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Abstract Background Extracellular vesicles (EVs) are a group of nanoscale cell-derived membranous structures secreted by all cell types, containing molecular cargoes involved in intercellular communication. EVs can be used to mimic “nature’s delivery system” to transport nucleic acids, peptides, lipids, and metabolites to target recipient cells. EVs offer a range of advantages over traditional synthetic carriers, thus paving the way for innovative drug delivery approaches that can be used in different diseases, including cancer. Here, by using breast cancer (BC) cells treated with the multi-kinase inhibitor sorafenib, we generated EVs enriched in specific non-coding RNAs (miR-23b-3p, miR-126-3p, and the long ncRNA GAS5) and investigated their potential impact on the aggressive properties of the BC in vitro and in vivo using zebrafish. Methods EVs were collected from 4 different BC cell lines (HCC1937, MDA-MB-231, MCF-7, and MDA-MB-453) and characterized by western blotting, transmission electron microscopy and nanoparticle tracking analysis. Levels of encapsulated miR-23b-3p, miR-126-3p, and GAS5 were quantified by ddPCR. The role of the EVs as carriers of ncRNAs in vivo was established by injecting MDA-MB-231 and MDA-MB-453 cells into zebrafish embryos followed by EV-based treatment of the xenografts with EVs rich in miR-23b-3p, miR-126-3p and GAS5. Results ddPCR analysis revealed elevated levels of miR-23b-3p, miR-126-3p, and GAS5, encapsulated in the EVs released by the aforementioned cell lines, following sorafenib treatment. The use of EVs as carriers of these specific ncRNAs in the treatment of BC cells resulted in a significant increase in the expression levels of the three ncRNAs along with the inhibition of cellular proliferation in vitro. In vivo experiments demonstrated a remarkable reduction of xenograft tumor area, suppression of angiogenesis, and decreased number of micrometastasis in the tails after administration of EVs enriched with these ncRNAs. Conclusions Our study demonstrated that sorafenib-induced EVs, enriched with specific tumor-suppressor ncRNAs, can effectively inhibit the aggressive BC characteristics in vitro and in vivo. Our findings indicate an alternative way to enrich EVs with specific tumor-suppressor ncRNAs by treating the cells with an anticancer drug and support the development of new potential experimental molecular approaches to target the aggressive properties of cancer cells.

Abstract Background The CapaCiTY programme includes three, multi-centre, randomised controlled trials aiming to develop an evidence based adult chronic constipation treatment pathway. The trials were conducted in the United Kingdom, National Health Service, aiming to recruit 808 participants from 26 March 2015 to 31 January 2019. Sites were selected based on their responses to site feasibility questionnaires (2014–2015), a common tool employed by sponsors to assess a site’s recruitment potential and ability to undertake the trial protocol. Failure to recruit the planned sample jeopardises reliability of results and wastes significant time and resources. The purpose of this study was to investigate barriers to recruitment in 2017. Methods We conducted site feasibility assessments with thirty-nine sites prior to trial commencement. Twenty-seven were selected to participate in the CapaCiTY programme, twelve were deemed unsuitable. We compared site contracted recruitment rates with actual recruitment rates and conducted a telephone survey and analysis from 5 July to 7 December 2017 (n = 24) to understand barriers to recruitment. Three sites declined to participate in the survey. Results At the time of survey, 15% of sites in the CapaCiTY programme were meeting recruitment targets, 85% were recruiting half or less of their target. Of these, 28% recruited no participants. The main barriers to recruitment were lack of resources, high workloads, lack of suitable participants and study design not being compatible with routine care. Despite multiple strategies employed to overcome these barriers, the trials were eventually stopped due to futility, recruiting only 34% of the programme sample size. Conclusions Improving the reliability of site feasibility assessments could potentially save a substantial amount in failed research investments and speed up the time to delivery of new treatments. We recommend 1) investment in training researchers in conducting and completing site feasibility; 2) funders to require pilot and feasibility data in grant applications, with an emphasis on patient and public involvement in trial design; 3) conducting site feasibility assessment at the pre-award stage; 4) development of a national database of sites’ previous trial recruitment performance; 5) data-driven site level assessment of recruitment potential. Trial registration ISRCTN11791740; 16/07/2015, ISRCTN11093872; 11/11/2015, ISRCTN11747152; 30/09/2015.

Abstract Background SMYD3 has been found implicated in cancer progression. Its overexpression correlates with cancer growth and invasion, especially in gastrointestinal tumors. SMYD3 transactivates multiple oncogenic mechanisms, favoring cancer development. Moreover, it was recently shown that SMYD3 is required for DNA restoration by promoting homologous recombination (HR) repair. Methods In cellulo and in vivo models were employed to investigate the role of SMYD3 in cancer chemoresistance. Analyses of SMYD3-KO cells, drug-resistant cancer cell lines, patients’ residual gastric or rectal tumors that were resected after neoadjuvant therapy and mice models were performed. In addition, the novel SMYD3 covalent inhibitor EM127 was used to evaluate the impact of manipulating SMYD3 activity on the sensitization of cancer cell lines, tumorspheres and cancer murine models to chemotherapeutics (CHTs). Results Here we report that SMYD3 mediates cancer cell sensitivity to CHTs. Indeed, cancer cells lacking SMYD3 functions showed increased responsiveness to CHTs, while restoring its expression promoted chemoresistance. Specifically, SMYD3 is essential for the repair of CHT-induced double-strand breaks as it methylates the upstream sensor ATM and allows HR cascade propagation through CHK2 and p53 phosphorylation, thereby promoting cancer cell survival. SMYD3 inhibition with the novel compound EM127 showed a synergistic effect with CHTs in colorectal, gastric, and breast cancer cells, tumorspheres, and preclinical colorectal cancer models. Conclusions Overall, our results show that targeting SMYD3 may be an effective therapeutic strategy to overcome chemoresistance.

Abstract Background This systematic review contributes to the understanding of the characteristics of built food environments that may be associated with choices of alternative protein foods (APF). Using the built food environment typology proposed by Downs et al., we investigated various environmental structures (e.g., supermarkets, other retailers, farmers’ markets, restaurants, schools, and online vendors) and the characteristics that may facilitate or hinder consumers’ choices. For example, facilitators and barriers may refer to the physical characteristics of environmental structures, food presentation practices, the organizational strategies or policies operating in the setting, or the actions that retailers or consumers engage in while selling, serving, choosing, trying, or purchasing APF in these environmental structures. Methods A systematic review (PROSPERO database preregistration; no. CRD42023388700) was conducted by searching 13 databases for peer-reviewed journals focusing on the fields of economics and business, agriculture, medical sciences, and social sciences. Data searches, coding, and quality evaluations were conducted by at least 2 researchers. A total of 31 papers (36 original studies) were included. The risk of bias was evaluated with the Joanna Briggs Institute quality evaluation tool, with 24 publications presenting low risk of bias. Results The findings indicate that perceived and actual availability facilitate consumers’ APF choices across a built food environment. Several barriers/facilitators were associated with APF choices in specific types of built food environments: the way food is presented in produce sections (supermarkets), consumer habits in terms of green and specialty shopping (grocery stores), and mismatches among retailer actions in regard to making APF available in one type of food environment structure (e-commerce) and consumers’ preferences for APF being available in other food environment structures (supermarkets, grocery stores). The effect of a barrier/facilitator may depend on the APF type; for example, social norms regarding masculinity were a barrier affecting plant-based APF choices in restaurants, but these norms were not a barrier affecting the choice of insect-based APF in restaurants. Conclusions Addressing barriers/facilitators identified in this review will help in developing environment-matching interventions that aim to make alternative proteins mainstream. Trial registration PROSPERO database registration: #CRD42023388700.

Abstract Background Based on the present global burden of oral diseases, unmet dental needs affect a more significant population worldwide. It is characterised by the need for dental care but receiving delayed or no care. The contributing factors include lack of knowledge about oral health, its consequences, and the availability of dental services. We need to find out the scale of the problem of unmet dental needs for the south Indian population. Therefore, the objective was to determine the relationship between the presence of oral disease and the quality of life-related to oral health using the OHIP-14 tool. Methods The unmet dental requirements of the south Indian population were determined using a cross-sectional questionnaire survey. Close-ended questions were used to obtain data from two investigators trained to record the answers from the patients. The data was collected using the OHIP-14 questionnaire, which consists of 14 items divided into seven domains with two questions each. Physical pain, psychological impairment, physical disability, psychological disability, social disability, and disability were all considered. An additional analysis of artificial neural network (ANN) was done. Results The response rate was 100 per cent. N = 1029 people replied to the questionnaire about their unmet dental needs. N = 497 (48.3%) were men, whereas N = 532 (51.7%) were women. The average age was 31.7811.72. As their current occupation, most of the included subjects (60.1%) were students. The respondents had no known personal habits and a mixed diet (94.93%). The average BMI was 24.022.59 (14-30.9). OHIP was present in 62.3% of the population. The average OHIP-14 severity score was 10.97. (8.54). The severity and degree of unmet dental need were substantial (p0.01) due to pain in the mouth/teeth/gums, malocclusion, and gum bleeding. The most common OHIP-14 domains affected by unmet oral needs were psychological discomfort, psychological limitation, social limitation, and feeling handicapped. The analysis of ANN revealed that high OHIP scores were primarily attributed to dental caries, poor oral health, and dental aesthetics. Conclusion The severity and degree of unmet dental needs were significant among the south Indian population. The most common oral health status that impacted OHIP-14 domains were pain, malocclusion, and bleeding gums. These patients were significantly impacted by psychological discomfort and social limitations and felt handicapped.

Abstract Background Activation of the Wnt pathway has been linked to colorectal cancer (CRC). Previous reports suggest that Wnt3a can activate p38. Besides, p38α feeds into the canonical Wnt/β-catenin pathway by inhibiting GSK3β through phosphorylation. Recently, we identified p38α as a new druggable member of β-catenin chromatin-associated kinase complexes in CRC. Methods The functional relationship between p38α and β-catenin was characterized in CRC cells, patient-derived CRC stem cells, patient-derived tumor intestinal organoids, and in vivo models (C57BL/6-APCMin/+ mice). The role of p38α in β-catenin transcriptional activity was assessed by pharmacological inhibition with ralimetinib. Results We used the GSK3β inhibitor TWS-119, which promotes the activation of Wnt signaling, to uncouple p38α nuclear/cytoplasmatic functions in the Wnt pathway. Upon GSK3β inhibition, nuclear p38α phosphorylates β-catenin at residues S111 and T112, allowing its binding to promoter regions of Wnt target genes and the activation of a transcriptional program implicated in cancer progression. If p38α is pharmacologically inhibited in addition to GSK3β, β-catenin is prevented from promoting target gene transcription, which is expected to impair carcinogenesis. Conclusions p38α seems to play a dual role as a member of the β-catenin destruction complex and as a β-catenin chromatin-associated kinase in CRC. This finding may help elucidate mechanisms contributing to human colon tumor pathogenesis and devise new strategies for personalized CRC treatment.

Abstract Background The prevalence of obesity is rising in all subregions of America, including Brazil. To understand the obesity problem in Brazil better, a possible approach could be to analyze its obesity trend by comparing it with the reality of a country that went previously through the epidemiological transition, such as the USA. In addition, the obesity trend must be analyzed in comparison with obesity risk factors trends, such as the physical inactivity (PI) trend. Our aim was comparatively to analyze the temporal trends of obesity between Brazil and the USA from the perspective of temporal trends of PI. Methods We conducted a temporal trend study based on data from national cross-sectional surveys: the VIGITEL (Surveillance System for Factors of Health Risk and Protection for Chronic Diseases by Telephone Survey) for Brazil and the BRFSS (Behavioral Risk Factor Surveillance System) for the USA, comparing the annual prevalence of obesity and PI between 2011 and 2021. For the analysis of each temporal variation, linear regressions were performed with the Prais-Winsten test, and Pearson’s correlation coefficient was conducted to correlate the trends of the same variables between countries and of different variables within each country. Results Considering the total sample, Brazil [coefficient (95%CI) 0.6 (0.4;0.7), p = 0.000] and the USA [coefficient (95%CI) 0.5 (0.5;0.6), p = 0.000] showed increasing trends in obesity. The tendency of PI was of stabilization in the two countries [Brazil: coefficient (95%CI) -0.03 (-0.3;0.2), p = 0.767 and USA coefficient (95%CI) -0.03 (-0.2;0.1), p = 0.584]. In addition, there was a correlation between obesity trends between Brazil and the USA (r = 0.971; p = 0.000), but there was no correlation between PI trends between the two countries, nor with obesity and PI trends within each country. Conclusions In the last decade, there was a trend towards increasing obesity and stabilization in PI, both in Brazil and the USA. However, there was no association between temporal trends in obesity and physical inactivity in both countries. Our data reinforce a call to action to prevent and control obesity, going with and beyond PI reduction.

Abstract Background COVID-19 pandemic represented a shock for healthcare systems. Italy was one of the first country to deal with a huge number of patients to be diagnosed, isolated, and treated with scarce evidence-based guidelines and resources. Several organizational and structural changes were needed to face the pandemic at local level. The article aims at studying the perceived impact of the newly implemented District Operation Centres (DOCs) of Local Health Authority (LHA) Roma 1 in managing active surveillance and home care of COVID-19 patients and their close contacts in cooperation with general practitioners (GPs). Methods A questionnaire, developed according to Delphi methodology, was validated by 7 experts and administered to a randomized sample of GPs and family paediatricians (FPs). All medical doctors selected received a phone interview between December 2020 and January 2021. The questionnaire investigated general characteristics of the sample, relations with DOC and its usefulness, and potential developments. A descriptive analysis was performed and inferential statistical tests were used to assess differences. Results In April 2020 the LHA Roma 1 implemented one DOCs in each local health district. 215 medical doctors were interviewed, reaching the sample target for health districts (80% CL and 10% MOE) and the whole LHA (90% CL and 5% MOE). Several aspects in the management of COVID-19 cases and close contacts of COVID-19 cases, and of the support of DOCs to GPs/FPs were investigated. More than 55% of the GPs and FPs interviewed found the DOCs useful and more than 78% would recommend a service DOC-like to other LHAs. The medical professionals interviewed would use DOCs in the future as support in treating vulnerable patients, utilizing digital health tools, enlisting specialist doctors, establishing networks, and facilitating professional counselling by nurses. Conclusions This study is an attempt to evaluate an organizational change happened during COVID-19 pandemic. DOCs were created to support GPs and FPs as a link between primary healthcare and public health. Although several difficulties were disclosed, DOCs’ experience can help to overcome the fragmentation of the systems and the duality between primary care and public health and make the system more resilient.

Abstract Background Triple negative breast cancers (TNBC) account for approximately 15% of all breast cancers and are associated with a shorter median survival mainly due to locally advanced tumor and high risk of metastasis. The current neoadjuvant treatment for TNBC consists of a regimen of immune checkpoint blocker and chemotherapy (chemo-ICB). Despite the frequent use of this combination for TNBC treatment, moderate results are observed and its clinical benefit in TNBC remains difficult to predict. Patient-derived tumor organoids (PDTO) are 3D in vitro cellular structures obtained from patient’s tumor samples. More and more evidence suggest that these models could predict the response of the tumor from which they are derived. PDTO may thus be used as a tool to predict chemo-ICB efficacy in TNBC patients. Method The TRIPLEX study is a single-center observational study conducted to investigate the feasibility of generating PDTO from TNBC and to evaluate their ability to predict clinical response. PDTO will be obtained after the dissociation of biopsies and embedding into extra cellular matrix. PDTO will be cultured in a medium supplemented with growth factors and signal pathway inhibitors. Molecular and histological analyses will be performed on established PDTO lines to validate their phenotypic proximity with the original tumor. Response of PDTO to chemo-ICB will be assessed using co-cultures with autologous immune cells collected from patient blood samples. PDTO response will finally be compared with the response of the patient to evaluate the predictive potential of the model. Discussion This study will allow to assess the feasibility of using PDTO as predictive tools for the evaluation of the response of TNBC patients to treatments. In the event that PDTO could faithfully predict patient response in clinically relevant time frames, a prospective clinical trial could be designed to use PDTO to guide clinical decision. This study will also permit the establishment of a living biobank of TNBC PDTO usable for future innovative strategies evaluation. Trial registration The clinical trial (version 1.2) has been validated by local research ethic committee on December 30th 2021 and registered at ClinicalTrials.gov with the identifier NCT05404321 on June 3rd 2022, version 1.2.

Abstract Background Molecular phylogenetics studies the evolutionary relationships among the individuals of a population through their biological sequences. It may provide insights about the origin and the evolution of viral diseases, or highlight complex evolutionary trajectories. A key task is inferring phylogenetic trees from any type of sequencing data, including raw short reads. Yet, several tools require pre-processed input data e.g. from complex computational pipelines based on de novo assembly or from mappings against a reference genome. As sequencing technologies keep becoming cheaper, this puts increasing pressure on designing methods that perform analysis directly on their outputs. From this viewpoint, there is a growing interest in alignment-, assembly-, and reference-free methods that could work on several data including raw reads data. Results We present phyBWT2, a newly improved version of phyBWT (Guerrini et al. in 22nd International Workshop on Algorithms in Bioinformatics (WABI) 242:23–12319, 2022). Both of them directly reconstruct phylogenetic trees bypassing both the alignment against a reference genome and de novo assembly. They exploit the combinatorial properties of the extended Burrows-Wheeler Transform (eBWT) and the corresponding eBWT positional clustering framework to detect relevant blocks of the longest shared substrings of varying length (unlike the k-mer-based approaches that need to fix the length k a priori). As a result, they provide novel alignment-, assembly-, and reference-free methods that build partition trees without relying on the pairwise comparison of sequences, thus avoiding to use a distance matrix to infer phylogeny. In addition, phyBWT2 outperforms phyBWT in terms of running time, as the former reconstructs phylogenetic trees step-by-step by considering multiple partitions, instead of just one partition at a time, as previously done by the latter. Conclusions Based on the results of the experiments on sequencing data, we conclude that our method can produce trees of quality comparable to the benchmark phylogeny by handling datasets of different types (short reads, contigs, or entire genomes). Overall, the experiments confirm the effectiveness of phyBWT2 that improves the performance of its previous version phyBWT, while preserving the accuracy of the results.

Abstract Background Video-assisted thoracic surgery (VATS) has become the standard for lung cancer diagnosis and treatment. However, this surgical technique requires specific and dedicated training. In the past 20 years, several simulator systems have been developed to promote VATS training. Advances in virtual reality may facilitate its integration into the VATS training curriculum. The present review aims to first provide a comprehensive overview of the simulators for thoracoscopic surgery, focused especially on simulators for lung lobectomy; second, it explores the role and highlights the possible efficacy of these simulators in the surgical trainee curriculum. Methods A literature search was conducted in the PubMed, EMBASE, Science Direct, Scopus and Web of Science databases using the following keywords combined with Boolean operators “AND” and “OR”: virtual reality, VR, augmented reality, virtual simulation, mixed reality, extended reality, thoracic surgery, thoracoscopy, VATS, video-assisted thoracoscopic surgery, simulation, simulator, simulators, training, and education. Reference lists of the identified articles were hand-searched for additional relevant articles to be included in this review. Results Different types of simulators have been used for VATS training: synthetic lung models (dry simulators); live animals or animal tissues (wet simulators); and simulators based on virtual or augmented reality. Their role in surgical training has been generally defined as useful. However, not enough data are available to ascertain which type is the most appropriate. Conclusions Simulator application in the field of medical education could revolutionize the regular surgical training curriculum. Further studies are required to better define their impact on surgeons’ training programs and, finally, on patients’ quality of care.

Abstract Background The conflict between Russia and Ukraine has strained the health systems of countries that welcome war refugees on all levels, from national to local. Despite the Public Health guidelines regarding assistance being published on the topic, the scientific literature currently lacks evidence on the experience of applying theory in practice. This study aims to describe evidence-based practices that were implemented and to provide a detailed description of emerging problems and solutions pertaining Ukrainian refugee assistance in the context of one of the biggest Local Health Authorities in Italy (LHA Roma 1). Methods LHA Roma 1 developed a strategic plan based on local expertise, national and international guidelines to ensure infectious disease prevention and control, as well as continuity of care for non-communicable diseases and mental health. Results The insertion of Ukrainian refugees in the National Health System through an identification code assignment and other services such as COVID-19 swab and vaccination were provided either in one of the three major assistance hubs or in local district level ambulatories spread throughout the LHA. Many challenges were faced during the implementation phase of the outlined practice guidelines, which required sensible and timely solutions. These challenges include the necessity of rapid resource provision, overcoming linguistic and cultural barriers, guaranteeing a standard of care across multiple sites and coordination of interventions. Public Private Partnerships, the creation of a centralized multicultural and multidisciplinary team and the mutually beneficial collaboration with the local Ukrainian community were essential to guarantee the success of all operations. Conclusions The experience of LHA Roma 1 helps shed light on the importance of leadership in emergency settings and how a dynamic relationship between policy and practice would allow each intervention to be modulated according to the local environment, to better realize the potential of local realities to provide appropriate health interventions to all those in need.

Abstract Malaria, chikungunya virus (CHIKV), and dengue virus (DENV) are endemic causes of fever among children in Kenya. The risks of infection are multifactorial and may be influenced by built and social environments. The high resolution overlapping of these diseases and factors affecting their spatial heterogeneity has not been investigated in Kenya. From 2014-2018, we prospectively followed a cohort of children from four communities in both coastal and western Kenya. Overall, 9.8% were CHIKV seropositive, 5.5% were DENV seropositive, and 39.1% were malaria positive (3521 children tested). The spatial analysis identified hot-spots for all three diseases in each site and in multiple years. The results of the model showed that the risk of exposure was linked to demographics with common factors for the three diseases including the presence of litter, crowded households, and higher wealth in these communities. These insights are of high importance to improve surveillance and targeted control of mosquito-borne diseases in Kenya.

Abstract Background The COVID-19 pandemic primarily impacted long-term care facilities by restricting visiting and circulation of visitors, affecting the quality of life (QoL) of older adults living in these institutions. Volunteer activities, essential for older adults’ daily life, were also interrupted and potentially negatively impacted the QoL of older adults, volunteers themselves, and also employees in these institutions. In this context, this study aims to evaluate the impact of the return of volunteer-led activities in a long-term care institution on the QoL of older adult residents, employees, and volunteers. Methods This study used a pre-test and post-test design within the same group. The first round of data collection was conducted before volunteer-led activities returned and the second round after 1 month of return. The instrument used to assess QoL was the EUROHIS-QoL-8 scale. This study was conducted within a nursing home in São Paulo, Brazil, created in 1937 by members of the Israeli community living in Brazil. Volunteer-led activities were part of residents’ daily life before the COVID-19 pandemic, when these activities were interrupted for about 20 months. A total of 79 individuals participated in both rounds (pre and post), of which: 29 residents, 27 volunteers, and 23 employees of the long-term care institution. Results Using a Wilcoxon signed-rank test, the analyses indicated improvements after the 1 month return in different QoL aspects for the three groups. Volunteers improved their personal relationships (Z − 2.332, p

Abstract Background The incidence of the highly morbid and potentially lethal gangrenous cholecystitis was reportedly increased during the COVID-19 pandemic. The aim of the ChoCO-W study was to compare the clinical findings and outcomes of acute cholecystitis in patients who had COVID-19 disease with those who did not. Methods Data were prospectively collected over 6 months (October 1, 2020, to April 30, 2021) with 1-month follow-up. In October 2020, Delta variant of SARS CoV-2 was isolated for the first time. Demographic and clinical data were analyzed and reported according to the STROBE guidelines. Baseline characteristics and clinical outcomes of patients who had COVID-19 were compared with those who did not. Results A total of 2893 patients, from 42 countries, 218 centers, involved, with a median age of 61.3 (SD: 17.39) years were prospectively enrolled in this study; 1481 (51%) patients were males. One hundred and eighty (6.9%) patients were COVID-19 positive, while 2412 (93.1%) were negative. Concomitant preexisting diseases including cardiovascular diseases (p

Abstract Discontinuation of antimicrobial stewardship programs (ASPs) and increased antibiotic use were described during SARS-CoV-2 pandemic. In order to measure COVID-19 impact on ASPs in a setting of high multidrug resistance organisms (MDRO) prevalence, a qualitative survey was designed. In July 2021, eighteen ID Units were asked to answer a questionnaire about their hospital characteristics, ASPs implementation status before the pandemic and impact of SARS-CoV-2 pandemic on ASPs after the 1st and 2nd pandemic waves in Italy. Nine ID centres (50%) reported a reduction of ASPs and in 7 cases (38.9%) these were suspended. After the early pandemic waves, the proportion of centres that restarted their ASPs was higher among the ID centres where antimicrobial stewardship was formally identified as a priority objective (9/11, 82%, vs 2/7, 28%). SARS-CoV-2 pandemic had a severe impact in ASPs in a region highly affected by COVID-19 and antimicrobial resistance but weaknesses related to the pre-existent ASPs might have played a role.

Abstract Background There is paucity of literature on why and how patients are intubated, and by whom, in Irish Emergency Departments (EDs). The aim of this pilot study was to characterise emergency airway management (EAM) of critically unwell patients presenting to Irish EDs. Methods A multisite prospective pilot study was undertaken from February 10 to May 10, 2020. This project was facilitated through the Irish Trainee Emergency Research Network (ITERN). All patients over 16 years of age requiring EAM were included. Eleven EDs participated in the project. Data recorded included patients’ demographics, indication for intubation, technique of airway management, medications used to facilitate intubation, level of training and specialty of the intubating clinician, number of attempts, success/complications rates and variation across centres. Results Over a 3-month period, 118 patients underwent 131 intubation attempts across 11 EDs. The median age was 57 years (IQR: 40–70). Medical indications were reported in 83% of patients compared to 17% for trauma. Of the 118 patients intubated, Emergency Medicine (EM) doctors performed 54% of initial intubations, while anaesthesiology/intensive care medicine (ICM) doctors performed 46%. The majority (90%) of intubating clinicians were at registrar level. Emergency intubation check lists, video laryngoscopy and bougie were used in 55, 53 and 64% of first attempts, respectively. The first pass success rate was 89%. Intubation complications occurred in 19% of patients. EM doctors undertook a greater proportion of intubations in EDs with > 50,000 attendance (65%) compared to EDs with

Abstract Background Migraine patients have musculoskeletal disorders and pain in the cervical. And, despite the pathophysiology demonstrating the relationship between migraine and the cervical spine, the effectiveness of craniocervical exercises in these patients has not been verified. So, the aimed of this study was verify the effectiveness of craniocervical muscle-strengthening exercise (CMSE) in reducing the frequency and intensity of headache in migraine patients. Methods A two-armed, parallel-group randomized controlled trial with a 3-month follow-up was performed. For eight weeks, the volunteers in the intervention group (n = 21) performed a protocol of CMSE, while those in the sham ultrasound group (n = 21) received the application of disconnected therapeutic ultrasound in the upper trapezius and guideline for home-stretching. The primary outcomes were the frequency and intensity of the headache. The secondary outcomes were questionnaires about migraine and neck disability, and satisfaction with the treatment, cervical range of motion, the pressure pain threshold, craniocervical flexion test (CCFT), cervical muscle strength and endurance test, and the cervical muscle activity during the physical tests. Results No differences were observed for the changes observed in primary outcomes after eight weeks and at the 3-months follow up (p > 0.05). For the secondary outcomes, craniocervical exercises improved the sensitivity of the frontal muscle (p = 0.040) and promoted a reduced amplitude of muscle activity of the anterior scalene and upper trapezius in the last stages of CCFT (p ≤ 0.010). There was also reduced muscle activity of the anterior scalene and splenius capitis in the endurance test (p ≤ 0.045), as evaluated by surface electromyography. Conclusion CMSE were insufficient in reducing the frequency and intensity of headache, improving the performance of the cervical muscles, or reducing migraine and neck pain-related disabilities. This was found despite a decreased electromyographic activity of the cervical muscles during the last stages of CCFT and increased median frequency during the endurance test. Trial registration Accession code RBR-8gfv5j , registered 28/11/2016 in the Registro Brasileiro de Ensaios Clínicos (ReBEC).

Abstract Background Intrahepatic Cholangiocarcinoma (iCCA) is characterized by a strong stromal reaction playing a role in tumor progression. Thymus cell antigen 1 (THY1), also called Cluster of Differentiation 90 (CD90), is a key regulator of cell–cell and cell–matrix interaction. In iCCA, CD90 has been reported to be associated with a poor prognosis. In an iCCA PDX model, we recently found that CD90 was downregulated in mice treated with the Notch γ-secretase inhibitor Crenigacestat. The study aims to investigate the role of CD90 in relation to the NOTCH pathway. Methods THY1/CD90 gene and protein expression was evaluated in human iCCA tissues and xenograft models by qRT-PCR, immunohistochemistry, and immunofluorescence. Notch1 inhibition was achieved by siRNA. THY1/CD90 functions were investigated in xenograft models built with HuCCT1 and KKU-M213 cell lines, engineered to overexpress or knockdown THY1, respectively. Results CD90 co-localized with EPCAM, showing its epithelial origin. In vitro, NOTCH1 silencing triggered HES1 and THY1 down-regulation. RBPJ, a critical transcriptional regulator of NOTCH signaling, exhibited putative binding sites on the THY1 promoter and bound to the latter, implying CD90 as a downstream NOTCH pathway effector. In vivo, Crenigacestat suppressed iCCA growth and reduced CD90 expression in the PDX model. In the xenograft model, Crenigacestat inhibited tumor growth of HuCCT1 cells transfected to overexpress CD90 and KKU-M213 cells constitutively expressing high levels of CD90, while not affecting the growth of HuCCT1 control cells and KKU-M213 depleted of CD90. In an iCCA cohort, patients with higher expression levels of NOTCH1/HES1/THY1 displayed a significantly shorter survival. Conclusions iCCA patients with higher NOTCH1/HES1/THY1 expression have the worst prognosis, but they are more likely to benefit from Notch signaling inhibition. These findings represent the scientific rationale for testing NOTCH1 inhibitors in clinical trials, taking the first step toward precision medicine for iCCA.

Abstract Introduction Transplantation among HIV positive patients may be a valuable therapeutic intervention. This study involves an HIV D+/R+ kidney–liver transplantation, where PBMC-associated HIV quasispecies were analyzed in donor and transplant recipients (TR) prior to transplantation and thereafter, together with standard viral monitoring. Methods The donor was a 54 year of age HIV infected woman: kidney and liver recipients were two HIV infected men, aged 49 and 61. HIV quasispecies in PBMC was analyzed by ultra-deep sequencing of V3 env region. During TR follow-up, plasma HIV-1 RNA, HIV-1 DNA in PBMC, analysis of proviral integration sites and drug-resistance genotyping were performed. Other virological and immunological monitoring included CMV and EBV DNA quantification in blood and CD4 T cell counts. Results Donor and TR were all ART-HIV suppressed at transplantation. Thereafter, TR maintained a nearly suppressed HIV-1 viremia, but HIV-1 RNA blips and the increase of proviral integration sites in PBMC attested some residual HIV replication. A transient peak in HIV-1 DNA occurred in the liver recipient. No major changes of drug-resistance genotype were detected after transplantation. CMV and EBV transient reactivations were observed only in the kidney recipient, but did not require specific treatment. CD4 counts remained stable. No intermixed quasispecies between donor and TR was observed at transplantation or thereafter. Despite signs of viral evolution in TR, HIV genetic heterogeneity did not increase over the course of the months of follow up. Conclusions No evidence of HIV superinfection was observed in the donor nor in the recipients. The immunosuppressive treatment administrated to TR did not result in clinical relevant viral reactivations.

Abstract Background Moving from the correlation between insulin-resistance and PCOS, metformin has been administered in some PCOS women improving ovulatory and metabolic functions and decreasing androgen levels. Inconsistency and unpredictability of response to metformin limit its extensive use. Aim of this study was to identify reliable predictors of response to metformin therapy for weight loss and reduction in plasma androgen levels using ANNs (artificial neural networks). Methods One hundred eight consecutive women with PCOS (ESHRE/ASRM 2003 Rotterdam criteria) treated with metformin 1500 mg/day, at inclusion and every 6 months underwent to a complete clinical, endocrine/metabolic assessment and ultrasonographic evaluation. Therapy outcomes were BMI reduction (≥1 kg/m2) in overweight/obese and free-androgen-index (FAI) decrease (≥1%) in hyperandrogenemic women. Semantic connectivity maps (SCMs) were obtained through Auto-CM, a fourth generation ANN, to compare patients’ baseline clinical features to the treatment outcomes. Multivariate logistic regression analysis was used to assess the major predictor in drop-out patients and the associated risk. Results At 6 months 54 out of 103 (52,4%) obese patients showed BMI reduction and 45 out of 89 (50,6%) hyperandrogenemic women showed FAI decrease. The further response rates at 12 months were 30,6 and 47%, respectively. SCMs showed a clear polarization for both the outcomes with elevated accuracy. Treatment responsiveness resulted strictly related to oligo-amenorrhea and hyperandrogenemia at baseline. In addition, lower serum testosterone levels at baseline were found to be the major predictor of treatment discontinuation. Conclusions In women with PCOS, menstrual pattern imbalance and ovarian androgens excess are the best predictors of metformin response. They may pave the way for a rethinking of the criteria for evaluating hyperandrogenism in order to better define the large population included in the diagnosis of PCOS. Baseline plasma testosterone level can serve as a sensitive marker to predict treatment compliance.

Abstract Family history of cancer (FHC) is a hallmark of cancer risk and an independent predictor of outcome, albeit with uncertain biologic foundations. We previously showed that FHC-high patients experienced prolonged overall (OS) and progression-free survival (PFS) following PD-1/PD-L1 checkpoint inhibitors. To validate our findings in patients with NSCLC, we evaluated two multicenter cohorts of patients with metastatic NSCLC receiving either first-line pembrolizumab or chemotherapy. From each cohort, 607 patients were randomly case–control matched accounting for FHC, age, performance status, and disease burden. Compared to FHC-low/negative, FHC-high patients experienced longer OS (HR 0.67 [95% CI 0.46–0.95], p = 0.0281), PFS (HR 0.65 [95% CI 0.48–0.89]; p = 0.0074) and higher disease control rates (DCR, 86.4% vs 67.5%, p = 0.0096), within the pembrolizumab cohort. No significant associations were found between FHC and OS/PFS/DCR within the chemotherapy cohort. We explored the association between FHC and somatic DNA damage response (DDR) gene alterations as underlying mechanism to our findings in a parallel cohort of 118 NSCLC, 16.9% of whom were FHC-high. The prevalence of ≥ 1 somatic DDR gene mutation was 20% and 24.5% (p = 0.6684) in FHC-high vs. FHC-low/negative, with no differences in tumor mutational burden (6.0 vs. 7.6 Mut/Mb, p = 0.6018) and tumor cell PD-L1 expression. FHC-high status identifies NSCLC patients with improved outcomes from pembrolizumab but not chemotherapy, independent of somatic DDR gene status. Prospective studies evaluating FHC alongside germline genetic testing are warranted.

Abstract Aims To compare the difference in the quality of life between temporomandibular disorders (TMD) patients and non-TMD subjects diagnosed with the Research Diagnostic Criteria for Temporomandibular Disorders (RDC/TMD) or the Diagnostic Criteria for Temporomandibular Disorders (DC/TMD). Methods Medical Literature Analysis and Retrieval System Online (MEDLINE), Excerpta Medica database (EMBASE) and Latin American and Caribbean Health Sciences Literature (LILACS) databases were searched in studies published in English and Portuguese. The search was performed by two independent reviewers in duplicate. A manual search and the gray literature were also included. The inclusion criteria were clinical studies that used the RDC/TMD axis I and quality of life with standard questionnaires in young and middle-aged adult population (18–55 years). The data were analyzed quantitatively by combining the results in a meta-analysis using forest plots. The measure of effect used was the standardized mean difference (SMD) in depression levels. The Newcastle–Ottawa Scale (NOS) was used to evaluate the quality of the studies. The publication bias was assessed by funnel plots. The initial search included 806 articles without duplications. Results Twenty-four articles were included in the final systematic review. Of these, 9 were included in the meta-analysis, where it was shown a statistically significant in all axis I groups: (a) global TMD—groups I, II and III combined, N = 3829, SMD (95% CI) = 1.06 (0.65–1.51), p = 0.000; (b) group I—muscle disorders, N = 3,056, SMD (95% CI) = 0.82 (0.45–1.18), p = 0.000; (c) group II—disc displacements, N = 3,184, SMD (95% CI) = 0.59 (0.26–0.91), p = 0.000; and (d) group III—arthralgia/arthritis/arthrosis, N = 2781, SMD (95% CI) = 0.98 (0.59–1.36), p = 0.000. When compared to controls. Conclusions Quality of life is affected in all axis I TMD patients, especially in groups I and III with higher pain intensity and disability as compared to group II.

Abstract Background The favourable safety profile and the increasing confidence with immune checkpoint inhibitors (ICIs) might have boosted their prescription in frail patients with short life expectancies, who usually are not treated with standard chemotherapy. Methods The present analysis aims to describe clinicians’ attitudes towards ICIs administration during late stages of life within a multicenter cohort of advanced cancer patients treated with single agent PD-1/PD-L1 checkpoint inhibitors in Italy. Results Overall, 1149 patients with advanced cancer who received single agent PD-1/PD-L1 checkpoint inhibitors were screened. The final study population consisted of 567 deceased patients. 166 patients (29.3%) had received ICIs within 30 days of death; among them there was a significantly higher proportion of patients with ECOG-PS ≥ 2 (28.3% vs 11.5%, p

Abstract Background Non-pharmacological approaches for the management of migraine exhibit low to moderate effectiveness due to a lack of high-quality randomized clinical trials. In fact, previous studies applied isolated techniques, which were not representative of common clinical practice. A multimodal approach for migraine may benefit these patients more than isolated approaches. This randomized clinical trial aims to determine the effectiveness of a multimodal protocol combining manual therapy, exercise, and therapeutic pain neuroscience education versus the application of manual therapy or pain neuroscience education alone in patients with migraine. Methods This clinical trial will include 75 individuals of both sexes, aged between 18 and 55 years, with migraine. Participants will be randomized into three groups: the therapeutic pain neuroscience education (TPNE; n = 25) group, the manual therapy (MT; n = 25) group, and the multimodal (MM; n = 25) group. The TPNE group will receive one orientation session on migraine and pain self-management, and recommendations for daily active stretching and walking, with subsequent therapist monitoring. The MT group will receive manual therapies targeting musculoskeletal disorders of the cervical spine. The MM group will receive manual therapies targeting musculoskeletal disorders of the cervical spine, active neck exercises, and therapeutic pain neuroscience education. The treatment period will last 12 weeks. The primary outcome will be the headache impact, measured using the Headache Impact Test (HIT-6). Secondary outcomes will include migraine frequency and intensity, cervical mobility and strength parameters, neck pain-related disability, kinesiophobia, cutaneous allodynia, pain-related catastrophizing, quality of life, and self-perception of change. All outcomes will be evaluated at the fourth, eighth, and twelfth weeks of the treatment period. Primary and secondary clinical outcomes, such as headache impact, frequency, and intensity, will also be evaluated at the 1-, 2-, and 4-month follow-ups. Discussion The results of this randomized clinical trial may provide high-quality clinical evidence of the effects of non-pharmacological treatment options for the management of migraine. Trial registration This study was registered under the access code RBR-7s22c75 in the Registro Brasileiro de Ensaios Clínicos (ReBEC) in December 2020.

Abstract Background To keep a high quality of assistance it is important for hospitals to invest in health technologies (HTs) that have the potential of improving health outcomes. Even though guidance exists on how HTs should be introduced, used and dismissed, there is a surprising gap in literature concerning the awareness of hospitals in the actual utilization of HTs. Methods We performed a systematic literature review of qualitative and quantitative studies aimed at investigating hospital contextual factors that influence the actual utilization of HTs. PubMed, Scopus, Web of Science, Econlit and Ovid Medline electronic databases were searched to retrieve articles published in English and Italian from January 2000 to January 2019. The quality of the included articles was assessed using the Critical Appraisal Skills Programme checklist for qualitative studies, Newcastle-Ottawa Scale for the cross-sectional studies and the Mixed Methods Appraisal Tool for mixed method studies. Results We included 33 articles, which were of moderate to high methodological quality. The included articles mostly addressed the contextual factors that impact the implementation of information and communication technologies (ICTs). Overall, for all HTs, the hospital contextual factors were part of four categories: hospital infrastructure, human resource management, financial resources and leadership styles. Conclusion Our systematic review reported that the contextual factors influencing the HTs utilization at hospital level are mainly explored for ICTs. Several factors should be considered when planning the implementation of a new HTs at hospital level. A potential publication bias might be present in our work, since we included articles published only in English and Italian Language, from January 2000 to January 2019. There remains a gap in the literature on the facilitators and barriers influencing the implementation and concrete utilization of medical and surgical HTs, suggesting the need for further studies for a better understanding.

Abstract Background Transmission of viral diseases (e.g., influenza A H1N1) via respiratory droplets takes place mainly in confined spaces, including in aircraft during commercial air travel. The adoption of hygiene measures may help to prevent disease spread aboard aircraft. This review summarizes the evidence on hand hygiene and the use of facemasks as viral disease prevention measures in aircraft. Methods A literature search was performed in the PubMed, Scopus, and Web of Science databases up to 10 June 2020, according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses criteria. A population, intervention, comparison, outcomes, and study design (PICOS) approach was used to define the review question. Results We included four studies published between 2007 and 2020, all targeting influenza virus disease, in the qualitative synthesis. Three studies used mathematical models to simulate single- or multiple-direction flights, and two of them showed that facemask (e.g., N95 respirator) use considerably reduced infection probability. In the third study, hand cleaning by 20 to 60% of people at any time in all airports (including on aircraft) reduced the measure of airports’ power to spread the disease across the globe by ~ 24 to 69%. The fourth study was a case-control study designed to trace an influenza outbreak in two flights during the 2009 influenza A H1N1 pandemic. The study showed that none (0%) of nine infected passengers compared to 15 (47%) of 32 healthy control passengers in the aircraft cabin during one of these flights wore a facemask (odds ratio, 0.0; 95% confidence interval, 0.0–0.7). In contrast, both case and control passengers appeared to be equally compliant in self-assessed hand hygiene. Conclusions Facemask use combined with hand hygiene may minimize the chance of droplet-transmitted virus spread by air travelers. Thus, it is necessary that hygiene measures become an integral part of standard procedures in commercial air travel.

Abstract Background Understanding the complexity of the vine plant’s response to water deficit represents a major challenge for sustainable winegrowing. Regulation of water use requires a coordinated action between scions and rootstocks on which cultivars are generally grafted to cope with phylloxera infestations. In this regard, a genome-wide association study (GWAS) approach was applied on an ‘ad hoc’ association mapping panel including different Vitis species, in order to dissect the genetic basis of transpiration-related traits and to identify genomic regions of grape rootstocks associated with drought tolerance mechanisms. The panel was genotyped with the GrapeReSeq Illumina 20 K SNP array and SSR markers, and infrared thermography was applied to estimate stomatal conductance values during progressive water deficit. Results In the association panel the level of genetic diversity was substantially lower for SNPs loci (0.32) than for SSR (0.87). GWAS detected 24 significant marker-trait associations along the various stages of drought-stress experiment and 13 candidate genes with a feasible role in drought response were identified. Gene expression analysis proved that three of these genes (VIT_13s0019g03040, VIT_17s0000g08960, VIT_18s0001g15390) were actually induced by drought stress. Genetic variation of VIT_17s0000g08960 coding for a raffinose synthase was further investigated by resequencing the gene of 85 individuals since a SNP located in the region (chr17_10,497,222_C_T) was significantly associated with stomatal conductance. Conclusions Our results represent a step forward towards the dissection of genetic basis that modulate the response to water deprivation in grape rootstocks. The knowledge derived from this study may be useful to exploit genotypic and phenotypic diversity in practical applications and to assist further investigations.

Abstract Background Reverse Transcription-Polymerase Chain Reaction (RT-PCR) for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-COV-2) diagnosis currently requires quite a long time span. A quicker and more efficient diagnostic tool in emergency departments could improve management during this global crisis. Our main goal was assessing the accuracy of artificial intelligence in predicting the results of RT-PCR for SARS-COV-2, using basic information at hand in all emergency departments. Methods This is a retrospective study carried out between February 22, 2020 and March 16, 2020 in one of the main hospitals in Milan, Italy. We screened for eligibility all patients admitted with influenza-like symptoms tested for SARS-COV-2. Patients under 12 years old and patients in whom the leukocyte formula was not performed in the ED were excluded. Input data through artificial intelligence were made up of a combination of clinical, radiological and routine laboratory data upon hospital admission. Different Machine Learning algorithms available on WEKA data mining software and on Semeion Research Centre depository were trained using both the Training and Testing and the K-fold cross-validation protocol. Results Among 199 patients subject to study (median [interquartile range] age 65 [46–78] years; 127 [63.8%] men), 124 [62.3%] resulted positive to SARS-COV-2. The best Machine Learning System reached an accuracy of 91.4% with 94.1% sensitivity and 88.7% specificity. Conclusion Our study suggests that properly trained artificial intelligence algorithms may be able to predict correct results in RT-PCR for SARS-COV-2, using basic clinical data. If confirmed, on a larger-scale study, this approach could have important clinical and organizational implications.

Abstract Background Deficiency of the natural antagonist of interleukin-1 was first described in 2009 and so far 20 patients has been reported. In Brazil just two cases have been reported both carrying the same homozygous 15 bp deletion. Blocking interleukin-1 has changed rate survival for DIRA patients. The use of anakinra and rilonacept has been reported safe and efficient, whereas the selective blockade of interleukin-1 beta, using the monoclonal antibody canakinumab has been reported in a single case only. Case presentation Here we report a case of a 7 years old Brazilian boy that presented with recurrent episodes of systemic inflammation with severe disabling osteomyelitis with mild pustular skin rash. A Next Generation Sequencing gene panel allowed to detect two pathogenic mutations in the IL1RN gene, described in compound heterozygosity. Corticosteroids was effective in controlling inflammation and anti-IL1 beta blocker triggered disease flare. Complete clinical control could be achieved using IL-1 receptor antagonist. Conclusions DIRA is a severe, life threatening autoinflammatory condition with low numbers of patients described all over the world. The mutation p.Asp72_Ile76del in IL1RN is presented in all Brazilian DIRA patients already described and p.Q45* (rs1019766125) is a new mutation affecting the IL1RN gene. Following the pathogenesis of DIRA, blocking both subunits of interleukin one as well as antagonizing the receptor using anakinra or rilonacept seems to be effective. There is just one report using canakinumab for the treatment of DIRA and this is the first report of disease flare using this drug.

Abstract Background Anticholinergic medication use is linked with increased cognitive decline, dementia, falls and mortality, and their use should be limited in older people. Here we estimate the prevalence of anticholinergic use in England’s older population in 1991 and 2011, and describe changes in use by participant’s age, sex, cognition and disability. Methods We compared data from participants aged 65+ years from the Cognitive Function and Ageing Studies (CFAS I and II), collected during 1990–1993 (N = 7635) and 2008–2011 (N = 7762). We estimated the prevalence of potent anticholinergic use (Anticholinergic Cognitive Burden [ACB] score = 3) and average anticholinergic burden (sum of ACB scores), using inverse probability weights standardised to the 2011 UK population. These were stratified by age, sex, Mini-Mental State Examination score, and activities of daily living (ADL) or instrumental ADL (IADL) disability. Results Prevalence of potent anticholinergic use increased from 5.7% (95% Confidence Interval [CI] 5.2–6.3%) of the older population in 1990–93 to 9.9% (9.3–10.7%) in 2008–11, adjusted odds ratio of 1.90 (95% CI 1.67–2.16). People with clinically significant cognitive impairment (MMSE [Mini Mental State Examination] 21 or less) were the heaviest users of potent anticholinergics in CFAS II (16.5% [95% CI 12.0–22.3%]). Large increases in the prevalence of the use medication with ‘any’ anticholinergic activity were seen in older people with clinically significant cognitive impairment (53.3% in CFAS I to 71.5% in CFAS II). Conclusions Use of potent anticholinergic medications nearly doubled in England’s older population over 20 years with some of the greatest increases amongst those particularly vulnerable to anticholinergic side-effects.

Abstract Objectives Hydroxychloroquine has shown to have antiviral activity in vitro against coronaviruses, specifically SARS-CoV-2. It is believed to block virus infection by increasing endosomal pH required for virus cell fusion and glycosylation of viral surface proteins. In addition to its antiviral activity, hydroxychloroquine has an immune-modulating activity that may synergistically enhance its antiviral effect in vivo, making it a potentially promising drug for the prevention and the cure of SARS-CoV-19. However, randomized controlled trials are needed to assess whether it can be used safely to treat COVID-19 patients or to prevent infection. The main objective of the present study is to evaluate the efficacy of hydroxychloroquine for (I) the prevention of COVID-19 or related symptoms in SARS-CoV-2-exposed subjects, such as as household members/contacts of COVID-19 patients and (II) the treatment of early-phase asymptomatic or paucisymptomatic COVID-19 patients. Trial design This is a controlled, open label, cluster-randomized, superiority trial with parallel group design. Subjects will be randomized either to receive hydroxychloroquine or to observation (2:1). Participants SARS-CoV-2-exposed subjects, including household members and/or contacts of COVID-19 patients and healthcare professionals (Group 1) or patients with COVID-19 (positive PCR test on a rhinopharyngeal or oropharyngeal swab for SARS-CoV-2), asymptomatic or paucisymptomatic in home situations who are not undergoing treatment with any anti COVID-19 medication (Group 2), will be enrolled. Paucisymptomatic patients are defined as patients with a low number of mild symptoms. All subjects must be aged ≥18 years, male or female, must be willing and able to give informed consent and must not have any contraindications to take hydroxychloroquine (intolerance or previous toxicity for hydroxychloroquine/chloroquine, bradycardia or reduction in heart rhythm with arrhythmia, ischemic heart disease, retinopathy, congestive heart failure with use of diuretics, favism or glucose-6-phosphate dehydrogenase (G6PD) deficiency, diabetes type 1, major comorbidities such as advanced chronic kidney disease or dialysis therapy, known history of ventricular arrhythmia, any oncologic/hematologic malignancy, severe neurological and mental illness, current use of medications with known significant drug-drug interactions, and known prolonged QT syndrome or current use of drugs with known QT prolongation). The study is monocentric and will be conducted at Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS. Subjects will be enrolled from a large epidemic region (North-Central Italy). The Public Health Departments of several Italian regions will collaborate by identifying potentially eligible subjects. Intervention and comparator The participants will be randomized (2:1 randomization) to receive either hydroxychloroquine (Arm A) or to Observation (Arm B). Hydroxychloroquine will be administered with the following schedule: Group1: A loading dose hydroxychloroquine 400 mg twice daily on day 1, followed by a weekly dose of hydroxychloroquine 200 mg twice daily on days 8, 15 and 22, for a total of one month of treatment. Group 2: A loading dose hydroxychloroquine 400 mg twice daily on day 1 followed by 200 mg twice daily for a total of 5-7 days. The comparator in this trial is observation given that currently neither treatment is administered to asymptomatic or paucisymptomatic subjects, nor prophylaxis is available for contacts. Hydroxychloroquine will be shipped to subjects within 24 hours of randomization. Given the extraordinary nature of the COVID-19 pandemic, only telephonic interviews will be carried out and electronic Patient Reported Outcomes (ePRO) completed. During treatment, each subject will be contacted every other day for the first week and weekly thereafter (Group 2) or weekly (Group 1) by a study physician to assess early onset of any COVID-19 symptom or any adverse reaction to hydroxychloroquine and to check subject compliance. Furthermore, all subjects will receive periodic ePROs which may be completed through smartphone or tablets to record drug self-administration and onset of any symptom or adverse event. All subjects will be followed up for a total of 6 months by periodic telephonic interviews and ePROs. Main outcomes The primary endpoint/outcome measure for this trial is: for Group 1, the proportion of subjects who become symptomatic and/or swab-positive in each arm within one month of randomization; for Group 2, the proportion of subjects who become swab-negative in each arm within 14 days of randomization. Randomization All household members and/or contacts of each COVID-19 index case, and the COVID-19 patient himself/herself, fulfilling all inclusion criteria will be grouped into a single cluster and this cluster will be randomized (2:1) to either arm A or arm B. Information on each subject will be recorded in specific data records. Randomization lists will be stratified according to the following factors regarding COVID-19 index cases: 1. COVID-19 risk level on the basis of province of residence (high vs. low/intermediate); 2. Index case is a healthcare professional (yes vs.no) 3. Index case with COVID-19 treatment (yes vs. no) An independent statistician not otherwise involved in the trial will generate the allocation sequence, and COVID-19 response teams will be unaware of the allocation of clusters. Randomization will be performed through an interactive web-based electronic data-capturing database. An Independent Data Monitoring Committee has been established. Blinding (masking) This study is open label. Numbers to be randomized (sample size) For Group 1, a sample size of about 2000 SARS-CoV-2-exposed subjects such as household members and/or contacts of COVID-19 patients will take part in the study. Assuming around 1.5-2.0 asymptomatic household members and/or contacts for each COVID-19 patient, we expect to identify approximately 1000-1300 COVID-19 index cases to be randomized. An interim analysis on efficacy is planned using standard alpha-spending function. For Group 2, sufficient power for primary objective (negative swab within 14 days of randomization) will be reached given a sample size of 300 asymptomatic or paucisymptomatic COVID-19 subjects in home situations not treated for COVID-19 (25%-30% of about 1000-1300 expected index cases). Since up to date reduced evidence about COVID-19 infection epidemiology, the continuous update of diagnostic and therapeutic approaches, the sample size estimation could be updated after a one third of population will be recruited and eventually modified according to a substantial protocol amendment. An interim analysis at 100 enrolled COVID-19 patients is planned. We have planned a Generalized Estimating Equation analysis, which is more efficient than a cluster level analysis, to take advantage of subject-specific covariates. The above reported sample size analysis is therefore to be considered conservative. Trial Status The current version of the PROTECT trial protocol is ‘Final version, 15 April 2020’. The study started on 9th May 2020. The first patient was enrolled on 14th May 2020. Recruitment is expected to last through September 2020. Trial registration The PROTECT trial is registered in the EudraCT database (no. 2020-001501-24) and in ClinicalTrials.gov ( NCT04363827 ), date of registration 24 April 2020. Full protocol The full PROTECT protocol is attached as an additional file, accessible from the Trials website (Additional file 1). In the interests of expediting dissemination of this material, the familiar formatting has been eliminated; this Letter serves as a summary of the key elements of the full protocol (Protocol final version, 15th April 2020). The study protocol has been reported in accordance with Standard Protocol Items: Recommendations for Clinical Interventional Trials (SPIRIT) guidelines (Additional file 2).

Abstract Background Vestibular symptoms and balance changes are common in patients with migraine, especially in the ones with aura and chronic migraine. However, it is not known if the balance changes are determined by the presence of vestibular symptoms or migraine subdiagnosis. Therefore, the aim of this study was to verify if the migraine subdiagnosis and/or the presence of vestibular symptoms can predict balance dysfunction in migraineurs. Methods The study included 49 women diagnosed with migraine with aura, 53 without aura, 51 with chronic migraine, and 54 headache-free women. All participants answered a structured questionnaire regarding migraine features and presence of vestibular symptoms, such as dizziness/vertigo. The participants performed the Modified Sensory Organization Test on an AMTI© force plate. The data were analysed using a linear mixed-effect regression model. Results The presence of vestibular symptoms did not predict postural sway, but the subdiagnosis was a significant predictor of postural sway. Migraine with aura patients exhibited more sway than migraine patients without aura when the surface was unstable. Additionally, we found high effect sizes (ES > 0.79) for postural sway differences between patients with chronic migraine or with aura compared to controls or migraine without aura, suggesting that these results are clinically relevant. Conclusions The subdiagnosis of migraine, instead of the presence of vestibular symptoms, can predict postural control impairments observed in migraineurs. This lends support to the notion that balance instability is related to the presence of aura and migraine chronicity, and that it should be considered even in patients without vestibular symptoms.

Abstract Background Migraine is a highly disabling condition and pharmacological treatment is the gold standard. However, several patients have also positive responses to the application of different manual techniques and therapeutic exercises in terms of reducing the intensity and frequency of migraine attacks. Nevertheless, the effects of a neck-specific exercise program have not yet been evaluated in these patients. Objective To determine the effectiveness of a neck-specific exercise program in reducing the intensity and frequency of migraine attacks as compared to a sham ultrasound group. Methods A two-armed, parallel-groups randomized controlled trial with 3 months of follow-up will be conducted. 42 individuals, both genders, aged between 18 and 55 years old with a medical diagnosis of migraine will be included. The intervention group will perform a protocol consisting of exercises for strengthening the muscles of the cervical spine. Participants within the sham ultrasound group will receive detuned ultrasound therapy in the upper trapezius muscle. Both groups will receive a weekly session for 8 weeks. The efficacy of each intervention will be measured by the frequency and intensity of migraine at a 3-months follow-up. Trial registration This study was registered under access code RBT-8gfv5j in the Registro Brasileiro de Ensaios Clínicos (ReBEC) in November 28, 2016. Conclusion This study will aim to determine the efficacy of a neck-specific exercise program in reducing the frequency and intensity of migraine attacks. If the results show that a neck-specific exercise program is effective in reducing the frequency and intensity of migraine attacks, therapists will have a low cost and easily applicable tool to treat migraine.

Abstract Background The Critical Assessment of Functional Annotation (CAFA) is an ongoing, global, community-driven effort to evaluate and improve the computational annotation of protein function. Results Here, we report on the results of the third CAFA challenge, CAFA3, that featured an expanded analysis over the previous CAFA rounds, both in terms of volume of data analyzed and the types of analysis performed. In a novel and major new development, computational predictions and assessment goals drove some of the experimental assays, resulting in new functional annotations for more than 1000 genes. Specifically, we performed experimental whole-genome mutation screening in Candida albicans and Pseudomonas aureginosa genomes, which provided us with genome-wide experimental data for genes associated with biofilm formation and motility. We further performed targeted assays on selected genes in Drosophila melanogaster, which we suspected of being involved in long-term memory. Conclusion We conclude that while predictions of the molecular function and biological process annotations have slightly improved over time, those of the cellular component have not. Term-centric prediction of experimental annotations remains equally challenging; although the performance of the top methods is significantly better than the expectations set by baseline methods in C. albicans and D. melanogaster, it leaves considerable room and need for improvement. Finally, we report that the CAFA community now involves a broad range of participants with expertise in bioinformatics, biological experimentation, biocuration, and bio-ontologies, working together to improve functional annotation, computational function prediction, and our ability to manage big data in the era of large experimental screens.

Abstract Background Drought is one of the most harmful abiotic stresses for plants, leading to reduced productivity of several economically important crops and, consequently, considerable losses in the agricultural sector. When plants are exposed to stressful conditions, such as drought and high salinity, they modulate the expression of genes that lead to developmental, biochemical, and physiological changes, which help to overcome the deleterious effects of adverse circumstances. Thus, the search for new specific gene promoter sequences has proved to be a powerful biotechnological strategy to control the expression of key genes involved in water deprivation or multiple stress responses. Results This study aimed to identify and characterize the GmRD26 promoter (pGmRD26), which is involved in the regulation of plant responses to drought stress. The expression profile of the GmRD26 gene was investigated by qRT-PCR under normal and stress conditions in Williams 82, BR16 and Embrapa48 soybean-cultivars. Our data confirm that GmRD26 is induced under water deficit with different induction folds between analyzed cultivars, which display different genetic background and physiological behaviour under drought. The characterization of the GmRD26 promoter was performed under simulated stress conditions with abscisic acid (ABA), polyethylene glycol (PEG) and drought (air dry) on A. thaliana plants containing the complete construct of pGmRD26::GUS (2.054 bp) and two promoter modules, pGmRD26A::GUS (909 pb) and pGmRD26B::GUS (435 bp), controlling the expression of the β-glucuronidase (uidA) gene. Analysis of GUS activity has demonstrated that pGmRD26 and pGmRD26A induce strong reporter gene expression, as the pAtRD29 positive control promoter under ABA and PEG treatment. Conclusions The full-length promoter pGmRD26 and the pGmRD26A module provides an improved uidA transcription capacity when compared with the other promoter module, especially in response to polyethylene glycol and drought treatments. These data indicate that pGmRD26A may become a promising biotechnological asset with potential use in the development of modified drought-tolerant plants or other plants designed for stress responses.

Abstract Background Hirschsprung Disease (HSCR) is a congenital defect of the intestinal innervations characterized by complex inheritance. Many susceptibility genes including RET, the major HSCR gene, and several linked regions and associated loci have been shown to contribute to disease pathogenesis. Nonetheless, a proportion of patients still remains unexplained. Copy Number Variations (CNVs) have already been involved in HSCR, and for this reason we performed Comparative Genomic Hybridization (CGH), using a custom array with high density probes. Results A total of 20 HSCR candidate regions/genes was tested in 55 sporadic patients and four patients with already known chromosomal aberrations. Among 83 calls, 12 variants were experimentally validated, three of which involving the HSCR crucial genes SEMA3A/3D, NRG1, and PHOX2B. Conversely RET involvement in HSCR does not seem to rely on the presence of CNVs while, interestingly, several gains and losses did co-occur with another RET defect, thus confirming that more than one predisposing event is necessary for HSCR to develop. New loci were also shown to be involved, such as ALDH1A2, already found to play a major role in the enteric nervous system. Finally, all the inherited CNVs were of maternal origin. Conclusions Our results confirm a wide genetic heterogeneity in HSCR occurrence and support a role of candidate genes in expression regulation and cell signaling, thus contributing to depict further the molecular complexity of the genomic regions involved in the Enteric Nervous System development. The observed maternal transmission bias for HSCR associated CNVs supports the hypothesis that in females these variants might be more tolerated, requiring additional alterations to develop HSCR disease.

Abstract Background Studies suggest that anticholinergic medication or benzodiazepine use could increase dementia risk. We tested this hypothesis using data from a UK cohort study. Methods We used data from the baseline (Y0), 2-year (Y2) and 10-year (Y10) waves of the Medical Research Council Cognitive Function and Ageing Study. Participants without dementia at Y2 were included (n = 8216). Use of benzodiazepines (including nonbenzodiazepine Z-drugs), anticholinergics with score 3 (ACB3) and anticholinergics with score 1 or 2 (ACB12) according to the Anticholinergic Cognitive Burden scale were coded as ever use (use at Y0 or Y2), recurrent use (Y0 and Y2), new use (Y2, but not Y0) or discontinued use (Y0, but not Y2). The outcome was incident dementia by Y10. Incidence rate ratios (IRR) were estimated using Poisson regression adjusted for potential confounders. Pre-planned subgroup analyses were conducted by age, sex and Y2 Mini-Mental State Examination (MMSE) score. Results Dementia incidence was 9.3% (N = 220 cases) between Y2 and Y10. The adjusted IRRs (95%CI) of developing dementia were 1.06 (0.72, 1.60), 1.28 (0.82, 2.00) and 0.89 (0.68, 1.17) for benzodiazepines, ACB3 and ACB12 ever-users compared with non-users. For recurrent users the respective IRRs were 1.30 (0.79, 2.14), 1.68 (1.00, 2.82) and 0.95 (0.71, 1.28). ACB3 ever-use was associated with dementia among those with Y2 MMSE> 25 (IRR = 2.28 [1.32–3.92]), but not if Y2 MMSE≤25 (IRR = 0.94 [0.51–1.73]). Conclusions Neither benzodiazepines nor ACB12 medications were associated with dementia. Recurrent use of ACB3 anticholinergics was associated with dementia, particularly in those with good baseline cognitive function. The long-term prescribing of anticholinergics should be avoided in older people.

Abstract Background One of the main issues in the automated protein function prediction (AFP) problem is the integration of multiple networked data sources. The UNIPred algorithm was thereby proposed to efficiently integrate —in a function-specific fashion— the protein networks by taking into account the imbalance that characterizes protein annotations, and to subsequently predict novel hypotheses about unannotated proteins. UNIPred is publicly available as R code, which might result of limited usage for non-expert users. Moreover, its application requires efforts in the acquisition and preparation of the networks to be integrated. Finally, the UNIPred source code does not handle the visualization of the resulting consensus network, whereas suitable views of the network topology are necessary to explore and interpret existing protein relationships. Results We address the aforementioned issues by proposing UNIPred-Web, a user-friendly Web tool for the application of the UNIPred algorithm to a variety of biomolecular networks, already supplied by the system, and for the visualization and exploration of protein networks. We support different organisms and different types of networks —e.g., co-expression, shared domains and physical interaction networks. Users are supported in the different phases of the process, ranging from the selection of the networks and the protein function to be predicted, to the navigation of the integrated network. The system also supports the upload of user-defined protein networks. The vertex-centric and the highly interactive approach of UNIPred-Web allow a narrow exploration of specific proteins, and an interactive analysis of large sub-networks with only a few mouse clicks. Conclusions UNIPred-Web offers a practical and intuitive (visual) guidance to biologists interested in gaining insights into protein biomolecular functions. UNIPred-Web provides facilities for the integration of networks, and supplies a framework for the imbalance-aware protein network integration of nine organisms, the prediction of thousands of GO protein functions, and a easy-to-use graphical interface for the visual analysis, navigation and interpretation of the integrated networks and of the functional predictions.

Abstract Background Familial Mediterranean Fever is a monogenic autoinflammatory disease, typically characterized by recurrent attacks of fever, serositis, aphthous of oral mucosa, erythema. “Pyogenic arthritis, pyoderma gangrenosum and acne syndrome” is a rare autoinflammatory disease with variable expression and typically involving joints and skin. Both the diseases are linked by the overproduction of IL-1. Case presentation We report on the case of two siblings affected by recurrent attacks of fever, oral aphthous stomatitis, abdominal pain, arthritis, undefined dermatitis at the hands, associated with increased AST, ALT, C-reactive protein, erythrocyte sedimentation rate, serum amyloid A, leucocytosis with neutrophilia. Infectious diseases were excluded. The genetic study for Familial Mediterranean Fever, tumor necrosis factor receptor-associated periodic syndrome, Mevalonate kinase deficiency, showed the homozygous mutation p.M680I of exon 10 in MEFV. Their parents were heterozygous for the same mutation p.M680I, however, the mother showed severe symptoms of FMF (recurrent attacks of fever, arthralgia and arthritis, abdominal pain, thoracic pain), the father showed recurrent pustulosis prevalent on the hands and limbs, with arthralgia and abdominal pain. Both the patients started colchicine, with an improvement in clinical manifestations and a reduction of serum amyloid A. For the atypical dermatologic signs present in the two siblings and in the father, the study of other autoinflammatory syndromes was performed with next generation sequencing and showed the heterozygous rare missense mutation of unknown significance: p.(Val408Ile) of PSTPIP1 gene in the two siblings and in the mother, the father was negative. Canakinumab treatment was started in the younger patient, with the resolution of the clinical symptoms and the normalization of serum amyloid A. Conclusions Further studies are needed to better describe the correlation between genotype and phenotype in patients with PAPA syndrome and with PAPA syndrome associated with FMF, considering that the presence of mutations in both genes may amplify clinical presentation and evolution of both diseases.

Abstract Background Autosomal dominant gain of function mutations in caspase recruitment domain family member 14 (CARD14) is a rare condition associated with plaque-type psoriasis, generalized pustular psoriasis, palmoplantar pustular psoriasis and pityriasis rubra pilaris. Recently, a new CARD14 –associated phenotype defined as CAPE (CARD14-associated papulosquamous eruption) with clinical features of both psoriasis and pityriasis rubra pilaris was reported. We describe a family carrying a novel heterozygous mutation in CARD14 gene, with childhood-onset erythrodermic psoriasis requiring an unusual extremely high dose (up to 2 mg/kg every 8 weeks) of ustekinumab to achieve disease remission. Case presentation We describe a large family with three pairs of twins presenting a clinical phenotype characterized by childhood-onset erythrodermic psoriasis; in some family members is also reported psoriatic arthritis. The two probands presented poor clinical response to topic and systemic therapy with antihistamine, steroid, retinoids, cyclosporine and etanercept. After exclusion of the most common genes associated to autoinflammatory diseases (IL36RN, IL1RN, MVK, TNFRSF1A, NLRP3, NLRP12, MEFV, NOD2, PSMB8, PSTPIP1, LPIN2) we approached a new gene search by subjecting to Whole Exome Sequencing (WES) analysis five members of the family. A novel heterozygous mutation (c.446 T > G, leading to the missense amino acid substitution p.L149R) in the exon 4 of the CARD14 gene was identified in all affected members. Increasing dosages (up to 2 mg/kg every 8 weeks) of ustekinumab, a human monoclonal antibody targeting interleukin-12 (IL-12) and interleukin-23 (IL-23), allowed the complete control of the clinical manifestations, with an evident reduction of circulating Th17 and Th22 CD4+ T cell subsets. Conclusions We describe the association of mutations of the CARD14 gene with an erythrodermic psoriasis pedigree, underlying the necessity to investigate CARD14 mutations in childhood-onset psoriasis cases and confirming the presence of CARD14 causative mutations also in erythrodermic psoriasis form, as recently reported. Also in pediatric age, ustekinumab represents a powerful therapeutic option for this rare condition, that is usually refractory to other treatments. In young children, high and frequent dosages allowed a complete control of the clinical manifestations without any severe side effects, with a long-term follow-up.

Abstract Background The demonstration of EGFR T790M gene mutation in plasma is crucial to assess the eligibility of Non Small Cell Lung Cancer (NSCLC) patients, who have acquired resistance to first or second generation Tyrosine Kinase Inhibitors (TKIs), to receive a subsequent treatment with osimertinib. Since circulating tumor DNA (ctDNA) is present in very low amounts in plasma, high sensitive and specific methods are required for molecular analysis. Improving sensitivity of T790M mutation detection in plasma ctDNA enables a larger number of NSCLC patients to receive the appropriate therapy without any further invasive procedure. Methods A tag-based next generation sequencing (NGS) platform capable of tagging rare circulating tumor DNA alleles was employed in this study for the identification of T790M mutation in 42 post-TKI NSCLC patients. Results Compared to Real Time PCR, tag-based NGS improved the T790M detection rate (42.85% versus 21.4%, respectively), especially in those cases with a low median mutation abundance (i.e. 0.24, range 0.07–0.78). Moreover, the tag-based NGS identified EGFR activating mutations more efficiently than Real Time PCR (85.7% versus 61.9% detection rate, respectively), particularly of the L858R variant type (0.06–0.75 mutation abundance range). Patients in whom the T790M mutation was detected in plasma, achieved an objective response to osimertinib (9/14, 64.28%). Conclusions Tag-based NGS represents an accurate and sensitive tool in a clinical setting for non-invasive assessment and monitoring of T790M variant in NSCLC patients.

Abstract Background Insect resistance in crops represents a main challenge for agriculture. Transgenic approaches based on proteins displaying insect resistance properties are widely used as efficient breeding strategies. To extend the spectrum of targeted pathogens and overtake the development of resistance, molecular evolution strategies have been used on genes encoding these proteins to generate thousands of variants with new or improved functions. The cotton boll weevil (Anthonomus grandis) is one of the major pests of cotton in the Americas. An α-amylase inhibitor (α-AIC3) variant previously developed via molecular evolution strategy showed inhibitory activity against A. grandis α-amylase (AGA). Results We produced in a few days considerable amounts of α-AIC3 using an optimised transient heterologous expression system in Nicotiana benthamiana. This high α-AIC3 accumulation allowed its structural and functional characterizations. We demonstrated via MALDI-TOF MS/MS technique that the protein was processed as expected. It could inhibit up to 100% of AGA biological activity whereas it did not act on α-amylase of two non-pathogenic insects. These data confirmed that N. benthamiana is a suitable and simple system for high-level production of biologically active α-AIC3. Based on other benefits such as economic, health and environmental that need to be considerate, our data suggested that α-AIC3 could be a very promising candidate for the production of transgenic crops resistant to cotton boll weevil without lethal effect on at least two non-pathogenic insects. Conclusions We propose this expression system can be complementary to molecular evolution strategies to identify the most promising variants before starting long-lasting stable transgenic programs.

Abstract Background CEA, CYFRA21-1 and NSE are tumor markers used for monitoring the response to chemotherapy in advanced adenocarcinoma, squamous cell carcinoma and small-cell lung cancer, respectively. Their role in cancer immunotherapy needs to be elucidated. Methods Patients with advanced non-small cell lung cancer (NSCLC) were treated with nivolumab 3 mg/kg every 2 weeks within the Italian Nivolumab Expanded Access Program. Blood samples were collected at baseline, at each cycle up to cycle 5 and then every two cycles until patient’s withdrawn from the study. All patients underwent a CT-scan after every 4 cycles of treatment and responses were classified according to RECIST 1.1. The biomarkers serum levels were measured with a chemiluminescent microparticle immunoassay for CEA and with an immuno radiometric assay for CYFRA21-1 and NSE. The markers values at baseline and after 4 cycles were used to analyze the relationship between their variation over baseline and the tumor response, evaluated as disease control rate (DCR: CR + PR + SD), and survival (PFS and OS). Results A total of 70 patients were evaluable for the analysis. Overall, a disease control was obtained in 24 patients (35.8%, 4 PR + 20 SD). After 4 cycles of nivolumab a CEA or CYFRA21-1 reduction ≥ 20% over the baseline was significantly associated with DCR (CEA, p = 0.021; CYFRA21-1, p

Abstract Background Several problems in network biology and medicine can be cast into a framework where entities are represented through partially labeled networks, and the aim is inferring the labels (usually binary) of the unlabeled part. Connections represent functional or genetic similarity between entities, while the labellings often are highly unbalanced, that is one class is largely under-represented: for instance in the automated protein function prediction (AFP) for most Gene Ontology terms only few proteins are annotated, or in the disease-gene prioritization problem only few genes are actually known to be involved in the etiology of a given disease. Imbalance-aware approaches to accurately predict node labels in biological networks are thereby required. Furthermore, such methods must be scalable, since input data can be large-sized as, for instance, in the context of multi-species protein networks. Results We propose a novel semi-supervised parallel enhancement of COSNet, an imbalance-aware algorithm build on Hopfield neural model recently suggested to solve the AFP problem. By adopting an efficient representation of the graph and assuming a sparse network topology, we empirically show that it can be efficiently applied to networks with millions of nodes. The key strategy to speed up the computations is to partition nodes into independent sets so as to process each set in parallel by exploiting the power of GPU accelerators. This parallel technique ensures the convergence to asymptotically stable attractors, while preserving the asynchronous dynamics of the original model. Detailed experiments on real data and artificial big instances of the problem highlight scalability and efficiency of the proposed method. Conclusions By parallelizing COSNet we achieved on average a speed-up of 180x in solving the AFP problem in the S. cerevisiae, Mus musculus and Homo sapiens organisms, while lowering memory requirements. In addition, to show the potential applicability of the method to huge biomolecular networks, we predicted node labels in artificially generated sparse networks involving hundreds of thousands to millions of nodes.

Abstract Background This study aimed to evaluate the survival rate of glass hybrid restorations placed under the atraumatic restorative treatment (ART) technique in first permanent molars affected by molar incisor hypomineralization (MIH). Methods Sixty teeth with severe MIH associated to carious dentin lesions without pulp involvement were included. Treatments were performed by one trained dentist using the ART approach and restored with a glass hybrid restorative system (Equia Forte, GC®) on school premises. Treatments were evaluated after 6 and 12 months by an independent examiner using the modified ART criterion. Data analysis involved descriptive statistics and actuarial success analysis. Results The sample comprised 24 (54.54%) girls and 20 (45.45%) boys with a mean age of 10.55 (±1.25) years. In regard to the number of surfaces involved in the restorations, 29 (48.3%) comprised one surface and 31 (51.7%) two or more surfaces. Considering cavity extent, 25 (41%) presented dentin cavitation without cusp weakness, 23 (37.7%) with large dentin cavitation with cusp weakness and 13 (21.3%) with large dentin cavitation with the breakdown of one or more cusps. Only 4 teeth required local anesthesia. A success rate of 98.3% after 6 and 12 months was observed, as only one restoration failed. The only failure occurred in a restoration involving three or more sur-faces presenting the breakdown of all cusps. Conclusion Restorations using a glass hybrid restorative system and performed in the field with the ART technique proved, after 12 months of evaluation, to be an effective approach to preserving first permanent molars affected by MIH. Trial registration REBEC-RBR-8drccq (17/06/15).

Abstract Background Survival advantage following trans-arterial chemoembolization (TACE) is variable in patients with hepatocellular carcinoma (HCC). We combined pre-TACE radiologic features to derive a novel prognostic signature in HCC. Methods A multi-institutional dataset of 98 patients was generated from two retrospective cohorts from United Kingdom (65%) and Italy (36%). The prognostic impact of a number baseline imaging parameters was assessed and factors significant on univariate analysis were combined to create a novel radiologic signature on multivariable analyses predictive of overall survival (OS) following TACE. Results Median OS was 15.4 months. Tumour size > 7 cm (p 7 cm, ITN and AEN identified patients with poor prognosis (p

Abstract Background Laparoscopic ventral mesh rectopexy (LVMR) is an established treatment for external full-thickness rectal prolapse. However, its clinical efficacy in patients with internal prolapse is uncertain due to the lack of high-quality evidence. Methods An individual level, stepped-wedge randomised trial has been designed to allow observer-blinded data comparisons between patients awaiting LVMR with those who have undergone surgery. Adults with symptomatic internal rectal prolapse, unresponsive to prior conservative management, will be eligible to participate. They will be randomised to three arms with different delays before surgery (0, 12 and 24 weeks). Efficacy outcome data will be collected at equally stepped time points (12, 24, 36 and 48 weeks). The primary objective is to determine clinical efficacy of LVMR compared to controls with reduction in the Patient Assessment of Constipation Quality of Life (PAC-QOL) at 24 weeks serving as the primary outcome. Secondary objectives are to determine: (1) the clinical effectiveness of LVMR to 48 weeks to a maximum of 72 weeks; (2) pre-operative determinants of outcome; (3) relevant health economics for LVMR; (4) qualitative evaluation of patient and health professional experience of LVMR and (5) 30-day morbidity and mortality rates. Discussion An individual-level, stepped-wedge, randomised trial serves the purpose of providing an untreated comparison for the active treatment group, while at the same time allowing the waiting-listed participants an opportunity to obtain the intervention at a later date. In keeping with the basic ethical tenets of this design, the average waiting time for LVMR (12 weeks) will be shorter than that for routine services (24 weeks). Trial registration ISRCTN registry, ISRCTN11747152. Registered on 30 September 2015. The trial was prospectively registered (first patient enrolled on 21 March 2016).

Abstract Background Our aim was to identify genomic regions via genome-wide association studies (GWAS) to improve the predictability of genetic merit in Holsteins for 10 calving and 28 body conformation traits. Animals were genotyped using the Illumina Bovine 50 K BeadChip and imputed to the Illumina BovineHD BeadChip (HD). GWAS were performed on 601,717 real and imputed single nucleotide polymorphism (SNP) genotypes using a single-SNP mixed linear model on 4841 Holstein bulls with breeding value predictions and followed by gene identification and in silico functional analyses. The association results were further validated using five scenarios with different numbers of SNPs. Results Seven hundred and eighty-two SNPs were significantly associated with calving performance at a genome-wise false discovery rate (FDR) of 5%. Most of these significant SNPs were on chromosomes 18 (71.9%), 17 (7.4%), 5 (6.8%) and 7 (2.4%) and mapped to 675 genes, among which 142 included at least one significant SNP and 532 were nearby one (100 kbp). For body conformation traits, 607 SNPs were significant at a genome-wise FDR of 5% and most of them were located on chromosomes 5 (30%), 18 (27%), 20 (13%), 6 (6%), 7 (5%), 14 (5%) and 13 (3%). SNP enrichment functional analyses for calving traits at a FDR of 1% suggested potential biological processes including musculoskeletal movement, meiotic cell cycle, oocyte maturation and skeletal muscle contraction. Furthermore, pathway analyses suggested potential pathways associated with calving performance traits including tight junction, oxytocin signaling, and MAPK signaling (P