Cellular and molecular bioengineering最新文献

Purpose: 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.

Methods: 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.

Results: 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.

Conclusions: 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.

Supplementary information: The online version contains supplementary material available at 10.1007/s12195-026-00922-0.

Background: 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.

Methods: 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.

Results: 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.

Conclusions: 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.

Supplementary information: The online version contains supplementary material available at 10.1007/s12195-026-00909-x.

Introduction: 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.

Methods: 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.

Results: 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% increase) at low shear. Both fibronectin and laminin demonstrated weak platelet adhesion, with thrombus formation decreasing as shear rate increased, and PDMP addition slightly increasing platelet adhesion, though without influence on thrombus dimensions.

Conclusion: Collagen exerts a dominant effect in platelet adhesion and aggregation across a wide range of shear, compared to fibrinogen, fibronectin and laminin. PDMPs alter platelet adhesion and resultant thrombus formation across all ECM proteins tested, likely through competitive occupation of platelet binding sites. Our findings underscore the complex interactions between ECM proteins, PDMPs, and shear forces under flow conditions.

Supplementary information: The online version contains supplementary material available at 10.1007/s12195-026-00920-2.

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.

Background: 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.

Methods: 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.

Results: 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.

Conclusions: 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.

Supplementary information: The online version contains supplementary material available at 10.1007/s12195-026-00915-z.

Background: 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.

Methods: 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.

Results: 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.

Conclusion: 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.

Supplementary information: The online version contains supplementary material available at 10.1007/s12195-026-00917-x.

Purpose: 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.

Methods: 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% strain, 1 Hz). Inhibitors (PF228 for FAK, PP1 for Src, ML-7 for cytoskeleton) were used to dissect signaling pathways.

Results: On soft and medium-stiffness substrates (8.9-49.4 kPa), cyclic stretching significantly enhanced cell migration (by 22.5-38.7%) and activated FAK/Src while suppressing RhoGDI α. High stiffness (99.5 kPa) rendered cells unresponsive to stretch (≤ 3.2% change). FAK and Src jointly regulated RhoGDI α, with FAK dominance under low stiffness. Cytoskeletal inhibition abolished mechanoresponses, confirming its role as a central mechanosensor.

Conclusion: The study establishes a biomimetic platform for dual mechanical stimulation and reveals stiffness-dependent synergy between tensile stress and matrix stiffness in promoting lung cancer migration via the FAK/Src-RhoGDI α axis. These findings provide mechanistic insights into mechanotransduction in lung cancer and suggest potential targets for inhibiting metastasis by disrupting mechanical signaling.

Supplementary information: The online version contains supplementary material available at 10.1007/s12195-026-00914-0.

Purpose: Tunneling nanotubes (TNTs) are membrane-based intercellular structures that support long-range communication between spatially separated cells. Although TNT-associated electrical responses have been observed under mechanical stimulation, whether these responses are linked to changes in cellular mechanical behavior remains unclear.

Methods: Mechanical stimulation was applied to one cell within TNT-connected C2C12 pairs. Traction force and cell spreading area in the connected cell were evaluated under different pharmacological conditions to investigate the involvement of calcium signaling, calcium channels, gap junction communication, and cellular contractility.

Results: Mechanical stimulation increased traction force and reduced cell spreading area in TNT-connected cells. These responses were suppressed by calcium depletion, inhibition of T-type calcium channels, or blockade of gap junction communication, but were not significantly affected by inhibition of L-type calcium channels.

Conclusions: These findings support a calcium-dependent association between TNT-mediated intercellular signaling and biomechanical regulation in connected C2C12 cells.

Supplementary information: The online version contains supplementary material available at 10.1007/s12195-026-00908-y.

Purpose: 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.

Methods: 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.

Results: 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.

Conclusions: 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.

Supplementary information: The online version contains supplementary material available at 10.1007/s12195-026-00911-3.