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Enhancing the removal of aggregate-prone toxic proteins is a rational therapeutic strategy for a number of neurodegenerative diseases, especially Huntington’s disease and various spinocerebellar ataxias. Ideally, such approaches should preferentially clear the mutant/misfolded species, while having minimal impact on the stability of wild-type/normally-folded proteins. Furthermore, activation of both ubiquitin-proteasome and autophagy-lysosome routes may be advantageous, as this would allow effective clearance of both monomeric and oligomeric species, the latter which are inaccessible to the proteasome. Here we find that compounds that activate the D1 ATPase activity of VCP/p97 fulfill these requirements. Such effects are seen with small molecule VCP activators like SMER28, which activate autophagosome biogenesis by enhancing interactions of PI3K complex components to increase PI(3)P production, and also accelerate VCP-dependent proteasomal clearance of such substrates. Thus, this mode of VCP activation may be a very attractive target for many neurodegenerative diseases. Several neurodegenerative diseases are characterized by the aggregation of cytoplasmic proteins. Here, the authors demonstrate that the small molecule SMER28 activates VCP, which enhances both autophagic and proteasomal clearance of aggregate-prone proteins.

The internal ribosome entry site (IRES), a highly conserved structured element of the hepatitis C virus (HCV) genomic RNA, is an attractive target for antiviral drugs. Here we show that benzimidazole inhibitors of the HCV replicon act by conformational induction of a widened interhelical angle in the IRES subdomain IIa, which facilitates the undocking of subdomain IIb from the ribosome and ultimately leads to inhibition of IRES-driven translation in HCV-infected cells.

Mass Spectrometry (MS) based chemoproteomics has recently become a main tool to identify and quantify cellular target protein interactions with ligands/drugs in drug discovery. The complexity associated with these new types of data requires scientists with a limited computational background to perform systematic data quality controls as well as to visualize the results derived from the analysis to enable rapid decision making. To date, there are no readily accessible platforms specifically designed for chemoproteomics data analysis. We developed a Shiny-based web application named DOSCHEDA (Down Stream Chemoproteomics Data Analysis) to assess the quality of chemoproteomics experiments, to filter peptide intensities based on linear correlations between replicates, and to perform statistical analysis based on the experimental design. In order to increase its accessibility, DOSCHEDA is designed to be used with minimal user input and it does not require programming knowledge. Typical inputs can be protein fold changes or peptide intensities obtained from Proteome Discover, MaxQuant or other similar software. DOSCHEDA aggregates results from bioinformatics analyses performed on the input dataset into a dynamic interface, it encompasses interactive graphics and enables customized output reports. DOSCHEDA is implemented entirely in R language. It can be launched by any system with R installed, including Windows, Mac OS and Linux distributions. DOSCHEDA is hosted on a shiny-server at https://doscheda.shinyapps.io/doscheda and is also available as a Bioconductor package (http://www.bioconductor.org/).

The ability to quantitatively probe diverse panels of proteins and their post-translational modifications (PTMs) across multiple samples would aid a broad spectrum of biological, biochemical and pharmacological studies. We report a novel, microarray analytical technology that combines immuno-affinity capture with Matrix Assisted Laser Desorption Ionization Mass Spectrometry (MALDI MS), which is capable of supporting highly multiplexed, targeted proteomic assays. Termed “Affinity-Bead Assisted Mass Spectrometry” (Affi-BAMS), this LC-free technology enables development of highly specific and customizable assay panels for simultaneous profiling of multiple proteins and PTMs. While affinity beads have been used previously in combination with MS, the Affi-BAMS workflow uses enrichment on a single bead that contains one type of antibody, generally capturing a single analyte (protein or PTM) while having enough binding capacity to enable quantification within approximately 3 orders of magnitude. The multiplexing capability is achieved by combining Affi-BAMS beads with different protein specificities. To enable screening of bead-captured analytes by MS, we further developed a novel method of performing spatially localized elution of targets from individual beads arrayed on a microscope slide. The resulting arrays of micro spots contain highly concentrated analytes localized within 0.5 mm diameter spots that can be directly measured using MALDI MS. While both intact proteins and protein fragments can be monitored by Affi-BAMS, we initially focused on applying this technology for bottom-up proteomics to enable screening of hundreds of samples per day by combining the robust magnetic bead-based workflow with the high throughput nature of MALDI MS acquisition. To demonstrate the variety of applications and robustness of Affi-BAMS, several studies are presented that focus on the response of 4EBP1, RPS6, ERK1/ERK2, mTOR, Histone H3 and C-MET to stimuli including rapamycin, H2O2, EPO, SU11274, Staurosporine and Vorinostat.

The internal ribosome entry site (IRES), a highly conserved structured element of the hepatitis C virus genomic RNA, is an attractive target for antiviral drugs. Here we show that benzimidazole inhibitors of the HCV replicon act by conformational induction of a widened interhelical angle in the IRES subdomain IIa which facilitates the undocking of subdomain IIb from the ribosome and ultimately leads to inhibition of IRES-driven translation in HCV-infected cells.

The mature left ventricular myocardium is arranged in a complex three-dimensional network of fibers that form a counterclockwise helix in the endocardial layer and a clockwise helix in the epicardial layer. There are no data in the literature on the development of left ventricular myocardium during the fetal life. The aims of this paper were to study the physiological maturation steps of the LV myocardium in fetuses from 17 to 40 gestational weeks, by means of speckle tracking applied to the endocardial and epicardial aspect of the left ventricle, and, to confirm our finds, through the histologic study of the myocardium of demised fetuses. We studied longitudinal endocardial and epicardial strain by echocardiography in 105 fetuses. Twenty non-diseased fetal hearts from autopsies were selected to assess the layer thickness and cardiac fiber orientation in relation to gestational age. Echocardiography showed a progressive increasing of epicardial/endocardial longitudinal strain ratio with gestational age (r=0.51; p<0.0001). The strain rate E/A ratio increased over time (r=0.27; p=0.018). Histological data revealed that during the same gestational period, the proportion of the epicardial layer increased fourfold, the mesocardiac layer decreased and the endocardial layer remained stable. We found an excellent correlation between the epicardial to endocardial strain ratio and epicardial to endocardial wall thickness (r=0.950, p<0.001). Left ventricular myocardium maturation begins early during fetal life. As the fetus develops, both the relative tissue volume and peak systolic strain rates shift together from the endocardium towards the epicardium. It is a slow process, completed late in fetal life.

Abstract Background Vaccine hesitancy is an emerging phenomenon in European countries and leads to decreasing trends in infant vaccine coverage. The aim of this study was to analyze the level of confidence and correct awareness about immunizations, which are crucial for the success of vaccination programmes. Methods As part of the NAVIDAD multicentre study, we examined vaccination confidence and complacency among a sample of 1820 pregnant women from 14 Italian cities. The questionnaire assessed the interviewee's knowledge, beliefs and misconceptions, as well as their socioeconomic status, information sources about vaccines and confidence in the Italian National Healthcare Service. Results Only 9% of women completely believed to the efficacy, necessity and safety of vaccinations. Almost 20% of them had misconceptions on most of the themes. There was a significant difference in the level of knowledge considering educational level: women with a high educational level have less probability of obtaining a low knowledge score (odds ratio (OR) 0.43 [95% confidence interval (CI) 0.34–0.54]). The level of knowledge was also influenced by the sources of information: women who received information from their general practitioner (GP) and from institutional websites had a significantly lower chance of having misconceptions (OR 0.74 [95% CI 0.58–0.96]; OR 0.59 [95% CI 0.46–0.74]). Finally, the results underlined the influence of trust in healthcare professional information on the likelihood of having misconceptions (OR 0.49 [95% CI 0.27–0.89]). Conclusions The data suggest the efficacy of GPs and institutional websites as a source of information to contrast misconceptions and underline the importance of confidence in the healthcare system to increase complacency and confidence in vaccines.

Despite the increasing trend in liver resections for non-colorectal non-neuroendocrine liver metastases (NCNNLM), the role of surgery for these liver malignancies is still debated. Registries are an essential, reliable tool for assessing epidemiology, diagnosis, and therapeutic approach in a single hub, especially when data are dispersive and inconclusive, as in our case. The dissemination of this preliminary survey would allow us to understand if the creation of an International Registry is a viable option, while still offering a snapshot on this issue, investigating clinical practices worldwide. The steering committee designed an online questionnaire with Google Forms, which consisted of 37 questions, and was open from October 5th, 2022, to November 30th, 2022. It was disseminated using social media and mailing lists of the Italian Society of Endoscopic Surgery and New Technologies (SICE), the Association of Italian Surgeons in Europe (ACIE), and the Spanish Chapter of the American College of Surgeons (ACS). Overall, 141 surgeons (approximately 18% of the total invitations sent) from 27 countries on four continents participated in the survey. Most respondents worked in general surgery units (62%), performing less than 50 liver resections/year (57%). A multidisciplinary discussion was currently performed to validate surgical indications for NCNNLM in 96% of respondents. The most commonly adopted selection criteria were liver resectability, RECIST criteria, and absence of extrahepatic disease. Primary tumors were generally of gastrointestinal (42%), breast (31%), and pancreaticobiliary origin (13%). The most common interventions were parenchymal-sparing resections (51% of respondents) of metachronous metastases with an open approach. Major post-operative complications (Clavien–Dindo > 2) occurred in up to 20% of the procedures, according to 44% of respondents. A subset analysis of data from high-volume centers (> 100 cases/year) showed lower post-operative complications and better survival. The present survey shows that NCNNLM patients are frequently treated by surgeons in low-volume hospitals for liver surgery. Selection criteria are usually based on common sense. Liver resections are performed mainly with an open approach, possibly carrying a high burden of major post-operative complications. International guidelines and a specific consensus on this field are desirable, as well as strategies for collaboration between high-volume and low-volume centers. The present study can guide the elaboration of a multi-institutional document on the optimal pathway in the management of patients with NCNNLM.