Biology subjects

Martins, T.

Publications and source records attributed to Martins, T..

13 recordsLinked to original sources

bioRxiv2026

CORONAVIRUS INFECTION ACCELERATES EXPERIMENTAL OSTEOPOROSIS THROUGH THE ACTIVATION OF OSTEOCLASTS

Coronavirus infection is increasingly associated with systemic inflammation and extrapulmonary sequelae, including bone loss. However, the impact of coronavirus infection on hosts with pre-existing osteoporosis remains poorly understood. Here, we investigated the effects of infection with the murine coronavirus MHV-A59 on bone remodeling in a mouse model of ovariectomy-induced osteoporosis. Female C57BL/6 mice underwent ovariectomy (OVX) or sham surgery and, after establishment of the osteoporotic phenotype (14 days post-surgery), were intranasally infected with MHV-A59. Clinical, inflammatory, and femur parameters were evaluated throughout acute and chronic phases of infection using histopathological, micro-computed tomography and biochemical analyses. In vitro infection assays with bone-marrow derived macrophages (BMDM) and BMDM-differentiated osteoclasts from Sham and OVX mice were also performed. MHV-A59 induced comparable lung inflammation in Sham and OVX mice. In contrast, infection accelerated bone resorption in OVX mice during the acute phase (5 dpi), whereas in non-osteoporotic animals, infection induced significant bone loss at later stages (60 dpi). These alterations were associated with increased osteoclast differentiation and activity and with elevated levels of osteometabolic and inflammatory mediators, including RANKL, CCL2, CCL3, CCL5, and TGF-{beta}1. In vitro, MHV-A59 infection enhanced the expression of osteoclast-associated markers, including Rank, Traf6, Nfac1, and cathepsin K, and inflammatory mediators by BMDM and osteoclasts. Importantly, these responses were significantly more pronounced in cells derived from OVX mice. Collectively, our findings demonstrate that coronavirus infection promotes bone resorption and accelerates early bone loss in the context of estrogen deficiency, highlighting enhanced osteoclast activity as a potential mechanism underlying this infection-induced phenotype.

cell biology↗

bioRxiv2026

CCR4+low-density neutrophils define a novel immunosuppressive subset associated with breast cancer progression and early mortality

Neutrophils are increasingly recognized as key players in cancer progression, yet their functional heterogeneity limits their therapeutic exploitation. In breast cancer (BC), elevated circulating and tumor-associated neutrophils have been associated with poor prognosis, and low-density neutrophils (LDN), particularly, have been linked to immunosuppression. However, the specific neutrophil subsets driving these effects remain undefined. Here, we identify a previously unrecognized CCR4-expressing neutrophil subset, strongly enriched among LDN, whose frequency rises with disease stage and correlates with faster progression and shorter survival. Serial blood sampling further showed that CCR4+LDN levels fluctuate in line with clinical course, with increases indicating near-term risk in metastatic patients. Our findings also reveal that this subset has an immunosuppressive signature more pronounced than that of the remaining LDN, migrates preferentially toward tumor-derived chemokines, and dampens T cell-driven tumor killing even when PD-1 is blocked. Additionally, in a pilot cohort of Triple-Negative BC patients on anti-PD-1 therapy, early changes in this population were associated with treatment outcome. Together, these results reveal a distinct neutrophil population that actively contributes to immune escape and tumor progression, establishing CCR4+LDN as a minimally invasive real-time biomarker of disease course and treatment response, and as a promising target for neutrophil-directed therapies in BC.

immunology↗

bioRxiv2026

SPARC mediates tumour-stroma intercellular communication through endosomal regulation of Delta and Notch signalling

Tumour progression relies on reciprocal communication between genetically altered cancer cells and surrounding stromal cells. While the genetic alterations that initiate tumorigenesis have been extensively studied, the dysregulated feedback signalling provided by co-opted stromal cells remains poorly understood. Here, we used a Drosophila cancer model to address this question and identified the matricellular protein SPARC as a mediator of tumour-stroma communication. SPARC is produced by mesenchymal cells and transferred into epithelial tumour cells, where it is internalized through the endocytic pathway. Following uptake, SPARC accumulates in Rab7 positive late endosomes and colocalize with the Notch ligand Delta. SPARC internalization promotes endosomal enlargement and reduces endosome dynamics. Increased SPARC levels in epithelial tumours indirectly attenuate Notch signalling activity through at least altered Delta trafficking. We further identify the N-terminal acidic domain of SPARC as specifically required for its targeting to Delta-associated endosomes. Together, our findings uncover a stromal feedback mechanism by which SPARC modulates Notch signalling through endosomal regulation during tumour development.

developmental biology↗

bioRxiv2025

Neutrophils matter: New clinical insights on their role in the progression of metastatic breast cancer

BackgroundMetastatic breast cancer (BC) remains a significant clinical challenge, needing innovative strategies to improve disease management and extend patient survival. Increased neutrophil levels have been observed in both peripheral blood and tumor tissues of patients with different types of cancer, often being associated with poor clinical outcomes. These findings suggest a crucial role for neutrophils in tumor progression, raising interest in neutrophil-based therapies. However, the functional and phenotypic heterogeneity of neutrophils complicates their therapeutic targeting. This study aims to investigate the clinical impact of immunosuppressive, protumor low-density neutrophils (LDN) in metastatic BC, comparing them with normal high-density neutrophils (HDN) to better understand their role in disease progression. MethodsLDN and HDN subpopulations were isolated from the blood of 151 BC patients (72 metastatic, 79 non-metastatic) using density gradient centrifugation. Their frequency, phenotype, and function were analyzed by flow cytometry and in vitro experiments. Correlations between LDN levels and clinical data from metastatic BC patients were evaluated, alongside individual longitudinal assessments. ResultsLDN accumulated significantly in the blood of BC patients, particularly in those with metastatic disease. Elevated LDN levels in these patients were associated with faster disease progression and reduced life expectancy, regardless of metastatic site. Longitudinal analysis revealed that higher LDN percentages often correlated with adverse clinical events, whereas lower levels of LDN were linked to stable disease. Functionally, LDN exhibited protumor properties, including elevated expressions of PD-L1 and MMP-9, contributing to immunosuppression and metastasis. Unlike HDN, which demonstrated cytotoxicity against tumor cells, LDN failed to reduce BC cell line viability in 3D co-cultures. Notably, BC cell lines exposed to LDN-conditioned medium showed increased invasive capacity and proliferation, while T cells cultured in the same medium displayed impaired activation, likely due to the effect of arginase. ConclusionOur results highlighted LDN and their secreted factors as major drivers of BC progression and increased aggressiveness. These findings suggest that incorporating LDN assessment into clinical practice could aid in identifying high-risk patients and enable more personalized treatment approaches. Furthermore, our data strengthen the relevance of targeting specific neutrophil subsets or their functions to improve metastatic BC management and patient outcomes.

immunology↗

bioRxiv2024

Enhancing HLA-DR in Cytotoxic T Lymphocytes is crucial for the development of efficient adoptive T cell Therapies for Breast Cancer

BackgroundDespite advances in breast cancer (BC) therapies, more effective interventions are needed, especially for chemotherapy-resistant tumors. Immune checkpoint inhibitors show promise for triple-negative breast cancer, but their effectiveness across all BC subtypes remains challenging. Therefore, novel strategies, including adoptive cellular therapy, employing patients own T lymphocytes expanded ex vivo, are under investigation. Previously, we demonstrated that cytotoxic T lymphocytes (CTLs) expressing high HLA-DR levels in the tumor microenvironment are associated with a good response to neoadjuvant chemotherapy (NACT), due to their pronounced anti-tumor properties compared to CTLs with low or no HLA-DR expression. In this paper, we demonstrated that HLA-DR expression in CTLs is crucial for efficient T lymphocytes-based therapies. MethodsTo clarify the role of HLA-DR in CTLs anti-tumor abilities, we performed in vitro and in vivo experiments. We also improved a protocol to expand ex vivo HLA-DR-expressing CTLs and employed a 3D co-culture platform to test the potential of different immune agents, namely an anti-PD1, anti-OX40, anti-VEGF and anti-CD137, on CTLs cytotoxicity against BC cells. Additionally, we conducted a bioinformatic analysis of scRNA-seq data of BC patients to better understand the modulation of HLA-DR expression in CTLs. ResultsOur findings revealed that CTLs require HLA-DR expression to eliminate tumor cells. Additionally, we unveiled that blocking HLA-DR or depleting CD4+ T cells compromised CTLs activation and cytotoxicity, suggesting antigen presentation by CTLs through HLA-DR, and CD4+ T cells, as probable mechanisms for CTLs increased anti-tumor immune response and treatment efficacy. We refined an ex vivo stimulation and cytokine supplementation protocol, observing that short-term stimulation increases HLA-DR expression while boosting CTLs functionality, unlike prolonged expansion. This result highlights the importance of prioritizing cell quality, over quantity, for therapy efficiency. Additionally, we verified that anti-PD-1 further increases HLA-DR levels in CTLs, enhancing their anti-tumor efficiency. Notably, an in silico analysis revealed that PD-1 in CTLs shares 34 co-expressed genes with HLA-DR, including several non-coding RNAs, suggesting a PD-1-mediated regulation of HLA-DR expression. ConclusionsGlobally, our findings underscore that heightening HLA-DR expression in CTLs, by combining anti-PD-1 with short-term stimulation, offers promise for improving T lymphocyte-based therapies for BC. Key MessageO_LIWhat is already known on this topic: While immunotherapy holds promise for breast cancer (BC), its success is still limited. Novel strategies are under investigation to improve outcomes across all BC subtypes. Previously we established a correlation between HLA-DR expression on CTLs and a positive response to neoadjuvant chemotherapy, likely due to enhanced anti-tumor properties of HLA-DR-expressing CTLs. However, the specific role of HLA-DR on CTLs and its implications for T lymphocyte-based therapies requires further investigation. C_LIO_LIWhat this study adds: This study revealed that HLA-DR expression is essential for CTLs to effectively eliminate tumor cells. Blocking HLA-DR or depleting CD4+ T cells impairs CTLs activation and cytotoxicity, indicating that HLA-DR-mediated antigen presentation and interaction with CD4+ T cell are crucial for CTLs function. The study also shows that combining ex vivo short-term stimulation and anti-PD-1 treatment increases HLA-DR levels in CTLs, enhancing their anti-tumor activity. C_LIO_LIHow this study might affect research, practice or policy: By emphasizing the importance of optimizing CTLs quality over quantity, this approach has the potential to improve the design of more efficient T lymphocyte-based therapies for BC. This could influence future research directions, clinical practices, and treatment policies, leading to improved therapeutic outcomes. C_LI

immunology↗

bioRxiv2024

Dynamic role of monocytes and meningeal macrophages in bacterial meningoencephalitis

Macrophages in the meninges, especially in the dura mater sheathing the brain from the skull, are involved in the immune defense of the central nervous system (CNS). However, their site-specific origin and function, both in steady state and in bacterial CNS infections are incompletely understood. Using an intravenous model of streptococcal meningoencephalitis that mimics hematogenous dissemination in humans, we found that bacteria accumulated predominantly in the leptomeninges and dura, whereas invasion into the brain parenchyma was rare. However, monocyte infiltration into the leptomeninges and parenchyma strongly correlated with disease severity. In the dura, infection triggered activation and loss of resident macrophages, followed by rapid engraftment of inflammatory monocytes that transiently replenished the dural macrophage niche. Under homeostasis, dural monocytes were supplied independently of CCR2 from adjacent skull bone marrow. In infection, however, this local reservoir was rapidly exhausted, and the markedly increased demand for monocytes required mobilization from peripheral bone marrow sources, revealing context-dependent heterogeneity in monocyte origin. Infection also reshaped ontogeny of this differential monocyte output, with an increase in Monocyte-Dendritic Cell Progenitor - derived monocytes (MDP-Mo). MDP-Mo exhibited enhanced MHC-II expression and persisted in the brain during the resolution phase together with CD4 T cells, suggesting a role in antigen presentation after bacterial clearance. Together, these findings reveal a highly dynamic and compartment-specific remodeling of monocyte ontogeny, recruitment, and differentiation across CNS borders during bacterial meningoencephalitis. These mechanisms may offer opportunities for therapeutic interventions in the future. One Sentence SummaryStreptococcal meningoencephalitis disrupts homeostatic, skull bone marrow-derived monocyte and macrophage trajectories in the dura.

immunology↗

bioRxiv2023

De-suppression of mesenchymal cell identities and variable phenotypic outcomes associated with knockout of Bbs1.

Bardet-Biedl syndrome (BBS) is an archetypal ciliopathy caused by dysfunction of primary cilia. BBS affects multiple tissues, including the kidney, eye and hypothalamic satiety response. Understanding pan-tissue mechanisms of pathogenesis versus those which are tissue specific, and gauging their associated inter-individual variation owing to genetic background and stochastic processes, is of paramount importance in syndromology. The BBSome is a membrane trafficking and intraflagellar transport (IFT) adaptor protein complex formed by 8 BBS proteins, including BBS1, which is the most commonly mutated gene in BBS. To investigate disease pathogenesis we generated a series of clonal renal collecting duct IMCD3 cell lines carrying defined biallelic nonsense or frameshift mutations in Bbs1, as well as a panel of matching wild-type CRISPR control clones. Using a phenotypic screen and an unbiased multi-omics approach we note significant clonal variability for all assays, emphasising the importance of analysing panels of genetically-defined clones. Our results suggest that BBS1 is required for suppression of mesenchymal cell identities as IMCD3 cell passage number increases. This was associated with a failure to express epithelial cell markers and tight junction formation, which was variable amongst clones. Transcriptomic analysis of hypothalamic preparations from BBS mutant mice, and BBS patient fibroblasts, suggested that dysregulation of epithelial-to-mesenchymal transition (EMT) genes is a general predisposing feature of BBS across tissues. Collectively this work suggests that the dynamic stability of the BBSome is essential for suppression of mesenchymal cell identities as epithelial cells differentiate.

genetics↗

bioRxiv2022

Immunotherapy of glioblastoma explants induces interferon-γ responses and immune cell rearrangements in tumor center, but not periphery

Recent therapeutic strategies for glioblastoma (GBM) aim at targeting immune tumor microenvironment (iTME) components to induce antitumoral immunity. A patient-tailored, ex vivo drug testing and response analysis platform for GBM would facilitate personalized therapy planning, provide insights into treatment-induced immune mechanisms in the iTME, and enable the discovery of biomarkers of therapy response and resistance. We cultured 47 GBM explants from tumor center and periphery from 7 patients in perfusion bioreactors to assess iTME responses to immunotherapy. Explants were exposed to antibodies blocking the immune checkpoints CD47, PD-1 or or their combination, and were analyzed by highly multiplexed microscopy (CODEX, co-detection by indexing) using an immune-focused 55-marker panel. Culture media were examined for changes of soluble factors including cytokines, chemokines and metabolites. CODEX enabled the spatially resolved identification and quantification of >850,000 single cells in explants, which were classified into 10 cell types by clustering. Explants from center and periphery differed significantly in their cell type composition, their levels of soluble factors, and their responses to immunotherapy. In a subset of explants, culture media displayed increased interferon-{gamma} levels, which correlated with shifts in immune cell composition within specific tissue compartments, including the enrichment of CD4+ and CD8+ T cells within an adaptive immune compartment. Furthermore, significant differences in the expression levels of functional molecules in innate and adaptive immune cell types were found between explants responding or not to immunotherapy. In non-responder explants, T cells showed higher expression of PD-1, LAG-3, TIM-3 and VISTA, whereas in responders, macrophages and microglia showed higher cathepsin D levels. Our study demonstrates that ex vivo immunotherapy of GBM explants enables an active antitumoral immune response within the tumor center in a subset of patients, and provides a framework for multidimensional personalized assessment of tumor response to immunotherapy.

cancer biology↗

bioRxiv2021

Angiopoietin 1 and integrin beta 1b are vital for zebrafish brain development

This study aimed at identifying the role of angiopoietin 1 (angpt1) in brain development, the mode of action of angpt1, and the main targets in the zebrafish brain. We investigated embryonic brain angiogenesis and neural development in the angpt1sa14264, itgb1bmi371, tekhu1667 mutant fish, and the effects of transgenic overexpression of angpt1 in the larval brain. Lack of angpt1 was associated with downregulation of tek and upregulation of itgb1b. We found deficiencies in the patterning of proliferation, the vascular network and reticulospinal neurons in the hindbrain, and selective deficiencies in specific neurotransmitter systems. In the angpt1sa14264 and itgb1bmi371 larval brains, using microangiography, retrograde labeling, and immunostaining, we demonstrated that the targeted destruction of angpt1sa14264 and itgb1bmi371 mutant fish caused severe irregular cerebrovascular development, aberrant hindbrain patterning, downregulation of neural proliferation, expansion of the radial glial progenitors, deficiencies of dopaminergic, histaminergic, and GABAergic populations in the larval brain. In contrast, the tekhu1667 mutants regularly grew with no such apparent phenotypes. Neurally overexpressed angpt1 promoted opposite effects by increasing the vascular branching, increasing cell proliferation, and neuronal progenitors. Notably, zebrafish angpt1 showed neurogenic activity independent of its typical receptor tek, indicating the novel role of a dual regulation by angpt1 in embryonic neurogenesis and angiogenesis in zebrafish. The results show that angpt1 and its interaction with itgb1b are crucial in zebrafish brain neuronal and vascular development and suggest that angpt1 through itgb1b can act as a neurogenic factor in the neural proliferation fate in the developing brain.

neuroscience↗

bioRxiv2021

Delta C2 Domain β1-2 loop contributes to robust Notch signalling

Accurate Notch signalling is critical for organism development and homeostasis. Fine-tuning of Notch-ligand interactions have substantial impact on signalling-outputs. Recent structural studies identified a conserved N-terminal C2 domain in human Notch ligands which conferred phospholipid binding in vitro. Here we show that Drosophila ligands adopt the same C2 domain structure with analogous variations in the loop regions, including the so-called {beta}1-2 loop that has been associated with phospholipid binding. Mutations in the {beta} 1-2 loop of Delta C2 domain retain Notch binding but have impaired ability to interact with phospholipids in vitro. To investigate its role in vivo we deleted five residues within the {beta} 1-2 loop of endogenous Delta by CRISPR/Cas9 gene editing. Strikingly, this change compromised ligand function. The modified Delta enhanced phenotypes produced by Delta loss of function alleles and suppressed that of Notch alleles. As the modified protein was present on the cell surface in normal amounts, these results argue that C2 domain phospholipid-binding is necessary for robust signalling in vivo where the balance of cis and trans ligand-receptor interactions is finely tuned.

developmental biology↗

bioRxiv2020

Improved 7 Tesla Transmit Field Homogeneity with Reduced Electromagnetic Power Deposition Using Coupled Tic Tac Toe Antennas

Recently cleared by the FDA, 7 Tesla (7T) MRI is a rapidly growing technology that can provide higher resolution and enhanced contrast in human MRI images. However, the increased operational frequency (~297 MHz) hinders its full potential since it causes inhomogeneities in the images and increases the power deposition in the tissues. This work describes the optimization of an innovative radiofrequency (RF) head coil coupled design, named Tic Tac Toe, currently used in large scale human MRI scanning at 7T; to date, this device was used in more than 1,300 patient/volunteer neuro 7T MRI scans. Electromagnetic simulations were performed for each of the coils antennas using the finite-difference time-domain method. Numerical optimizations were used to combine the calculated electromagnetic fields produced by these antennas, based on the superposition principle, and successfully produced homogeneous magnetic field distributions at low levels of power deposition in the tissues. The simulations were then successfully validated in-vivo using the Tic Tac Toe RF head coil system on a 7T MRI scanner.

bioengineering↗

bioRxiv2020

Twists and turns of the salicylate catabolism in Aspergillus terreus

In fungi, salicylate catabolism was believed to proceed only through the catechol branch of the 3-oxoadipate pathway, as shown e.g. in Aspergillus nidulans. However, the observation of a transient accumulation of gentisate upon cultivation of Aspergillus terreus in salicylate media questions this concept. To address this we have run a comparative analysis of the transcriptome of these two species after growth in salicylate using acetate as a control condition. The results revealed the high complexity of the salicylate metabolism in A. terreus with the concomitant positive regulation of several pathways for the catabolism of aromatic compounds. This included the unexpected joint action of two pathways: the nicotinate and the 3-hydroxyanthranilate, possibly crucial for the catabolism of aromatics in this fungus. New genes participating in the nicotinate metabolism are here proposed, whereas the 3-hydroxyanthranilate catabolic pathway in fungi is described for the first time. The transcriptome analysis showed also for the two species an intimate relationship between salicylate catabolism and secondary metabolism. This study emphasizes that the central pathways for the catabolism of aromatic hydrocarbons in fungi hold many mysteries yet to be discovered. IMPORTANCEAspergilli are versatile cell factories used in industry for production of organic acids, enzymes and pharmaceutical drugs. To date, organic acids bio-based production relies on food substrates. These processes are currently being challenged to switch to renewable non-food raw materials; a reality that should inspire the use of lignin derived aromatic monomers. In this context, Aspergilli emerge at the forefront of future bio-based approaches due to their industrial relevance and recognized prolific catabolism of aromatic compounds. Notwithstanding considerable advances in the field, there are still important knowledge gaps in the central catabolism of aromatic hydrocarbons in fungi. Here, we disclosed a novel central pathway, defying previous established ideas on the central metabolism of the aromatic amino acid tryptophan in Ascomycota. We also observed that the catabolism of the aromatic salicylate greatly activated the secondary metabolism, furthering the significance of using lignin derived aromatic hydrocarbons as a distinctive biomass source.

genetics↗

bioRxiv2019

Self-supervised retinal thickness prediction enables deep learning from unlabeled data to boost classification of diabetic retinopathy

Access to large, annotated samples represents a considerable challenge for training accurate deep-learning models in medical imaging. While current leading-edge transfer learning from pre-trained models can help with cases lacking data, it limits design choices, and generally results in the use of unnecessarily large models. We propose a novel, self-supervised training scheme for obtaining high-quality, pre-trained networks from unlabeled, cross-modal medical imaging data, which will allow for creating accurate and efficient models. We demonstrate this by accurately predicting optical coherence tomography (OCT)-based retinal thickness measurements from simple infrared (IR) fundus images. Subsequently, learned representations outperformed advanced classifiers on a separate diabetic retinopathy classification task in a scenario of scarce training data. Our cross-modal, three-staged scheme effectively replaced 26,343 diabetic retinopathy annotations with 1,009 semantic segmentations on OCT and reached the same classification accuracy using only 25% of fundus images, without any drawbacks, since OCT is not required for predictions. We expect this concept will also apply to other multimodal clinical data-imaging, health records, and genomics data, and be applicable to corresponding sample-starved learning problems.

bioinformatics↗