Factors Associated with Prospective Acceptability and Preferences for Unified Transdiagnostic Cognitive-Behavioral Treatments and Group Therapy in the Portuguese General Population.

Group transdiagnostic cognitive-behavioral therapy (CBT) offers a promising solution for limited mental health access in Portugal. Understanding barriers to patient adherence is crucial for successful implementation. This study aimed to characterize the prospective acceptability and preferences for unified transdiagnostic CBT and group therapy in the Portuguese general population and explore their correlates. A sample of 243 participants (18-88 years old), recruited online, completed an online survey collecting information on sociodemographic and clinical characteristics, acceptability of transdiagnostic CBT treatments, specifically of Unified Protocol (UP), acceptability of group therapy, therapeutic format preferences, beliefs about group therapy and help-seeking attitudes. Most participants were receptive to and perceived as useful both unified transdiagnostic CBT and group therapy. Overall, participants presented significantly more favorable attitudes than unfavorable attitudes toward unified transdiagnostic CBT and group therapy (p < .001). Multivariate analyses revealed that (1) favorable attitudes toward transdiagnostic treatments were negatively associated with being employed and positively associated with living in an urban area, and higher efficacy scores; (2) unfavorable attitudes toward transdiagnostic treatments were positively associated with being married/cohabitating and negatively associated with vulnerability scores; (3) being female, living in an urban area, and higher efficacy and myth scores emerged as positive predictors of favorable attitudes toward group therapy; and (4) efficacy and vulnerability scores and help-seeking propensity emerged as negative predictors of unfavorable attitudes toward group therapy. These findings highlight the importance of delineating strategies to increase knowledge and acceptance of unified transdiagnostic CBT and group therapy in the Portuguese population, addressing specific individual characteristics.

Analysis of Live Single Cells by Confocal Microscopy and High-Resolution Mass Spectrometry to Study Drug Uptake, Metabolism, and Drug-Induced Phospholipidosis.

The analysis of large numbers of cells from a population results in information that does not reflect differences in cell phenotypes. Individual variations in cellular drug uptake, metabolism, and response to drug treatment may have profound effects on cellular survival and lead to the development of certain disease states, drug persistence, and resistance. Herein, we present a method that combines live cell confocal microscopy imaging with high-resolution mass spectrometry to achieve absolute cell quantification of the drug amiodarone (AMIO) and its major metabolite, N-desethylamiodarone (NDEA), in single liver cells (HepG2 and HepaRG cells). The method uses a prototype system that integrates a confocal microscope with an XYZ stage robot to image and automatically sample selected cells from a sample compartment, which is kept under growth conditions, with nanospray tips. Besides obtaining the distributions of AMIO and NDEA cell concentrations across a population of individual cells, as well as variabilities in drug metabolism, the effect of these on phospholipidosis and cell morphology was studied. The method was suited to identify subpopulations of cells that metabolized less drug and to correlate cell drug concentrations with cell phospholipid content, cell volume, sphericity, and other cell phenotypic features. Using principal component analysis (PCA), the treated cells could be clearly distinguished from vehicle control cells (0 µM AMIO) and HepaRG cells from HepG2 cells. The potential of using multidimensional and multimodal information collected from single cells to build predictive models for cell classification is demonstrated.

Selection of the Activated Carbon Type for the Treatment of Landfill Leachate by Fenton-Adsorption Process.

Sanitary landfill leachates usually have characteristics that depend on the region where they are generated and according to the age of the landfill, which is why a unique treatment for their sanitation has not been found. However, the adsorption preceded by the Fenton process has been proven to be highly efficient at removing contaminants. In this study, the adsorptive capacity of two types of activated carbon, granular and powdered, was analyzed to determine which was more efficient in the adsorption stage in the Fenton-adsorption process. Likewise, its behavior was analyzed using three isotherm models (Langmuir, Freundlich and Temkin), testing the raw leachate and the Fenton-treated one with both carbons. The adsorption that is carried out on the carbons is better adjusted to the Freundlich and Temkin models. It concludes that multilayers, through the physical adsorption, carry out the adsorption of pollutants on the surface of the carbons. The results show that, statistically, granular activated carbon is more efficient at removing chemical oxygen demand (COD), and powdered activated carbon removes color better. Finally, an adsorption column was designed for the Fenton-adsorption process that was able to remove 21.68 kgCOD/kg carbon. Removal efficiencies for color and COD were >99%.

Development of an HPLC-based guanosine monophosphate kinase assay and application to Plasmodium vivax guanylate kinase.

The development of a high-performance liquid chromatography (HPLC)-based method, for guanosine monophosphate kinase activity assays, is presented. The method uses the intrinsic UV absorption (at 260 nm) of substrates and products of the enzymatic reaction (GMP, ATP, ADP and GDP) to unambiguously determine percent conversion of substrate into product. It uses a commercially available C18 column which can separate reaction samples by elution under isocratic conditions in 12 min per run. The kinetics of the forward reaction catalyzed by Plasmodium vivax guanylate kinase (PvGK), a potential drug target against malaria, was determined. The relative concentrations of the two substrates (GMP and ATP) have a distinct effect on reaction velocity. Kinetic analyses showed the PvGK-catalyzed reaction to be associated with atypical kinetics, where substrate inhibition kinetics and non-Michaelis-Menten (sigmoidal) kinetics were found with respect to GMP and ATP, respectively. Additionally, the method was used in inhibition assays to screen twenty fragment-like compounds. The assays were robust and reproducible, with a signal window of 3.8 and a Z' factor of 0.6. For the best inhibitor, an IC50 curve was generated.

Self-criticism, negative automatic thoughts and postpartum depressive symptoms: the buffering effect of self-compassion.

Objective: This study aimed to explore the direct and indirect effects of self-criticism on postpartum depressive symptoms, through postpartum cognitions, and analyse the moderating role of self-compassion in this relationship. Background: Self-criticism and self-compassion are associated with postpartum depression. However, further research is needed to understand how these mechanisms operate in the development/maintenance of depressive symptoms. Methods: 686 women in the postpartum period (up to 12 months after birth) recruited in-person and online answered a cross-sectional survey. Results: The effect of self-criticism on postpartum depressive symptoms occurred sequentially, increasing the frequency of negative automatic thoughts and subsequently the metacognitive appraisal of these thoughts. Self-compassion had a moderating effect only on the relation between self-criticism and postpartum cognitions. The effect of self-criticism on postpartum cognitions decreased in the presence of higher self-compassion. Conclusion: This study emphasises the negative effect of a self-critical thinking style and of a negative appraisal of thought's content on depression symptoms. Moreover, this work underscores the buffering role of self-compassion in the relationship between self-criticism and postpartum cognitions. These results highlight the need to address the reduction of self-criticism and the promotion of self-compassion strategies to deal with postpartum cognitions, in order to prevent and treat postpartum depressive symptoms.

Native Mass Spectrometry in Fragment-Based Drug Discovery.

The advent of native mass spectrometry (MS) in 1990 led to the development of new mass spectrometry instrumentation and methodologies for the analysis of noncovalent protein-ligand complexes. Native MS has matured to become a fast, simple, highly sensitive and automatable technique with well-established utility for fragment-based drug discovery (FBDD). Native MS has the capability to directly detect weak ligand binding to proteins, to determine stoichiometry, relative or absolute binding affinities and specificities. Native MS can be used to delineate ligand-binding sites, to elucidate mechanisms of cooperativity and to study the thermodynamics of binding. This review highlights key attributes of native MS for FBDD campaigns.

Dietary Silicon Deficiency Does Not Exacerbate Diet-Induced Fatty Lesions in Female ApoE Knockout Mice.

BACKGROUND: Dietary silicon has been positively linked with vascular health and protection against atherosclerotic plaque formation, but the mechanism of action is unclear. OBJECTIVES: We investigated the effect of dietary silicon on 1) serum and aorta silicon concentrations, 2) the development of aortic lesions and serum lipid concentrations, and 3) the structural and biomechanic properties of the aorta. METHODS: Two studies, of the same design, were conducted to address the above objectives. Female mice, lacking the apolipoprotein E (apoE) gene, and therefore susceptible to atherosclerosis, were separated into 3 groups of 10-15 mice, each exposed to a high-fat diet (21% wt milk fat and 1.5% wt cholesterol) but with differing concentrations of dietary silicon, namely: silicon-deprived (-Si; <3-µg silicon/g feed), silicon-replete in feed (+Si-feed; 100-µg silicon/g feed), and silicon-replete in drinking water (+Si-water; 115-µg silicon/mL) for 15-19 wk. Silicon supplementation was in the form of sodium metasilicate (feed) or monomethylsilanetriol (drinking water). RESULTS: The serum silicon concentration in the -Si group was significantly lower than in the +Si-feed (by up to 78%; P < 0.003) and the +Si-water (by up to 84%; P < 0.006) groups. The aorta silicon concentration was also lower in the -Si group than in the +Si-feed group (by 65%; P = 0.025), but not compared with the +Si-water group. There were no differences in serum and aorta silicon concentrations between the silicon-replete groups. Body weights, tissue wet weights at necropsy, and structural, biomechanic, and morphologic properties of the aorta were not affected by dietary silicon; nor were the development of fatty lesions and serum lipid concentrations. CONCLUSIONS: These findings suggest that dietary silicon has no effect on atherosclerosis development and vascular health in the apoE mouse model of diet-induced atherosclerosis, contrary to the reported findings in the cholesterol-fed rabbit model.

Treatment patterns and preferences of people living with HIV starting or switching antiretroviral therapy: Real-world evidence from Portugal.

BACKGROUND: There is a lack of up-to-date real-life evidence on antiretroviral therapy (ART) strategies among people living with HIV (PLWH) in Portugal. This study aimed to describe the treatment strategy used in PLWH either initiating or switching ART. METHODS: Non-interventional, cross-sectional, multicenter study carried out between December 2019 and October 2021 in Portugal. RESULTS: A total of 237 PLWH were included in this study, 171 of whom were ART-experienced and 66 were ART-naïve. The study showed that triple regimens were the most common ART strategy and integrase strand transfer inhibitors-based therapy was the most frequently used therapeutic class in both ART-naïve and ART-experienced PLWH. Nevertheless, about a third of PLWH who started a triple regimen transitioned to a dual regimen. Patient-reported outcomes revealed high HIV literacy and similar ART preferences in both groups. CONCLUSIONS: This real-world study showed that triple regimens were the most widely used ART strategy, even after the European AIDS Clinical Society guidelines introduced the recommendation of a dual regimen for naïve patients. The cohorts of this study presented a high level of HIV literacy at the time of inclusion. Our findings highlighted that taking pills only once a day is considered a very important feature for most patients.

Determinants of HIV late presentation among men who have sex with men in Portugal (2014-2019): who's being left behind?

Introduction: HIV late presentation (LP) remains excessive in Europe. We aimed to analyze the factors associated with late presentation in the MSM population newly diagnosed with HIV in Portugal between 2014 and 2019. Methods: We included 391 newly HIV-1 diagnosed Men who have Sex with Men (MSM), from the BESTHOPE project, in 17 countrywide Portuguese hospitals. The data included clinical and socio-behavioral questionnaires and the viral genomic sequence obtained in the drug resistance test before starting antiretrovirals (ARVs). HIV-1 subtypes and epidemiological surveillance mutations were determined using different bioinformatics tools. Logistic regression was used to estimate the association between predictor variables and late presentation (LP). Results: The median age was 31 years, 51% had a current income between 501-1,000 euros, 28% were migrants. 21% had never been tested for HIV before diagnosis, with 42.3% of MSM presenting LP. 60% were infected with subtype B strains. In the multivariate regression, increased age at diagnosis, higher income, lower frequency of screening, STI ever diagnosed and higher viral load were associated with LP. Conclusion: Our study suggests that specific subgroups of the MSM population, such older MSM, with higher income and lower HIV testing frequency, are not being targeted by community and clinical screening services. Overall, targeted public health measures should be strengthened toward these subgroups, through strengthened primary care testing, expanded access to PrEP, information and promotion of HIV self-testing and more inclusive and accessible health services.

Regeneration of granular activated carbon clogged in the treatment of leachates.

Landfill leachates are highly contaminated liquids and complex to treat. Two of the processes which are promising for the treatment are the advanced oxidation and adsorption methods. With the combination of the Fenton and adsorption methods, practically all the organic load of leachates can be removed; however, this combination of processes is limited due to the soon clogging of adsorbent material, which leads to high operation costs. In the present work, the results of the regeneration of clogged activated carbon are shown after the application of the Fenton/adsorption process in leachates. This research consisted of four stages: sampling and leachate characterization, clogging of the carbon through the Fenton/adsorption process, carbon regeneration through the oxidative Fenton process, and lastly, evaluation of regenerated carbon adsorption through jar and column tests. In the experiments, HCl 3 M was used, and different concentration of hydrogen peroxide (0.15 M, 0.2 M, 0.25 M) were tested at different times (16 h and 30 h). The activated carbon regeneration through the Fenton process and the optimal peroxide dosage was 0.15 M for 16 h. The regeneration efficiency was obtained from comparing the adsorption efficiency between regenerated and virgin carbon, reaching 98.27% and can be applied up to 4 times without losing regeneration efficiency. These results prove that it is possible to restore the clogged activated carbon adsorption capacity during the Fenton/adsorption process.

Antibacterial benzofuran neolignans and benzophenanthridine alkaloids from the roots of Zanthoxylum capense.

Two new 2-arylbenzofuran neolignans and a new benzophenanthridine alkaloid, together with six known benzophenanthridine alkaloids, namely, decarine, norchelerythrine, dihydrochelerythrine, 6-acetonyldihydrochelerythrine, tridecanonchelerythrine, and 6-acetonyldihydronitidine, have been isolated from the MeOH extract of the roots of Zanthoxylum capense. Their structures were elucidated by means of spectroscopic techniques including 2D NMR experiments. All the isolated compounds were evaluated for their in vitro antibacterial activity against gram-positive and gram-negative bacteria. Some compounds showed significant inhibitory activity against Staphylococcus aureus ATCC 6538 with MIC values ranging from 12.5 to 50 µg/mL.

HIV-1-Transmitted Drug Resistance and Transmission Clusters in Newly Diagnosed Patients in Portugal Between 2014 and 2019.

Objective: To describe and analyze transmitted drug resistance (TDR) between 2014 and 2019 in newly infected patients with HIV-1 in Portugal and to characterize its transmission networks. Methods: Clinical, socioepidemiological, and risk behavior data were collected from 820 newly diagnosed patients in Portugal between September 2014 and December 2019. The sequences obtained from drug resistance testing were used for subtyping, TDR determination, and transmission cluster (TC) analyses. Results: In Portugal, the overall prevalence of TDR between 2014 and 2019 was 11.0%. TDR presented a decreasing trend from 16.7% in 2014 to 9.2% in 2016 (p for-trend = 0.114). Multivariate analysis indicated that TDR was significantly associated with transmission route (MSM presented a lower probability of presenting TDR when compared to heterosexual contact) and with subtype (subtype C presented significantly more TDR when compared to subtype B). TC analysis corroborated that the heterosexual risk group presented a higher proportion of TDR in TCs when compared to MSMs. Among subtype A1, TDR reached 16.6% in heterosexuals, followed by 14.2% in patients infected with subtype B and 9.4% in patients infected with subtype G. Conclusion: Our molecular epidemiology approach indicates that the HIV-1 epidemic in Portugal is changing among risk group populations, with heterosexuals showing increasing levels of HIV-1 transmission and TDR. Prevention measures for this subpopulation should be reinforced.

Development of a high-throughput AlphaScreen assay measuring full-length LRRK2(G2019S) kinase activity using moesin protein substrate.

Mutations within the LRRK2 (leucine-rich repeat kinase 2) gene predispose humans to develop late-onset Parkinson's disease (PD). The most prevalent of these mutations, G2019S, has been shown to increase LRRK2 kinase activity. Therefore, the discovery of small molecule inhibitors of LRRK2(G2019S) through high-throughput screening (HTS) may provide a novel therapeutic strategy for treating PD. Current biochemical assays monitoring the activity of LRRK2(G2019S) either are radioactive or use short peptidic substrates. Here we describe the development and optimization of a novel HTS AlphaScreen assay for measuring the catalytic activity of full-length LRRK2(G2019S) using its putative physiological protein substrate moesin. The high sensitivity of this optimized 384-well assay allowed the use of enzyme concentrations as low as 0.75nM. The estimated apparent K(m) value for adenosine triphosphate (6 microM) using the glutathione S-transferase-moesin substrate was much lower than the one previously reported using LRRKtide, a synthetic peptide derived from moesin. Testing of nonselective kinase inhibitors (staurosporine, H-1152, and Y-27632) generated potencies consistent with published data. Finally, robotic validation of the assay yielded an average Z' factor of 0.80. Overall, these results indicate that the present HTS AlphaScreen assay might provide a more relevant biochemical approach for the discovery of novel LRRK2(G2019S) inhibitors.

Oligodendrocytes Provide Antioxidant Defense Function for Neurons by Secreting Ferritin Heavy Chain.

An evolutionarily conserved function of glia is to provide metabolic and structural support for neurons. To identify molecules generated by glia and with vital functions for neurons, we used Drosophila melanogaster as a screening tool, and subsequently translated the findings to mice. We found that a cargo receptor operating in the secretory pathway of glia was essential to maintain axonal integrity by regulating iron buffering. Ferritin heavy chain was identified as the critical secretory cargo, required for the protection against iron-mediated ferroptotic axonal damage. In mice, ferritin heavy chain is highly expressed by oligodendrocytes and secreted by employing an unconventional secretion pathway involving extracellular vesicles. Disrupting the release of extracellular vesicles or the expression of ferritin heavy chain in oligodendrocytes causes neuronal loss and oxidative damage in mice. Our data point to a role of oligodendrocytes in providing an antioxidant defense system to support neurons against iron-mediated cytotoxicity.

Neuropilin-1 facilitates SARS-CoV-2 cell entry and infectivity.

The causative agent of coronavirus disease 2019 (COVID-19) is the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). For many viruses, tissue tropism is determined by the availability of virus receptors and entry cofactors on the surface of host cells. In this study, we found that neuropilin-1 (NRP1), known to bind furin-cleaved substrates, significantly potentiates SARS-CoV-2 infectivity, an effect blocked by a monoclonal blocking antibody against NRP1. A SARS-CoV-2 mutant with an altered furin cleavage site did not depend on NRP1 for infectivity. Pathological analysis of olfactory epithelium obtained from human COVID-19 autopsies revealed that SARS-CoV-2 infected NRP1-positive cells facing the nasal cavity. Our data provide insight into SARS-CoV-2 cell infectivity and define a potential target for antiviral intervention.

Enabling drug discovery and development through single-cell imaging.

INTRODUCTION: Single-cell imaging-based assays are an area of active and growing investment in drug discovery and development. This approach offers researchers the capability to interrogate rare subpopulations of cells with minimal sample consumption and multiplexed readouts. Recent technological advances in the optical interrogation and manipulation of single cells have substantially increased the throughput and sensitivity of these assays. Areas covered: In this review, the authors focus on three classes of single-cell imaging-based analyses: single-cell microscopy combined with microfluidics, mass spectrometric imaging for subcellular compound localization, and imaging mass cytometry (IMC). They provide an overview of each technology and recent examples of their utility in advancing drug discovery, based on the potential for scalability, multiplexing, and capability to generate definitive data on cellular heterogeneity and target engagement. Expert opinion: Understanding target engagement and heterogeneity at the single-cell level will enable the development of safer and more effective therapies, particularly for new modalities like CAR-T cell therapies and gene editing approaches (AAV, CRISPR). Successful adoption of new single-cell imaging-based approaches in drug discovery will require tandem investment in advanced computational analysis and bioinformatic approaches, due to the complexity and multivariate nature of single-cell imaging data.

Programmable DNA switches and their applications.

DNA switches are ideally suited for numerous nanotechnological applications, and increasing efforts are being directed toward their engineering. In this review, we discuss how to engineer these switches starting from the selection of a specific DNA-based recognition element, to its adaptation and optimisation into a switch, with applications ranging from sensing to drug delivery, smart materials, molecular transporters, logic gates and others. We provide many examples showcasing their high programmability and recent advances towards their real life applications. We conclude with a short perspective on this exciting emerging field.

Fragment-Based Screening of a Natural Product Library against 62 Potential Malaria Drug Targets Employing Native Mass Spectrometry.

Natural products are well known for their biological relevance, high degree of three-dimensionality, and access to areas of largely unexplored chemical space. To shape our understanding of the interaction between natural products and protein targets in the postgenomic era, we have used native mass spectrometry to investigate 62 potential protein targets for malaria using a natural-product-based fragment library. We reveal here 96 low-molecular-weight natural products identified as binding partners of 32 of the putative malarial targets. Seventy-nine (79) fragments have direct growth inhibition on Plasmodium falciparum at concentrations that are promising for the development of fragment hits against these protein targets. This adds a fragment library to the published HTS active libraries in the public domain.

Novel Mechanism for Buffering Dietary Salt in Humans: Effects of Salt Loading on Skin Sodium, Vascular Endothelial Growth Factor C, and Blood Pressure.

High dietary sodium intake triggers increased blood pressure (BP). Animal studies show that dietary salt loading results in dermal Na+ accumulation and lymphangiogenesis mediated by VEGF-C (vascular endothelial growth factor C), both attenuating the rise in BP. Our objective was to determine whether these mechanisms function in humans. We assessed skin electrolytes, BP, and plasma VEGF-C in 48 healthy participants randomized to placebo (70 mmol sodium/d) and slow sodium (200 mmol/d) for 7 days. Skin Na+ and K+ concentrations were measured in mg/g of wet tissue and expressed as the ratio Na+:K+ to correct for variability in sample hydration. Skin Na+:K+ increased between placebo and slow sodium phases (2.91±0.08 versus 3.12±0.09; P=0.01). In post hoc analysis, there was a suggestion of a sex-specific effect, with a significant increase in skin Na+:K+ in men (2.59±0.09 versus 2.88±0.12; P=0.008) but not women (3.23±0.10 versus 3.36±0.12; P=0.31). Women showed a significant increase in 24-hour mean BP with salt loading (93±1 versus 91±1 mm Hg; P<0.001) while men did not (96±2 versus 96±2 mm Hg; P=0.91). Skin Na+:K+ correlated with BP, stroke volume, and peripheral vascular resistance in men but not in women. No change was noted in plasma VEGF-C. These findings suggest that the skin may buffer dietary Na+, reducing the hemodynamic consequences of increased salt, and this may be influenced by sex.

Optimization of Electrospray Ionization by Statistical Design of Experiments and Response Surface Methodology: Protein-Ligand Equilibrium Dissociation Constant Determinations.

Electrospray ionization mass spectrometry (ESI-MS) binding studies between proteins and ligands under native conditions require that instrumental ESI source conditions are optimized if relative solution-phase equilibrium concentrations between the protein-ligand complex and free protein are to be retained. Instrumental ESI source conditions that simultaneously maximize the relative ionization efficiency of the protein-ligand complex over free protein and minimize the protein-ligand complex dissociation during the ESI process and the transfer from atmospheric pressure to vacuum are generally specific for each protein-ligand system and should be established when an accurate equilibrium dissociation constant (KD) is to be determined via titration. In this paper, a straightforward and systematic approach for ESI source optimization is presented. The method uses statistical design of experiments (DOE) in conjunction with response surface methodology (RSM) and is demonstrated for the complexes between Plasmodium vivax guanylate kinase (PvGK) and two ligands: 5'-guanosine monophosphate (GMP) and 5'-guanosine diphosphate (GDP). It was verified that even though the ligands are structurally similar, the most appropriate ESI conditions for KD determination by titration are different for each. Graphical Abstract ᅟ.