Biology subjects

Sousa, C.

Publications and source records attributed to Sousa, C..

10 recordsLinked to original sources

bioRxiv2026

The immunogenic 33-mer gliadin peptide attenuates flagellin-induced TLR5/NF-κB signaling

Wheat gluten is the main trigger of gluten-related disorders such as celiac disease, in which the proteolysis-resistant, 33-mer peptide is a key immunodominant factor. It has been reported that the 33-mer forms oligomers and thin-plate superstructures that accumulate in the Caco-2 intestinal epithelial cell line, although their biological effects remain only partially understood. Here, we screened the effects of these nanostructures on Toll-like receptors in Caco-2 cells to investigate their effect on the early innate immune response. Our findings showed a specific upregulation of TLR5 mRNA in response to 33-mer gliadin superstructures, although no inflammatory response was detected based on CXCL1 and CXCL8 levels. Moreover, immunofluorescence analysis revealed partial co-localization with TLR5. We further found that the 33-mer superstructures attenuate flagellin-induced TLR5/NF{kappa}B signaling in reporter cells, similar to a blocking anti-hTLR5-IgA2 antibody. Molecular docking suggested that the 33-mer peptide occupies the concave region of TLR5 as well as 54% of the TLR5-TLR5 dimerization interface. The interference with the TLR5 dimerization domain could explain the experimental attenuation of flagellin-mediated TLR5 activation. Given that TLR5 contributes not only to inflammation but also to intestinal homeostasis, our findings suggest that the formation of 33-mer nanostructures upon its accumulation may increase the local concentration of 33-mer, interfering with TLR5-mediated mucosal immune responses. These findings provide a rationale for investigating the role of TLR5 and 33-mer aggregates in clinical samples from patients with celiac disease and other gluten-related disorders.

immunology↗

bioRxiv2026

Anopheles azevedoi (Ribeiro, 1969) in Angola: New geographical records, molecular characterization, and insecticide susceptibility profile of a commonly misidentified species

BACKGROUNDAngola ranks among the five countries with the highest malaria burden globally. The Ministry of Health in Angola has consistently partnered with international donors to sustain entomological surveillance and vector control strategies in a context of high malaria burden. METHODSVector surveillance was carried out in Luanda, Benguela, Namibe and Cuanza Sul provinces from 2016-2022. Collected adult mosquitoes were tested to assess the presence of Plasmodium parasites and determine blood sources. Larvae collections provided live material to test insecticide susceptibility in local Anopheles populations. Taxonomic determination of mosquitoes was based on external morphology and confirmed with molecular assays. The presence of Anopheles azevedoi was confirmed through morphology and genetic sequences, and errors in the original species determination were detected, discussed and corrected. OBJECTIVESThe study aimed to update the geographical range of Anopheles azevedoi in Angola and monitor the species susceptibility to public health insecticides. FINDINGS and MAIN CONCLUSIONSWe report on populations of Anopheles azevedoi occurring along the western coast of Angola, a highly abundant species with anthropophilic behavior in urban areas. Anopheles azevedoi is widely resistant to pyrethroids and DDT but fully susceptible to chlorfenapyr. We contribute with COI and ITS-2 barcoding sequences for future species identification and explain the reasons for which this species has been for long misidentified in Angola.

zoology↗

bioRxiv2026

Decellularized Small Intestine for Full Thickness Burn Wound Treatment

Treating extensive full-thickness burn wounds remains difficult in clinical practice because available donor skin is often limited, the risk of infection is high, and many standard dressings do not perform well when defects are large or structurally complex. These limitations have shifted attention to decellularized extracellular matrix (dECM) scaffolds, which can provide physical coverage while preserving biochemical cues that may support tissue repair. Based on this rationale, we designed a decellularization method that improves reagent penetration to produce a full-thickness porcine decellularized small intestine (dSI) scaffold for use in burn wound coverage. The protocol removed most cellular material while leaving low levels of detergent residue, and it maintained the native three-layer structure of the intestinal wall. Most key ECM components, such as collagen and glycosaminoglycans, were also retained. In this study, the dSI showed several properties relevant to burn care, capacity to absorb large amounts of fluid, water vapor transmission rates similar to those reported for skin, and resisted microbial penetration in vitro. From a mechanical standpoint, the scaffold retained anisotropic behaviour and remained stable under cyclic loading. This pattern indicates that it could withstand repeated deformation instead of acting like a fragile membrane. Degradation tests under enzymatic and oxidative conditions indicate that the material breaks down in a controlled way over a period that appears consistent with typical wound-healing timelines. In vitro assays indicated that the scaffold was cytocompatible, as human dermal fibroblasts and keratinocytes both attached to its surface and continued to proliferate. Cell responses differed depending on surface orientation, suggesting that preserved intestinal layers may shape cell behaviour in ways that are often missing in thinner or more uniform matrices. Overall, full-thickness dSI appears to act as a biologically active scaffold and shows mechanical properties that exceed those of many currently used burn dressings.

bioengineering↗

bioRxiv2024

Lack of Synergistic Outcomes with Roflumilast Combined with Levetiracetam or Adipose Stem Cell Secretome After Spinal Cord Injury

The Spinal Cord Injury (SCI) pathophysiology is highly complex, contributing to a poor prognosis and lack of effective treatments. Previously, we demonstrated that Roflumilast (Rof), leads to functional recovery when tested in a SCI contusion model. However, it is unlikely that Rof treatment on its own could fully restore the spinal cord. Therefore, our objective was to scrutinize the synergistic effects of combining Rof with neuroprotective approaches. Herein we tested two therapies, firstly, Rof combined with Levetiracetam (Lev), and in a second phase, the complementary interplay between Rof and Adipose Stem Cells secretome (Sec). We induced SCI using a weight drop trauma model at the T8 level. Functional recovery was assessed according to the Basso, Beattie, and Bresnahan scale, Activity Box Test, Motor Swimming Test, and Von Frei test. Results indicate that the unilateral use of Rof, Lev, or Sec was effective in promoting functional recovery. However, the combination of Rof + Lev or Rof + Sec did not lead to an improvement in functional outcomes when compared to standalone treatments. Moreover, the combination of Rof + Sec actually led to worst functional outcome than the single treatments. Further studies are needed to find a combinatorial treatment that can lead to superior therapeutic effects with potential clinical application.

neuroscience↗

bioRxiv2024

Dbp1 is a low performance paralog of RNA helicase Ded1 that drives impaired translation and heat stress response

Ded1 and Dbp1 are paralogous conserved RNA helicases that enable translation initiation in yeast. Ded1 has been heavily studied but the role of Dbp1 is poorly understood. We find that the expression of these two helicases is controlled in an inverse and condition-specific manner. In meiosis and other long-term starvation states, Dbp1 expression is upregulated and Ded1 is downregulated, whereas in mitotic cells, Dbp1 expression is extremely low. Inserting the DBP1 ORF in place of the DED1 ORF cannot replace the function of Ded1 in supporting translation, partly due to inefficient mitotic translation of the DBP1 mRNA, dependent on features of its ORF sequence but independent of codon optimality. Global measurements of translation rates and 5 leader translation, activity of mRNA-tethered helicases, ribosome association, and low temperature growth assays show that--even at matched protein levels--Ded1 is more effective than Dbp1 at activating translation, especially for mRNAs with structured 5 leaders. Ded1 supports halting of translation and cell growth in response to heat stress, but Dbp1 lacks this function, as well. These functional differences in the ability to efficiently mediate translation activation and braking can be ascribed to the divergent, disordered N- and C-terminal regions of these two helicases. Altogether, our data show that Dbp1 is a "low performance" version of Ded1 that cells employ in place of Ded1 under long-term conditions of nutrient deficiency.

genomics↗

bioRxiv2021

An efficient urine peptidomics workflow identifies chemically defined dietary gluten peptides from patients with celiac disease

Celiac disease (CeD) is an autoimmune disorder induced by consuming gluten proteins from wheat, barley, and rye. Glutens resist gastrointestinal proteolysis, resulting in peptides that elicit inflammation in patients with CeD. Despite well-established connections between glutens and CeD, chemically defined, bioavailable peptides produced from dietary proteins have never been identified from humans in an unbiased manner. This is largely attributable to technical challenges, impeding our knowledge of potentially diverse peptide species that encounter the immune system. Here, we developed a novel liquid chromatographic-mass spectrometric workflow for untargeted sequence analysis of the urinary peptidome. We detected 679 distinct dietary peptides, of which ~35% have a CeD-relevant T cell epitope and ~5% are known to stimulate innate immune responses. Remarkably, gluten peptides from patients with CeD qualitatively and quantitatively differ from controls. Our results provide a new foundation for understanding gluten immunogenicity, improving CeD management, and characterizing the dietary and urinary peptidomes.

biochemistry↗

bioRxiv2020

Neurodegeneration and neuroinflammation are linked, but independent of alpha-synuclein inclusions, in a seeding/spreading mouse model of Parkinson disease

A key process of neurodegeneration in Parkinson disease (PD) is the transneuronal spreading of alpha-synuclein. Alpha-synuclein is a presynaptic protein that is implicated in the pathogenesis of PD and other synucleinopathies, where it forms, upon intracellular aggregation, pathological inclusions. Other hallmarks of PD include neurodegeneration and microgliosis in susceptible brain regions. Whether it is primarily transneuronal spreading of alpha-synuclein particles, inclusion formation, or other mechanisms, such as inflammation, that cause neurodegeneration in PD is unclear. We used spreading/aggregation of alpha-synuclein induced by intracerebral injection of alpha-synuclein preformed fibrils into the mouse brain to address this question. We performed quantitative histological analysis for alpha-synuclein inclusions, neurodegeneration, and microgliosis in different brain regions, and a gene expression profiling of the ventral midbrain, at two different timepoints after disease induction. We observed significant neurodegeneration and microgliosis in brain regions not only with, but also without alpha-synuclein inclusions. We also observed prominent microgliosis in injured brain regions that did not correlate with neurodegeneration nor with inclusion load. In longitudinal gene expression profiling experiments, we observed early and unique alterations linked to microglial mediated inflammation that preceded neurodegeneration, indicating an active role of microglia in inducing neurodegeneration. Our observations indicate that alpha-synuclein inclusion formation is not the major driver in the early phases of PD-like neurodegeneration, but that diffusible, oligomeric alpha-synuclein species, which induce unusual microglial reactivity, play a key role in this process. Our findings uncover new features of alpha-synuclein induced pathologies, in particular microgliosis, and point to the necessity of a broader view of the process of prion-like spreading of that protein.

neuroscience↗

bioRxiv2019

Regulation of nerve growth and patterning by cell surface protein disulphide isomerase

Contact repulsion of growing axons is an essential mechanism for spinal nerve patterning. In birds and mammals the embryonic somites generate a linear series of impenetrable barriers, forcing axon growth cones to traverse one half of each somite as they extend towards their body targets. This study shows that protein disulphide isomerase provides a key component of these barriers, mediating contact repulsion at the cell surface in half-somites. Repulsion is reduced both in vivo and in vitro by a range of methods that inhibit enzyme activity. The activity is critical in initiating a nitric oxide/S-nitrosylation-dependent signal transduction pathway that regulates the growth cone cytoskeleton. Rat forebrain grey matter extracts contain a similar activity, and the enzyme is expressed at the surface of cultured human astrocytic cells and rat cortical astrocytes. We suggest this system is co-opted in the brain to counteract and regulate aberrant nerve terminal growth.

developmental biology↗

bioRxiv2019

Quantifying genetic regulatory variation in human populations improves transcriptome analysis in rare disease patients

Transcriptome data holds substantial promise for better interpretation of rare genetic variants in basic research and clinical settings. Here, we introduce ANalysis of Expression VAriation (ANEVA) to quantify genetic variation in gene dosage from allelic expression (AE) data in a population. Application to GTEx data showed that this variance estimate is robust across datasets and is correlated with selective constraint in a gene. We next used ANEVA variance estimates in a Dosage Outlier Test (ANEVA-DOT) to identify genes in an individual that are affected by a rare regulatory variant with an unusually strong effect. Applying ANEVA-DOT to AE data form 70 Mendelian muscular disease patients showed high accuracy in detecting genes with pathogenic variants in previously resolved cases, and lead to one confirmed and several potential new diagnoses in cases previously unresolved. Using our reference estimates from GTEx data, ANEVA-DOT can be readily incorporated in rare disease diagnostic pipelines to better utilize RNA-seq data.\n\nOne Sentence SummaryNew statistical framework for modelling allelic expression characterizes genetic regulatory variation in populations and informs diagnosis in rare disease patients

genomics↗

bioRxiv2019

On the front line of Klebsiella pneumoniae surface structures understanding: establishment of Fourier Transform Infrared (FT-IR) spectroscopy as a capsule typing method

Genomics-based population analysis of multidrug resistant (MDR) Klebsiella pneumoniae (Kp) motivated a renewed interest on capsule (K) types given their importance as evolutionary and virulence markers of clinically relevant strains. However, there is a gap between genotypic based predictions and information on capsular polysaccharide structure and composition. We used molecular genotypic, comparative genomics, biochemical and phenotypic data on the cps locus to support the usefulness of Fourier-Transform Infrared (FT-IR) spectroscopy as a phenotypic approach for K-type characterization and identification. The approach was validated with a collection of representative MDR Kp isolates from main lineages/Clonal Groups (CGs) involved in local or nationwide epidemics in 6 European and South American countries. FT-IR-based K-type assignments were compared with those obtained by genotypic methods and WGS (cps operon), and further complemented with data on the polysaccharide composition and structure of known K-types. We demonstrate that our FT-IR-based spectroscopy approach can discriminate all 21 K-types identified with a resolution comparable (or even higher) to that provided by WGS, considered gold-standard methodology. Besides contributing to enlighten K-type diversity among a significant MDR Kp collection, the specific associations between certain K-types and Kp lineages identified in different geographic regions over time support the usefulness of our FT-IR-based approach for strain typing. Additionally, we demonstrate that FT-IR discriminatory ability is correlated with variation on the structure/composition of known K-types and, supported on WGS data, we were able to predict the sugar composition and chemical structure of new KL-types. Our data revealed an unprecedent resolution at a quick and low-cost rate of Kp K-types at the phenotypic level. Our FT-IR spectroscopy-based approach might be extremely useful not only as a cost-effective Kp typing tool, but also to improve our understanding on sugar-based coating structures of high relevance for strain evolution and host adaptation.

developmental biology↗