In the absence of a reliable cure, Osteoarthritis (OA), treatment targets symptomatic pain and inflammation. However, chronically administered drugs often result in adverse side effects. In order to develop an improved treatment, our studies focused on controlled dual drug delivery. Bupivacaine (BUP), a local anesthetic, is used to manage pain directly and dexamethasone (DEX), a glucocorticoid, reduces inflammation. Key obstacles of both BUP and DEX administration, is that at high concentrations both drugs may be toxic, while at low concentrations they can provide pain relief and control inflammation for only a short time. We developed nanoparticles composed of Poly(lactic- co-glycolic acid) (PLGA), a biodegradable polymer, to prolong the release of lower doses of BUP and DEX for at least 30 days. Using the nanoparticle (NP) diffusion profiles, loading capacity, and optimal dose for our target cell types, we tested our dual NP therapy on chondrocytes, macrophages, and in baseline and activated co-cultures in order to compare bolus and NP drug effects either alone or in combination. We found that the combined nanoparticle co-therapy maintained cell viability and reduced inflammation more than either DEX nanoparticle (DexNP) or BUP nanoparticle (BupNP) therapy alone, especially in co-culture, and can therefore ultimately be incorporated into longer term therapeutic protocols. Pathway analysis enabled an approach to understanding the potential synergy between the two drugs. Overall, the macrophage, chondrocyte, and co-culture studies indicate that DualNP therapy maintains viability while reducing IL-8 secretion more than either DexNP, BupNP, and bolus DUAL in all conditions. While further analysis is needed, due to its versatility, our approach can be further developed to potentially improve an array of clinical outcomes.
Fluorescence lifetime imaging microscopy (FLIM) is an advanced microscopy technique that enables label-free, non-destructive optical metabolic imaging (OMI) of biological materials at spatiotemporal resolution. Accordingly, FLIM has emerged as a widely used platform for multi-scale characterization of the tumor microenvironment (TME), providing valuable fundamental insights into cancer biology and demonstrating great promise for clinical utility. The surrounding host tissue, or peritumor microenvironment (pTME), has received considerably less attention in this area, however, despite being recognized as a molecularly distinct tissue region compared to both malignant and normal tissue. In this Review, we describe the application of FLIM for monitoring the metabolic TME and its relatively minimal use in probing the pTME. Notably, within the context of the TME, the ability of FLIM to exploit metabolic heterogeneity has been extensively leveraged to characterize diverse tumor-associated and malignant cell populations and states, perform drug screening and monitoring in vitro and in vivo, and non-invasively distinguish tumor from tumor-adjacent tissue regions. By comparison, only a handful of studies have utilized FLIM (mostly at the in vitro scale) to explore and analyze the pTME, with these studies suggesting that tumor-peritumor cross-talk promotes metabolic changes in peritumoral cell types. Furthermore, the biological significance of the peritumor is frequently overlooked in most diagnostic studies, as it is frequently classified as normal rather than treated as a distinct, potentially tumor-supportive compartment. Thus, evaluating the contribution and clinical relevancy of the pTME to tumor progression and treatment response represents an exciting, yet not fully realized, opportunity for FLIM. Owing to its successful and extensive application in interrogating the TME, we propose that the demonstrated capabilities of FLIM can and should be readily extended to the pTME. Doing so has the potential to advance the field, both in our fundamental understanding of cancer progression and our efforts to improve patient outcomes.
Lymphocyte recruitment to inflamed intestines in inflammatory bowel disease (IBD) critically depends on shearstress-regulated integrin α4β7/MAdCAM-1 interactions. However, the mechano-chemical coupling between the chemokine CCL25 and the intracellular adaptor Kindlin-3 in activating α4β7 under physiological flow remains unclear. Utilizing parallel-plate flow chamber assays combined with fluorescence microscopy and molecular perturbation, wedemonstrate that CCL25 triggers robust integrin α4β7 activation and stable lymphocyte adhesion only under physiological shear stress, a process regulated by Kindlin-3. Mechanistically, Kindlin-3 acts as a force transmission hub, enabling CCL25 signaling to enhance α4β7-mediatedadhesion strengthening. Knockdown of Kindlin-3 specifi cally enhanced the force-sensitized, CCL25-induced activationof α4β7. This study reveals, for the first time, that Kindlin-3 is crucial for translating chemokine (CCL25) signals intobiomechanical activation of integrin α4β7 under shear flow. Our findings establish Kindlin-3 as a critical mechano-regulator of chemokine-induced integrin activation during lymphocyte homing, providing novel mechanistic insightsinto IBD pathogenesis and highlighting this pathway as a promising target for anti-adhesion therapeutics.
Increased extracellular matrix stiffness is a defining mechanical feature of solid tumors, yet how it regulates extracellular vesicle-mediated intercellular communication remains poorly understood in three-dimensional tumor microenvironments. Here, we demonstrate that ECM stiffness mechanistically regulates extracellular vesicle (EV) cargo loading in oral squamous cell carcinoma spheroids. Using a tunable three-dimensional spheroid culture platform, we show that increased matrix stiffness enriches tumorigenic and metastatic non-coding RNA transcripts in EVs. At a functional level, stiffness-primed EVs influence recipient spheroid growth by modulating proliferation and apoptosis. Notably, our study reveals that EVs are enriched in parental biomolecular cargo, including the mechanosensitive Piezo1 ion channel and adhesion and stemness molecule CD44. Protein expression and small RNA sequencing analyses confirm the incorporation of these components into spheroid-derived EVs in a stiffness-independent manner. Together, our findings identify ECM stiffness as a mechanistic regulator of EV composition and establish EVs as biomechanical signaling vectors that further influence cell proliferation in three-dimensional microenvironments.
NF2-related schwannomatosis (NF2-SWN) is a devastating genetic disorder characterized by the development of bilateral vestibular schwannomas (VSs), which are histologically benign tumors that cause hearing loss and vestibular dysfunction. Currently, there are no FDA-approved pharmacologic therapies for VS or VS-associated hearing loss, representing a major unmet medical need for patients with NF2-SWN. Although immunotherapy has transformed the treatment of many solid malignancies, its efficacy in VS has not been comprehensively evaluated. We employed VS mouse models that faithfully recapitulate tumor-induced hearing loss and ataxia to evaluate the therapeutic efficacy of anti-PD-1 (αPD-1) immunotherapy alone and in combination with losartan, an angiotensin II type 1 receptor blocker known to reduce tumor extracellular matrix density and normalize tumor vascular perfusion. Tumor growth, survival, hearing function, and neurological outcomes were assessed. Losartan significantly enhanced the intratumoral delivery of αPD-1 antibody and increased immune effector cell infiltration in VS. Consequently, combined losartan and αPD-1 therapy more effectively inhibited tumor growth and prolonged survival compared with monotherapy. In parallel, losartan attenuated inflammatory signaling, thereby reducing neuro-edema. As a result, the combination treatment more effectively prevented tumor-induced hearing loss and alleviated ataxia in the VS mouse model. Targeting the fibrotic tumor microenvironment with losartan sensitizes vestibular schwannomas to immunotherapy and improves neurological outcomes. These findings provide critical preclinical evidence supporting a novel combination strategy using ICIs for NF2-SWN and offer important insights for the design of future clinical trials aimed at suppressing VS growth while preserving hearing and neurological function.
Cartilage is an avascular tissue with a limited capacity for self-regeneration. Traditional autologous cartilage transplantation is incapable of fulfilling the increasing demand for repair of various cartilage tissue lesions. The advent of 3D bioprinting technology provides an opportunity to repair articular cartilage defects through the construction of organized living structures composed of biomaterials and cells. This technology can mimic natural cartilage by allowing control of cell distribution, and the modulation of mechanical and chemical properties with high precision. This review provides insight into the current developments in 3D bioprinting for cartilage tissue engineering. Recent studies on 3D-bioprinted cartilage tissue constructs and various bioinks are highlighted. The advantages and limitations of commonly used natural, synthetic, and composite bioinks in terms of printability, mechanical properties, and biological performance in bioprinting of anatomically shaped constructs (nasal, auricle, knee, and tracheal cartilage) are discussed. Furthermore, outlooks and challenges of 3D bioprinting of complex constructs with variable mechanical and biological properties are provided. Based on recent progress, it is expected that 3D bioprinting will lead to improved 3D tissue-engineered constructs for regeneration and repair of cartilage tissue.
Hydrogels are widely used in the design of tissue substitutes because of their ability to mimic the extracellular matrix (ECM). Their mechanical cues critically influence the cellular response, making accurate characterization essential. However, it remains challenging due to their intricate nature. This study computationally evaluates the hyperelastic properties of next-generation hydrogels of high biomedical interest, including basal membrane extract and decellularized liver matrices, as well as structural proteins. We present a combined framework based on Bayesian optimization and statistical analyses that go beyond classical least-squares fitting, leveraging rheological experimental data. It defines each hyperelastic strain-energy density function, and addresses both intra- and inter-sample variability. This approach quantifies uncertainty and reveals the natural variability that deterministic models overlook, and it also enables quantification of coefficient variation with composition. Validation against experimental data shows computational fits of 5
Inadequate angiogenesis in obesogenic adipose tissue (AT) has been implicated in disrupted adipogenesis and metabolic disorders. While several regulators of AT angiogenesis have been identified, our understanding of the quantitative changes in angiogenic signaling proteins during obesity progression remains incomplete, particularly regarding sex-specifi c responses. This study sought to identify the dysregulated elements within the Vascular Endothelial Growth Factor (VEGF) and Platelet-Derived Growth Factor (PDGF) systems during obesity progression. We employ a mouse model, comprising both male and female mice, to investigate the changes in the VEGF/PDGF concentration and their receptor distribution in gonadal AT during short- and long-term weight gain and weight loss. Female mice preserve adipose tissue angiogenic signatures during obesity progression, including sustained upregulation of endothelial VEGFR1 protein, maintenance of VEGF-A levels and endothelial cell populations, and differential regulation of PDGF levels, compared to males. These sex-specifi c patterns correlate with improved adipose expandability and may contribute to the more metabolically healthy obesity phenotype commonly observedin females These sex-specifi c patterns correlate with improved adipose expandability and may contribute to the more metabolically healthy obesity phenotype commonly observed in females. Our quantitative profi ling also lays the groundwork for developing computational models of VEGF/PDGF signaling networks in AT, allowing for the simulation of complex biological interactions and the prediction of therapeutic outcomes.
Nonviral gene delivery using DNA vectors is widely used in cell engineering, vaccination, and gene therapy, but delivery efficiency remains lower than those of viral vectors and mRNA-based approaches, partly due to inefficient nuclear entry, as transfected DNA must enter the nucleus for transcription. Therefore, a mechanistic understanding of nuclear entry pathways is essential for developing strategies to improve the efficiency. This review evaluated mechanistic studies of DNA nuclear entry in mammalian cells, mathematical models of intracellular DNA trafficking, quantitative analyses of DNA nuclear accumulation and transgene expression, and strategies to enhance nuclear delivery of DNA. Two mechanistically distinct pathways for DNA nuclear entry have been reported: enclosure upon nuclear envelope reformation in divided cells, and active transport through nuclear pore complexes (NPCs). Various strategies have been developed to enhance nuclear import through these pathways; however, their effectiveness depends on multiple factors, including cell type, delivery methods, and cell cycle status. Although DNA vectors are significantly larger than the nominal inner diameter of NPCs, they may traverse NPCs through deformation and interactions with nuclear transport proteins. Quantitative studies show that DNA nuclear accumulation is time dependent and heterogeneous within the same population. Nuclear entry plays a key role in determining efficiency of nonviral gene delivery. Advances in mechanistic studies, quantitative modeling, and imaging-based analyses have improved our understanding of intracellular DNA trafficking and nuclear accumulation. Integrating these insights with delivery strategies that enhance nuclear access while preserving the cellular machinery required for transgene expression will be critical for developing more efficient and reliable nonviral DNA delivery systems for therapeutic and biotechnological applications.
The extracellular matrix (ECM) is a dynamic fiber environment containing structural information that significantly impacts cell behavior. Recent experimental work has demonstrated that cells also have significant capacity to remodel their microenvironment, often resulting in ECM heterogeneity. We present an open-source molecular dynamics platform that simulates cell-mediated remodeling wherein cells plasticly remodel their microenvironment and respond to induced structural heterogeneity over multiple retraction cycles. The model applies a coarse-grained discrete fiber approach to cell-mediated remodeling. The ECM, represented by a bead-spring polymer, allows proximity-mediated fiber-fiber interactions, representing fiber crosslinking and entanglement. A simulated cell interprets the heterogeneity of its local microenvironment with variable sensitivity and exhibits anisotropic behavior informed by its microenvironment. The cells generate tractors, representing pseudopods, that bind to the ECM and retract towards the cell surface, causing ECM displacement. The cell detaches from the ECM by deleting tractors and allowing the ECM to relax before re-interpreting its surroundings and repeating this process. Metrics of ECM remodeling (fiber densification, orientation, bond strain) and cell morphology were recorded throughout the simulation. The model was extended to a cell remodeling fiber networks with different levels of pre-existing alignment. The addition of plasticity in the model enables measurable remodeling: increasing fiber density close to the cell, reorienting fibers radially, and increasing residual fiber bond strain over time. These patterns of remodeling were consistent with previously published experimental results. In initially unaligned fiber networks, cell remodeling resulted in ECM heterogeneity that depended on distance from the cell surface and alignment with the cell’s primary axis. In aligned networks, pre-alignment and sensitivity synergized to increase the heterogeneity of the remodeled networks at further distances from the cell surface. These findings suggest that cell-ECM feedback mechanisms contribute to heterogeneous remodeling patterns and illustrate that pre-existing alignment impacts remodeling patterns far from the cell. Further, the model presented herein provides a novel modular platform for further investigations into cell-ECM sensing and ECM remodeling heterogeneity.
Critical-sized craniofacial defects pose a significant clinical challenge, prompting the investigation of novel regenerative strategies. While mesenchymal stem cells (MSCs) and extracellular vesicles (EVs) hold promise, the optimal cell source and EV efficacy for craniofacial bone regeneration remain unclear. This study compares adipose-derived stem cells (ASCs), bone marrow-derived stem cells (BMSCs), and their derived EVs to address this gap in a critical-sized calvarial defect model. EVs from BMSCs and ASCs were isolated via ultracentrifugation and size exclusion chromatography. Nanoparticle tracking analysis and bicinchoninic acid assay quantified yield and protein, respectively. Transmission electron microscopy and Western blotting verified EV morphology and markers. In vitro, osteogenic potential of BMSCs and ASCs treated with their respective EVs was assessed using alkaline phosphatase activity assay, viability assays, and mineralization staining. In vivo, bone regeneration was compared in a rat critical-sized calvarial defect model treated with BMSCs, BMSC-derived EVs, and a combination of BMSC and BMSC-derived EVs. EVs isolated by ultracentrifugation yielded superior numbers of particles compared to size exclusion chromatography. In vitro, BMSC-derived EVs enhanced osteogenic differentiation of BMSCs, whereas ASC-derived EVs inhibited proliferation and osteogenesis of BMSCs. Although BMSC-derived EVs induced osteogenic phenotype in ASCs, osteoinductive efficiency was low, along with reduced cell proliferation. In vivo, both BMSC-derived EVs and BMSCs individually promoted bone regeneration compared to vehicle controls. Notably, the combination of BMSCs and BMSC-derived EVs demonstrated a significantly superior healing within the bone defect. Ultracentrifugation is the preferred method for isolating EVs for clinical translation. BMSC-derived EVs are the optimal source for craniofacial bone regeneration compared to ASC-derived EVs, as they exhibited superior osteogenic potential and promoted bone regeneration. Moreover, BMSC-derived EVs combined with BMSCs demonstrated a synergistic effect that further improved bone regeneration. Notably, mismatching origins of MSC and MSC-derived EVs could dysregulate the cellular function of MSCs, potentially compromising the regenerative outcomes. Our study highlights that the proper matching of cell and EVs sources is imperative for optimizing therapeutic efficacy in craniofacial bone regeneration.
Platelet adhesion and aggregation on exposed vascular extracellular matrix (ECM) is critical for haemostasis, with dysregulation and inappropriate thrombus formation associated with cardiovascular disease. While collagen is recognised as vital in these processes, the role of other ECM proteins is less understood. Platelet-derived microparticles (PDMPs), small vesicles released by activated platelets, similarly influence haemostasis, though their modulatory effect on ECM substrates is also unclear. We investigated platelet adhesion and aggregation on various ECM proteins–collagen, fibrinogen, fibronectin, and laminin–and examined the modulatory role of PDMPs under physiologic and pathologic flow conditions. Whole blood, alone or enriched with PDMPs, was perfused through microfluidic channels coated with ECM substrates at varying shear rates. Fluorescence imaging assessed platelet behaviour measuring surface coverage, number of thrombi, mean thrombus area and thrombus height. Collagen exhibited greatest overall thrombus formation, versus the other ECM substrates, with platelet adhesion and aggregation increasing at higher shear rates. Addition of PDMPs significantly reduced thrombus area (p < 0.01) and height (p < 0.05–0.01) at all shear rates. Fibrinogen supported stable but smaller thrombi, with PDMP addition resulting in increased platelet adhesion (23
The nucleolus, traditionally viewed as the site of ribosome biogenesis, is now recognized as a multifunctional organelle involved in stress sensing, genome regulation, and cellular homeostasis. In parallel, mechanical cues arising from neighboring cells, extracellular matrix, and fluid flow have emerged as key regulators of development, tissue remodeling, and disease initiation and progression.These biophysical cues are transmitted to the nucleus through integrin, LINC complex-dependent pathways that modulate nuclear architecture, lamin organization, and chromatin structure. Because the nucleolus is embedded in a mechanically responsive environment and interacts directly or indirectly with the surrounding components, it is increasingly considered a potential target for mechanical cues. However, direct experimental evidence linking defined mechanical cues to nucleolar structure, phase behavior, and function remains limited. This review summarizes current knowledge on the emerging interface between nucleolus and mechanobiology, highlighting potential molecular pathways that connect force transmission to the nuclear response, and outlines unanswered questions relevant to development and disease.
Immaturity of stem cell-derive cardiomyocytes limits their use in tissue engineering applications. Macrophage contributions to the development of cardiomyocytes have not yet been fully established. While some recent studies have added macrophages to stem cell-derived models of the human myocardium, these previous approaches do not replicate the early colonization of the heart. Due to their importance in regulating cardiomyocyte metabolism, we hypothesized that developmentally informed addition of macrophages to cardiomyocytes would improve cardiomyocyte maturity. We generated cardiomyocytes and embryonic-like macrophages from a single cell line. Macrophages were added to developing cardiomyocytes 8, 16, and 19 days after induction of differentiation, based on changes in cardiomyocyte media formulation at these stages. Cardiomyocytes were cultured until 30 days post differentiation, where they were prepared for analysis. Metabolism was measured through Seahorse Mitochondria Stress assays. Corresponding changes in subcellular structures were measured through high-resolution microscopy. Mitophagy in cardiomyocytes was measured through the fluorescent reporter mtKeima. Addition of macrophages to cardiomyocytes 8 days after the induction of differentiation results in a significant increase in cardiomyocyte basal and maximal metabolism. Developing cardiomyocytes shed lowly polarized mitochondria, which are taken up by macrophages. As a result, cardiomyocytes adopt an adjusted mitochondria network architecture featuring less interconnected mitochondria. Mitophagy flux measurements show that cardiomyocytes develop more active mitophagy programs while in coculture with macrophages. Pharmacological inhibition of mitophagy reveals that this interaction is dependent on macrophage MerTK-mediated reception of cardiomyocyte-derived mitochondria material. These results improve our understanding of the responsibility of macrophages in the development of cardiomyocyte metabolism. We establish interactions between macrophages and developing cardiomyocytes as essential to produce more mature cardiomyocytes and physiologically relevant models of the human myocardium.
The secretory output from mesenchymal stem cells (MSCs) have emerged as promising therapeutics with extracellular vesicles (EVs) gaining prominence due to solution stability and optimal size for overcoming biological barriers during delivery. However, reproducible and scalable production of EVs for therapeutic use remains a challenge in biotechnology. Here we demonstrate optimization of EV production from MSCs using soft hydrogel microcarriers. Gelatin methacryloyl (GelMA) hydrogels were prepared at a range of concentrations for the culture of two sources of MSCs: adipose derived stem cells (ADSCs) and induced pluripotent stem cell derived MSCs (iMSCs). The mechanical properties of the hydrogels were evaluated using shear rheology. EVs were isolated and analyzed for physical and biological characteristics using electron microscopy, nanoparticle tracking, proteomics, and functional assays for wound healing and angiogenesis. Both cell types were responsive to hydrogel stiffness (0.3-16 KPa), showing optimal EV secretion from cultures on 10 KPa hydrogels, with a further 18-fold increase when formulated as microcarriers compared to traditional monolayer culture. Proteomics analysis and functional assays revealed that EVs from microcarrier culture displayed increased wound healing and regenerative properties. This study demonstrates the advantages of hydrogel microcarriers in the production of cell-derived products, with optimized design parameters to guide scaleup and translation to manufacturing, in support of biotechnology and biomedical applications.
Pancreatic ductal adenocarcinoma (PDAC) develops within a biomechanically abnormal tumor microenvironment, characterized by a dense stroma and elevated compressive forces. While extracellular matrix stiffness has been extensively studied, the impact of compressive forces on immune regulation and tumor–immune interactions remains poorly understood. We integrated two complementary bioengineered compression models, a 2D transmembrane pressure device and confined 3D spheroids, with bulk transcriptomic and Liquid Chromatography–Mass Spectrometry (LC-MS)–based exometabolomic profiling to examine how mechanical compression shapes macrophage behaviour and tumor–immune crosstalk. Controlled compressive stress (0–8 mmHg) was applied to macrophages, tumor cells, and tumor–macrophage cocultures, followed by pathway analysis, functional assays, and multi-omic integration. Mechanical compression activated conserved mechanotransduction pathways in macrophages, including PI3K/Akt and MAPK/SAPK signaling, and induced transcriptional programs associated with inflammatory activation consistent with an M1-like macrophage phenotype and cytoskeletal remodelling. In parallel, compressed tumor cells adopted an immunomodulatory state marked by increased expression of immunosuppressive cytokines and macrophage checkpoint signals. When tumor cells and macrophages were simultaneously exposed to compression, functional assays revealed a shift of macrophages toward immunosuppressive phenotypes. Bulk RNA sequencing identified cell-type–specific transcriptional responses converging on metabolic pathways, while LC-MS exometabolomics revealed compression-dependent enrichment of extracellular nucleotide metabolites in tumor–macrophage cocultures. These findings identify compressive stress as a critical regulator of immune suppression and tumor–immune metabolic coupling in PDAC, highlighting mechanical forces as important drivers of immune dysfunction in mechanically constrained tumors.
To investigate the synergistic effects of matrix stiffness and cyclic tensile stress—key mechanical cues in the lung tumor microenvironment—on lung cancer cell migration, and to elucidate the underlying mechanotransduction pathways. A novel cell-stretch coupling device was developed by integrating tunable P(SBMA-co-AAm) hydrogels (8.9, 49.4, and 99.5 kPa) with stretchable PDMS chambers via an interpenetrating polymer network. A549 lung cancer cells expressing FRET-based biosensors for FAK, Src, and RhoGDI α were subjected to cyclic stretching (10 or 20
Vestibular schwannoma (VS) progressively stiffens and remodels its extracellular matrix (ECM) during growth. However, how mechanical confinement and adhesive ECM signaling regulate schwannoma behavior in vitro remain incompletely defined. Human Nf2-/- schwannoma and primary VS cells established from fresh surgical specimens were cultured in complementary 3D hydrogel platforms. Biochemically inert agarose hydrogels spanning physiologic to pathologic stiffnesses created a non-adhesive mechanical confinement environment, while type I collagen hydrogels modeled an adhesive environment with a dense, fibrillar matrix characteristic of fibrotic tumor ECM. Hydrogel stiffness was quantified by rheology. Cell viability, proliferation, morphology, mechanosensitive signaling, and ECM remodeling were quantified. Mechanical stress was relieved by enzymatic degradation. YAP activity was pharmacologically inhibited, and transforming growth factor-β (TGF-β) was used to induce collagen remodeling. Under non-adhesive confinement, increasing stiffness suppressed schwannoma reduced cell spreading, decreased N-cadherin expression, and increased nuclear YAP localization. Stress relief reversed YAP activation while enabling enhanced proliferative recovery and increased N-cadherin–associated adhesion. Cells under increased confinement exhibited increased sensitivity to YAP inhibition, indicating confinement-dependent reliance on mechanotransduction. In contrast, adhesive ECM conditions supported active matrix remodeling with increasing stiffness, including elevated activities of MMP9 and phosphorylated focal adhesion kinase (pFAK). TGF-β induced both collagen disorganization and SMAD3 nuclear localization, which was attenuated by YAP inhibition. Mechanical confinement and ECM composition drive distinct, context-dependent adaptation programs in VS. As stiffness increases, cells in non-adhesive environments adopt a reversible, YAP-associated stress response, while an adhesive ECM shifts behavior toward matrix remodeling and cell adhesion-driven signaling.
