations for inclusion using agreed criteria. Studies were assessed for quantitative data in addition to study design and purpose. A meta-analysis was performed using a random-effects model of studies with appropriate data. RESULTS: The search resulted in 1979 titles for review. Of these, 24 studies were eligible for inclusion in the review and data from five studies were used in the meta-analysis. Studies with published data of operative experience at completion of surgical training originated from the USA (19), UK (2), the Netherlands (1), Spain (1) and Thailand (1). Mean total operative experience in training varied from 783 procedures in Thailand to 1915 in the UK. Meta-analysis produced a mean pooled estimate of 1366 (95 per cent c.i. 1026 to 1707) procedures per trainee at completion of training. There was marked heterogeneity between studies (I2 = 99·6 per cent). CONCLUSION: There is a lack of robust data describing the operative experiences of general surgical trainees outside the USA. The number of surgical procedures performed by general surgeons in training varies considerably across the world.

Patient-reported outcomes during and after definitive chemoradiotherapy for oesophageal cancer.

BACKGROUND: Limited data describe patient-reported outcomes (PROs) of localised oesophageal cancer treated with definitive chemoradiotherapy(CRT). The phase 2/3 SCOPE-1 trial assessed the effectiveness of CRT±cetuximab. The trial for the first time provided an opportunity to describe PROs from a multi-centre group of patients treated with CRT that are presented here. METHODS: Patients undergoing CRT±cetuximab within the SCOPE-1 trial (258 patients from 36 UK centres) completed generic-, disease- and treatment-specific health-related quality of life (HRQL) questionnaires (EORTC QLQ-C30, QLQ-OES18, Dermatology Life-Quality Index (DLQI)) at baseline and at 7, 13, 24, 52 and 104 weeks. Mean EORTC functional scale scores (>15 point change significant), DLQI scores (>4 point change significant) and proportions of patients (>15% significant) with 'minimal' or 'severe' symptoms are presented. RESULTS: Questionnaire response rates were good. At baseline, EORTC functional scores were high (>75%) and few symptoms were reported except for severe problems with fatigue, insomnia and eating-related symptoms (e.g., appetite loss, dysphagia, dry mouth) in both groups(>15%). Functional aspects of health deteriorated and symptoms increased with treatment and by week 13 global quality of life, physical, role and social function significantly deteriorated and more problems with fatigue, dyspnoea, appetite loss and trouble with taste were reported. Recovery occurred by 6 months (except severe fatigue and insomnia in >15% of patients) and maintained at follow-up with no differences between groups. CONCLUSIONS: CRT for localised oesophageal cancer has a significant detrimental impact on many aspects of HRQL; however, recovery is achieved by 6 months and maintained with the exception of persisting problems with severe fatigue and insomnia. The data suggest that the HRQL recovery after definitive CRT is quicker, and there is little lasting deficit compared with treatment including surgery. These data need to be compared with HRQL data from studies evaluating treatments including surgery for oesophageal cancer.

Effective disinfection of rough rice using infrared radiation heating.

The objective of this study was to investigate the effect of infrared (IR) heating and tempering treatments on disinfection of Aspergillus flavus in freshly harvested rough rice and storage rice. Rice samples with initial moisture contents (IMCs) of 14.1 to 27.0% (wet basis) were infected with A. flavus spores before the tests. The infected samples were heated by IR radiation to 60°C in less than 1 min, and then samples were tempered at 60°C for 5, 10, 20, 30, 60, or 120 min. High heating rates and corresponding high levels of moisture removal were achieved using IR heating. The highest total moisture removal was 5.3% for the fresh rice with an IMC of 27.0% after IR heating and then 120 min of tempering. IR heating followed by tempering for 120 min resulted in 2.5- and 8.3-log reductions of A. flavus spores in rough rice with the lowest and highest IMCs, respectively. To study the effect on disinfection of rewetting dried storage rice, the surface of the dry rice was rewetted to achieve IMCs of 14.7 to 19.4% (wet basis). The rewetting process for the dry rice had a significant effect on disinfection. IR heating followed by tempering for 60 min resulted in 7.2-log reductions in A. flavus on rewetted rough rice. The log-linear plus tail model was applied to estimate the tempering time needed to achieve a 5-log reduction of A. flavus in rice of different IMCs. At least 30 and 20 min of tempering were needed for fresh rice and rewetted rice, respectively, with the highest IMCs. The recommended conditions of simultaneous disinfection and drying for fresh rice was IR heating to 60°C followed by tempering for 120 min and natural cooling, resulting in a final MC of 16.5 to 22.0%, depending on the IMC. For the rewetted dry rice with an IMC of 19.4%, the recommended condition for disinfection and drying involved only 20 min of tempering. The final MC of the sample was 13.8%, which is a safe MC for storage rice.

Addressing bioterrorism concerns: options for investigating the mechanism of action of Staphylococcus aureus enterotoxin B.

Staphylococcal enterotoxin B (SEB) is of concern to military and civilian populations as a bioterrorism threat agent. It is a highly potent toxin produced by Staphylococcus aureus and is stable in storage and under aerosolisation; it is able to produce prolonged highly incapacitating illness at very low-inhaled doses and death at elevated doses. Concerns regarding SEB are compounded by the lack of effective medical countermeasures for mass treatment of affected populations. This article considers the mechanism of action of SEB, the availability of appropriate experimental models for evaluating the efficacy of candidate medical countermeasures with particular reference to the need to realistically model SEB responses in man and the availability of candidate countermeasures (with an emphasis on commercial off-the-shelf options). The proposed in vitro approaches would be in keeping with Dstl's commitment to reduction, refinement and replacement of animal models in biomedical research, particularly in relation to identifying valid alternatives to the use of nonhuman primates in experimental studies.

Study of optimal extraction conditions for achieving high yield and antioxidant activity of tomato seed oil.

UNLABELLED: Value of tomato seed has not been fully recognized. The objectives of this research were to establish suitable processing conditions for extracting oil from tomato seed by using solvent, determine the impact of processing conditions on yield and antioxidant activity of extracted oil, and elucidate kinetics of the oil extraction process. Four processing parameters, including time, temperature, solvent-to-solid ratio and particle size were studied. A second order model was established to describe the oil extraction process. Based on the results, increasing temperature, solvent-to-solid ratio, and extraction time increased oil yield. In contrast, larger particle size reduced the oil yield. The recommended oil extraction conditions were 8 min of extraction time at temperature of 25 °C, solvent-to-solids ratio of 5/1 (v/w) and particle size of 0.38 mm, which gave oil yield of 20.32% with recovery rate of 78.56%. The DPPH scavenging activity of extracted oil was not significantly affected by the extraction parameters. The inhibitory concentration (IC(50) ) of tomato seed oil was 8.67 mg/mL which was notably low compared to most vegetable oils. A 2nd order model successfully described the kinetics of tomato oil extraction process and parameters of extraction kinetics including initial extraction rate (h), equilibrium concentration of oil (C(s) ), and the extraction rate constant (k) could be precisely predicted with R(2) of at least 0.957. PRACTICAL APPLICATION: The study revealed that tomato seed which is typically treated as a low value byproduct of tomato processing has great potential in producing oil with high antioxidant capability. The impact of processing conditions including time, temperature, solvent-to-solid ratio and particle size on yield, and antioxidant activity of extracted tomato seed oil are reported. Optimal conditions and models which describe the extraction process are recommended. The information is vital for determining the extraction processing conditions for industrial production of high quality tomato seed oil.

Using in-vivo fluorescence imaging in personalized cancer diagnostics and therapy, an image and treat paradigm.

The major goal in developing drugs targeting specific tumor receptors, such as Monoclonal AntiBodies (MAB), is to make a drug compound that targets selectively the cancer-causing biomarkers, inhibits their functionality, and/or delivers the toxin specifically to the malignant cells. Recent advances in MABs show that their efficacy depends strongly on characterization of tumor biomarkers. Therefore, one of the main tasks in cancer diagnostics and treatment is to develop non-invasive in-vivo imaging techniques for detection of cancer biomarkers and monitoring their down regulation during the treatment. Such methods can potentially result in a new imaging and treatment paradigm for cancer therapy. In this article we have reviewed fluorescence imaging approaches, including those developed in our group, to detect and monitor Human Epidermal Growth Factor 2 (HER2) receptors before and during therapy. Transition of these techniques from the bench to bedside is the ultimate goal of our project. Similar approaches can be used potentially for characterization of other cancer related cell biomarkers.

R-Ras interacts with filamin a to maintain endothelial barrier function.

The molecular mechanisms regulating vascular barrier integrity remain incompletely elucidated. We have previously reported an association between the GTPase R-Ras and repeat 3 of Filamin A (FLNa). Loss of FLNa has been linked to increased vascular permeability. We sought to determine whether FLNa's association with R-Ras affects endothelial barrier function. We report that in endothelial cells endogenous R-Ras interacts with endogenous FLNa as determined by co-immunoprecipitations and pulldowns with the FLNa-GST fusion protein repeats 1-10. Deletion of FLNa repeat 3 (FLNaΔ3) abrogated this interaction. In these cells FLNa and R-Ras co-localize at the plasma membrane. Knockdown of R-Ras and/or FLNa by siRNA promotes vascular permeability, as determined by TransEndothelial Electrical Resistance and FITC-dextran transwell assays. Re-expression of FLNa restored endothelial barrier function in cells lacking FLNa whereas re-expression of FLNaΔ3 did not. Immunostaining for VE-Cadherin in cells with knocked down R-Ras and FLNa demonstrated a disorganization of VE-Cadherin at adherens junctions. Loss of R-Ras and FLNa or blocking R-Ras function via GGTI-2133, a selective R-Ras inhibitor, induced vascular permeability and increased phosphorylation of VE-Cadherin (Y731) and Src (Y416). Expression of dominant negative R-Ras promoted vascular permeability that was blocked by the Src inhibitor PP2. These findings demonstrate that maintaining endothelial barrier function is dependent upon active R-Ras and association between R-Ras and FLNa and that loss of this interaction promotes VE-Cadherin phosphorylation and changes in downstream effectors that lead to endothelial leakiness.

A phase II trial of continuous 5-fluorouracil in recurrent or metastatic transitional cell carcinoma of the urinary tract.

AIMS: To assess the activity of a continuous infusion of 5-fluorouracil in patients with recurrent locally advanced or metastatic transitional cell carcinoma of the urinary tract. MATERIALS AND METHODS: Eight centres within the UK entered 50 patients into the study. Twenty-four weeks of continuously infused 5-fluorouracil, 300mg/m(2)/day through a mini-pump, were planned. The primary outcome was tumour response at 8 weeks after the start of treatment. RESULTS: The median age of the patients was 68 years and 37 (80.4%) had a World Health Organization performance status of 0 or 1. The overall response rate at 8 weeks, according to the response evaluation criteria in solid tumors (RECIST) criteria in 46 evaluable patients, was 15% (95% confidence interval 5-26%) and 20% (95% confidence interval 8-31%) when assessments at all time points were included. The median progression-free survival was 1.9 months (95% confidence interval 1.8-2.7 months) and the median overall survival was 6.5 months (95% confidence interval 4.1-8.5 months). The most frequent grade 3/4 toxicities were mucositis and diarrhoea (each in 6.5% of patients) and nausea/vomiting and hand-foot syndrome (each in 4.3% of patients). CONCLUSIONS: Continuous infusional 5-fluorouracil has activity in transitional cell carcinoma of the urinary tract. Prolonged fluoropyrimidine administration may be a useful component of future combination regimens for this disease, particularly in patients with poor renal function.