Biology subjects

Martins, A. M.

Publications and source records attributed to Martins, A. M..

4 recordsLinked to original sources

bioRxiv2026

Coverslip Hypoxia High-Content Screening (CH-HCS): A 2D Imaging Platform for Spatially Resolved Analysis of CAR-T Function Under Oxygen Gradients

Hypoxia within the tumor microenvironment profoundly limits the efficacy of immune and cellular therapies, yet most in vitro cytotoxicity assays neglect spatial oxygen heterogeneity. We developed Coverslip Hypoxia High-Content Screening (CH-HCS), a simple, scalable 2D co-culture platform that enables quantitative, region-resolved evaluation of CAR-T cell activity across controlled oxygen gradients within a single well. In CH-HCS, a 5 mm glass coverslip placed over tumor-stroma co-cultures restricts oxygen diffusion, generating concentric hypoxia-normoxia zones in standard 96-well plates. Fluorescently labeled CD19+ Raji or CD19- K562 tumor cells, stromal cells, and anti-CD19 CAR-T cells were analyzed using multiparametric fluorescence imaging with an ImageXpress Micro XLS system coupled to custom CellProfiler-KNIME pipelines, enabling segmentation of spatial Regions of Interest--InnerCore, OuterCore, Periphery, and Outside--and single-cell quantification of tumor death (SYTOX Green) and T-cell morphodynamics. The platform reproducibly established oxygen gradients that strongly shaped cellular behavior: CAR-T cytotoxicity and motility were maximal in normoxic regions but markedly suppressed within hypoxic cores, whereas effector cell survival increased under low oxygen. Unlike bulk cytotoxicity assays, CH-HCS directly visualizes spatial functional heterogeneity within the same well, allowing simultaneous comparison of matched hypoxic and normoxic compartments. Together, CH-HCS provides a cost-effective, high-throughput, and physiologically relevant tool for preclinical screening of CAR-T products and therapeutic strategies aimed at overcoming hypoxia-driven immune resistance at the tumor-stroma interface. Key PointO_LISpatially resolved quantification: Simultaneous measurement of cellular behavior in matched hypoxic and normoxic compartments within the same well. C_LIO_LIPhysiologically relevant gradients: Coverslip geometry generates reproducible oxygen diffusion profiles that emulate the in vivo tumor-stroma interface. C_LIO_LIMultiparametric single-cell readouts: High-content imaging coupled with automated segmentation provides data on viability, cytotoxicity, and effector morphology. C_LIO_LIHigh-throughput and low-cost: Fully compatible with 96-well formats, enabling parallel pharmacological or genetic screening without specialized hypoxia chambers. C_LIO_LIGeneralizable design: Applicable beyond CAR-T assays to study stromal adaptation, drug resistance, or immune suppression across controlled oxygen gradients. C_LI

cancer biology↗

bioRxiv2025

Dormant viral pathways underlie space-induced neural senescence: a neuroprotective strategy for spaceflight and neurological diseases

Long-duration spaceflight is associated with neurological symptoms in astronauts, yet the underlying molecular mechanisms remain unclear. Using human brain organoids cultured aboard the International Space Station, we analyzed three independent spaceflights to demonstrate that exposure to the space environment triggers Space-Induced Neural Senescence (SINS), characterized by chromatin remodeling, mitochondrial dysfunction, and activation of viral-like transcriptional programs in the absence of infection. Multi-omics analyses identified upregulation of endogenous LINE-1 (L1) retroelements, whose activity was markedly enhanced in organoids lacking MECP2, a known L1 repressor implicated in Rett syndrome. The resulting accumulation of cytoplasmic L1 DNA elicited an IL-6-mediated inflammatory and neurotoxic response, which was reversed by reverse transcriptase inhibitors (RTi) such as lamivudine or stavudine. Parallel preclinical experiments in Mecp2-deficient mice confirmed that RTi treatment restored neuronal morphology, synaptogenesis, function, cognition, and survival. These findings reveal that the space environment reactivates dormant genomic retroelements, providing an unexpected mechanistic insight into astronaut neurobiology and identifying a potential therapeutic strategy for both space-induced and terrestrial neurological conditions. Our pioneering study demonstrates the value of space-enabling research in accelerating drug discovery and disease treatment on Earth.

neuroscience↗

bioRxiv2025

Patient-derived organoids to study glycosylation dynamics during gastric disease

BACKGROUND AND AIMSAberrant cellular glycosylation remains a key event that accompanies and actively sustains gastric neoplastic transformation. Patient-derived organoids (PDOs) have recently emerged as a promising ex vivo model to study human gastric disorders. Since the PDOs glycosylation landscape remains unknown, this study aims to evaluate PDOs as potential avatars of in vivo tissue glycosylation profiles in the gastric context. METHODSFresh gastric mucosa samples derived from non-tumoral obese patients (n=11), adjacent tumor mucosa samples (n=29), and tumor tissue samples derived from gastric cancer (GC) patients (n=30) were used to establish a biobank of gastric PDOs (n=56). The N- and O-glycophenotypes of normal, adjacent, and tumor PDOs and respective in vivo tissues were thoroughly characterized by immunostaining. Additionally, a comparative glycan analysis was performed over time, upon PDO biobanking and xenografting in mice. The binding of two Helicobacter pylori (H. pylori) isogenic strains with distinct glycan-binding affinities was assessed in parental gastric mucosa tissues and compared with the respective PDOs before and after modulation of their glycan landscape. RESULTSOur results show that PDOs mimic different phenotypes of the carcinogenic cascade and recapitulate parental gastric tissues glycosylation profile. Tumor PDOs recapitulate the inter- and intra-heterogeneity features observed in GC, which is maintained over time, upon biobanking and xenografting. We demonstrated that the expression of type I and type II Lewis antigens is dynamically controlled by PDOs differentiation status, which results in differential binding to H. pylori strains displaying distinct glycan-binding adhesins, mirroring the gastric epithelium tissue interactions. CONCLUSIONSThis study established PDOs as invaluable ex vivo tools to study the complex glycan dynamics in both gastric physiological and pathological settings.

cell biology↗

bioRxiv2023

B12-dependent virioplankton demonstrate interseasonal dynamics and associate with a diversity of pelagic bacterioplankton

Through infection and lysis of their coexisting bacterial hosts, viruses impact the biogeochemical cycles sustaining globally significant pelagic oceanic ecosystems. Currently, little is known of the ecological interactions between lytic viruses and their bacterial hosts underlying these biogeochemical impacts at ecosystem scales. This study focused on populations of lytic viruses carrying the B12- dependent Class II monomeric ribonucleotide reductase (RNR) gene, ribonucleotide triphosphate reductase (RTPR), documenting seasonal changes in pelagic virioplankton and bacterioplankton using amplicon sequences of RTPR and the 16S rRNA gene, respectively. Amplicon sequence libraries were analyzed using compositional data analysis tools that account for the compositional nature of these data. Both virio- and bacterioplankton communities responded to environmental changes typically seen across seasonal cycles as well as shorter term upwelling-downwelling events. Defining RTPR-carrying viral populations according to major phylogenetic clades proved a more robust means of exploring virioplankton ecology than operational taxonomic units defined by percent sequence homology. Virioplankton RTPR populations showed positive associations with a broad phylogenetic diversity of bacterioplankton including dominant taxa within pelagic oceanic ecosystems such as Prochlorococcus and SAR11. Temporal changes in RTPR-virioplankton, occurring as both free viruses and within infected cells, indicated possible viral-host pairs undergoing sustained infection and lysis cycles throughout the seasonal study. Phylogenetic relationships inferred from RTPR sequences mirrored ecological patterns in virio- and bacterioplankton populations demonstrating possible genome to phenome associations for an essential viral replication gene.

ecology↗