Biology subjects

Samitier, J.

Publications and source records attributed to Samitier, J..

8 recordsLinked to original sources

bioRxiv2026

Computationally guided design of a metastasis-on-a-chip platform for quantitative evaluation of chemotactic cues in developmental cancers

Metastatic dissemination is initiated by tumor cells interpreting spatially organized biochemical and biophysical cues that remain difficult to reproduce using conventional migration assays. Here, we developed a computationally guided metastasis-on-a-chip (MET-on-a-chip) platform based on the concept of the Minimally Functional Unit (MFU), in which only the biological components required to answer a defined experimental question are incorporated. The platform consists of two independent culture chambers connected through an array of confined microchannels that permits diffusion of soluble factors while constraining tumor cell migration. Rather than relying on empirical optimization, finite-element COMSOL simulations were first used to predict molecular transport, define growth factor loading conditions, identify biologically relevant exposure regions, and guide the rational design of the microfluidic assay. Computational predictions were experimentally validated using 70-kDa FITC-dextran diffusion and VEGF release studies, confirming the formation of stable spatial concentration gradients across the microfluidic platform. The simulations further demonstrated that both growth factor loading and cell positioning relative to the predicted gradients critically influenced assay performance, leading to the optimization of the platform through spatial reconfiguration of the tumor compartment. Using the optimized configuration, we compared the migratory responses of neuroblastoma, Ewing sarcoma, and osteosarcoma cells to vascular (VEGF-A165) and lymphatic (VEGF-C) chemotactic cues. VEGF-C significantly increased migration through the microchannel array in Ewing sarcoma and osteosarcoma cells, whereas VEGF-A165 produced no significant effect. In contrast, neuroblastoma cells exhibited minimal migration under either condition, revealing tumor-specific differences in responsiveness to VEGF signaling. Together, these findings establish a computationally guided workflow for the rational design of metastasis-on-a-chip assays, in which predictive modeling informs experimental design before biological validation. By substantially reducing empirical trial-and-error while enabling quantitative control over growth factor exposure, this strategy provides a robust framework for developing minimally functional microphysiological systems capable of dissecting individual steps of the metastatic cascade under experimentally defined conditions. Translational Impact StatementMetastatic dissemination remains one of the greatest clinical challenges in pediatric oncology, yet experimental models capable of quantitatively evaluating early migratory events remain limited. The computationally guided MET-on-a-Chip workflow presented here provides a human-relevant platform in which soluble microenvironmental cues can be systematically investigated under controlled and predictive conditions. Although demonstrated here using VEGF-A165 and VEGF-C, the platform can be readily adapted to study virtually any chemotactic factor, cytokine, extracellular vesicle population, or therapeutic candidate involved in metastatic dissemination. The modular MFU design allows biological complexity to be incorporated progressively as dictated by the scientific question, providing a flexible framework for future applications. In the longer term, this workflow could be combined with patient-derived tumor cells, organoids, or biopsy material to investigate patient-specific metastatic behavior and evaluate anti-metastatic therapeutic strategies in a personalized setting. Beyond identifying pro-migratory signaling pathways, the platform may serve as a preclinical tool to prioritize compounds capable of preventing tumor cell dissemination before evaluation in more complex animal models or clinical studies.

bioengineering↗

bioRxiv2026

3D neuroblastoma models expose divergent responses to magnetic hyperthermia and photothermal therapy

PurposeHyperthermia is emerging as an adjunct strategy in pediatric oncology, yet its translation is limited by poor understanding of how different modalities impact complex tumor microenvironments. Neuroblastoma (NB), the most common extracranial solid tumor in children, displays profound heterogeneity that hampers therapeutic predictability. Here, we performed the first systematic head-to-head comparison of photothermal therapy (PTT) and magnetic hyperthermia (MH) in tissue-engineered NB (TE-NB) models. MethodsTE-NB scaffolds incorporating five NB cell lines were loaded with magnetic nanoparticles (MNPs) and subjected to PTT (808-nm laser, 130 W/cm2, 10 min) or MH (285 kHz, 20 mT, 60 min). Constructs were analyzed at 24 h, 48 h, and 5 d post-treatment for DNA content, cell viability, proliferation (Ki67 immunohistochemistry), and apoptosis (caspase-3/7 staining). ResultsMH produced consistent MNP-dependent heating with minimal background, while PTT was dominated by nonspecific medium absorption. Both modalities modulated proliferation within 24 h, but effects varied sharply by cell line and scaffold region, reflecting microenvironmental heterogeneity. By 48 h, PTT often triggered paradoxical increases in proliferation, whereas MH disrupted scaffold integrity, reduced DNA content, and suppressed Ki67 expression. Notably, neither modality induced sustained caspase-3/7 activation, indicating that cytotoxicity proceeds via non-apoptotic pathways. ConclusionOur findings position MH as a superior modality for uniform heating and proliferation control in 3D NB models, but also highlight that hyperthermia should be considered a context-dependent modulator rather than a binary cytotoxic agent. By integrating patient-specific TE-NB platforms, "precision hyperthermia" could define individualized therapeutic windows, optimize combinations with pro-apoptotic or immunomodulatory agents, and accelerate translation of hyperthermia strategies for children with NB.

bioengineering↗

bioRxiv2024

Cell-derived matrices mechanics as a functional read-out in Collagen VI-related Congenital Muscular Dystrophies

Atomic force microscopy-force spectroscopy (AFM-FS) applied to biological samples can provide information on the micro- and nanoscale mechanical properties of tissues, which are a determinant of cellular behavior. Mechanical properties are largely determined by the structure and composition of the extracellular matrix (ECM), for which several in vitro models have been developed, including cell-derived matrices (CDMs). CDMs are decellularized natural ECMs assembled by cells, that closely mimic the in vivo stromal fiber organization and molecular content. Here we applied AFM-FS to evaluate the nanomechanical properties of CDMs obtained from primary skin fibroblast cultures of patients affected by collagen VI-related congenital muscular dystrophies (COL6-RDs). COL6-RDs are a set of neuromuscular conditions caused by pathogenic variants in any of the three major COL6 genes, which result in deficiency or dysfunction of the COL6 incorporated into the ECM of connective tissues. Current diagnosis includes the genetic confirmation of the disease and categorization of the phenotype based on maximum motor ability, as no direct correlation exists between genotype and phenotype of COL6-RDs. The development of new tools able to identify phenotype traits can significantly contribute to the diagnosis and prognosis of COL6-RDs. We describe differences in the elastic modulus (E) among CDMs deriving from patients with different clinical phenotypes, as well as the restoration of E in CDMs obtained from genetically edited cells. Results anticipate the potential of the nanomechanical analysis of CDMs as a complementary clinical tool, providing phenotypic information about COL6-RDs and their response to gene therapies.

biophysics↗

bioRxiv2022

BBB-on-a-chip with Integrated micro-TEER for permeability evaluation of multi-functionalized gold nanorods against Alzheimer's disease

BackgroundThe lack of predictive models that mimic the blood-brain barrier (BBB) hinders the development of effective drugs for neurodegenerative diseases. Animal models behave differently from humans, are expensive and have ethical constraints. Organ-on-a-chip (OoC) platforms offer several advantages to resembling physiological and pathological conditions in a versatile, reproducible, and animal-free manner. In addition, OoC give us the possibility to incorporate sensors to determine cell culture features such as trans-endothelial electrical resistance (TEER). Here, we developed a BBB-on-a-chip (BBB-oC) platform with a TEER measurement system in close distance to the barrier used for the first time for the evaluation of the permeability performance of GNR-PEG-Ang2/D1 for Alzheimers disease. GNR-PEG-Ang2/D1 is a therapeutic nanosystem previously developed by us consisting of gold nanorods (GNR) functionalized with polyethylene glycol (PEG), angiopep-2 peptide (Ang2) to overcome the BBB and the D1 peptide as beta amyloid fibrillation inhibitor, finally obtaining GNR-PEG-Ang2/D1 which showed to be useful for disaggregation of the amyloid in in vitro and in vivo models. In this work, we evaluated its cytotoxicity, permeability, and some indications of its impact on the brain endothelium by employing an animal-free device based on neurovascular human cells. ResultsIn this work, we fabricated a BBB-oC with human astrocytes, pericytes and endothelial cells and a TEER measuring system (TEER-BBB-oC) integrated at a micrometric distance of the endothelial barrier. The characterization displayed a neurovascular network and the expression of tight junctions in the endothelium. We produced GNR-PEG-Ang2/D1 and determined its non-cytotoxic range (0,05-0,4 nM) for plated cells included in the BBB-oC and confirmed its harmless effect at the highest concentration (0.4 nM) in the microfluidic device. The permeability assays revealed that GNR-PEG-Ang2/D1 cross the BBB and this entry is facilitated by Ang2 peptide. Parallel to the permeability analysis of GNR-PEG-Ang2/D1, an interesting behavior of the TJs expression was observed after its administration probably related to the ligands on the nanoparticle surface. ConclusionBBB-oC with TEER integrated setup was proven as a functional and throughput platform to evaluate the brain permeability performance of nanotherapeutics in a physiological environment with human cells, putting forward a viable alternative to animal experimentation.

bioengineering↗

bioRxiv2022

Substrate adhesion determines migration during mesenchymal cell condensation in chondrogenesis

Effective cartilage development relies on the successful formation of mesenchymal cell condensates. Mesenchymal condensation is a prevalent morphogenetic transition, which involves the upregulation of the adhesive extracellular glycoprotein fibronectin (FN). During condensation, there is an active directional migration of cells from the surrounding loose mesenchyme towards regions of increasing matrix adherence (the condensation centers). In this study, we live imaged the first 40 h of mesenchymal condensation during chondrogenesis on nanopatterns of the cell-adhesive peptide arginine-glycine-aspartic acid (RGD), present in FN. Results show cell-substrate adhesions modulate both single-cell and collective cell migration during mesenchymal condensation. Single cell tracking analysis showed that substrate adhesion determines the migration mode, protrusion formation and the directionality of the cell movement. Cells on the more adhesive nanopatterns presented traits among amoeboid and mesenchymal modes of migration facilitating a more directional movement and reducing contact inhibition of locomotion (CIL), which allows merging and condensation. Inhibition experiments demonstrated that neural cadherin (N-Cad) is required in cell-cell interactions, enabling cells to coordinate their movement and directionality in a multicellular environment and to maintain the group cohesiveness during migration. Altogether, this contributes to create a sufficiently dynamic scenario, in which there is a balance between cell-substrate and cell-cell adhesions for condensates to grow. Our results provide a framework for the regulation of single and collective cell migration during mesenchymal condensation, through nanoscale cell-substrate adherence. Summary statementThe fine tuning of substrate adherence through nanopatterning allows control of mesenchymal cell migration and determines condensation during chondrogenesis in vitro.

biophysics↗

bioRxiv2021

Substrate ligand density modulates gap junction intercellular communication during mesenchymal cell condensation

To unveil the influence of cell-matrix adhesions in the establishment of gap junction intercellular communication (GJIC) during cell condensation in chondrogenesis. Materials & MethodsPreviously developed nanopatterns of the cell adhesive ligand arginine-glycine-aspartic acid (RGD) were used as cell culture substrates to control cell adhesion at the nanoscale. We conducted in vitro chondrogenesis of mesenchymal stem cells on the nanopatterns. We evaluated cohesion and GJIC in cell condensates. ResultsMechanical stability and GJIC are enhanced by a nanopattern configuration in which 90% of the surface area presents adhesion sites separated less than 70 nm, thus providing an onset for cell signaling. ConclusionCell-matrix adhesions regulate GJIC of mesenchymal cell condensates during in vitro chondrogenesis from a threshold configuration at the nanoscale.

bioengineering↗

bioRxiv2020

Real-time Ratiometric Imaging of Micelles Assembly State in a Microfluidic Cancer-on-a-chip

The performance of supramolecular nanocarriers as drug delivery systems depends on their stability in the complex and dynamic biological media. After administration, nanocarriers are challenged by confronting different barriers such as shear stress and proteins present in blood, endothelial wall, extracellular matrix and eventually cancer cell membranes. While early disassembly will result in a premature drug release, extreme stability of the nanocarriers can lead to poor drug release and low efficiency. Therefore, comprehensive understanding of the stability and assembly state of supramolecular carriers in each stage of delivery is a key factor for the rational design of these systems. One of the key challenges is that current 2D in vitro models do not provide exhaustive information, as they do not fully recapitulate the 3D tumor microenvironment. This deficiency of the 2D models complexity is the main reason for the differences observed in vivo when testing the performance of supramolecular nanocarriers. Herein, we present a real-time monitoring study of self-assembled micelles stability and extravasation, combining spectral confocal microscopy and a microfluidic tumor-on-a-chip. The combination of advanced imaging and a reliable organ-on-a-chip model allow us to track micelle disassembly by following the spectral properties of the amphiphiles in space and time during the crucial steps of drug delivery. The spectrally active micelles were introduced under flow and their position and conformation followed during the crossing of barriers by spectral imaging, revealing the interplay between carrier structure, micellar stability and extravasation. Integrating the ability of the micelles to change their fluorescent properties when disassembled, spectral confocal imaging and 3D microfluidic tumor blood vessel-on-a-chip, resulted in the establishment of a robust testing platform, suitable for real-time imaging and evaluation of supramolecular drug delivery carriers stability.

bioengineering↗

bioRxiv2020

Sequential combinations of chemotherapeutic agents with BH3 mimetics to treat rhabdomyosarcoma and avoid resistance.

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in childhood and adolescence. Refractory/relapsed RMS patients present a bad prognosis that combined with the lack of specific biomarkers difficult the development of new therapies. We here utilize dynamic BH3 Profiling (DBP), a functional predictive biomarker that measures net changes in mitochondrial apoptotic signaling, to identify anti-apoptotic adaptations upon treatment. We use this information to guide the use of BH3 mimetics to specifically inhibit BCL-2 pro-survival proteins, defeat resistance and avoid relapse to therapy. Indeed, we found that BH3 mimetics that selectively target BCL-xL and MCL-1 synergistically enhance the effect of the clinically used chemotherapeutic agents vincristine and doxorubicin in RMS cells. We validated this strategy in vivo using a RMS patient-derived xenograft (PDX) model and observed a reduction on tumor growth with a tendency to its stabilization with the sequential combination of vincristine and the MCL-1 inhibitor S63845. Finally, we identified the molecular mechanism by which RMS cells acquire resistance to vincristine: through the anti-apoptotic protein MCL-1 for which we observed an enhanced binding between MCL-1 and BID after drug exposure, which is suppressed by the sequential addition of S63845. In conclusion, our findings validate the use of DBP as a functional assay to predict treatment effectiveness in RMS and provide a rationale for BH3 mimetic combination with chemotherapeutic agents to avoid tumor resistance, improve treatment efficiency and decrease undesired secondary effects.

cancer biology↗