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Martins, J.

Publications and source records attributed to Martins, J..

14 recordsLinked to original sources

Quantitative live imaging reveals region-dependent medulloblastoma disseminationin the zebrafish brain

Medulloblastoma is the most common malignant pediatric brain tumor, and relapse is frequently driven by dissemination within the central nervous system. Yet the early dynamics of tumor spread remain difficult to resolve in vivo. Here, we orthotopically transplanted human medulloblastoma cells into larval zebrafish and followed individual xenografts by longitudinal high-resolution confocal imaging. ONS-76 showed particularly robust dissemination, with hindbrain tumors spreading into the developing cerebellum in 81.5% of larvae, more than fourfold the frequency observed for forebrain or midbrain tumors. Disseminating cells extended along brain-surface and leptomeningeal-like regions and frequently associated with cerebral vasculature. Quantitative analysis of tumor area, perimeter, compactness, convex hull area, and fractal dimension captured changes in growth and spatial complexity over time. Dasatinib reduced tumor expansion and dissemination-related features in a concentration-dependent manner, while higher doses revealed host toxicity. These findings establish a powerful in vivo model for resolving medulloblastoma dissemination and therapeutic response.

cancer biology↗

Annexin A1 as a key modulator of lung inflammation during coronavirus infections

Exacerbated inflammation is a major contributor to tissue damage and mortality in infectious diseases, including SARS-CoV-2. The resolution phase of inflammation is critical for restoring tissue homeostasis following an injury. Annexin A1 (AnxA1) is a ubiquitous protein that plays a fundamental role in the resolution of inflammation, including in preclinical models of infectious disease. Here, we investigated the role of AnxA1 in coronavirus infection and its potential as a host-targeted therapeutic strategy against SARS-CoV-2. Wild-type (WT) and AnxA1 knockout (AnxA1KO) mice were intranasally infected with the murine betacoronavirus MHV-3 to study the endogenous role of AnxA1. Immunohistochemistry and Western blot analyses in the lungs of MHV-3-infected mice revealed increased AnxA1 expression and its cleavage, which was associated with neutrophilic infiltration (Ly6G+ cells) mainly in peribronchiolar and perivascular regions. AnxA1-deficient mice exhibited higher neutrophilic infiltration and lung damage, alongside increased CXCL1 production in the lungs, when compared to WT-infected mice. In a murine model of SARS-CoV-2 infection in K18-hACE2 mice, we found increased AnxA1 cleavage associated with lung inflammation. Treatment of SARS-CoV-2-infected K18-hACE2 mice with the AnxA1-mimetic peptide, Ac2-26, reduced lung damage and lethality, without altering the host ability to deal with viral replication. Notably, Ac2-26-treated mice exhibited similar levels of protection to that afforded by the nucleotide analogue Remdesivir, following SARS-CoV-2 infection. Our findings highlight the protective role of the endogenous AnxA1 in mitigating coronavirus-induced lung inflammation and underscore the therapeutic potential of AnxA1 mimetic Ac2-26 as a host-targeted therapy against SARS-CoV-2.

immunology↗

Mapping functional traces of opioid memories in the rat brain

Addiction to psychoactive substances is a maladaptive learned behavior. Contexts surrounding drug use integrate this aberrant mnemonic process and hold strong relapse triggering ability. Here we asked where context and salience might be concurrently represented in the brain, and found circuitry hubs specific to morphine-related contextual information storage. Starting with a classical rodent morphine-conditioned place preference (CPP) apparatus, we developed a CPP protocol that allows stimuli presentation inside a magnetic resonance imaging scanner, to allow the investigation of whole brain activity during retrieval of drug-context paired associations, as well as resting state functional connectivity under the effect of morphine conditioning. Using fMRI we found context-specific responses to stimulus onset in multiple brain regions, namely limbic, sensory, and striatal. Furthermore, we found increased functional connectivity of lateral septum with regions within and beyond a proposed limbic network, and of the lateral habenula with hippocampal CA1 region, in response to repeated pairings of drug and context. Subsequent exposure to either morphine or saline-conditioned contexts led to significant, context-specific, functional interconnectivity among amygdala, lateral habenula, and lateral septum. Resting-state connectivity of the lateral habenula and amygdala, and that during saline-paired context presentation significantly predicted inter-individual CPP score differences. In sum, our findings show that drug- and saline-paired contexts form distinct memory traces in overlapping functional brain microcircuits, and intrinsic connectivity of habenula, septum, and amygdala likely influences the maladaptive contextual learning in response to opioids. We identify functional mechanisms involved in the acquisition and retrieval of drug-related memories that might be behind the relapse-triggering ability of opioid-associated sensory/contextual cues. Graphical abstractTo investigate the brain-wide regional activity underlying drug addiction Gomes-Ribeiro et al. have developed a rodent morphine-induced CPP (conditioned place preference) protocol whose contextual cues can be consistently presented inside an MRI apparatus. The authors found a common circuitry supporting the neural mechanisms responsible for memorizing an association between distinct contexts, and morphine or the absence thereof, that includes regions involved in affect, reward, contextual perception and memory, with subtle, intriguing, functional specificities ultimately underlying the storage of distinct, individual, CPP memory engrams across the different animals. Animals that are more prone to strong emotional responses (as measured by the baseline resting state amygdala and habenula functional connectivity), exhibit a kind of neural circuit priming effect, and thus develop stronger connectivity patterns in anticipation of stronger morphine addiction behavior. These findings could help to clarify the inter-individual sensitivity to opioids in humans, since despite responding positively to the effects of opioids, many humans do not develop an addiction. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=138 SRC="FIGDIR/small/552221v2_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@1cd4989org.highwire.dtl.DTLVardef@96c76corg.highwire.dtl.DTLVardef@feadd0org.highwire.dtl.DTLVardef@4547b6_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

Local neuroplasticity in adult glaucomatous visual cortex

The degree to which the adult human visual cortex retains the ability to functionally adapt to damage at the level of the eye remains ill-understood. Previous studies on cortical neuroplasticity primarily focused on the consequences of foveal visual field defects (VFD), yet these findings may not generalize to peripheral defects such as occur in glaucoma. Moreover, recent findings on neuroplasticity are often based on population receptive field (pRF) mapping, but interpreting these results is complicated in the absence of appropriate control conditions. Here, we used fMRI-based neural modeling to assess putative changes in pRFs associated with glaucomatous VFD. We compared the fMRI-signals and pRF estimates in participants with glaucoma to those of controls with case-matched simulated VFD. We found that the amplitude of the fMRI-signal is reduced in glaucoma compared to control participants and correlated with disease severity. Furthermore, while coarse retinotopic structure is maintained in all participants with glaucoma, we observed local pRF shifts and enlargements in early visual areas, relative to control participants. These differences imply that the adult brain retains local neuroplasticity. This finding has translational relevance, as it is consistent with VFD masking, which prevents glaucoma patients from noticing their VFD and seeking timely treatment.

neuroscience↗

SARS-CoV-2 infection of human brain microvascular endothelial cells leads to inflammatory activation through NF-κB non-canonical pathway and mitochondrial remodeling

Neurological effects of COVID-19 and long-COVID-19 as well as neuroinvasion by SARS-CoV-2 still pose several questions and are of both clinical and scientific relevance. We described the cellular and molecular effects of the human brain microvascular endothelial cells (HBMECs) in vitro infection by SARS-CoV-2 to understand the underlying mechanisms of viral transmigration through the Blood-Brain Barrier. Despite the low to non-productive viral replication, SARS-CoV-2-infected cultures displayed increased apoptotic cell death and tight junction protein expression and immunolocalization. Transcriptomic profiling of infected cultures revealed endothelial activation via NF-{kappa}B non-canonical pathway, including RELB overexpression, and mitochondrial dysfunction. Additionally, SARS-CoV-2 led to altered secretion of key angiogenic factors and to significant changes in mitochondrial dynamics, with increased mitofusin-2 expression and increased mitochondrial networks. Endothelial activation and remodeling can further contribute to neuroinflammatory processes and lead to further BBB permeability in COVID-19.

cell biology↗

Micro-aggregation of a pristine grassland soil selects for bacterial and fungal communities and changes in nitrogen cycling potentials

Microbial analysis at the micro scale of soil is essential to the overall understanding of microbial organization and interactions, and necessary for a better understanding of soil ecosystem functioning. While bacterial communities have been extensively described, little is known about the organization of fungal communities as well as functional potentials at scales relevant to microbial interactions. Fungal and bacterial communities and changes in nitrogen cycling potentials in the pristine Rothamsted Park Grass soil (bulk soil) as well as in its particle size sub-fractions (PSFs; > 250 m, 250-63 m, 63-20 m, 20-2 m, < 2 m and supernatant) were studied. The potential for nitrogen reduction was found elevated in bigger aggregates. The relative abundance of Basidiomycota deceased with decreasing particle size, Ascomycota showed an increase and Mucoromycota became more prominent in particles less than 20 m. Bacterial community structures changed below 20 m at the scale where microbes operate. Strikingly, only members of two bacterial and one fungal phyla (Proteobacteria, Bacteroidota and Ascomycota, respectively) were washed-off the soil during fractionation and accumulated in the supernatant fraction where most of the detected bacterial genera (e.g., Pseudomonas, Massilia, Mucilaginibacter, Edaphobaculum, Duganella, Janthinobacterium and Variovorax) were previously associated with exopolysaccharide production and biofilm formation. Overall, the applied method shows potential to study soil microbial communities at micro scales which might be useful in studies focusing on the role of specific fungal taxa in soil structure formation as well as research on how and by whom biofilm-like structures are distributed and organized in soil. ImportanceIntensive exploitation of soils has led to increasing environmental concerns such as pollution, erosion, emission of greenhouse gases and, in general, the weakening of its ecosystem services that are mainly regulated by microbial activity. Microbial activity and metabolism drive the formation of soil aggregates, ranging in size from a few micrometres to several millimetres. Understanding biological mechanisms related to aggregate size classes can provide insight into large-scale processes, but most research has focused on macroaggregates. Here, we investigated the microbial community and its functional changes at these smaller scales that are clearly more relevant for assessing microbial activity. We demonstrated that fungal communities are more sensitive to bigger size classes than bacteria, suggesting their dominant role in soil structure formation and turnover. We also identified preferential niches for reductive processes within the nitrogen cycle and a selection of specific taxa by analysing the water used for the wet-fractionation approach.

ecology↗

Plasmodium falciparum Acetyl-CoA Synthetase is essential for parasite intraerythrocytic development and chromatin modification

The malaria parasite Plasmodium falciparum possesses a unique Acetyl-CoA Synthetase (PfACS) which provides acetyl moieties for different metabolic and regulatory cellular pathways. We characterized PfACS and studied its role focusing on epigenetic modifications using the var gene family as reporter genes. For this, mutant lines to modulate plasmodial ACS expression by degron-mediated protein degradation or ribozyme induced transcript decay were created. Additionally, an ACS inhibitor was tested for its effectiveness and specificity in interfering with PfACS. The knockdown of PfACS or its inhibition led to impaired parasite growth. Decreased levels of PfACS also led to differential histone acetylation patterns, altered variant gene expression and concomitantly decreased cytoadherence of infected red blood cells containing knocked-down parasites. Further, ChIP analysis revealed the presence of PfACS in many loci in ring stage parasites, underscoring its involvement in the regulation of chromatin. Due to its significant differences to human ACS, PfACS seems an interesting target for drug development.

microbiology↗

Effect of Initial pH and Organic Matter Concentration on Production of Volatile Organic Acids Through Acidogenesis of Sugarcane Vinasse

Sugarcane vinasse is an industrial liquid waste generated in great amounts in Brazilian ethanol industries. Nowadays its main use occurs at sugarcane crops, where vinasse is applied as a nutrient source for fertirrigation. However, continued use of vinasse in soil can cause several environmental impacts. So, aiming to provide a more environmentally friendly destination to the effluent, the goal of this work was to investigate the acidogenesis using a synthetic vinasse as substrate, focusing on the effects of initial pH and Chemical Oxygen Demand (COD) on short chain organic acids (SCOAs) concentrations. Synthetic vinasse was prepared at laboratory taking some real sugarcane vinasse composition given in previous works as references. So, major contribution presented here is the investigation on obtaining high added-value SCOAs from a simulated effluent. Cattle manure sludge was utilized as inoculum to promote the conversion of carbohydrate (sucrose, Suc) in synthetic vinasse into SCOAs in batch reactors during a total incubation time of 72 h. Acidogenesis profiles have shown that concentration of lactic acid (HLa) was prevailing among all metabolites, indicating that process followed through an essentially lactic route. Furthermore, considerable concentrations of propionic, acetic and isobutyric acids were also verified at some specific operation times, while solventogenesis was not detected at all. The greatest peak of lactate content was 4.96 g HLa L-1, observed under initial pH 6.0 and 25 g COD L-1, at 16 h. Maximum of lactate productivity was 332.10 g HLa L-1 h-1 at 8 h, associated to a yield of 189.14 g HLa (g Suc)-1, under initial pH 7.5 and 20 g COD L-1.

bioengineering↗

Acidogenic Fermentation of Sugarcane Vinasse and Glucose Using Two Different Anaerobic Digestion Inocula

Vinasse is an agroindustrial wastewater obtained at large amounts in countries such as Brazil, which is one of the world greatest sugar and ethanol producers from sugarcane. Despite its most common use is as nutrient for fertirrigation in sugarcane crops nowadays, alternative processing has been intensely sought in last decades for vinasse aiming primarily the production of energy carriers and high-added value chemicals. One of these alternatives is the anaerobic digestion promoted by bacteria, which results in several products of economic importance. In view of these information, this work presents an investigation of acidogenic fermentation by C. acetobutylicum ATCC 824 and C. beijerinckii ATCC 25752 was conducted in batch reactors by using synthetic vinasse and aqueous glucose solution as substrates. Evaluation of effects of microorganism and carbon source was focused mainly on the production of short chain organic acids (SCOAs), although other metabolites, such as ethanol, were also detected. Carbohydrates were effectively consumed by bacteria, so remaining concentrations of sucrose and glucose (Suc and Glu) in both substrates were considerably low at the end of tests. Lactic acid (HLa) was the prevailing metabolite in all experiments, determining the trends observed for total SCOAs content as a function of incubation time, while other acids were generated in much lesser amounts. Moreover, ethanol (EtOH) attained notably higher concentrations in batch reactors containing glucose as substrate, which showed more relevant production of this chemical. Maximum of 14.1 g HLa L-1 was quantified for lactic acid at incubation time 96 h in reactor using synthetic vinasse and C. beijerinckii (VB), linked to a productivity P = 146.46 mg HLa L-1 h-1 and a yield Y = 1108.03 mg HLa (g Sue)-1. For ethanol, maximum was 1.6 g EtOH L-1, occurring in reactor containing glucose and C. acetobutylicum (GA) also at incubation time 90 h, resulting in P = 13.60 mg EtOH L-1 h-1 and Y = 73.04 mg EtOH (g Glu)-1. Finally, kinetic models were adjusted to experimental data of carbohydrate content, lactate concentration and optical density (indicative of microorganism growth), showing good prediction capability as suggested by values of fit quality parameters such as Residual Sum of Squares (RSS) and R2.

bioengineering↗

Norepinephrine and glucocorticoids modulate chronic unpredictable stress-induced increase in the type 2 CRF and glucocorticoid receptors in brain structures related to the HPA axis activation.

The stress response is multifactorial and enrolls circuitries to build a coordinated reaction, leading to behavioral, endocrine, and autonomic changes. These changes are mainly related to the hypothalamus-pituitary-adrenal (HPA) axis activation and the organisms integrity. However, when self-regulation is ineffective, stress becomes harmful and predisposes the organism to pathologies. The chronic unpredictable stress (CUS) is a widely used experimental model since it induces physiological and behavioral changes and better mimics the stressors variability encountered in daily life. Corticotropin-releasing factor (CRF) and glucocorticoid (GCs) are deeply implicated in the CUS-induced physiological and behavioral changes. Nonetheless, the CUS modulation of CRF receptors and GR and the norepinephrine role in extra-hypothalamic brain areas were not well explored. Here, we show that 14-days of CUS induced a long-lasting HPA axis hyperactivity evidenced by plasmatic corticosterone increase and adrenal gland hypertrophy, which was dependent on both GCs and NE release induced by each stress session. CUS also increased CRF2 mRNA expression and GR protein levels in fundamental brain structures related to HPA regulation and behavior, such as the lateral septal nucleus intermedia part (LSI), ventromedial hypothalamic nucleus (VMH), and central nucleus of the amygdala (CeA). We also showed that NE participates in the CUS-induced increase in CRF2 and GR levels in the LSI, reinforcing the locus coeruleus (LC) involvement in the HPA axis modulation. Despite the CUS-induced molecular changes in essential areas related to anxiety-like behavior, this phenotype was not observed in CUS animals 24 h after the last stress session. HighlightsO_LICUS persistently increased plasma CORT levels via GCs and NE signaling. C_LIO_LICUS persistently increased CRF2 mRNA in extra-hypothalamic brain areas. C_LIO_LICUS increased GR protein levels in brain regions related to GCs release control. C_LIO_LINE and GCs participate in the CUS-induced increase in CRF2 and GR levels. C_LIO_LILSI could be the brain nucleus that dictates the fine-tuned response of CUS. C_LIO_LICUS animals did not present anxiety-like behavior. C_LI

neuroscience↗

Visual field reconstruction using fMRI-based techniques

PurposeTo evaluate the accuracy and reliability of functional magnetic resonance imaging (fMRI)-based techniques to assess the integrity of the visual field (VF). MethodsWe combined fMRI and neurocomputational models, i.e conventional population receptive field (pRF) mapping and a new advanced pRF framework "micro-probing" (MP), to reconstruct the visual field representations of different cortical areas. To demonstrate their scope, both approaches were applied in healthy participants with simulated scotomas (SS) and participants with glaucoma. For the latter group we compared the VFs obtained with standard automated perimetry (SAP) and via fMRI. ResultsUsing SS, we found that the fMRI-based techniques can detect absolute defects in VFs that are larger than 3 deg, in single participants, and based on 12 minutes of fMRI scan time. Moreover, we found that MP results in a less biased estimation of the preserved VF. In participants with glaucoma, we found that fMRI-based VF reconstruction detected VF defects with a correspondence to SAP that was decent, reflected by the positive correlation between fMRI-based sampling density and SAP-based contrast sensitivity loss (SAP) r2=0.44, p=0.0002.This correlation was higher for our new approach (MP) compared to that for the conventional pRF analysis. ConclusionsfMRI-based reconstruction of the VF enables the evaluation of vision loss and provides useful details on the properties of the visual cortex. Translational RelevancefMRI-based VF reconstruction provides an objective alternative to detect VF defects. It may either complement SAP, or could provide VF information in patients unable to perform SAP.

neuroscience↗

Off-target glycans encountered along the synthetic biology route towards humanized N-glycans in Pichia pastoris

BackgroundThe glycosylation pathways of several eukaryotic protein expression hosts are being engineered to enable the production of therapeutic glycoproteins with humanized application-customized glycan structures. In several expression hosts, this has been quite successful, but one caveat is that the new N-glycan structures inadvertently might be substrates for one or more of the multitude of endogenous glycosyltransferases in such heterologous background. This then results in the formation of novel, undesired glycan structures, which often remain insufficiently characterized. ResultsWhen expressing mouse interleukin-22 (mIL-22) in a Pichia pastoris (syn. Komagataella phaffi) GlycoSwitchM5 strain which had been optimized to produce Man5GlcNAc2 N-glycans, glycan profiling revealed two major species: Man5GlcNAc2 and an unexpected, partially -mannosidase-resistant structure. A detailed structural analysis using exoglycosidase sequencing, mass spectrometry, linkage analysis and NMR, revealed that this novel glycan was Man5GlcNAc2 modified with a Glc-1,2-Man{beta}-1,2-Man{beta}-1,3-Glc-1,3-R tetra-saccharide. Also the biosynthetic intermediates of this off-target modification were detected. Expression of a Golgi-targeted GlcNAc Transferase-I strongly inhibited the formation of this novel modification, resulting in more homogeneous modification with the targeted GlcNAcMan5GlcNAc2 structure. We have also observed the off-target glycan on other glycoproteins produced in the GlycoSwitchM5 strain. This illustrates the intricacies of Golgi glycosylation pathways and cautions that the use of glyco-engineered expression host cells should always be accompanied by detailed glycan analysis of the particular therapeutic proteins being produced. ConclusionsOur findings reinforce accumulating evidence that robustly customizing the N-glycosylation pathway in Pichia pastoris to produce particular human-type structures is still an incompletely solved synthetic biology challenge, which will require further innovation to enable safe glycoprotein pharmaceutical production.

synthetic biology↗

DENTAL CARIES AND ORAL HEALTH BEHAVIOR ASSESSMENT AMONG PORTUGUESE ADOLESCENTS

IntroductionIt is during the school phase that children and adolescents consolidate healthy behaviors, which will contribute to the decrease of diseases, especially in the reduction of dental caries. The main objective of the present study was to assess the decayed, missing and filled deciduous and permanent teeth index and oral health behaviors among Portuguese adolescents.\n\nMaterials and methodsAn observational cross-sectional study was designed including a sample of 694 adolescents between the ages of 12 and 18 years old from five public schools in the Viseu and Guarda districts, Portugal. After a self-administered questionnaire was filled out by the participants, a clinical examination was carried out in order to assess oral status and dental caries identification. A descriptive analysis of the variables was performed using the Chi-square, Mann-Whitney and Kruskal-Wallis tests (p<0.05).\n\nResultsThe decayed, missing and filled permanent teeth index was 2.91{+/-}2.9 and the decayed, missing and filled deciduous teeth index was 1.10{+/-}1.4. Of the total sample, 73% consumed sugary food on a daily basis, 54.7% drank bottled water, 50.1% considered oral health good, 70.8% did not report pain in the last 12 months, but noticed gingival bleeding (51.5%). Most adolescents (79.4%) brushed their teeth daily and 60% did not use dental floss. Of the total sample, 96.4% had a dental appointment in the last 12 months, being 46.4% due to prevention treatments. The high decayed, missing and filled deciduous teeth index was associated with low maternal scholarship, male gender and living in a rural residence area (p<0.05). Adolescents who brush their teeth daily presented a good perception about their oral health (p<0.001).\n\nConclusionsPortuguese adolescents presented a low decayed, missing and filled deciduous and permanent teeth index index. The decayed, missing and filled deciduous teeth index was associated with sociodemographic factors. Oral hygiene habits were associated with self-perception of oral health. It is suggested that oral health promotion and prevention programs should be improved in schools in order to reduce the risks of oral disease development.

scientific communication and education↗

HABIT - a webserver for interactive T cell neoepitope discovery

Neoepitopes generated by amino acid variants specifically found in pathogens or cancer cells are gaining momentum in immunotherapy development. HABIT (HLA Binding InTelligence) is a web platform designed to generate and analyse machine learning-based T cell epitope predictions for improved neoepitope discovery. Availability and ImplementationHABIT is available at http://habit.evobiomed.com. Peptide-HLA binding predictions by NetMHCpan 4.0 and NetMHCIIpan 3.1 were implemented in a web application for interactive data exploration using shiny package powered by RStudio. Contactnosorio@med.uminho.pt Supplementary informationSupplementary data and video tutorials are available online.

bioinformatics↗