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Fibroblast growth factors (FGFs) act as proangiogenic and mitogenic cytokines in several cancers, including multiple myeloma (MM). Indeed, corrupted FGF autocrine and paracrine secretion induces an aberrant activation of the FGF receptor (FGFR) signaling sustaining cancer cell spreading and resistance to pharmacological treatments. Thus, FGF traps may represent a promising anti-cancer strategy to hamper the ligand-dependent activation of the FGF/FGFR system. We previously identified NSC12 as the first orally available small molecule FGF trap able to inhibit the growth and progression of several FGF-dependent tumor models. NSC12 is a pregnenolone derivative carrying a 1,1-bis-trifluoromethyl-1,3-propanediol chain in position 17 of the steroid nucleus. Investigation of structure-activity relationships (SARs) provided more potent and specific NSC12 steroid derivatives and highlighted that the C17-side chain is pivotal for the FGF trap activity. Here, a scaffold hopping approach allowed to obtain two FGF trap compounds (22 and 57) devoid of the steroid nucleus and able to efficiently bind FGF2 and to inhibit FGFR activation in MM cells. Accordingly, these compounds exert a potent anti-tumor activity on MM cell lines both in vitro and in vivo and on MM patient-derived primary cells, strongly affecting the survival of both proteasome-inhibitor sensitive and resistant MM cells. These results propose a new therapeutic option for relapsed/refractory MM patients and set the bases for the development of novel FGF traps prone to chemical diversification to be used in the clinic for the treatment of those tumors in which the FGF/FGFR system plays a pivotal role, including MM.

Abstract The EGFR tyrosine kinase inhibitor osimertinib has been approved for the first-line treatment of EGFR-mutated Non-Small Cell Lung Cancer (NSCLC) patients. Despite its efficacy, patients develop resistance. Mechanisms of resistance are heterogeneous and not fully understood, and their characterization is essential to find new strategies to overcome resistance. Ceramides are well-known regulators of apoptosis and are converted into glucosylceramides (GlcCer) by glucosylceramide synthase (GCS). A higher content of GlcCers was observed in lung pleural effusions from NSCLC patients and their role in osimertinib-resistance has not been documented. The aim of this study was to determine the therapeutic potential of inhibiting GCS in NSCLC EGFR-mutant models resistant to osimertinib in vitro and in vivo. Lipidomic analysis showed a significant increase in the intracellular levels of glycosylceramides, including GlcCers in osimertinib resistant clones compared to sensitive cells. In resistant cells, the GCS inhibitor PDMP caused cell cycle arrest, inhibition of 2D and 3D cell proliferation, colony formation and migration capability, and apoptosis induction. The intratumoral injection of PDMP completely suppressed the growth of OR xenograft models. This study demonstrated that dysregulation of ceramide metabolism is involved in osimertinib-resistance and targeting GCS may be a promising therapeutic strategy for patients progressed to osimertinib.

Abstract Circulating monocytes participate in pain chronification but the molecular events that cause their deployment are unclear. Using a mouse model of hyperalgesic priming (HP), we show that monocytes enable progression to pain chronicity through a mechanism that requires transient activation of the hydrolase, N-acylethanolamine acid amidase (NAAA), and the consequent suppression of NAAA-regulated lipid signaling at peroxisome proliferator-activated receptor-α (PPAR-α). Inhibiting NAAA in the 72 hours following administration of a priming stimulus prevented HP. This effect was phenocopied by NAAA deletion and depended on PPAR-α recruitment. Mice lacking NAAA in CD11b+ cells – monocytes, macrophages, and neutrophils – were resistant to HP induction. Conversely, mice overexpressing NAAA or lacking PPAR-α in the same cells were constitutively primed. Depletion of monocytes, but not resident macrophages, generated mice that were refractory to HP. The results identify NAAA-regulated signaling in monocytes as a control node in the induction of HP and, potentially, the transition to pain chronicity.

ABSTRACT The human organism is inhabited by trillions of microorganisms, known as microbiota, which are considered to exploit a pivotal role in the regulation of host health and immunity. Recent investigations have suggested a relationship between the composition of the human microbiota and COVID-19 infection, highlighting a possible role of bacterial communities in the modulation of the disease severity. In this study, we performed a shotgun metagenomics analysis to explore and compare the nasopharyngeal microbiota of 38 hospitalized Italian patients with and without COVID-19 infection during the third and fourth pandemic waves. In detail, the metagenomic analysis combined with specific correlation analyses suggested a positive association of several microbial species, such as S. parasanguinis and P. melaninogenica, with the severity of COVID-19 infection. Furthermore, the comparison of the microbiota composition between the nasopharyngeal and their respective fecal samples highlighted an association between these different compartments represented by a sharing of several bacterial species. Additionally, lipidomic and deep-shotgun functional analyses of the fecal samples suggested a metabolic impact of the microbiome on the host’s immune response, indicating the presence of key metabolic compounds in COVID-19 patients, such as lipid oxidation end products, potentially related to the inflammatory state. Conversely, the patients without COVID-19 displayed enzymatic patterns associated with the biosynthesis and degradation of specific compounds like lysine (synthesis) and phenylalanine (degradation) that could positively impact disease severity and contribute to modulating COVID-19 infection. IMPORTANCE The human microbiota is reported to play a major role in the regulation of host health and immunity, suggesting a possible impact on the severity of COVID-19 disease. This preliminary study investigated the possible correlation between nasopharyngeal microbiota and COVID-19 infection. In detail, the analysis of the nasopharyngeal microbiota of hospitalized Italian patients with and without COVID-19 infection suggested a positive association of several microbial species with the severity of the disease and highlighted a sharing of several bacteria species with the respective fecal samples. Moreover, the metabolic analyses suggested a possible impact of the microbiome on the host's immune response and the disease severity.

Selective ligands of the CB2 receptor are receiving considerable attention due to their potential as therapeutic agents for a variety of diseases. Recently, 7-hydroxy-5-oxopyrazolo[4,3-b]pyridine-6-carboxamide derivatives were shown to act at the CB2 receptor either as agonists or as inverse agonists/antagonists in vitro and to have anti-osteoarthritic activity in vivo. In this article, we report the synthesis, pharmacological profile, and molecular modeling of a series of twenty-three new 7-hydroxy-5-oxopyrazolo[4,3-b]pyridine-6-carboxamides with the aim of further developing this new class of selective CB2 ligands. In addition to these compounds, seven other analogs that had been previously synthesized were included in this study to better define the structure–activity relationship (SAR). Ten of the new compounds studied were found to be potent and selective ligands of the CB2 receptor, with Ki values ranging from 48.46 to 0.45 nM and CB1/CB2 selectivity indices (SI) ranging from >206 to >4739. In particular, compounds 54 and 55 were found to be high-affinity CB2 inverse agonists that were not active at all at the CB1 receptor, whereas 57 acted as an agonist. The functional activity profile of the compounds within this structural class depends mainly on the substitution pattern of the pyrazole ring.

N-acylethanolamine acid amidase (NAAA) is an N-terminal nucleophile (Ntn) hydrolase that catalyses the intracellular deactivation of the endogenous analgesic and anti-inflammatory agent palmitoylethanolamide (PEA). NAAA inhibitors counteract this process and exert marked therapeutic effects in animal models of pain, inflammation and neurodegeneration. While it is known that NAAA preferentially hydrolyses saturated fatty acid ethanolamides (FAEs), a detailed profile of the relationship between catalytic efficiency and fatty acid-chain length is still lacking. In this report, we combined enzymatic and molecular modelling approaches to determine the effects of acyl chain and polar head modifications on substrate recognition and hydrolysis by NAAA. The results show that, in both saturated and monounsaturated FAEs, the catalytic efficiency is strictly dependent upon fatty acyl chain length, whereas there is a wider tolerance for modifications of the polar heads. This relationship reflects the relative stability of enzyme-substrate complexes in molecular dynamics simulations.

Heparanase is a validated target in cancer therapy and a potential target for several inflammatory pathologies. A ligand-based virtual screening of commercial libraries was performed to expand the chemical space of small-molecule inhibitors. The screening was based on similarity with known inhibitors and was performed in several runs, starting from literature compounds and progressing through newly discovered inhibitors. Among the fifty-five tested compounds, nineteen had IC50 values lower than 5 µM and some showed remarkable potencies. Importantly, tere- and isophthalamides derivatives belong to new structural classes of heparanase inhibitors and some of them showed enzyme affinities (61 and 63, IC50 = 0.32 and 0.12 µM, respectively) similar to those of the most potent small-molecule inhibitors reported so far. Docking studies provided a comprehensive binding hypothesis shared by compounds with significant structural diversity. The most potent inhibitors reduced cell invasiveness and inhibited the expression of proangiogenic factors in tumour cell lines.

The transcriptional regulators YAP (Yes-associated protein) and TAZ (transcriptional co-activator with PDZ-binding motif) are the major downstream effectors in the Hippo pathway and are involved in cancer progression through modulation of the activity of TEAD (transcriptional enhanced associate domain) transcription factors. To exploit the advantages of drug repurposing in the search of new drugs, we developed a similar approach for the identification of new hits interfering with TEAD target gene expression. In our study, a 27-member in-house library was assembled, characterized, and screened for its cancer cell growth inhibition effect. In a secondary luciferase-based assay, only seven compounds confirmed their specific involvement in TEAD activity. IA5 bearing a p-quinoid structure reduced the cytoplasmic level of phosphorylated YAP and the YAP–TEAD complex transcriptional activity and reduced cancer cell growth. IA5 is a promising hit compound for TEAD activity modulator development.

UniPR129, an L-β-homotryptophan conjugate of the secondary bile acid lithocholic acid (LCA), acts as an effective protein-protein interaction (PPI) inhibitor of the Eph–ephrin system but suffers from a poor oral bioavailability in mice. To improve UniPR129 bioavailability, a metabolic soft spot, i.e., the 3α-hydroxyl group on the LCA steroidal ring, was functionalized to 3-hydroxyimine. In vitro metabolism of UniPR129 and 3-hydroxyimine derivative UniPR500 was compared in mouse liver subcellular fractions, and main metabolites were profiled by high resolution (HR-MS) and tandem (MS/MS) mass spectrometry. In mouse liver microsomes (MLM), UniPR129 was converted into several metabolites: M1 derived from the oxidation of the 3-hydroxy group to 3-oxo, M2–M7, mono-hydroxylated metabolites, M8–M10, di-hydroxylated metabolites, and M11, a mono-hydroxylated metabolite of M1. Phase II reactions were only minor routes of in vitro biotransformation. UniPR500 shared several metabolic pathways with parent UniPR129, but it showed higher stability in MLM, with a half-life (t1/2) of 60.4 min, if compared to a t1/2 = 16.8 min for UniPR129. When orally administered to mice at the same dose, UniPR500 showed an increased systemic exposure, maintaining an in vitro valuable pharmacological profile as an EphA2 receptor antagonist and an overall improvement in its physico-chemical profile (solubility, lipophilicity), if compared to UniPR129. The present work highlights an effective strategy for the pharmacokinetic optimization of aminoacid conjugates of bile acids as small molecule Eph–ephrin antagonists.

Fibroblast Growth Factor Receptors (FGFR1–4) have a critical role in the progression of several human cancers, including Squamous Non-Small-Cell Lung Cancer (SQCLC). Both non-selective and selective reversible FGFR inhibitors are under clinical investigation for the treatment of patients with tumors harboring FGFR alterations. Despite their potential efficacy, the clinical development of these drugs has encountered several challenges, including toxicity, and the appearance of drug resistance. Recent efforts have been directed at development of irreversible FGFR inhibitors, which have the potential to exert superior anti-proliferative activity in tumors carrying FGFR alterations. With this in mind, we synthetized, and investigated a set of novel inhibitors possessing a warhead potentially able to covalently bind a cysteine in the P-loop of FGFR. Among them, the chloroacetamide UPR1376 resulted able to irreversible inhibit FGFR1 phosphorylation in FGFR1 over-expressing cells generated from SQCLC SKMES-1 cells. In addition, this compound inhibited cell proliferation in FGFR1-amplified H1581 cells with a potency higher than the reversible inhibitor BGJ398 (infigratinib), while sparing FGFR1 low-expressing cells. The anti-proliferative effects of UPR1376 were demonstrated in both 2D and 3D systems and were associated with the inhibition of MAPK and AKT/mTOR signaling pathways. UPR1376 inhibited cell proliferation also in two BGJ398-resistant cell clones generated from H1581 by chronic exposure to BGJ398, although at concentrations higher than those effective in the parental cells, likely due to the persistent activation of the MAPK pathway associated to NRAS amplification. Combined blockade of FGFR1 and MAPK signaling, by UPR1376 and trametinib respectively, significantly enhanced the efficacy of UPR1376, providing a means of circumventing resistance to FGFR1 inhibition. Our findings suggest that the insertion of a chloroacetamide warhead on a suitable scaffold, as exemplified by UPR1376, is a valuable strategy to develop a novel generation of FGFR inhibitors for the treatment of SQCLC patients with FGFR alterations.

N-anilinoethylamides are a class of melatoninergic agents with the aniline portion mimicking the indole ring of the natural ligand and the ethylamide chain reproducing that of melatonin. The simplest compound in this class, N-{2-[(3-methoxyphenyl)methylamino]ethyl}acetamide (UCM793), has nanomolar binding affinity for MT1 and MT2 membrane receptors. To explore the effect of chain conformation on receptor binding, a methyl group was inserted on the methylene alpha or beta to the amide nitrogen and conformational equilibria were investigated by NMR spectroscopy and molecular dynamics simulations. Receptor affinity was conserved only for the beta-methyl derivative, which also showed significant stereoselectivity, with the (S) enantiomer being the eutomer. Molecular dynamics simulations, validated by NMR spectroscopy, showed that the beta-methyl group affects the conformational preferences of the ethylamide chain. Docking into the receptor crystal structure provides a rationale for the observed chiral recognition, suggesting that the (S)-beta-methyl group favors the conformation that better fits the receptor binding site.

The EphA2 receptor and its ephrin-A1 ligand form a key cell communication system, which has been found overexpressed in many cancer types and involved in tumor growth. Recent medicinal chemistry efforts have identified bile acid derivatives as low micromolar binders of the EphA2 receptor. However, these compounds suffer from poor physicochemical properties, hampering their use in vivo. The identification of compounds able to disrupt the EphA2-ephrin-A1 complex lacking the bile acid scaffold may lead to new pharmacological tools suitable for in vivo studies. To identify the most promising virtual screening (VS) protocol aimed at finding novel EphA2 antagonists, we investigated the ability of both ligand-based and structure-based approaches to retrieve known EphA2 antagonists from libraries of decoys with similar molecular properties. While ligand-based VSs were conducted using UniPR129 and ephrin-A1 ligand as reference structures, structure-based VSs were performed with Glide, using the X-ray structure of the EphA2 receptor/ephrin-A1 complex. A comparison of enrichment factors showed that ligand-based approaches outperformed the structure-based ones, suggesting ligand-based methods using the G-H loop of ephrin-A1 ligand as template as the most promising protocols to search for novel EphA2 antagonists.

The Eph–ephrin system plays a critical role in tumor growth and vascular functions during carcinogenesis. We had previously identified cholanic acid as a competitive and reversible EphA2 antagonist able to disrupt EphA2-ephrinA1 interaction and to inhibit EphA2 activation in prostate cancer cells. Herein, we report the synthesis and biological evaluation of a set of cholanic acid derivatives obtained by conjugation of its carboxyl group with a panel of naturally occurring amino acids with the aim to improve EphA2 receptor inhibition. Structure-activity relationships indicate that conjugation of cholanic acid with linear amino acids of small size leads to effective EphA2 antagonists whereas the introduction of aromatic amino acids reduces the potency in displacement studies. The b-alanine derivative 4 was able to disrupt EphA2-ephrinA1 interaction in the micromolar range and to dose-dependently inhibit EphA2 activation on PC3 cells. These findings may help the design of novel EphA2 antagonists active on cancer cell lines.

Overcoming the emergence of acquired resistance to clinically approved epidermal growth factor receptor (EGFR) inhibitors is a major challenge in the treatment of advanced non–small cell lung cancer (NSCLC). The aim of this study was to investigate the effects of a series of novel compounds affecting viability of NSCLC NCI-H1975 cells (carrying the EGFR T790M mutation). The inhibition of the autophosphorylation of EGFR occurred at nanomolar concentrations and both UPR1282 and UPR1268 caused a significant induction of apoptosis. Targeting of EGFR and downstream pathways was confirmed by a peptide substrate array, which highlighted the inhibition of other kinases involved in NSCLC cell aggressive behavior. Accordingly, the drugs inhibited migration (about 30% vs. control), which could be, in part, explained also by the increase of E-cadherin expression. Additionally, we observed a contraction of the volume of H1975 spheroids, associated with the reduction of the cancer stem–like cell hallmark CD133. The activity of UPR1282 was retained in H1975 xenograft models where it determined tumor shrinkage (P < .05) and resulted well tolerated compared to canertinib. Of note, the kinase activity profile of UPR1282 on xenograft tumor tissues showed overlapping results with respect to the activity in H1975 cells, unraveling the inhibition of kinases involved in pivotal proliferation and invasive signaling pathways. In conclusion, UPR1282 and UPR1268 are effective against various processes involved in malignancy transformation and progression and may be promising compounds for the future treatment of gefitinib-resistant NSCLCs.

Palmitoylethanolamide (PEA) has antinflammatory and antinociceptive properties widely exploited in veterinary and human medicine, despite its poor pharmacokinetics. Looking for prodrugs that could progressively release PEA to maintain effective plasma concentrations, we prepared carbonates, esters and carbamates at the hydroxyl group of PEA. Chemical stability (pH 7.4) and stability in rat plasma and liver homogenate were evaluated by in vitro assays. Carbonates and carbamates resulted too labile and too resistant in plasma, respectively. Ester derivatives, prepared by conjugating PEA with various amino acids, allowed to modulate the kinetics of PEA release in plasma and stability in liver homogenate. L-Val-PEA, with suitable PEA release in plasma, and D-Val-PEA, with high resistance to hepatic degradation, were orally administered to rats and plasma levels of prodrugs and PEA were measured at different time points. Both prodrugs showed significant release of PEA, but provided lower plasma concentrations than those obtained with equimolar doses of PEA. Amino-acid esters of PEA are a promising class to develop prodrugs, even if they need further chemical optimization.

The N-terminal nucleophile (Ntn) hydrolases are a superfamily of enzymes specialized in the hydrolytic cleavage of amide bonds. Even though several members of this family are emerging as innovative drug targets for cancer, inflammation, and pain, the processes through which they catalyze amide hydrolysis remains poorly understood. In particular, the catalytic reactions of cysteine Ntn-hydrolases have never been investigated from a mechanistic point of view. In the present study, we used free energy simulations in the quantum mechanics/molecular mechanics framework to determine the reaction mechanism of amide hydrolysis catalyzed by the prototypical cysteine Ntn-hydrolase, conjugated bile acid hydrolase (CBAH). The computational analyses, which were confirmed in water and using different CBAH mutants, revealed the existence of a chair-like transition state, which might be one of the specific features of the catalytic cycle of Ntn-hydrolases. Our results offer new insights on Ntn-mediated hydrolysis and suggest possible strategies for the creation of therapeutically useful inhibitors.

Abstract A genomic database of all Earth’s eukaryotic species could contribute to many scientific discoveries; however, only a tiny fraction of species have genomic information available. In 2018, scientists across the world united under the Earth BioGenome Project (EBP), aiming to produce a database of high-quality reference genomes containing all ~1.5 million recognized eukaryotic species. As the European node of the EBP, the European Reference Genome Atlas (ERGA) sought to implement a new decentralised, equitable and inclusive model for producing reference genomes. For this, ERGA launched a Pilot Project establishing the first distributed reference genome production infrastructure and testing it on 98 eukaryotic species from 33 European countries. Here we outline the infrastructure and explore its effectiveness for scaling high-quality reference genome production, whilst considering equity and inclusion. The outcomes and lessons learned provide a solid foundation for ERGA while offering key learnings to other transnational, national genomic resource projects and the EBP.

The diverse physiography of the Portuguese land and marine territory, spanning from continental Europe to the Atlantic archipelagos, has made it an important repository of biodiversity throughout the Pleistocene glacial cycles, leading to a remarkable diversity of species and ecosystems. This rich biodiversity is under threat from anthropogenic drivers, such as climate change, invasive species, land use changes, overexploitation, or pathogen (re)emergence. The inventory, characterisation, and study of biodiversity at inter- and intra-specific levels using genomics is crucial to promote its preservation and recovery by informing biodiversity conservation policies, management measures, and research. The participation of researchers from Portuguese institutions in the European Reference Genome Atlas (ERGA) initiative and its pilot effort to generate reference genomes for European biodiversity has reinforced the establishment of Biogenome Portugal. This nascent institutional network will connect the national community of researchers in genomics. Here, we describe the Portuguese contribution to ERGA’s pilot effort, which will generate high-quality reference genomes of six species from Portugal that are endemic, iconic, and/or endangered and include plants, insects, and vertebrates (fish, birds, and mammals) from mainland Portugal or the Azores islands. In addition, we outline the objectives of Biogenome Portugal, which aims to (i) promote scientific collaboration, (ii) contribute to advanced training, (iii) stimulate the participation of institutions and researchers based in Portugal in international biodiversity genomics initiatives, and (iv) contribute to the transfer of knowledge to stakeholders and engaging the public to preserve biodiversity. This initiative will strengthen biodiversity genomics research in Portugal and fuel the genomic inventory of Portuguese eukaryotic species. Such efforts will be critical to the conservation of the country’s rich biodiversity and will contribute to ERGA’s goal of generating reference genomes for European species.

Background: The incidence of Legionnaires’ disease (LD) is increasing steadily in Europe. Its early diagnosis by general practitioners (GPs) is crucial for better patient outcomes. Study objectives: This study assessed Belgian GPs’ knowledge about LD and the accessibility of diagnostic tests in their practices. Methods: A specifically designed questionnaire was distributed to actively practicing GPs, including primary care trainees, between 31 January 2022 and 13 March 2022. This survey targeted approximately 4200 GPs with an estimated population catchment of 30% of the actively working Belgian GPs. Results: The response rate was estimated at 3%. Over 70% of the GPs correctly identified the LD occurrence peak, major risk factors, and clinical manifestations. While 62% of participants preferred the Legionella pneumophila urinary antigen test (UAT) as a primary diagnostic method, 75% were unsure about its availability within their laboratories and 82% had not prescribed it in the last year. Finally, 76% expressed a desire for additional information on this topic. Conclusions: Belgian GPs should evaluate the possibility of conducting UAT testing in their laboratories to enhance LD case management and improve their preparedness. Furthermore, initiatives should be implemented to improve communication between specialists and GPs and develop educational programs directed at Belgian GPs.

Bone loss is a major burden for society and impacts people’s health all over the world. In a changing world looking toward a more conscious use of raw materials, efforts are being made to increasingly consider new promising biomaterials that account for, on one side, the ability to provide specific functional biological activities and, on the other, the feature of being well tolerated. In this regard, the use of phenolic compounds in the field of bone-related bioengineering shows a rising interest in the development of medical solutions aimed at taking advantage of the multiple beneficial properties of these plant molecules. In this work, the anti-bacterial and anti-inflammatory power of a biphasic calcium phosphate synthetic bone filler coated with a mixture of phenolic compounds was investigated by evaluating the minimal inhibitory concentration (MIC) value against Streptococcus mutans and Porphyromonas gingivalis and the expression of genes involved in inflammation and autophagy by real-time reverse transcription polymerase chain reaction (RT-qPCR) on J774a.1 murine macrophage cells. Results show a MIC of 0.8 μg/mL, a neat anti-inflammatory effect, and induction of autophagy key genes compared to a ceramic bone filler. In conclusion, functionalization with a polyphenol-rich extract confers to a ceramic bone filler anti-bacterial and anti-inflammatory properties.

The Ralstonia species (RB) and Burkholderia species (BB) are bacteria responsible for nosocomial infections in frail patients such as hemodialyzed (HD) patients. Here, we report how we managed an outbreak caused by RB and BB that occurred in a dialysis unit. From the 7th to the 16th of April 2021, an infection due to RB and BB occurred in 7 out of 39 (17.9%) HD patients with central venous catheter (CVC). Disinfectants, CVC-lock therapy solutions, water by reverse osmosis unit (ROW) and dialysis concentrates were cultured, including the biofilm from the loading plastic tubes (LPTs) that connect the hemodialysis consoles (HCs) to the ROW delivery line. The antibiotic treatment was successful for all patients. RB and BB were isolated in the biofilm of 11/37 LPTs. Three out of 11 positive LPTs were associated with the infected patients. The ROW delivery line was modified to provide a whole disinfection with the HCs connected, avoiding the risk of new contamination of the LPTs. A filtration module of 0.01 mm was added prior to the ROW delivery line. Our experience suggests that outbreaks sustained by unusual bacteria such as RB and BB should be promptly investigated to treat the infected patients with the appropriate therapy and to identify the possible source of infection, making the needful changes to achieve a safer dialysis unit.

Abstract Randomization-based neural networks have gained wide acceptance in the scientific community owing to the simplicity of their algorithm and generalization capabilities. Random vector functional link (RVFL) networks and their variants are a class of randomization-based neural networks. RVFL networks have shown promising results in classification, regression, and clustering problems. For real-world applications, learning algorithms that can train with new samples over previous results are necessary because of to the constant generation of problems related to large-scale datasets. Various online sequential algorithms, commonly involving an initial learning phase followed by a sequential learning phase, have been proposed to address this issue. This paper presents a training algorithm based on multiple online sequential random vector functional link (OS-RVFL) networks for large-scale databases using a shared memory architecture. The training dataset is distributed among p OS-RVFL networks, which are trained in parallel using p threads. Subsequently, the test dataset samples are classified using each trained OS-RVFL network. Finally, a frequency criterion is applied to the results obtained from each OS-RVFL network to determine the final classification. Additionally, an equation was derived to reasonably predict the total training time of the proposed algorithm based on the learning time in the initial phase and the time scaling factor compared to the sequential learning phase. The results demonstrate a drastic reduction in training time because of data distribution and an improvement in accuracy because of the adoption of the frequency criterion.

Due to their similar boiling points, separation of benzene and cyclohexane mixtures is among the current challenging processes faced by the petrochemical industry. As recently assessed, the soft imine‐based covalent organic framework [(TAM)(BDA)2] (COF‐300; TAM = tetrakis(4‐aminophenyl)methane, BDA = terephthaldehyde) possesses higher affinity for benzene than cyclohexane in both static conditions at 298 K and dynamic conditions in the range of 298–348 K. As shown in this contribution, in situ powder X‐ray diffraction while dosing benzene and cyclohexane vapors in the range of 0.01–4.74 bar on the narrow‐pore form of COF‐300 confirmed the coherent switchability of its framework, unveiling the progressive formation of different intermediate‐ and large‐pore forms. In addition, a basket of otherwise inaccessible key crystallochemical details—“on/off” structural‐feature changes cooperating to adsorption, primary adsorption sites, and host–guest and guest–guest interactions—was successfully retrieved. Overall, these findings allowed to shed light on the framework dynamics underneath the previously observed selectivity toward benzene over cyclohexane, completing this case of study and providing relevant information for the design of new‐generation adsorbents for this applicative context.

The emergence of novel ecosystems, characterized by shifts in species composition and interactions, abiotic conditions and altered ecosystem functioning, is a major and inevitable challenge to contemporary ecosystem management. This study examines the ecological implications of Trachycarpus fortunei, a non-native evergreen palm, currently involved in a biome shift from deciduous temperate forest to evergreen laurophyllous forest in the Southern European Alps. Specifically, we investigated the effects of T. fortunei encroachment in peri-urban forests on flying and ground-dwelling arthropod communities as well as on leaf-litter decomposition. We found that the presence of T. fortunei altered arthropod community composition, mostly by reducing the number of herbivore species. This effect was likely driven by the lower quality of palm leaves as a food source compared to deciduous, dicotyledonous tree leaves and by a lower plant richness in the herb and shrub layer. Furthermore, we observed higher rates of leaf-litter decomposition associated with increasing abundance of young palm trees, which was not explained by predictors commonly associated with litter decomposition (i.e., detritivore abundance, litter depth and air temperature). Hence, the slow decay of palm leaves appears to be counterbalanced by the favourable conditions for litter decomposition within dense palm stands. Overall, our findings indicate that high densities of the non-native palm species (T. fortunei) impacts herbivore arthropod communities in the Southern European Alps, providing first evidence of possible effects on ecosystem functioning of this ongoing biome shift. These outcomes are integral components of the broader process of laurophyllisation in the study region, a phenomenon linked to climate warming and land use change, encompassing both native and non-native species.

IntroductionSystemic sclerosis (SSc) is a complex autoimmune connective tissue disease characterized by microvascular damage, immune system reactivity and progressive fibrosis of skin and internal organs. Interstitial lung disease is the leading cause of death for SSc patients (SSc-ILD), and the process of lung fibrosis involves also circulating monocytes and alveolar macrophages.MethodsCurrent study aimed to identify monocyte/macrophage phenotypes in lung and peripheral blood of SSc-ILD patients by immunostaining and flow cytometry, respectively. Single immunostaining was performed using primary antibodies against CD68 (pan-macrophage marker), CD80, CD86, TLR4 (M1 markers), CD163, CD204, and CD206 (M2 markers). Flow cytometry analysis included the evaluation of CD45, CD14, CD16 (monocyte lineage), CD1c (dendritic lineage), together with M1 and M2 activation markers on circulating monocytes. Protein synthesis of TLR4 and M2 markers was also investigated in cultured monocytes-derived macrophages (MDMs) from SSc-ILD patients by Western Blotting.ResultsLung samples were obtained from 9 SSc-ILD patients (50 ± 9 years old) and 5 control non-SSc patients without lung fibrosis (58 ± 23 years old). Alveolar macrophages (CD68+ cells) showed a significantly higher positivity of M1 and M2 markers in SSc-ILD lung samples than in controls (p

The case law of the European Court of Justice (CJEU), despite confirming in theory the importance of res judicata, has regularly affirmed the need to overcome stability of national judges’ opinions formally qualified as final if that is needed to ensure effective consumers protection. Against this scenario, the Italian Supreme Court (Cass., sez. un. 6 April 2023, n. 9479), seems to have initiated a dialogue with the CJEU, mainly aimed at giving actuation of the principles enacted by the European Court but nevertheless with some apparent revindications of autonomy. The variety of solutions adopted by national judges and the creative effort of the Italian Supreme Court seem to confirm that an intervention by European or national legislators could be required at this point on the matter.

Autoimmune encephalitis (AE) is an immune-mediated condition that induces brain inflammation due to several neural-specific autoantibodies. The main triggering and predisposing factors are infections, genetics, the use of immune checkpoint inhibitors and tumors. We report a case of a 57-year-old male with a biopsy-confirmed Langerhans cell histiocytosis (LCH) and a concomitant anti-LGI1 encephalitis discussing a possible relationship in the pathogenesis of these phenomena. Only sporadic cases of AE developing in the context of histiocytic neoplasms have been described and there are no other reports on the relationship between LGI1-AE and LCH worldwide due to small number of cases.

Abstract Detection of high glycated hemoglobin (A1c) is associated with worse postoperative outcomes, including predisposition to develop systemic and local infectious events. Diabetes and infectious Outcomes in Cardiac Surgery (DOCS) study is a retrospective case–control study aimed to assess in DM and non-DM cardiac surgery patients if a new screening and management model, consisting of systematic A1c evaluation followed by a specialized DM consult, could reduce perioperative infections and 30-days mortality. Effective July 2021, all patients admitted to the cardiac surgery of IRCCS ISMETT were tested for A1c. According to the new protocol, glucose values of patients with A1c ≥ 6% or with known diabetes were monitored. The diabetes team was activated to manage therapy daily until discharge or provide indications for the diagnostic-therapeutic process. Propensity score was used to match 573 patients managed according to the new protocol (the Screen+ Group) to 573 patients admitted before July 2021 and subjected to the traditional management (Screen−). Perioperative prevalence of infections from any cause, including surgical wound infections (SWI), was significantly lower in the Screen+ as compared with the Screen− matched patients (66 [11%] vs. 103 [18%] p = 0.003). No significant difference was observed in 30-day mortality. A1c analysis identified undiagnosed DM in 12% of patients without known metabolic conditions. In a population of patients undergoing cardiac surgery, systematic A1c evaluation at admission followed by specialist DM management reduces perioperative infectious complications, including SWI. Furthermore, A1c screening for patients undergoing cardiac surgery unmasks unknown DM and enhances risk stratification.

Abstract Blue-green infrastructure (BGI), combining semi-natural and engineered elements, offers multifaceted benefits like stormwater management, water purification, heat mitigation, and habitat provision. However, current BGI designs prioritize engineering goals, overlooking its ecological potential. Here we advocate for integrating engineering and ecological objectives into BGI design to enhance performance and biodiversity. Through an interdisciplinary literature review, we emphasize the importance of species diversity, abundance, and ecological processes, to improve engineering performance and resilience, and lower management costs. We emphasize the importance of interdisciplinary collaboration to navigate trade-offs between engineering and ecological objectives, ultimately enabling us to engineer both for and with biodiversity.

Abstract Globe-LFMC 2.0, an updated version of Globe-LFMC, is a comprehensive dataset of over 280,000 Live Fuel Moisture Content (LFMC) measurements. These measurements were gathered through field campaigns conducted in 15 countries spanning 47 years. In contrast to its prior version, Globe-LFMC 2.0 incorporates over 120,000 additional data entries, introduces more than 800 new sampling sites, and comprises LFMC values obtained from samples collected until the calendar year 2023. Each entry within the dataset provides essential information, including date, geographical coordinates, plant species, functional type, and, where available, topographical details. Moreover, the dataset encompasses insights into the sampling and weighing procedures, as well as information about land cover type and meteorological conditions at the time and location of each sampling event. Globe-LFMC 2.0 can facilitate advanced LFMC research, supporting studies on wildfire behaviour, physiological traits, ecological dynamics, and land surface modelling, whether remote sensing-based or otherwise. This dataset represents a valuable resource for researchers exploring the diverse LFMC aspects, contributing to the broader field of environmental and ecological research.

This work explores the feasibility of adopting an LSI produced C/SiC composite to build an aeronautical Auxiliary Power Unit system muffler. The study first covers the experimental material characterization through tensile and compressive tests. The material properties are characterized also after exposing the samples to an oxidizing atmosphere, that is typical for the proposed application. The material response is characterized by significant non-linearities and a pseudo plastic response, which were numerically modeled using a Drucker-Prager model. The detailed design of the muffler is described and verified, for different loading conditions, using a Finite Element model. Finally, a full-scale prototype is produced and assembled, thus proving the technological feasibility of the design. The manufacturing phase required to study and understand the phenomena that were leading to defects in the proposed closed axial symmetric shape, and to implement suitable technological solutions in order to get an acceptable prototype.

Abstract Decades of research on multifunctional Green Infrastructure (GI) has yet to translate into holistic implementation in the built environment. This oversight stems from assumptions that many ecosystem services occur passively and thus potential synergies are overlooked during planning and design. This study offers specific guidance for coordinating GI planning, design, and construction by examining the current state of academic literature on these aspects. It identifies 15 GI elements (e.g., green roofs) and 15 objectives (e.g., biodiversity) to collectively consider before implementation. The literature tends to isolate discussions of “engineered” GI elements with water-related objectives, while more “natural” GI are linked to biodiversity and human well-being. Coordinating across GI objectives and elements remains imperative, but evaluating too many options risks a paradox of choice. This study recommends short-term adherence to principles of adaptive design and, in the long-term, reemphasizes multifunctionality assessments, inter and transdisciplinary collaboration, and political will.

Abstract Background While cutaneous melanomas are well-documented, primary melanoma of the lung (PMML), particularly with endobronchial origin, remains rare and poorly characterized. This case report addresses gaps in understanding by presenting a comprehensive case of a 71-year-old male with primary endobronchial melanoma and conducting a systematic review of PMML cases. Case Presentation The patient, a former smoker, presented with dyspnea, cough, and hemoptysis. Imaging revealed left lung atelectasis and a suspicious nodule. Bronchoscopy identified an endobronchial mass, subsequently treated with argon plasma coagulation and resection. Biopsy confirmed melanoma. Extensive examinations ruled out a primary skin lesion. Despite initial treatment, recurrence led to pneumonectomy. Histopathology confirmed melanoma. The patient received treatment with pembrolizumab and ipilimumab, but with poor clinical benefit. Conclusions Primary endobronchial melanoma is a rare entity, comprising 0.01% of lung tumors. This case underscores diagnostic challenges and emphasizes histological criteria to distinguish primary from metastatic lesions. The pathogenesis remains unclear, with theories proposing foetal melanocyte migration or squamous metaplasia. Prognosis varies, necessitating radical surgical extirpation. A systematic review revealed diverse outcomes, supporting the need for further research. In conclusion, endobronchial melanoma involves an endoscopic and surgical management, but evolving therapies, such as immunotherapy, may reshape treatment paradigms. This case contributes to our understanding of PMML, guiding future research and clinical management. As therapeutic options evolve, continued research is crucial to refine our understanding and improve outcomes for this rare malignancy.

Abstract Background Abiotic stresses in plants include all the environmental conditions that significantly reduce yields, like drought and heat. One of the most significant effects they exert at the cellular level is the accumulation of reactive oxygen species, which cause extensive damage. Plants possess two mechanisms to counter these molecules, i.e. detoxifying enzymes and non-enzymatic antioxidants, which include many classes of specialized metabolites. Sunflower, the fourth global oilseed, is considered moderately drought resistant. Abiotic stress tolerance in this crop has been studied using many approaches, but the control of specialized metabolites in this context remains poorly understood. Here, we performed the first genome-wide association study using abiotic stress-related specialized metabolites as molecular phenotypes in sunflower. After analyzing leaf specialized metabolites of 450 hybrids using liquid chromatography-mass spectrometry, we selected a subset of these compounds based on their association with previously known abiotic stress-related quantitative trait loci. Eventually, we characterized these molecules and their associated genes. Results We putatively annotated 30 compounds which co-localized with abiotic stress-related quantitative trait loci and which were associated to seven most likely candidate genes. A large proportion of these compounds were potential antioxidants, which was in agreement with the role of specialized metabolites in abiotic stresses. The seven associated most likely candidate genes, instead, mainly belonged to cytochromes P450 and glycosyltransferases, two large superfamilies which catalyze greatly diverse reactions and create a wide variety of chemical modifications. This was consistent with the high plasticity of specialized metabolism in plants. Conclusions This is the first characterization of the genetic control of abiotic stress-related specialized metabolites in sunflower. By providing hints concerning the importance of antioxidant molecules in this biological context, and by highlighting some of the potential molecular mechanisms underlying their biosynthesis, it could pave the way for novel applications in breeding. Although further analyses will be required to better understand this topic, studying how antioxidants contribute to the tolerance to abiotic stresses in sunflower appears as a promising area of research.

Abstract Understanding the mechanisms that drive TDP-43 pathology is integral to combating amyotrophic lateral sclerosis (ALS), frontotemporal lobar degeneration (FTLD) and other neurodegenerative diseases. Here we generated a longitudinal quantitative proteomic map of the cortex from the cytoplasmic TDP-43 rNLS8 mouse model of ALS and FTLD, and developed a complementary open-access webtool, TDP-map ( https://shiny.rcc.uq.edu.au/TDP-map/ ). We identified distinct protein subsets enriched for diverse biological pathways with temporal alterations in protein abundance, including increases in protein folding factors prior to disease onset. This included increased levels of DnaJ homolog subfamily B member 5, DNAJB5, which also co-localized with TDP-43 pathology in diseased human motor cortex. DNAJB5 over-expression decreased TDP-43 aggregation in cell and cortical neuron cultures, and knockout of Dnajb5 exacerbated motor impairments caused by AAV-mediated cytoplasmic TDP-43 expression in mice. Together, these findings reveal molecular mechanisms at distinct stages of ALS and FTLD progression and suggest that protein folding factors could be protective in neurodegenerative diseases.

Summary: The amnion is a thin layer of fetal origin in contact with the amniotic fluid which plays a key role at the feto-maternal interface during pregnancy. Here, we present a protocol for isolation of human and Rhesus macaque amnion cells. We describe steps for tissue dissection, cell isolation for flow cytometry analysis, and RNA isolation for RNA sequencing library preparation and analysis. This protocol can provide insights into altered immunological pathways during intrauterine infections to develop new therapeutic strategies.For complete details on the use and execution of this protocol, please refer to Presicce et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Agrivoltaics is presented as a possible solution to the need for new sources of renewable energies, also responding to the increasing demand for feed/food and energy in a strongly efficient and sustainable manner. To this aim, agrivoltaics proposes to combine agricultural and renewable energy production on the same land using photovoltaic technology. The performance of this new production model strongly depends on the interaction between the two systems, agricultural and photovoltaic. In that sense, one of the most important aspects to consider are the effects of the shadows of the photovoltaic panels on the crop land. Overall, the experiment clearly indicated that a fourth cycle of escarole is possible under the PVs of agrivoltaics. Both fresh weight and size of the salad bowls were significantly increased by the shade provided by the PVs. Escarole appeared to be very tolerant to the shade and commercial yields were boosted, compared to full sun treatments, even under extended shade conditions. Such an effect can be likely explained by an overall amelioration of the water status in shaded plots. Therefore, a further study of the behavior of escarole under agrivoltaic conditions will be desirable.