Flavonoid-Rich Fruit Intake in Midlife and Late-Life and Associations with Risk of Dementia: The Framingham Heart Study.

BACKGROUND: Fruits are an important source of flavonoids, and greater intake of dietary flavonoids in older adults has been shown to be associated with decreased risk of dementia. It is unclear whether this relationship is similar or different between younger adults and older adults. OBJECTIVES: We examined for associations between midlife and late-life intake of flavonoid-rich fruits and incident dementia. We hypothesized that greater total cumulative intake of flavonoid-rich fruits in midlife and late-life adults would be associated with reduced risk of all-cause dementia. DESIGN: Longitudinal, cohort study design. SETTING: Framingham Heart Study, which is a longitudinal, multi-generational community-based cohort based in Framingham, Massachusetts, USA. PARTICIPANTS: Participants from the Framingham Heart Study Offspring cohort were included (n = 2,790) who attended the fifth core exam between 1991 to 1995, and were dementia-free and at least 45 years of age at that time, as well as had valid food frequency questionnaires from the fifth to ninth core exams. MEASUREMENTS: Consumption of fruits with high flavonoid content or are important contributors to overall flavonoid intake was collected via food frequency questionnaire. Flavonoid-rich fruits from the food frequency questionnaire included raisins or grapes, prunes, bananas, fresh apples or pears, apple juice or cider, oranges, orange juice, grapefruit, grapefruit juice, strawberries, blueberries, and peaches, apricots, or plums. Dementia ascertainment was based on a multidisciplinary consensus committee, and included all-cause dementia and Alzheimer's disease dementia diagnoses based on research criteria. Cox models were used to examine associations between cumulative fruit intake and incident dementia, stratified by midlife (45-59 years; n = 1,642) and late-life (60-82 years; n = 1,148). RESULTS: Greater cumulative total fruit intake in midlife, but not late-life, was significantly associated with a 44% decreased risk of all-cause dementia (HR = 0.56; 95% CI = 0.32 - 0.98; p = 0.044). Decreased risk of all-cause dementia was also associated with higher intake of apples or pears in midlife and late-life, as well as higher intake of raisins or grapes in midlife only, and higher intake of oranges, grapefruit, blueberries, and peaches, apricots, or plums in late-life only. CONCLUSIONS: Among participants from the Framingham Heart Study, greater overall consumption of flavonoid-rich fruits in midlife was associated with reduced risk of dementia, though intake of specific fruits in midlife and late-life may have a protective role against developing dementia. These findings may help to inform future recommendations on when dietary interventions may be most beneficial to healthy brain aging across the life course.

Immune microenvironment modulation following neoadjuvant therapy for oesophageal adenocarcinoma: a translational analysis of the DEBIOC clinical trial.

BACKGROUND: The Dual Erb B Inhibition in Oesophago-gastric Cancer (DEBIOC) trial reported an acceptable safety profile for neoadjuvant oxaliplatin and capecitabine (Xelox) ± AZD8931 in oesophageal adenocarcinoma (OAC) but limited efficacy. We evaluated the impact of neoadjuvant Xelox ± AZD8931, a novel small-molecule inhibitor with equipotent activity against epidermal growth factor receptor (EGFR), human epidermal growth factor receptor (HER)2 and HER3, on biological pathways using a unique software-driven solution. PATIENTS AND METHODS: Transcriptomic profiles from 25 pre-treatment formalin-fixed paraffin-embedded OAC biopsies and 18 matched resection specimens, treated with Xelox + AZD8931 (n = 16) and Xelox alone (n = 9), were analysed using the Almac claraT total mRNA report analysing 92 gene signatures, 100 unique single-gene drug targets and 7337 single genes across 10 hallmarks of cancer. Gene-set enrichment analysis (GSEA) was utilised to investigate pathways governing pathological response. Tumour-infiltrating lymphocytes (TILs) were assessed digitally using the QuPath software. RESULTS: Hierarchical clustering identified three molecular subgroups classified by activation of innate immune signalling. The immune-high subgroup was associated with HER2 positivity, increased pathological response and a marked reduction in immune signalling and TILs following neoadjuvant therapy. The immune-low cluster was predominantly HER2/EGFR-negative, and EGFR positivity was associated with the immune-mixed subgroup. Treatment with neoadjuvant therapy induced common resistance mechanisms, such as angiogenesis and epithelial-mesenchymal transition signalling, and a reduction in DNA repair signatures. Addition of AZD8931 was associated with reduction of expression of EGFR, HER2 and AKT pathways and also promoted an immunosuppressive microenvironment. GSEA showed that patients with a pathological response to treatment had increased immune signalling, whereas non-responders to neoadjuvant therapy were enriched for nucleotide repair and cellular growth through the action of E2F transcription factors. CONCLUSION: OAC may be subdivided into three immune-related subgroups which undergo modulation in response to neoadjuvant therapy with marked suppression of the immune microenvironment in HER2-positive/immune-high tumours.

Perspectives on improving photosynthesis to increase crop yield.

Improving photosynthesis, the fundamental process by which plants convert light energy into chemical energy, is a key area of research with great potential for enhancing sustainable agricultural productivity and addressing global food security challenges. This perspective delves into the latest advancements and approaches aimed at optimizing photosynthetic efficiency. Our discussion encompasses the entire process, beginning with light harvesting and its regulation and progressing through the bottleneck of electron transfer. We then delve into the carbon reactions of photosynthesis, focusing on strategies targeting the enzymes of the Calvin-Benson-Bassham (CBB) cycle. Additionally, we explore methods to increase carbon dioxide (CO2) concentration near the Rubisco, the enzyme responsible for the first step of CBB cycle, drawing inspiration from various photosynthetic organisms, and conclude this section by examining ways to enhance CO2 delivery into leaves. Moving beyond individual processes, we discuss two approaches to identifying key targets for photosynthesis improvement: systems modeling and the study of natural variation. Finally, we revisit some of the strategies mentioned above to provide a holistic view of the improvements, analyzing their impact on nitrogen use efficiency and on canopy photosynthesis.

Rigid bronchoscopy: a consultant survey.

INTRODUCTION: Inhalation of foreign bodies represents a potentially fatal emergency in both adults and children. Chest x-ray, in isolation, is neither sensitive nor specific. Rigid bronchoscopy represents the gold standard to diagnose and retrieve paediatric foreign bodies. Cases are encountered infrequently, creating anxieties about their management. Little is known about the confidence in, and maintenance of, rigid bronchoscopy skills by ear, nose and throat teams. METHODS: A 15-question survey was completed by 50 practising otolaryngology consultants in England. RESULTS: Results show that almost 40% of otolaryngology consultants covering rigid bronchoscopy have not performed bronchoscopy in more than 5 years. Consultants raised concerns about the anaesthetic support and the speed of equipment assembly. Questions on clinical practice showed disparities in practice in the same scenario. CONCLUSIONS: The authors advocate addressing many of the issues raised by the study with a greater availability of simulation courses and regular scheduled intradepartmental teaching days for all professionals involved. National guidelines on criteria for transfer to tertiary centres would improve the consistency of practice.

The influence of spinal venous blood pressure on cerebrospinal fluid pressure.

In Alligator mississippiensis the spinal dura is surrounded by a venous sinus; pressure waves can propagate in the spinal venous blood, and these spinal venous pressures can be transmitted to the spinal cerebrospinal fluid (CSF). This study was designed to explore pressure transfer between the spinal venous blood and the spinal CSF. At rest the cardiac-related CSF pulsations are attenuated and delayed, while the ventilatory-related pulsations are amplified as they move from the spinal venous blood to the spinal CSF. Orthostatic gradients resulted in significant alterations of both cardiac- and ventilatory-related CSF pulsations. Manual lateral oscillations of the alligator's tail created pressure waves in the spinal CSF that propagated, with slight attenuation but no delay, to the cranial CSF. Oscillatory pressure pulsations in the spinal CSF and venous blood had little influence on the underlying ventilatory pulsations, though the same oscillatory pulsations reduced the ventilatory- and increased the cardiac-related pulsations in the cranial CSF. In Alligator the spinal venous anatomy creates a more complex pressure relationship between the venous and CSF systems than has been described in humans.

Impact of pod and seed photosynthesis on seed filling and canopy carbon gain in soybean.

There is a limited understanding of the carbon assimilation capacity of nonfoliar green tissues and its impact on yield and seed quality since most photosynthesis research focuses on leaf photosynthesis. In this study, we investigate the photosynthetic efficiency of soybean (Glycine max) pods and seeds in a field setting and evaluate its effect on mature seed weight and composition. We demonstrate that soybean pod and seed photosynthesis contributes 13% to 14% of the mature seed weight. Carbon assimilation by soybean pod and seed photosynthesis can compensate for 81% of carbon loss through the respiration of the same tissues, and our model predicts that soybean pod and seed photosynthesis contributes up to 9% of the total daily carbon gain of the canopy. Chlorophyll fluorescence (CF) shows that the operating efficiency of photosystem II in immature soybean seeds peaks at the 10 to 100 mg seed weight stage, while that of immature pods peaks at the 75 to 100 mg stage. This study provides quantitative information about the efficiency of soybean pod and seed photosynthesis during tissue development and its impact on yield.

Sensitivity and performance of three novel quantitative assays of SARS-CoV-2 nucleoprotein in blood.

To assess if SARS-CoV-2 (COVID-19) systemic disease can be determined by available nucleoprotein assays, we compared the performance of three commercial SARS-CoV-2 nucleoprotein (N) assays in plasma. A total of 272 plasma samples collected in the period November-December 2021 were analyzed by the methods Simoa SARS CoV-2 N Protein Advantage Kit [Quanterix Simoa], Solsten SARS-CoV-2 Antigen enzyme immunosorbent assay (ELISA) [Solsten ELISA], and Elecsys SARS-CoV-2 Antigen electrochemiluminescence immunoassay [Elecsys ECLIA]. Additionally, a dilution series of inactivated virus culture was analyzed by the three assays. The SARS CoV-2 PCR-status was not known for the patients. Linear correlation in the pairwise correlation between assays as well as linearity of dilution series of inactivated virus culture was estimated by Spearman score. Sensitivity and specificity were estimated by pairwise comparison. The three assays showed poor agreement on patient samples with regards to concentration. Performance on virus culture was excellent but with different level of detection (LOD). Positive vs negative results show comparable sensitivity and specificity of Quanterix Simoa and Solsten ELISA, with a higher LOD in Elecsys ECLIA and thus lower sensitivity and high specificity. N by all tested assays can be used as a marker for systemic COVID-19 disease.

Psychological impact of lung cancer screening using a novel antibody blood test followed by imaging: the ECLS randomized controlled trial.

BACKGROUND: The Early CDT®-Lung antibody blood test plus serial computed tomography scans for test-positives (TPGs) reduces late-stage lung cancer presentation. This study assessed the psychological outcomes of this approach. METHODS: Randomized controlled trial (n = 12 208) comparing psychological outcomes 1-12 months post-recruitment in a subsample (n = 1032) of TPG, test-negative (TNG) and control groups (CG). RESULTS: Compared to TNG, TPG had lower positive affect (difference between means (DBM), 3 months (3m: -1.49 (-2.65, - 0.33)), greater impact of worries (DBM 1m: 0.26 (0.05, 0.47); 3m: 0.28 (0.07, 0.50)), screening distress (DBM 1m: 3.59 (2.28, 4.90); 3m: 2.29 (0.97, 3.61); 6m: 1.94 (0.61, 3.27)), worry about tests (odds ratio (OR) 1m: 5.79 (2.66, 12.63) and more frequent lung cancer worry (OR 1m: 2.52 (1.31, 4.83); 3m: 2.43 (1.26, 4.68); 6m: 2.87 (1.48, 5.60)). Compared to CG, TPG had greater worry about tests (OR 1m: 3.40 (1.69, 6.84)). TNG had lower negative affect (log-transformed DBM 3m: -0.08 (-0.13, -0.02)), higher positive affect (DBM 1m: 1.52 (0.43, 2.61); 3m: 1.43 (0.33, 2.53); 6m: 1.27 (0.17, 2.37)), less impact of worries (DBM 3m: -0.27 (-0.48, -0.07)) and less-frequent lung cancer worry (OR 3m: 0.49 (0.26, 0.92)). CONCLUSIONS: Negative psychological effects in TPG and positive effects in TNG were short-lived and most differences were small.

Evaluation of a novel heparin-iloprost-based antithrombotic formulation blood collection tube for clinical usage.

BACKGROUND: The objective of our study was to evaluate a single blood collection tube with a novel antithrombotic formulation to measure both hematological, biochemical, and d-dimer analytes. METHODS: Paired samples of gold standard blood tubes (EDTA, lithium heparin, sodium citrate) and a new antithrombotic formulation blood tube were collected from 187 patients. The new antithrombotic tube is a lithium heparin tube preloaded with a liquid form of prostacyclin analog. The novel tube was tested on seventeen hematological parameters and smears against EDTA, on fourteen biochemical parameters against lithium heparin and on d-dimer against sodium citrate. RESULTS: All correlation coefficients were close to 0.99. The Bland-Altman analyses presented a satisfactory correspondence for all analytes. All the hematological examinations demonstrated comparable results between EDTA and the novel formulation, except for platelet counts analyzed by impedance method, but not by fluorescence. We detected lower mean platelet volume with/without outliers (5.06%)/(5.13%) in the novel formulation and increased mean corpuscular hemoglobin concentration (2.55%). All the biochemistry analytes demonstrated comparable results between lithium heparin and the novel tube. d-dimer showed comparable results between citrated blood and the novel formulation after dilution correction. CONCLUSIONS: We describe a novel antithrombotic formulation tube with the potential to be introduced into clinical laboratories for simultaneous analysis of thirty-two blood analytes.

Long-Term Comparison of 7 SARS-CoV-2 Antibody Assays in the North Zealand Covid-19 Cohort.

BACKGROUND: Throughout the coronavirus disease 2019 (Covid-19) pandemic numerous severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibody assays have been approved through Emergency Use Authorization and require further evaluation of sensitivity and specificity in clinical laboratory settings prior to implementation. METHODS: We included 1733 samples from 375 PCR-confirmed SARS-CoV-2-positive individuals of the North Zealand Covid-19 Cohort in an 8-month period. We investigated diagnostic sensitivity and specificity against consensus and PCR and interassay agreement over time for 5 SARS-CoV-2 immunoassays [Roche-nucleocapsid (NC)-total, Roche-receptor binding domain (RBD)-total, Siemens-RBD-IgG, Siemens-RBD-total, Thermo Fisher Scientific (TFS)-RBD-IgG] commercially available on automated platforms and 2 ELISA assays (TFS-RBD-total, Wantai-RBD-total). RESULTS: Early interassay discrepancy in up to 49% of samples decreased steadily during the first 18 days. By day 18, all assays had reached a plateau between 82.3% and 90.5% seropositivity compared to PCR. Assays ranked by closest agreement with the consensus model beyond day 18 (sensitivity/specificity against consensus) were as follows: Roche-RBD-total, 99.8%/100.0%; Wantai-RBD-total, 99.8%/99.7%; Roche-NC-total, 97.8%/100.0%; Siemens-RBD-total, 98.0%/98.7%; TFS-RBD-total, 96.9%/99.7%; TFS-RBD-IgG, 91.5%/100.0%; and Siemens-RBD-IgG, 94.6%/89.9%. We found that 7.8% of PCR-positive patients remained seronegative in all assays throughout the study. CONCLUSIONS: All included assays had sensitivities against consensus >90% past day 18. For the current recommended use of antibody assays to detect former, undocumented Covid-19, our data suggest the use of total antibody assays rather than IgG-specific assays due to higher long-term sensitivity. Finally, a nonresponding subpopulation of 7.8% in our cohort with persistent seronegative results raises concern of a possible substantial number of people with continued low protection following natural SARS-CoV-2 infection.

Cochlear implant eligibility in an adult hearing aid population: a multi-perspective service evaluation of a patient referral pathway at a British district general hospital.

OBJECTIVES: To quantify patient eligibility for cochlear implantation following National Institute for Health and Care Excellence 2019 guidelines (TA566) over five years at our institution, and identify factors influencing patients' decisions surrounding cochlear implantation referral. METHODS: A multi-perspective service evaluation was conducted at a district general hospital, comprising cochlear implantation eligible patients. The main outcome measures were: eligibility numbers for 2014-2019, comparing application of TA566 versus 2009 (TA166) guidelines; and patient interview transcripts and questionnaires. RESULTS: There was a 259 per cent average increase in cochlear implantation eligibility from 2014 to 2019. Most patients' thresholds were 80 dB HL or more at 3 kHz and 4 kHz. There are several cochlear implantation barriers, including patient-centred issues (e.g. health-related anxieties, implantation misperceptions) and external barriers (difficulty getting to regional implant centres). Motivating factors for cochlear implantation include improved quality of life and access to local cochlear implantation services. CONCLUSION: The TA566 guidelines have increased cochlear implantation eligibility, putting pressure on cochlear implantation centres and referring hospitals. Current referral systems have external and patient-centred implantation barriers. British cochlear implantation delivery may need rethinking to meet increasing populational demands and improve accessibility for those most vulnerable to these barriers.

Characterization of the public transit air microbiome and resistome reveals geographical specificity.

BACKGROUND: The public transit is a built environment with high occupant density across the globe, and identifying factors shaping public transit air microbiomes will help design strategies to minimize the transmission of pathogens. However, the majority of microbiome works dedicated to the public transit air are limited to amplicon sequencing, and our knowledge regarding the functional potentials and the repertoire of resistance genes (i.e. resistome) is limited. Furthermore, current air microbiome investigations on public transit systems are focused on single cities, and a multi-city assessment of the public transit air microbiome will allow a greater understanding of whether and how broad environmental, building, and anthropogenic factors shape the public transit air microbiome in an international scale. Therefore, in this study, the public transit air microbiomes and resistomes of six cities across three continents (Denver, Hong Kong, London, New York City, Oslo, Stockholm) were characterized. RESULTS: City was the sole factor associated with public transit air microbiome differences, with diverse taxa identified as drivers for geography-associated functional potentials, concomitant with geographical differences in species- and strain-level inferred growth profiles. Related bacterial strains differed among cities in genes encoding resistance, transposase, and other functions. Sourcetracking estimated that human skin, soil, and wastewater were major presumptive resistome sources of public transit air, and adjacent public transit surfaces may also be considered presumptive sources. Large proportions of detected resistance genes were co-located with mobile genetic elements including plasmids. Biosynthetic gene clusters and city-unique coding sequences were found in the metagenome-assembled genomes. CONCLUSIONS: Overall, geographical specificity transcends multiple aspects of the public transit air microbiome, and future efforts on a global scale are warranted to increase our understanding of factors shaping the microbiome of this unique built environment.

Conventional landmark palpation vs. preprocedural ultrasound for neuraxial analgesia and anaesthesia in obstetrics - a systematic review and meta-analysis with trial sequential analyses.

The aim of this systematic review and meta-analysis was to examine the efficacy, time taken and the safety of neuraxial blockade performed for obstetric patients with the assistance of preprocedural ultrasound, in comparison with the landmark palpation method. The bibliographic databases Central, CINAHL, EMBASE, Global Health, MEDLINE, Scopus and Web of Science were searched from inception to 13 February 2020 for randomised controlled trials that included pregnant women having neuraxial procedures with preprocedural ultrasound as the intervention and conventional landmark palpation as the comparator. For continuous and dichotomous outcomes, respectively, we calculated the mean difference using the inverse-variance method and the risk ratio with the Mantel-Haenszel method. In all, 22 trials with 2462 patients were included. Confirmed by trial sequential analysis, preprocedural ultrasound increased the first-pass success rate by a risk ratio (95%CI) of 1.46 (1.16-1.82), p = 0.001 in 13 trials with 1253 patients. No evidence of a difference was found in the total time taken between preprocedural ultrasound and landmark palpation, with a mean difference (95%CI) of 50.1 (-13.7 to 113.94) s, p = 0.12 in eight trials with 709 patients. The quality of evidence was graded as low and very low, respectively, for these co-primary outcomes. Sub-group analysis underlined the increased benefit of preprocedural ultrasound for those in whom the neuraxial procedure was predicted to be difficult. Complications, including postpartum back pain and headache, were decreased with preprocedural ultrasound. The adoption of preprocedural ultrasound for neuraxial procedures in obstetrics is recommended and, in the opinion of the authors, should be considered as a standard of care, in view of its potential to increase efficacy and reduce complications without significant prolongation of the total time required.

Influenza vaccine failure in the tropics: a retrospective cohort study of waning effectiveness.

Influenza vaccine effectiveness (VE) wanes over the course of a temperate climate winter season but little data are available from tropical countries with year-round influenza virus activity. In Singapore, a retrospective cohort study of adults vaccinated from 2013 to 2017 was conducted. Influenza vaccine failure was defined as hospital admission with polymerase chain reaction-confirmed influenza infection 2-49 weeks after vaccination. Relative VE was calculated by splitting the follow-up period into 8-week episodes (Lexis expansion) and the odds of influenza infection in the first 8-week period after vaccination (weeks 2-9) compared with subsequent 8-week periods using multivariable logistic regression adjusting for patient factors and influenza virus activity. Records of 19 298 influenza vaccinations were analysed with 617 (3.2%) influenza infections. Relative VE was stable for the first 26 weeks post-vaccination, but then declined for all three influenza types/subtypes to 69% at weeks 42-49 (95% confidence interval (CI) 52-92%, P = 0.011). VE declined fastest in older adults, in individuals with chronic pulmonary disease and in those who had been previously vaccinated within the last 2 years. Vaccine failure was significantly associated with a change in recommended vaccine strains between vaccination and observation period (adjusted odds ratio 1.26, 95% CI 1.06-1.50, P = 0.010).

RX-5902, a novel ß-catenin modulator, potentiates the efficacy of immune checkpoint inhibitors in preclinical models of triple-negative breast Cancer.

BACKGROUND: Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype with limited systemic treatment options. RX-5902 is a novel anti-cancer agent that inhibits phosphorylated-p68 and thus attenuates nuclear ß-catenin signaling. The purpose of this study was to evaluate the ability of ß-catenin signaling blockade to enhance the efficacy of anti-CTLA-4 and anti-PD-1 immune checkpoint blockade in immunocompetent, preclinical models of TNBC. METHODS: Treatment with RX-5902, anti-PD-1, anti-CTLA-4 or the combination was investigated in BALB/c mice injected with the 4 T1 TNBC cell line. Humanized BALB/c-Rag2nullIl2rγnullSIRPαNOD (hu-CB-BRGS) mice transplanted with a human immune system were implanted with MDA-MB-231 cells. Mice were randomized into treatment groups according to human hematopoietic chimerism and treated with RX-5902, anti-PD-1 or the combination. At sacrifice, bone marrow, lymph nodes, spleen and tumors were harvested for flow cytometry analysis of human immune cells. RESULTS: The addition of RX-5902 to CTLA-4 or PD-1 inhibitors resulted in decreased tumor growth in the 4 T1 and human immune system and MDA-MB-231 xenograft models. Immunologic analyses demonstrated a significant increase in the number of activated T cells in tumor infiltrating lymphocytes (TILs) with RX-5902 treatment compared to vehicle (p < 0.05). In the RX-5902/nivolumab combination group, there was a significant increase in the percentage of CD4+ T cells in TILs and increased systemic granzyme B production (p < 0.01). CONCLUSIONS: Conclusions: RX-5902 enhanced the efficacy of nivolumab in a humanized, preclinical model of TNBC. Several changes in immunologic profiles were noted in mice treated with RX-5902 and the combination, including an increase in activated TILs and a decrease in human myeloid populations, that are often associated with immunosuppression in a tumor microenvironment. RX-5902 also was shown to potentiate the effects of checkpoint inhibitors of CTLA4 and the PD-1 inhibitor in the 4 T-1 murine TNBC model. These findings indicate that RX-5902 may have important immunomodulatory, as well as anti-tumor activity, in TNBC when combined with a checkpoint inhibitor.

Patients' attitudes towards cost feedback to doctors to prevent unnecessary testing: a qualitative focus group study.

OBJECTIVES: There is a need to improve efficiency in healthcare delivery without compromising quality of care. One approach is the development and evaluation of behavioural strategies to reduce unnecessary use of common tests. However, there is an absence of evidence on patient attitudes to the use of such approaches in the delivery of care. Our objective was to explore patient acceptability of a nudge-type intervention that aimed to modify blood test requests by hospital doctors. STUDY DESIGN: Single-centre qualitative study. METHODS: The financial costs of common blood tests were presented to hospital doctors on results reports for 1 year at a hospital. Focus group discussions were conducted with recent inpatients at the hospital using a semi-structured question schedule. Discussions were transcribed and analysed using qualitative content analysis to identify and prioritise common themes explaining attitudes to the intervention approach. RESULTS: Three focus groups involving 17 participants were conducted. Patients were generally apprehensive about the provision of blood test cost feedback to doctors. Attitudes were organised around themes representing beliefs about blood tests, the impact on doctors and their autonomy, and beliefs about unnecessary testing. Patients thought that blood tests were important, powerful and inexpensive, and cost information could place doctors under additional pressure. CONCLUSION: The findings identify predominantly positive beliefs about testing and negative attitudes to the use of financial costs in the decision-making of hospital doctors. Public discussion and education about the possible overuse of common tests may allow more resources to be allocated to evidence-based healthcare, by reducing the perception that such strategies to improve healthcare efficiency negatively impact on quality of care.

RX-3117 (Fluorocyclopentenyl-Cytosine)-Mediated Down-Regulation of DNA Methyltransferase 1 Leads to Protein Expression of Tumor-Suppressor Genes and Increased Functionality of the Proton-Coupled Folate Carrier.

(1) Background: RX-3117 (fluorocyclopentenyl-cytosine) is a cytidine analog that inhibits DNA methyltransferase 1 (DNMT1). We investigated the mechanism and potential of RX-3117 as a demethylating agent in several in vitro models. (2) Methods: we used western blotting to measure expression of several proteins known to be down-regulated by DNA methylation: O6-methylguanine-DNA methyltransferase (MGMT) and the tumor-suppressor genes, p16 and E-cadherin. Transport of methotrexate (MTX) mediated by the proton-coupled folate transporter (PCFT) was used as a functional assay. (3) Results: RX-3117 treatment decreased total DNA-cytosine-methylation in A549 non-small cell lung cancer (NSCLC) cells, and induced protein expression of MGMT, p16 and E-cadherin in A549 and SW1573 NSCLC cells. Leukemic CCRF-CEM cells and the MTX-resistant variant (CEM/MTX, with a deficient reduced folate carrier) have a very low expression of PCFT due to promoter hypermethylation. In CEM/MTX cells, pre-treatment with RX-3117 increased PCFT-mediated MTX uptake 8-fold, and in CEM cells 4-fold. With the reference hypomethylating agent 5-aza-2'-deoxycytidine similar values were obtained. RX-3117 also increased PCFT gene expression and PCFT protein. (4) Conclusion: RX-3117 down-regulates DNMT1, leading to hypomethylation of DNA. From the increased protein expression of tumor-suppressor genes and functional activation of PCFT, we concluded that RX-3117 might have induced hypomethylation of the promotor.

Core outcome set for uncomplicated acute appendicitis in children and young people.

BACKGROUND: Research studies to inform clinical practice and policy in children and young people with appendicitis are hampered by inconsistent selection and reporting of outcomes. The aim of this study was to develop a core outcome set for reporting all studies of uncomplicated acute appendicitis in children and young people. METHODS: Systematic literature reviews, qualitative interviews with parents and patients treated for uncomplicated acute appendicitis, and a Study-Specific Advisory Group informed a long list of outcomes. Outcomes were then prioritized by stakeholders based in the UK (patients, parents, and paediatric and general surgeons) in an online three-round Delphi consensus process, followed by face-to-face consensus meetings. RESULTS: A long list of 40 items was scored by 147 key stakeholders in the first Delphi round, of whom 90 completed the two subsequent Delphi rounds. The final core outcome set comprises 14 outcomes: intra-abdominal abscess, reoperation (including interventional radiology procedure), readmission to hospital, bowel obstruction, wound infection, antibiotic failure, wound complication, negative appendicectomy, recurrent appendicitis, death, patient stress/psychological distress, length of hospital stay, time away from full activity and child's quality of life. CONCLUSION: A core outcome set comprising 14 outcomes across five key domains has been developed for reporting studies in children and young people with uncomplicated acute appendicitis. Further work is required to determine how and when to measure these outcomes.

ANTECEDENTES: Los estudios de investigación que sirvan de base para la práctica clínica y la política en niños y adultos jóvenes con apendicitis se ven obstaculizados por inconsistencias en la selección y descripción de los resultados. El objetivo de este estudio fue desarrollar un conjunto central de resultados para todos los estudios de apendicitis aguda no complicada en niños y adultos jóvenes. MÉTODOS: Para establecer una lista de resultados se efectuaron revisiones sistemáticas de la literatura, entrevistas cualitativas con padres y pacientes tratados por apendicitis aguda no complicada, y consulta con un Grupo de Asesoramiento Específico para el Estudio. Seguidamente, los resultados se priorizaron de acuerdo con los intereses de las partes interesadas (pacientes, padres, y cirujanos pediátricos y generales) en el Reino Unido a través de un proceso de consenso Delphi de tres rondas en Internet, seguido de reuniones personales de consenso. RESULTADOS: Un total de 147 participantes puntuaron una larga lista de 40 ítems en la primera ronda Delphi, de los cuales 90 completaron las dos rondas Delphi subsiguientes. El conjunto final incluye 14 resultados: absceso intra-abdominal, reoperación (incluyendo procedimientos radiológicos intervencionistas), reingreso, obstrucción intestinal, infección de herida, otras complicaciones de la herida, fracaso del tratamiento con antibióticos, apendicectomía blanca, apendicitis recidivante, muerte, estrés del paciente/sufrimiento psicológico, duración de la estancia hospitalaria, tiempo alejado de todas sus actividades y calidad de vida. CONCLUSIÓN: Se ha desarrollado un conjunto central de resultados que incluye 14 resultados en cinco dominios clave para la descripción de estudios en niños y adultos jóvenes con apendicitis aguda no complicada. Se requieren más trabajos para determinar cómo y cuándo conviene medir estos resultados.

Targeted PMP22 TATA-box editing by CRISPR/Cas9 reduces demyelinating neuropathy of Charcot-Marie-Tooth disease type 1A in mice.

Charcot-Marie-Tooth 1A (CMT1A) is the most common inherited neuropathy without a known therapy, which is caused by a 1.4 Mb duplication on human chromosome 17, which includes the gene encoding the peripheral myelin protein of 22 kDa (PMP22). Overexpressed PMP22 protein from its gene duplication is thought to cause demyelination and subsequently axonal degeneration in the peripheral nervous system (PNS). Here, we targeted TATA-box of human PMP22 promoter to normalize overexpressed PMP22 level in C22 mice, a mouse model of CMT1A harboring multiple copies of human PMP22. Direct local intraneural delivery of CRISPR/Cas9 designed to target TATA-box of PMP22 before the onset of disease, downregulates gene expression of PMP22 and preserves both myelin and axons. Notably, the same approach was effective in partial rescue of demyelination even after the onset of disease. Collectively, our data present a proof-of-concept that CRISPR/Cas9-mediated targeting of TATA-box can be utilized to treat CMT1A.