Biology subjects

Moreira, A.

Publications and source records attributed to Moreira, A..

8 recordsLinked to original sources

bioRxiv2025

Quantitative Proteomics Links Mitochondrial Dysfunction to Metabolic Changes and Epithelial Differentiation Defects in Hyperoxia-Exposed Neonatal Airway Cells

Premature infants often require supplemental oxygen therapy, however, exposure to supraphysiological oxygen (hyperoxia) can disrupt normal lung development and contribute to bronchopulmonary dysplasia (BPD). Mitochondrial dysfunction is increasingly recognized as a contributor to hyperoxia-induced BPD. However, the effects of hyperoxia on mitochondrial function and mucociliary differentiation in the developing upper airway epithelium remain poorly understood. This study tested the hypothesis that hyperoxia impairs neonatal airway mucociliary differentiation by disrupting mitochondrial bioenergetic function. Neonatal tracheal airway epithelial cells (nTAECs) from term infants (n=5) were cultured in a 3D air-liquid interface (ALI) model and exposed to 60% O2 during the mid-phase of differentiation (ALI day 7-14). Cellular phenotype was assessed using immunofluorescence staining and gene expression analyses. Mitochondrial function was evaluated through Seahorse metabolic flux analysis, and global protein changes were characterized by quantitative proteomics. Hyperoxia exposure significantly impaired terminal epithelial differentiation, characterized by reductions in ciliated and goblet cells. Seahorse assay revealed a decrease in baseline oxygen consumption and mitochondrial ATP production, accompanied by a compensatory increase in glycolytic ATP production. Quantitative proteomics identified disruption of mitochondrial Complex I as a central feature of the hyperoxic response. Downstream proteomic pathway analyses further confirmed the metabolic shift from mitochondrial to glycolytic ATP production and demonstrated altered epithelial differentiation pathways, including NOTCH and TGF-{beta} signaling. These findings reveal that moderate hyperoxia impairs mitochondrial bioenergetics and alters metabolic programming, leading to disrupted mucociliary differentiation. Future in vivo studies should evaluate mitochondrial oxidative fitness as a therapeutic target in neonatal lung disease. NEW & NOTEWORTHYWe report that moderate hyperoxia during a critical window of mucociliary differentiation disrupts terminal maturation in neonatal airway epithelial cells cultured in a 3D model. Hyperoxia induced mitochondrial bioenergetic dysfunction and metabolic reprogramming, with proteomic analysis identifying Complex I disruption as a key driver of impaired differentiation. Overall, these findings reveal a previously underrecognized link between mitochondrial bioenergetics and airway epithelial development, positioning metabolic dysfunction as an early trigger of hyperoxia-induced neonatal airway injury.

developmental biology↗

bioRxiv2024

Deep learning uncovers histological patterns of YAP1/TEAD activity related to disease aggressiveness in cancer patients.

Over the last decade, Hippo signaling has emerged as a major tumor-suppressing pathway. Its dysregulation is associated with abnormal expression of YAP1 and TEAD-family genes. Recent works have highlighted the role of YAP1/TEAD activity in several cancers and its potential therapeutic implications. Therefore, identifying patients with a dysregulated Hippo pathway is key to enhancing treatment impact. Although recent studies have derived RNAseq-based signatures, there remains a need for a reproducible and cost-effective method to measure the pathway activation. In recent years, deep learning applied to histology slides have emerged as an effective way to predict molecular information from a data modality available in clinical routine. Here, we trained models to predict YAP1/TEAD activity from H&E-stained histology slides in multiple cancers. The robustness of our approach was assessed in seven independent validation cohorts. Finally, we showed that histological markers of disease aggressiveness were associated with dysfunctional Hippo signaling.

bioinformatics↗

bioRxiv2023

Single nucleus transcriptome of a "Super RV" shows increased insulin and angiogenesis signaling

The right ventricle (RV) is one of the four pumping chambers of the heart, pumping blood to the lungs. In severe forms of congenital heart disease and pulmonary hypertension, the RV is made to pump into the systemic circulation. Such systemic RVs typically display early failure due to pressure overload. In rare cases a systemic RV persists into later decades of life - colloquially called a Super RV. Here we present the single-nucleus transcriptome of a systemic RV from a 60-year-old with congenitally corrected transposition of great arteries (ccTGA). Our data shows two specific signaling pathways enriched in the ccTGA RV myocardium. First, we show increased insulin like growth factor (IGF1) signaling within the systemic RV myocardium: there is increased expression of the main receptor IGFR1 within the cardiomyocytes, and IGF1 ligands within the cardiofibroblasts and macrophages. Second, we find increased VEGF and Wnt9 ligand expression in cardiomyocytes and increased VEGF1R and Wnt9 receptors in endothelial cells, which are implicated in angiogenesis. We show that increased insulin and angiogenesis signaling are potentially beneficial RV adaptations to increased pressure overload. This study of an adult systemic RV provides an important framework for understanding RV remodeling to systemic pressures in congenital heart disease and pulmonary hypertension.

developmental biology↗

bioRxiv2023

Unravelling the regulatory function of a short 3'UTR sequence in zebrafish and in human cells

The mechanisms of gene expression regulation are essential for cell identity and function, and their disruption usually leads to human disease. The 3 untranslated region (UTR) of mRNA contains important regulatory elements of gene expression, including upstream sequence elements (USEs) that are cis-regulatory sequences localized upstream of polyA signals (PAS). One of the best functionally characterised USEs is located in the 3UTR of the Drosophilas polo gene, which disruption leads to critical phenotypes in adult flies. In this work we found that the USE of the Drosophilas polo gene (DplUSE) is also found in in the 3UTR of vertebrate genes, including zebrafish, mouse and human genes, showing higher levels of conservation than the whole 3UTR sequence. Using reporter assays, we show that DplUSE is able to increase gene expression in vitro in human cell lines and in vivo in zebrafish embryos. Importantly, in humans, the DplUSE containing genes are enriched for genes associated to serious diseases such as Congenital abnormalities and Malignant neoplasms, illustrating the potential of this sequence to modulate genes with relevant biological functions and related with human health. Concomitantly, when sequestering the molecular machinery that operates at the DplUSE using a dominant negative strategy, we show that this is enough to dysregulate DplUSE containing genes in human cells and disrupt proper embryo development in zebrafish. Aiming to understand the molecular mechanism operating at the DplUSE, we identified three RNA binding proteins (RBP) that specifically bind to the DplUSE in vertebrates. Importantly, one of such RBPs is PTBP1, the vertebrate orthologue of the fruit flys RBP Heph, that was demonstrated to be required for the DplUSE function in Drosophila. To test if PTBP1is essential for the DplUSE function, as observed in Drosophila, we depleted PTBP1 from human cells and observed a downregulation of the expression of DplUSE containing genes, demonstrating that the molecular mechanisms that operate at DplUSE are ultra-conserved. Finally, we explored if variants in DplUSE consensus could be associated to human disease. We found a reported single nucleotide polymorphism (SNP; rs3087967) that is associated with malignant tumor of colon and generates an ectopic consensus of DplUSE in the 3 UTR of the tumorigenic POU2AF2/C11orf53 gene. We further show that this ectopic DplUSE motif causes a gain-of-function in vivo in zebrafish gut cells, suggesting its involvement in colon cancer development. These results show that a short motif present in the 3UTR of genes from phylogenetically distant bilaterians, from fruit flies to humans, control genes expression through an ultra-conserved mechanism involving RBPs binding and its dysregulation might impact in human disease.

molecular biology↗

bioRxiv2023

Impacts of urban stressors on freshwater biofilms

Aquatic ecosystems and their communities are exposed to numerous stressors of various natures (chemical and physical), which impacts are often poorly documented. In urban areas, the use of biocides such as dodecyldimethylbenzylammonium chloride (DDBAC) and their subsequent release in wastewater result in their transfer to urban aquatic ecosystems. DDBAC is known to be toxic to most aquatic organisms. Artificial light at night (ALAN) is another stressor that is increasing globally, especially in urban areas. ALAN may have a negative impact on photosynthetic cycles of periphytic biofilms, which in turn may result in changes in their metabolic functioning. Moreover, studies suggest that exposure to artificial light could increase the biocidal effect of DDBAC on biofilms. The present study investigates the individual and combined effects of DDBAC and/or ALAN on the functioning and structure of photosynthetic biofilms. We exposed biofilms in artificial channels to a nominal concentration of 30 mg.L-1 of DDBAC and/or ALAN for 10 days. ALAN modified DDBAC exposure, decreasing concentrations in the water but not accumulation in biofilms. DDBAC had negative impacts on biofilm functioning and structure. Photosynthetic activity was inhibited by > 90% after 2 days of exposure, compared to the controls, and did not recover over the duration of the experiment. Biofilm composition was also impacted, with a marked decrease in green algae and the disappearance of microfauna under DDBAC exposure. The integrity of algal cells was compromised where DDBAC exposure altered the chloroplasts and chlorophyll content. Impacts on autotrophs were also observed through a shift in lipid profiles, in particular a strong decrease in glycolipid content was noted. We found no significant interactive effect of ALAN and DDBAC on the studied endpoints. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=47 SRC="FIGDIR/small/544504v2_ufig1.gif" ALT="Figure 1"> View larger version (13K): org.highwire.dtl.DTLVardef@15e217borg.highwire.dtl.DTLVardef@e81f3forg.highwire.dtl.DTLVardef@15b9c08org.highwire.dtl.DTLVardef@1610534_HPS_FORMAT_FIGEXP M_FIG C_FIG

ecology↗

bioRxiv2023

Pro-inflammatory polarization and colorectal cancer modulate alternative and intronic polyadenylation in primary human macrophages

Macrophages are essential cells of the immune system that alter their inflammatory profile depending on their microenvironment. Alternative polyadenylation in the 3UTR (3UTR-APA) and intronic polyadenylation (IPA) are mechanisms that modulate gene expression, in particular in cancer and activated immune cells. Yet, how polarization and colorectal cancer (CRC) cells microenvironment affect 3UTR-APA and IPA in primary human macrophages remains unknown. Here, primary human monocytes were isolated from healthy donors, differentiated and polarized into a pro-inflammatory state and ChrRNA-Seq and 3RNA-Seq were performed to quantify gene expression and characterize new 3UTR-APA and IPA mRNA isoforms. Our results show that polarization of human macrophages from naive to a pro-inflammatory state causes a marked increase both in proximal polyA site selection in the 3UTR and in IPA events, in genes relevant for macrophage functions. Additionally, we found a negative correlation between differential gene expression and IPA during pro-inflammatory polarization of primary human macrophages. As macrophages are abundant immune cells in the CRC microenvironment that either promote or abrogate cancer progression, we investigated how indirect exposure to CRC cells affects macrophage gene expression and 3UTR-APA and IPA mRNA events. Co-culture with CRC cells alters the inflammatory phenotype of macrophages, increases the expression of pro-tumoral genes and induce 3UTR-APA alterations. Notably, some of these gene expression differences were also found in tumour-associated macrophages of CRC patients, indicating that they are physiological relevant. Upon macrophage pro-inflammatory polarization SRSF12 is the pre-mRNA processing gene that is most upregulated. After SRSF12 knockdown in M1 macrophages there is a global downregulation of gene expression, in particular in genes involved in gene expression regulation and in immune responses. Our results reveal new 3UTR-APA and IPA mRNA isoforms produced during pro-inflammatory polarization of primary human macrophages and CRC co-culture that may be used in the future as diagnostic or therapeutic tools.

molecular biology↗

bioRxiv2020

Topoisomerase 1 inhibition therapy protects against SARS-CoV-2-induced inflammation and death in animal models.

The ongoing pandemic caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is currently affecting millions of lives worldwide. Large retrospective studies indicate that an elevated level of inflammatory cytokines and pro-inflammatory factors are associated with both increased disease severity and mortality. Here, using multidimensional epigenetic, transcriptional, in vitro and in vivo analyses, we report that Topoisomerase 1 (Top1) inhibition suppresses lethal inflammation induced by SARS-CoV-2. Therapeutic treatment with two doses of Topotecan (TPT), a FDA-approved Top1 inhibitor, suppresses infection-induced inflammation in hamsters. TPT treatment as late as four days post-infection reduces morbidity and rescues mortality in a transgenic mouse model. These results support the potential of Top1 inhibition as an effective host-directed therapy against severe SARS-CoV-2 infection. TPT and its derivatives are inexpensive clinical-grade inhibitors available in most countries. Clinical trials are needed to evaluate the efficacy of repurposing Top1 inhibitors for COVID-19 in humans.

microbiology↗

bioRxiv2019

MMP2 As An Independent Prognostic Stratifier In Oral Cavity Cancers

BackgroundAround 25% of oral cavity squamous cell carcinoma (OCSCC) are not controlled by standard of care. Identifying those patients could offer them possibilities for intensified and personalized regimen. However, there is currently no validated biomarker for OCSCC patient selection in a pre-treatment setting.\n\nPatients and methodsOur objectives were to determine a robust and independent predictive biomarker for disease related death in OCSCC treated with standard of care. Tumor and juxtatumor secretome were analyzed in a prospective discovery cohort of 37 OCSCC treated by primary surgery. Independent biomarker validation was performed by RTqPCR in a retrospective cohort of 145 patients with similar clinical features. An 18-gene signature (18G) predictive of the response to PD-1 blockade was evaluated in the same cohort..\n\nResultsAmong 29 deregulated molecules in a secretome analysis, we identified soluble MMP2 as a prognostic biomarker. In our validation cohort (n=145), high levels of MMP2 and CD276, and low levels of CXCL10 and STAT1 mRNA were associated with poor prognosis in univariate analysis (Kaplan-Meier). MMP2 (p = 0.001) and extra-nodal extension (ENE) (p = 0.006) were independent biomarkers of disease-specific survival (DSS) in multivariate analysis, and defined prognostic groups with 5-year DSS ranging from 36% (MMP2highENE+) to 88% (MMP2lowENE-). The expression of 18G was similar in the different prognostic groups, suggesting comparable responsiveness to anti-PD-1.\n\nConclusionHigh levels of MMP2 was an independent and validated prognostic biomarker, which may be used to select poor prognosis patients for intensified neoadjuvant or adjuvant regimens.

cancer biology↗