ION Thallos-HTL: a fluorescent thallium indicator that enables cell-selective and localizable thallium flux assays.

Ion channels are essential proteins for all organisms. Electrophysiology is a useful and commonly employed method to study ion channels, however there is a need for operationally simpler, cost-effective and higher throughput techniques to study ion channel functions in their native environments. Fluorescent ion indicators, such as Fluo-4 and Thallos, have been used for decades to study ion channel activity by measuring the flux of ions through channels of interest. In this work, we present ION Thallos-HTL, a thallium indicator that can be localized using HaloTag technology. This novel indicator enables specific labeling of cells and intracellular compartments in live cells and responds to changes in thallium concentration within these environments. We demonstrate the utility of ION Thallos-HTL by conducting a thallium flux assay using high-throughput instrumentation in a mixed cell population where some cells are expressing HaloTag and some are not.

Proteometabolomics of initial and recurrent glioblastoma highlights an increased immune cell signature with altered lipid metabolism.

BACKGROUND: There is an urgent need to better understand the mechanisms associated with the development, progression, and onset of recurrence after initial surgery in glioblastoma (GBM). The use of integrative phenotype-focused -omics technologies such as proteomics and lipidomics provides an unbiased approach to explore the molecular evolution of the tumor and its associated environment. METHODS: We assembled a cohort of patient-matched initial (iGBM) and recurrent (rGBM) specimens of resected GBM. Proteome and metabolome composition were determined by mass spectrometry-based techniques. We performed neutrophil-GBM cell coculture experiments to evaluate the behavior of rGBM-enriched proteins in the tumor microenvironment. ELISA-based quantitation of candidate proteins was performed to test the association of their plasma concentrations in iGBM with the onset of recurrence. RESULTS: Proteomic profiles reflect increased immune cell infiltration and extracellular matrix reorganization in rGBM. ASAH1, SYMN, and GPNMB were highly enriched proteins in rGBM. Lipidomics indicates the downregulation of ceramides in rGBM. Cell analyses suggest a role for ASAH1 in neutrophils and its localization in extracellular traps. Plasma concentrations of ASAH1 and SYNM show an association with time to recurrence. CONCLUSIONS: We describe the potential importance of ASAH1 in tumor progression and development of rGBM via metabolic rearrangement and showcase the feedback from the tumor microenvironment to plasma proteome profiles. We report the potential of ASAH1 and SYNM as plasma markers of rGBM progression. The published datasets can be considered as a resource for further functional and biomarker studies involving additional -omics technologies.

Review of an international pilot project to adapt the Canadian Adult Obesity Clinical Practice Guideline.

Background: The evidence-based Canadian Adult Obesity Clinical Practice Guideline (CPG) released in August 2020 were developed through a systematic literature review and patient-oriented research process. This CPG is considered a paradigm shift for obesity care as it introduced a new obesity definition that is based on health not body size, incorporates lived experiences of people affected by obesity, and addresses the pervasive weight bias and stigma that patients face in healthcare systems. The purpose of this pilot project was to assess the feasibility of adapting the Canadian CPG in Chile and Ireland. Methods: An International Clinical Practice Guideline Adaptation Committee was established to oversee the project. The project was conducted through four interrelated phases: 1) planning and preparation; 2) pilot project application process; 3) adaptation; and 4) launch, dissemination, and implementation. Ireland used the GRADE-ADAPTE framework and Chile used the GRADE-ADOLOPMENT approach. Results: Chile and Ireland developed their adapted guidelines in one third of the time it took to develop the Canadian guidelines. In Ireland, 18 chapters, which underpin the 80 key recommendations, were contextually adapted. Chile adopted 18 chapters and 76 recommendations, adapted one recommendation, and developed 12 new recommendations.. Conclusion: The pilot project demonstrated it is feasible to adapt the Canadian CPG for use in other countries with different healthcare systems, languages, and cultural contexts, while retaining the Canadian CPG's key principles and values such as the treatment of obesity as a chronic disease, adoption of new clinical assessment approaches that go beyond anthropometric measurements, elimination of weight bias and stigma, shifting obesity care outcomes to improved health and well-being rather than weight loss alone, and the use of patient-centred, collaborative and shared-decision clinical care approaches.

Systemic toxicity of snake venom metalloproteinases: Multi-omics analyses of kidney and blood plasma disturbances in a mouse model.

Snakebite envenomation is classified as a Neglected Tropical Disease. Bothrops jararaca venom induces kidney injury and coagulopathy. HF3, a hemorrhagic metalloproteinase of B. jararaca venom, participates in the envenomation pathogenesis. We evaluated the effects of HF3 in mouse kidney and blood plasma after injection in the thigh muscle, mimicking a snakebite. Transcriptomic analysis showed differential expression of 31 and 137 genes related to kidney pathology after 2 h and 6 h, respectively. However, only subtle changes were observed in kidney proteome, with differential abundance of 15 proteins after 6 h, including kidney injury markers. N-terminomic analysis of kidney proteins showed 420 proteinase-generated peptides compatible with meprin specificity, indicating activation of host proteinases. Plasma analysis revealed differential abundance of 90 and 219 proteins, respectively, after 2 h and 6 h, including coagulation-cascade and complement-system components, and creatine-kinase, whereas a semi-specific search of N-terminal peptides indicated activation of endogenous proteinases. HF3 promoted host reactions, altering the gene expression and the proteolytic profile of kidney tissue, and inducing plasma proteome imbalance driven by changes in abundance and proteolysis. The overall response of the mouse underscores the systemic action of a hemorrhagic toxin that transcends local tissue damage and is related to known venom-induced systemic effects.

SNAP-tagging live cells via chelation-assisted copper-catalyzed azide-alkyne cycloaddition.

SNAP-tag is a single-turnover enzyme that has become a powerful tool, hence a popular choice, of targeted cellular protein labeling. Three SNAP-tag substrates that carry the copper-chelating 2-picolyl azide moiety are prepared, one of which has an unconventional 5-pyridylmethyl-substituted guanine structure, rather than the usual benzylguanine that is optimized to be accepted by SNAP-tag. All three substrates are effective in transferring a 2-picolyl azide moiety to SNAP-tag in live cells under conventional labeling conditions (30-minute incubation of cells with labeling reagents at 37 °C under 5% CO2). Live cells that are decorated with chelating azido groups on the extracellular side of membranes undergo copper-catalyzed azide-alkyne cycloaddition (CuAAC) with an ethynyl-functionalized fluorophore to accomplish membrane protein labeling by a fluorescent dye. The chelation-assisted CuAAC labeling step is rapid (<1 minute) with a relatively low dose of the copper catalyst (20 µM), and consequently exerts no ill effect on the labeled cells. A SNAP-tag substrate that carries a non-chelating azide moiety, on the other hand, fails to produce satisfactory labeling under the same constraints. The rapid, live cell-compatible SNAP-tag/chelation-assisted CuAAC two-step method expands the utility of SNAP-tag in protein labeling applications.

Orthogonal Versatile Interacting Peptide Tags for Imaging Cellular Proteins.

Genetic tags are transformative tools for investigating the function, localization, and interactions of cellular proteins. Most studies today are reliant on selective labeling of more than one protein to obtain comprehensive information on a protein's behavior in situ. Some proteins can be analyzed by fusion to a protein tag, such as green fluorescent protein, HaloTag, or SNAP-Tag. Other proteins benefit from labeling via small peptide tags, such as the recently reported versatile interacting peptide (VIP) tags. VIP tags enable observations of protein localization and trafficking with bright fluorophores or nanoparticles. Here, we expand the VIP toolkit by presenting two new tags: TinyVIPER and PunyVIPER. These two tags were designed for use with MiniVIPER for labeling up to three distinct proteins at once in cells. Labeling is mediated by the formation of a high-affinity, biocompatible heterodimeric coiled coil. Each tag was validated by fluorescence microscopy, including observation of transferrin receptor 1 trafficking in live cells. We verified that labeling via each tag is highly specific for one- or two-color imaging. Last, the self-sorting tags were used for simultaneous labeling of three protein targets (i.e., TOMM20, histone 2B, and actin) in fixed cells, highlighting their utility for multicolor microscopy. MiniVIPER, TinyVIPER, and PunyVIPER are small and robust peptide tags for selective labeling of cellular proteins.

The serum proteome of VA-ECMO patients changes over time and allows differentiation of survivors and non-survivors: an observational study.

BACKGROUND: Veno-arterial extracorporeal membrane oxygenation (VA-ECMO) is applied in patients with refractory hemodynamic failure. Exposure of blood components to high shear stress and the large extracorporeal surfaces in the ECMO circuit trigger a complex inflammatory response syndrome and coagulopathy which are believed to worsen the already poor prognosis of these patients. Mass spectrometry-based proteomics allow a detailed characterization of the serum proteome as it provides the identity and concentration of large numbers of individual proteins at the same time. In this study, we aimed to characterize the serum proteome of patients receiving VA-ECMO. METHODS: Serum samples were collected on day 1 and day 3 after initiation of VA-ECMO. Samples underwent immunoaffinity based depletion for the 14 most abundant serum proteins, in-solution digestion and PreOmics clean-up. A spectral library was built with multiple measurements of a master-mix sample using variable mass windows. Individual samples were measured in data independent acquisition (DIA) mode. Raw files were analyzed by DIA-neural network. Unique proteins were log transformed and quantile normalized. Differential expression analysis was conducted with the LIMMA-R package. ROAST was applied to generate gene ontology enrichment analyses. RESULTS: Fourteen VA-ECMO patients and six healthy controls were recruited. Seven patients survived. Three hundred and fifty-one unique proteins were identified. One hundred and thirty-seven proteins were differentially expressed between VA-ECMO patients and controls. One hundred and forty-five proteins were differentially expressed on day 3 compared to day 1. Many of the differentially expressed proteins were involved in coagulation and the inflammatory response. The serum proteomes of survivors and non-survivors on day 3 differed from each other according to partial least-squares discriminant analysis (PLS-DA) and 48 proteins were differentially expressed. Many of these proteins have also been ascribed to processes in coagulation and inflammation (e.g., Factor IX, Protein-C, Kallikrein, SERPINA10, SEMA4B, Complement C3, Complement Factor D and MASP-1). CONCLUSION: The serum proteome of VA-ECMO patients displays major changes compared to controls and changes from day 1 until day 3. Many changes in the serum proteome are related to inflammation and coagulation. Survivors and non-survivors can be differentiated according to their serum proteomes using PLS-DA analysis on day 3. Our results build the basis for future studies using mass-spectrometry based serum proteomics as a tool to identify novel prognostic biomarkers. TRIAL REGISTRATION: DRKS00011106.

Proteome alterations in human autopsy tissues in relation to time after death.

Protein expression is a primary area of interest for routine histological diagnostics and tissue-based research projects, but the limitations of its post-mortem applicability remain largely unclear. On the other hand, tissue specimens obtained during autopsies can provide unique insight into advanced disease states, especially in cancer research. Therefore, we aimed to identify the maximum post-mortem interval (PMI) which is still suitable for characterizing protein expression patterns, to explore organ-specific differences in protein degradation, and to investigate whether certain proteins follow specific degradation kinetics. Therefore, the proteome of human tissue samples obtained during routine autopsies of deceased patients with accurate PMI (6, 12, 18, 24, 48, 72, 96 h) and without specific diseases that significantly affect tissue preservation, from lungs, kidneys and livers, was analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). For the kidney and liver, significant protein degradation became apparent at 48 h. For the lung, the proteome composition was rather static for up to 48 h and substantial protein degradation was detected only at 72 h suggesting that degradation kinetics appear to be organ specific. More detailed analyses suggested that proteins with similar post-mortem kinetics are not primarily shared in their biological functions. The overrepresentation of protein families with analogous structural motifs in the kidney indicates that structural features may be a common factor in determining similar postmortem stability. Our study demonstrates that a longer post-mortem period may have a significant impact on proteome composition, but sampling within 24 h may be appropriate, as degradation is within acceptable limits even in organs with faster autolysis.

Targeted and explorative profiling of kallikrein proteases and global proteome biology of pancreatic ductal adenocarcinoma, chronic pancreatitis, and normal pancreas highlights disease-specific proteome remodelling.

Pancreatic ductal adenocarcinoma (PDAC) represents one of the most aggressive and lethal malignancies worldwide with an urgent need for new diagnostic and therapeutic strategies. One major risk factor for PDAC is the pre-indication of chronic pancreatitis (CP), which represents highly inflammatory pancreatic tissue. Kallikreins (KLKs) are secreted serine proteases that play an important role in various cancers as components of the tumor microenvironment. Previous studies of KLKs in solid tumors largely relied on either transcriptomics or immunodetection. We present one of the first targeted mass spectrometry profiling of kallikrein proteases in PDAC, CP, and normal pancreas. We show that KLK6 and KLK10 are significantly upregulated in PDAC (n=14) but not in CP (n=7) when compared to normal pancreas (n=16), highlighting their specific intertwining with malignancy. Additional explorative proteome profiling identified 5936 proteins in our pancreatic cohort and observed disease-specific proteome rearrangements in PDAC and CP. As such, PDAC features an enriched proteome motif for extracellular matrix (ECM) and cell adhesion while there is depletion of mitochondrial energy metabolism proteins, reminiscent of the Warburg effect. Although often regarded as a PDAC hallmark, the ECM fingerprint was also observed in CP, alongside with a prototypical inflammatory proteome motif as well as with an increased wound healing process and proteolytic activity, thereby possibly illustrating tissue autolysis. Proteogenomic analysis based on publicly accessible data sources identified 112 PDAC-specific and 32 CP-specific single amino acid variants, which among others affect KRAS and ANKHD1. Our study emphasizes the diagnostic potential of kallikreins and provides novel insights into proteomic characteristics of PDAC and CP.

Dynamic Delirium Prediction in the Intensive Care Unit using Machine Learning on Electronic Health Records.

Delirium is a syndrome of acute brain failure which is prevalent amongst older adults in the Intensive Care Unit (ICU). Incidence of delirium can significantly worsen prognosis and increase mortality, therefore necessitating its rapid and continual assessment in the ICU. Currently, the common approach for delirium assessment is manual and sporadic. Hence, there exists a critical need for a robust and automated system for predicting delirium in the ICU. In this work, we develop a machine learning (ML) system for real-time prediction of delirium using Electronic Health Record (EHR) data. Unlike prior approaches which provide one delirium prediction label per entire ICU stay, our approach provides predictions every 12 hours. We use the latest 12 hours of ICU data, along with patient demographic and medical history data, to predict delirium risk in the next 12-hour window. This enables delirium risk prediction as soon as 12 hours after ICU admission. We train and test four ML classification algorithms on longitudinal EHR data pertaining to 16,327 ICU stays of 13,395 patients covering a total of 56,297 12-hour windows in the ICU to predict the dynamic incidence of delirium. The best performing algorithm was Categorical Boosting which achieved an area under receiver operating characteristic curve (AUROC) of 0.87 (95% Confidence Interval; C.I, 0.86-0.87). The deployment of this ML system in ICUs can enable early identification of delirium, thereby reducing its deleterious impact on long-term adverse outcomes, such as ICU cost, length of stay and mortality.

Obesity in adults: Clinical practice guideline adapted for Chile. / Obesidad en adultos: guía de práctica clínica adaptada para Chile.

Introduction: The Chilean Society of Bariatric and Metabolic Surgery, together with other scientific societies, led a process for adapting the Canadian clinical practice guideline for obesity in adults for Chile. The aim of the Canadian guideline, among its main objectives, was to propose changes in obesity management using a chronic disease framework and focusing on improving patient-centered health outcomes, rather than focusing on weight loss alone. Methods: A group of 58 healthcare professionals applied the GRADE-Adolopment method to analyze and adapt the original recommendations and to create de novo recommendations. New recommendations were developed through a systematic review of the evidence using the Epistemonikos database and based on the GRADE-Evidence to Decision (EtD) framework. Results: Seventy-six (76) of the 80 original recommendations were adopted, one recommendation was adapted, and 12 new recommendations were created. Conclusions: The adaptation process reduced the time needed to develop a Chilean clinical practice guideline for the management of obesity in adults. The change in obesity management approaches towards non-stigmatizing and patient-centered strategies focused on improving health outcomes and not solely on weight reduction is universal and it is possible to apply this approach in different countries and contexts.

Introducción: La Sociedad Chilena de Cirugía Bariátrica y Metabólica, junto a otras sociedades científicas, lideró el proceso de adaptación de la guía de práctica clínica de obesidad en adultos para Chile, tomando como base las directrices desarrolladas para Canadá. La guía canadiense buscó, entre sus principales objetivos, proponer cambios en el enfoque del manejo de la obesidad como una enfermedad crónica y para mejorar los desenlaces de salud centrados en los pacientes, en lugar de enfocarse en la pérdida de peso como principal y único objetivo. Métodos: Se convocó a un grupo de 58 profesionales para el desarrollo del proyecto, quienes revisaron y utilizaron el método para el análisis de las recomendaciones originales y desarrollo de recomendaciones . Para la elaboración de nuevas recomendaciones, se llevó a cabo una búsqueda de revisiones sistemáticas en la base de datos Epistemonikos, y se utilizó metodología GRADE y el marco para la evaluación de la evidencia y la descripción de la recomendación. Resultados: Se adoptaron 76 de las 80 recomendaciones de la guía canadiense, se adaptó una recomendación y se desarrollaron 12 preguntas nuevas con sus respectivas recomendaciones. Conclusiones: Conclusiones.

Obesidad en adultos: guía de práctica clínica adaptada para Chile / Obesity in adults: Clinical practice guideline adapted for Chile

Introducción La Sociedad Chilena de Cirugía Bariátrica y Metabólica, junto a otras sociedades científicas, lideró el proceso de adaptación de la guía de práctica clínica de obesidad en adultos para Chile, tomando como base las directrices desarrolladas para Canadá. La guía canadiense buscó, entre sus principales objetivos, proponer cambios en el enfoque del manejo de la obesidad como una enfermedad crónica y para mejorar los desenlaces de salud centrados en los pacientes, en lugar de enfocarse en la pérdida de peso como principal y único objetivo. Métodos Se convocó a un grupo de 58 profesionales para el desarrollo del proyecto, quienes revisaron y utilizaron el método para el análisis de las recomendaciones originales y desarrollo de recomendaciones . Para la elaboración de nuevas recomendaciones, se llevó a cabo una búsqueda de revisiones sistemáticas en la base de datos Epistemonikos, y se utilizó metodología GRADE y el marco para la evaluación de la evidencia y la descripción de la recomendación. Resultados Se adoptaron 76 de las 80 recomendaciones de la guía canadiense, se adaptó una recomendación y se desarrollaron 12 preguntas nuevas con sus respectivas recomendaciones. Conclusiones El proceso de adaptación permitió acortar el tiempo necesario para elaborar una guía de práctica clínica en obesidad del adulto para nuestro país. El cambio en el enfoque hacia una aproximación sin estigma y centrada en la salud y no en el peso, es universal y posible de aplicar en diferentes países y contextos.

Introduction The Chilean Society of Bariatric and Metabolic Surgery, together with other scientific societies, led a process for adapting the Canadian clinical practice guideline for obesity in adults for Chile. The aim of the Canadian guideline, among its main objectives, was to propose changes in obesity management using a chronic disease framework and focusing on improving patient-centered health outcomes, rather than focusing on weight loss alone. Methods A group of 58 healthcare professionals applied the GRADE-Adolopment method to analyze and adapt the original recommendations and to create de novo recommendations. New recommendations were developed through a systematic review of the evidence using the Epistemonikos database and based on the GRADE-Evidence to Decision (EtD) framework. Results Seventy-six (76) of the 80 original recommendations were adopted, one recommendation was adapted, and 12 new recommendations were created. Conclusions The adaptation process reduced the time needed to develop a Chilean clinical practice guideline for the management of obesity in adults. The change in obesity management approaches towards non-stigmatizing and patient-centered strategies focused on improving health outcomes and not solely on weight reduction is universal and it is possible to apply this approach in different countries and contexts.

Obesidad en adultos: guía de práctica clínica adaptada para Chile / Obesity in adults: clinical practice guideline adapted for Chile

La Sociedad Chilena de Cirugía Bariátrica y Metabólica, junto a otras sociedades científicas, lideró el proceso de adaptación de la guía de práctica clínica de obesidad en adultos para Chile, tomando como base las directrices desarrolladas para Canadá. La guía canadiense buscó, entre sus principales objetivos, proponer cambios en el enfoque del manejo de la obesidad como una enfermedad crónica y para mejorar los desenlaces de salud centrados en los pacientes, en lugar de enfocarse en la pérdida de peso como principal y único objetivo. Se convocó a un grupo de 58 profesionales para el desarrollo del proyecto, quienes revisaron y utilizaron el método para el análisis de las recomendaciones originales y desarrollo de recomendaciones . Para la elaboración de nuevas recomendaciones, se llevó a cabo una búsqueda de revisiones sistemáticas en la base de datos Epistemonikos, y se utilizó metodología GRADE y el marco para la evaluación de la evidencia y la descripción de la recomendación. Se adoptaron 76 de las 80 recomendaciones de la guía canadiense, se adaptó una recomendación y se desarrollaron 12 preguntas nuevas con sus respectivas recomendaciones. El proceso de adaptación permitió acortar el tiempo necesario para elaborar una guía de práctica clínica en obesidad del adulto para nuestro país. El cambio en el enfoque hacia una aproximación sin estigma y centrada en la salud y no en el peso, es universal y posible de aplicar en diferentes países y contextos.

Krüppel-like factor 7 influences translation and pathways involved in ribosomal biogenesis in breast cancer.

BACKGROUND: Ribosomal biogenesis and ribosomal proteins have attracted attention in the context of tumor biology in recent years. Instead of being mere translational machineries, ribosomes might play an active role in tumor initiation and progression. Despite its importance, regulation of ribosomal biogenesis is still not completely understood. METHODS: Using Gene Set Enrichment Analysis of RNA sequencing and proteomical mass spectrometry data in breast cancer cells expressing Krüppel-like factor 7 (KLF7), we identified processes altered by this transcription factor. In silico analyses of a cohort of breast cancer patients in The Cancer Genome Atlas confirmed our finding. We further verified the role of KLF7 the identified ribosomal processes in in vitro assays of mammary carcinoma cell lines and analyses of breast cancer patients' tissue slices. RESULTS: We identified the transcription factor Krüppel-like factor 7 (KLF7) as a regulator of ribosomal biogenesis and translation in breast cancer cells and tissue. Highly significant overlapping processes related to ribosomal biogenesis were identified in proteomics and transcriptomics data and confirmed in patients' breast cancer RNA Seq data. Further, nucleoli, the sites of ribosomal biogenesis, were morphologically altered and quantitatively increased in KLF7-expressing cells. Pre-rRNA processing was identified as one potential process affected by KLF7. In addition, an increase in global translation independent from proliferation and transcription was observed upon exogenous KLF7 expression in vitro. Importantly, in a cohort of breast cancer patients, KLF7-expression levels correlated with aggressiveness of the intrinsic breast cancer subtype and tumor grading. Moreover, KLF7 correlated with nucleolar characteristics in human breast tumor tissue, indicating a role for KLF7 in ribosomal biogenesis. CONCLUSION: In mammary carcinoma, KLF7 is involved in ribosomal biogenesis. Alterations of ribosomal biogenesis has far reaching quantitative and qualitative implications for the proteome of the cancer cells. This might influence the aggressiveness of cancer cells.

Profiling the serum proteome during Schistosoma mansoni infection in the BALB/c mice: A focus on the altered lipid metabolism as a key modulator of host-parasite interactions.

Schistosomiasis represents a condition in which every aspect of the disease, starting from skin invasion of the cercariae to egg laying by adult worms, incites a tissue response from the vertebrate host. This response, whether acute or chronic, leads to the appearance of reporter molecules of tissue injury in bodily fluids that could be surveyed as markers for disease diagnosis, status and prognosis. In this scenario, the serum proteome associated with a schistosome infection remains poorly explored; particularly by the use of high-throughput mass spectrometric instrumentation. In this study, we aimed to comparatively examine the serum proteome of control versus infected BALB/c mice, spanning the interval between the onset of egg laying and the peak of the acute phase of infection. Compositional analysis of the sera, using one dimensional reversed-phase fractionation of tryptic peptides coupled to mass spectrometry, allowed identification of 453 constituents. Among these, over 30% (143 molecules) were differentially present comparing sera from infected and non-infected mice, as revealed by quantitative label-free shotgun approach. The majority of proteins exhibiting altered levels was categorised as belonging to immune response (acute phase-related proteins) followed by those linked to lipid transport and metabolism. Inspection of the lipid profile from control and infected individuals demonstrated more pronounced and significant alterations in triglycerides, VLDL and HDL fractions (p<0,001), attesting for a disturbance in circulating lipid molecules, and suggesting a key role in host-parasite interactions. Our findings provide a global view of the serum proteome in the context of experimental schistosomiasis during the acute phase of infection. It contributes by listing key molecules that could be monitored to inform on the associated inflammatory disease status. We hope it will shed light into uncovered aspects of the Schistosoma mansoni parasitism in the vertebrate host, particularly those related to modulation of the lipid metabolism mediating immune responses.

Discrete protein metric (DPM): A new image similarity metric to calculate accuracy of deep learning-generated cell focal adhesion predictions.

Understanding cell behaviors can provide new knowledge on the development of different pathologies. Focal adhesion (FA) sites are important sub-cellular structures that are involved in these processes. To better facilitate the study of FA sites, deep learning (DL) can be used to predict FA site morphology based on limited microscopic datasets (e.g., cell membrane images). However, calculating the accuracy score of these predictions can be challenging due to the discrete/point pattern like nature of FA sites. In the present work, a new image similarity metric, discrete protein metric (DPM), was developed to calculate FA prediction accuracy. This metric measures differences in distribution (d), shape/size (s), and angle (a) of FA sites between predicted and ground truth microscopy images. Performance of the DPM was evaluated by comparing it to three other commonly used image similarity metrics: Pearson correlation coefficient (PCC), feature similarity index (FSIM), and Intersection over Union (IoU). A sensitivity analysis was performed by comparing changes in each metric value due to quantifiable changes in FA site location, number, aspect ratio, area, or orientation. Furthermore, accuracy score of DL-generated predictions was calculated using all four metrics to compare their ability to capture variation across samples. Results showed better sensitivity and range of variation for DPM compared to the other metrics tested. Most importantly, DPM had the ability to determine which FA predictions were quantitatively more accurate and consistent with qualitative assessments. The proposed DPM hence provides a method to validate DL-generated FA predictions and has the potential to be used for investigation of other sub-cellular protein aggregates relevant to cell biology.

An adapted passive model of anti-MPO dependent crescentic glomerulonephritis reveals matrix dysregulation and is amenable to modulation by CXCR4 inhibition.

Anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitides (AAV) are severe inflammatory disorders that often involve focal necrotizing glomerulonephritis (FNGN) and consequent glomerular scarring, interstitial fibrosis, and chronic kidney disease. Robust murine models of scarring in FNGN that may help to further our understanding of deleterious processes are still lacking. Here, we present a murine model of severe FNGN based on combined administration of antibodies against the glomerular basement membrane (GBM) and myeloperoxidase (MPO), and bacterial lipopolysaccharides (LPS), that recapitulates acute injury and was adapted to investigate subsequent glomerular and interstitial scarring. Hematuria without involvement of other organs occurs consistently and rapidly, glomerular necrosis and crescent formation are evident at 12 days, and consequent glomerular and interstitial scarring at 29 days after initial treatment. Using mass-spectrometric proteome analysis, we provide a detailed overview of matrisomal and cellular changes in our model. We observed increased expression of the matrisome including collagens, fibronectin, tenascin-C, in accordance with human AAV as deduced from analysis of gene expression microarrays and tissue staining. Moreover, we observed tissue infiltration by neutrophils, macrophages, T cells and myofibroblasts upon injury. Experimental inhibition of CXCR4 using AMD3100 led to a sustained histological presence of fibrin extravasate, reduced chemokine expression and leukocyte activation, but did not markedly affect ECM composition. Altogether, we demonstrate an adapted FNGN model that enables the study of matrisomal changes both in disease and upon intervention, as exemplified via CXCR4 inhibition.

Sex-related differences of early cardiac functional and proteomic alterations in a rat model of myocardial ischemia.

BACKGROUND: Reduced cardiovascular risk in premenopausal women has been the focus of research in recent decades. Previous hypothesis-driven experiments have highlighted the role of sex hormones on distinct inflammatory responses, mitochondrial proteins, extracellular remodeling and estrogen-mediated cardioprotective signaling pathways related to post-ischemic recovery, which were associated with better cardiac functional outcomes in females. We aimed to investigate the early, sex-specific functional and proteomic changes following myocardial ischemia in an unbiased approach. METHODS: Ischemia was induced in male (M-Isch) and female (F-Isch) rats with sc. injection of isoproterenol (85 mg/kg) daily for 2 days, while controls (M-Co, F-Co) received sc. saline solution. At 48 h after the first injection pressure-volume analysis was carried out to assess left ventricular function. FFPE tissue slides were scanned and analyzed digitally, while myocardial proteins were quantified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) using isobaric labeling. Concentrations of circulating steroid hormones were measured with LC-MS/MS. Feature selection (PLS and PLS-DA) was used to examine associations among functional, proteomic and hormonal datasets. RESULTS: Induction of ischemia resulted in 38% vs 17% mortality in M-Isch and F-Isch respectively. The extent of ischemic damage to surviving rats was comparable between the sexes. Systolic dysfunction was more pronounced in males, while females developed a more severe impairment of diastolic function. 2224 proteins were quantified, with 520 showing sex-specific differential regulation. Our analysis identified transcriptional, cytoskeletal, contractile, and mitochondrial proteins, molecular chaperones and the extracellular matrix as sources of disparity between the sexes. Bioinformatics highlighted possible associations of estrogens and their metabolites with early functional and proteomic alterations. CONCLUSIONS: Our study has highlighted sex-specific alterations in systolic and diastolic function shortly after ischemia, and provided a comprehensive look at the underlying proteomic changes and the influence of estrogens and their metabolites. According to our bioinformatic analysis, inflammatory, mitochondrial, chaperone, cytoskeletal, extracellular and matricellular proteins are major sources of intersex disparity, and may be promising targets for early sex-specific pharmacologic interventions.

Creencias y búsqueda de la salud en la sociedad española contemporánea / Beliefs and health pursuit in the present Spanish society

El presente artículo pretende abordar la relación entre creencias religiosas y espirituales y búsqueda de la salud que existe en la sociedad española de los últimos tiempos. Se puede observar cómo las prácticas médicas institucionales coexisten junto al curanderismo y otras manifestaciones de la medicina popular que tienen relación con la religiosidad tradicional y junto a nuevas formas de buscar la salud mediante la espiritualidad New Age, de inspiración principalmente oriental. En la sociedad española actual conviven diversas formas de acercarse a la salud mediante múltiples prácticas culturales, por lo que la medicina oficial y la enfermería comparten pacientes con formas tradicionales y nuevas de creencia que pretenden alcanzar la sanación. (AU)

This article tries to deal with the relationship between the religious and spiritual beliefs and the health pursuit that exist in the Spanish society lately. We can observe how institutional medical practices coexist with quackery and other manifestations of popular medicine which are related to traditional religiosity and whith other ways of searching for health through spirituality New Age which has, mainly, an eastern inspiration. In the present Spanish society coexist different ways of approaching health through multiple cultural practices, that is why official medicine and nursery share patients with traditional and new ways of believing that try to reach healing. (AU)