Matrix Stiffness Research Articles

DDDR-29. MATRIX STIFFNESS INDUCED PHENOTYPE IN BRAIN METASTATIC BREAST CANCER CELLS DETERMINES RESPONSE TO THERAPY

Abstract Breast cancer is the most prevalent form of cancer, representing 12.5% of all cancer cases diagnosed worldwide. Breast cancer cells can metastasize to distant organs (i.e., brain) and remain dormant for extended periods of time. This aspect makes it very difficult to treat these cells as they exhibit reduced growth as well as resistance to therapy. Despite advances in our understanding of breast cancer metastasis, the specific mechanisms that enable dormant cancer cells to resist therapeutics, particularly in brain metastatic breast cancer (BMBC), remain unclear. Herein, we employed brain tissue mimetic hyaluronic acid (HA) hydrogels of varying stiffness (i.e., ~0.4 kPa vs. ~4.5 kPa) to investigate the response to chemo or targeted therapy in dormant versus proliferative BMBC cells. It was revealed that BMBC cells grown on soft HA hydrogels (~0.4 kPa) displayed a non-proliferative (i.e., dormant) phenotype and exhibited resistance to Paclitaxel (PTX) or Lapatinib (LAP). However, BMBC cells grown on stiff HA hydrogels (~4.5 kPa) displayed a proliferative phenotype and sensitivity to PTX or LAP. Furthermore, we found that resistance to therapy was due to the enhanced expression of the serum/glucocorticoid regulated kinase 1 (SGK1) gene, mediated, in part, via the p38 mitogen-activated protein kinase (MAPK) pathway. Consequently, SGK1 inhibition using a SGK inhibitor in BMBC cells cultured on soft HA hydrogels resulted in a dormant-to-proliferative switch as well as response to PTX or LAP. In sum, our study demonstrated that matrix stiffness plays a major role in the dormancy-associated therapy response, mediated, in part, via the p38/SGK1 pathway.

A collagenase-decorated Cu-based nanotheranostics: remodeling extracellular matrix for optimizing cuproptosis and MRI in pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC), characterized by a dense extracellular matrix (ECM), presents significant therapeutic challenges due to its poor prognosis and high resistance to chemotherapy. Current chemodrugs and diagnostic agents largely fail to cross the barrier posed by the ECM, which severely limits the PDAC theranostics. This study introduces a novel theranostic strategy using thioether-hybridized hollow mesoporous organosilica nanoparticles (dsMNs) for the co-delivery of copper (Cu) and disulfiram (DSF), aiming to induce cuproptosis in PDAC cells. Our approach leverages the ECM-degrading enzyme collagenase, integrated with dsMNs, to enhance drug penetration by reducing matrix stiffness. Furthermore, the innovative use of a pancreatic cancer cell membrane coating on the nanoparticles enhances tumor targeting and stability (dsMCu-D@M-Co). The multifunctional platform not only facilitates deep drug penetration and triggers cuproptosis effectively but also utilizes the inherent properties of Cu to serve as a T1-weighted magnetic resonance imaging (MRI) contrast agent. In vitro and in vivo assessments demonstrate significant tumor size reduction in PDAC-bearing mice, highlighting the dual functionality of our platform in improving therapeutic efficacy and diagnostic precision. This integrated strategy represents a significant advancement in the management of PDAC, offering a promising new direction for overcoming one of the most lethal cancers.Graphical

Endothelin-1 receptor blockade impairs invasion patterns in engineered 3D high-grade serous ovarian cancer tumoroids.

High-grade serous ovarian cancer (HG-SOC), accounting for 70-80% of ovarian cancer deaths, is characterized by a widespread and rapid metastatic nature, influenced by diverse cell types, cell-cell interactions, and acellular components of the tumor microenvironment (TME). Within this tumor type, autocrine and paracrine activation of the endothelin-1 receptors (ET-1R), expressed in tumor cells and stromal components, drives metastatic progression. The lack of three-dimensional models that faithfully recapitulate the unique HG-SOC TME has been the bottleneck in performing drug screening for personalized medicine. Herein, we developed HG-SOC tumoroids by engineering a dense central artificial cancer mass (ACM) containing HG-SOC cells, nested within a compressed hydrogel recapitulating the stromal compartment comprising type I collagen, laminin, fibronectin, and stromal cells (fibroblasts and endothelial cells). ET-1-stimulated HG-SOC cells in the tumoroids showed an altered migration pattern and formed cellular aggregates, mimicking micrometastases that invaded the stroma. Compared to control cells, ET-1-stimulated tumoroids showed a higher number of invasive bodies, which were reduced by treatment with the dual ET-1 receptor (ET-1R) antagonist macitentan. In addition, ET-1 increased the size of the invading aggregates compared to control cells. This study establishes an experimental 3D multicellular model eligible for mechanical research, investigating the impact of matrix stiffness and TME interactions, which will aid drug screening to guide therapeutic decisions in HG-SOC patients.

Sex Differences in Response to Diet Enriched with Glutathione Precursors in the Aging Heart.

Common features of the aging heart are dysregulated metabolism, inflammation, and fibrosis. Elevated oxidative stress is another hallmark of cardiac aging that can exacerbate each of these conditions. We hypothesize that by increasing natural antioxidant levels (glutathione), we will improve cardiac function. Twenty-one-month-old mice were fed Glycine and N-Acetyl Cysteine (GlyNAC) (glutathione precursors)-supplemented or control diets for 12 weeks. Heart function was monitored longitudinally, and the exercise performance was determined at the end of the study. We found that the GlyNAC diet was beneficial for old male but not old female mice, leading to an increase of Ndufb8 expression (a subunit of the mitochondrial respiratory chain complex), and higher enzymatic activity for CPT1b and CrAT, two carnitine acyltransferases that are critical to cardiomyocyte metabolism. Although no quantifiable change of collagen turnover was detected, hearts from GlyNAC-fed old males exhibited a slight but significant enrichment in Fmod, a protein that can inhibit collagen fibril formation, possibly reducing extracellular matrix (ECM) stiffness and thus improving diastolic function. Cardiac diastolic function was modestly improved in males but not females, and surprisingly GlyNAC-fed female mice showed a decline in exercise performance. In summary, our work supports the concept that aged male and female hearts are phenotypically different. These basic differences may affect the response to pharmacological and diet interventions, including antioxidants.

Cellular Alignment and Matrix Stiffening Induced Changes in Human Induced Pluripotent Stem Cell Derived Cardiomyocytes.

Biological processes are inherently dynamic, necessitating biomaterial platforms capable of spatiotemporal control over cellular organization and matrix stiffness for accurate study of tissue development, wound healing, and disease. However, most in vitro platforms remain static. In this study, a dynamic biomaterial platform comprising a stiffening hydrogel is introduced and achieved through a stepwise approach of addition followed by light-mediated crosslinking, integrated with an elastomeric substrate featuring strain-responsive lamellar surface patterns. Employing this platform, the response of human induced pluripotent stem cell-derived cardiomyocytes (hIPSC-CMs) is investigated to dynamic stiffening from healthy to fibrotic tissue stiffness. The results demonstrate that culturing hIPSC-CMs on physiologically relevant healthy stiffness significantly enhances their function, as evidenced by increased sarcomere fraction, wider sarcomere width, significantly higher connexin-43 content, and elevated cell beating frequency compared to cells cultured on fibrotic matrix. Conversely, dynamic matrix stiffening negatively impacts hIPSC-CM function, with earlier stiffening events exerting a more pronounced hindering effect. These findings provide valuable insights into material-based approaches for addressing existing challenges in hIPSC-CM maturation and have broader implications across various tissue models, including muscle, tendon, nerve, and cornea, where both cellular alignment and matrix stiffening play pivotal roles in tissue development and regeneration.

Poroelastic Response of a Fractured Rock to Hydrostatic Pressure Oscillations

AbstractPoroelastic coupling between fractures and the surrounding rock is important to numerous applications in geosciences. We measure the in‐situ fluid pressure and local strain response of a fractured carbonate sample to hydrostatic pressure oscillations. A linear poroelastic model that represents the rock sample is parameterized using X‐ray imaging and ultrasonic wave transmission measurements. The numerical solution, based on Biot's quasistatic equations, is consistent with the measured frequency dependent dispersion of the apparent bulk modulus of the background matrix and the in‐situ pore pressure response, which is caused by fluid pressure diffusion from the compliant fractures into the stiffer matrix. The observed fluid pressure diffusion is causally related to the numerically quantified intrinsic attenuation at seismic frequencies, which is a major contributor to the dissipation of seismic waves. Our analysis supports the use of a simple approximation of fractures as compliant and planar inclusions in numerical simulations based on linear poroelasticity.

Anti-fibrotic effects of the sirt6-activator mdl-800 in cardiac stromal cells

Abstract Background Epigenetic changes are among the molecular substrates of cellular stress responses and tissue remodeling in heart failure progression. The sirtuin family of NAD+-dependent histone and non-histone protein deacetylases exert a protective anti-fibrotic role by negatively modulating the inflammatory response and by positive regulation of autophagy-related genes. Sirtuin6 (SIRT6) stands out as a key cardiac protector that can mitigate aging-induced cardiomyocyte senescence and cardiac hypertrophy. Cardiac stromal cells (CSCs) play a pivotal role in fibrosis and remodeling, regulating the composition and stiffness of the extracellular matrix (ECM). Autophagy enhancement has anti-fibrotic effects in CSCs, but the specific role of SIRT6 modulation in CSCs remains unexplored. Purpose To investigate the biological effects of MDL-800, a synthetic allosteric SIRT6 activator, on the stress response and fibrotic activation of CSCs. Methods CSCs were isolated from 4-week-old C57Bl6J mice by explant culture and stimulated with 10ng/ml of TGF-beta1 (TGFb1) up to 72h to induce fibrotic activation. CSCs were treated with MDL-800 up to 10µM and up to 72h, analyzed for cell viability, gene and protein expression levels. Results TGFb1 treatment led to a 0.64-fold reduction in SIRT6 gene expression after 24h. The MTS assay was used to assess a viability dose-response to MDL-800 up to 72h of treatment. The non-toxic concentrations of 2.5µM and 5µM were selected for further experiments (2.5µM vs CTR 0.97-fold and 5µM vs CTR 0.92-fold, normalized OD vs t0). Co-treatment of 5µM MDL-800 with TGFb1 for 72h resulted in a dose-dependent decrease in the expression of TGFb1-induced fibroblast activation markers, as confirmed by immunofluorescence staining for aSMA (TGFb1 vs CTR 2.21-fold, p<0.05; 5µM vs TGFb1 0.46-fold, p<0.05, N=4; mean fluorescence intensity). The anti-fibrotic effect mediated by MDL-800 was further validated by real-time PCR indicating a reduction in the mRNA expression of multiple fibrotic markers, including ACTA2 (TGFb1 vs CTR 1.67-fold, 5µM vs TGFB 0.29-fold; N=2), COL1a1 (TGFb1 vs CTR 1.32-fold, 5µM vs TGFb1 0.87-fold; N=2) COL3a1 (TGFb1 vs CTR 1.06-fold, 5µM vs TGFb1 0.43-fold; N=2) and POSTN (TGFb1 vs CTR 6.37-fold, 5µM vs TGFb1 0.23-fold; N=2). Treatment with 5µM MDL-800 demonstrated an increase in SIRT6 gene expression, suggesting a transcriptional modulation (TGFb1 vs CTR 0.95-fold, 5µM vs TGFb1 1.68-fold; N=2). Western Blot analyses showed an increase in protein levels of the autophagosome marker LC3-II following treatment with 5µM MDL-800 (TGFb1 vs CTR 0.95-fold; 5µM vs TGFb1 3.01-fold), suggesting that autophagy enhancement may be a potential mechanism for the observed anti-fibrotic effects. Conclusion MDL-800 treatment in CSCs is associated with a dose-dependent reduction of TGFb1-induced fibrotic markers and to increased levels of the autophagy marker, suggesting its potential use as an anti-fibrotic molecule for the heart.

Chemomechanical regulation of EZH2 localization controls epithelial-mesenchymal transition.

The methyltransferase enhancer of zeste homolog 2 (EZH2) regulates gene expression and aberrant EZH2 expression and signaling can drive fibrosis and cancer. However, it is not clear how chemical and mechanical signals are integrated to regulate EZH2 and gene expression. We show that culture of cells on stiff matrices in concert with transforming growth factor (TGF)-β1 promotes nuclear localization of EZH2 and an increase in the levels of the corresponding histone modification, H3K27me3, thereby regulating gene expression. EZH2 activity and expression are required for TGFβ1- and stiffness-induced increases in H3K27me3 levels as well as for morphological and gene expression changes associated with epithelial-mesenchymal transition (EMT). Inhibition of Rho associated kinase (ROCK) or myosin II signaling attenuates TGFβ1-induced nuclear localization of EZH2 and decreases H3K27me3 levels in cells cultured on stiff substrata, suggesting that cellular contractility, in concert with a major cancer signaling regulator TGFβ1, modulates EZH2 subcellular localization. These findings provide a contractility-dependent mechanism by which matrix stiffness and TGFβ1 together mediate EZH2 signaling to promote EMT.

Synthetic hydrogel substrate for human induced pluripotent stem cell definitive endoderm differentiation

Human induced pluripotent stem cells (hiPSCs) can give rise to multiple lineages derived from three germ layers, endoderm, mesoderm and ectoderm. Definitive endoderm (DE) cell types and tissues have great potential for regenerative medicine applications. Current hiPSC differentiation protocols focus on the addition of soluble factors; however, extracellular matrix properties are known to also play a role in dictating cell fate. Matrigel™ is the gold standard for DE differentiation, but this xenogeneic, poorly defined basement membrane extract limits the clinical translatability of DE-derived tissues. Here we present a fully defined PEG-based hydrogel substrate to support hiPSC-derived DE differentiation. We screened hydrogel formulations presenting different adhesive peptides and matrix stiffness. Our results demonstrate that presenting a short peptide, cyclic RGD, on the engineered PEG hydrogel supports the transition from undifferentiated hiPSCs to DE using a serum-free, commercially available kit. We show that increasing substrate stiffness (G’ = 1.0–4.0 kPa) results in an increased linear response in DE differentiation efficiency. We also include a temporal analysis of the expression of integrin and syndecan receptors as the hiPSCs undergo specification towards DE lineage. Finally, we show that focal adhesion kinase activity regulates hiPSC growth and DE differentiation efficiency. Overall, we present a fully defined matrix as a synthetic alternative for Matrigel™ supporting DE differentiation.

Mitochondrial mechanotransduction through MIEF1 coordinates the nuclear response to forces.

Tissue-scale architecture and mechanical properties instruct cell behaviour under physiological and diseased conditions, but our understanding of the underlying mechanisms remains fragmentary. Here we show that extracellular matrix stiffness, spatial confinements and applied forces, including stretching of mouse skin, regulate mitochondrial dynamics. Actomyosin tension promotes the phosphorylation of mitochondrial elongation factor 1 (MIEF1), limiting the recruitment of dynamin-related protein 1 (DRP1) at mitochondria, as well as peri-mitochondrial F-actin formation and mitochondrial fission. Strikingly, mitochondrial fission is also a general mechanotransduction mechanism. Indeed, we found that DRP1- and MIEF1/2-dependent fission is required and sufficient to regulate three transcription factors of broad relevance-YAP/TAZ, SREBP1/2 and NRF2-to control cell proliferation, lipogenesis, antioxidant metabolism, chemotherapy resistance and adipocyte differentiation in response to mechanical cues. This extends to the mouse liver, where DRP1 regulates hepatocyte proliferation and identity-hallmark YAP-dependent phenotypes. We propose that mitochondria fulfil a unifying signalling function by which the mechanical tissue microenvironment coordinates complementary cell functions.

A Photopolymerizable Hyaluronic Acid-Collagen Model of the Invasive Glioma Microenvironment with Interstitial Flow.

Glioblastoma recurrence is a major hindrance to treatment success and is driven by the invasion of glioma stem cells (GSCs) into healthy tissue that are inaccessible to surgical resection and are resistant to existing chemotherapies. Tissue-level fluid movement, or interstitial fluid flow (IFF), regulates GSC invasion in a manner dependent on the tumor microenvironment (TME), highlighting the need for model systems that incorporate both IFF and the TME. We present an accessible method for replicating the invasive TME in glioblastoma: a hyaluronan-collagen I hydrogel composed of human GSCs, astrocytes, and microglia seeded in a tissue culture insert. Elevated IFF can be represented by applying a fluid pressure head to the hydrogel. Additionally, this model can be tuned to replicate inter- or intra-patient differences in cellular ratios, flow rates, or matrix stiffnesses. Invasion can be quantified, while gels can be harvested for a variety of outcomes, including GSC invasion, flow cytometry, protein or RNA extraction, or imaging.

Planar cell polarity zebrafish models of congenital scoliosis reveal novel underlying defects in notochord morphogenesis.

Congenital scoliosis (CS) is a type of vertebral malformation whose etiology remains elusive. The notochord is pivotal for vertebrae development but its role in CS is still understudied. Zebrafish knockout of ptk7a, a planar cell polarity (PCP) gene that is essential for convergence and extension (C&E) of the notochord, developed congenital scoliosis-like vertebral malformations (CVM). Maternal zygotic ptk7a mutants displayed severe C&E defects of the notochord. Excessive apoptosis occurred in the malformed notochord, causing a significantly reduced number of vacuolated cells, and compromising the mechanical properties of the notochord. The latter manifested as a less stiff extracellular matrix along with a significant reduction in the number of the caveolae and severely loosened intercellular junctions in the vacuolated region. These defects led to focal kinks, abnormal mineralization, and CVM exclusively at the anterior spine. Loss of function of another PCP gene, vangl2, also revealed excessive apoptosis in the notochord associated with CVM. This study suggests a new model for CS pathogenesis that is associated with defects in notochord C&E and highlights an essential role of PCP signaling in vertebrae development.

Gastric Cancer Actively Remodels Mechanical Microenvironment to Promote Chemotherapy Resistance via MSCs-Mediated Mitochondrial Transfer.

Chemotherapy resistance is the main reason of treatment failure in gastric cancer (GC). However, the mechanism of oxaliplatin (OXA) resistance remains unclear. Here, we demonstrate that extracellular mechanical signaling plays crucial roles in OXA resistance within GC. We selected OXA-resistant GC patients and analyzed tumor tissues by single-cell sequencing, and found that the mitochondrial content of GC cells increased in a biosynthesis-independent manner. Moreover, we found that the increased mitochondria of GC cells were mainly derived from mesenchymal stromal cells (MSCs), whichcould repair the mitochondrial function and reduce the levels of mitophagy in GC cells, thus leading to OXA resistance. Furthermore, we investigated the underlying mechanism and found that mitochondrial transfer was mediated by mechanical signals of the extracellular matrix (ECM). After OXA administration, GC cells actively secreted ECM in the tumor microenvironment (TEM), increasing matrix stiffness of the tumor tissues, which promoted mitochondria to transfer from MSCs to GC cells via microvesicles (MVs). Meanwhile, inhibiting the mechanical-related RhoA/ROCK1 pathway could alleviate OXA resistance in GC cells. In summary, these results indicate that matrix stiffness could be used as an indicator to identify chemotherapy resistance, and targeting mechanical-related pathway could effectively alleviate OXA resistance and improve therapeutic efficacy.

YAP/TAZ drives Notch and angiogenesis mechanoregulation in silico

Endothelial cells are key players in the cardiovascular system. Among other things, they are responsible for sprouting angiogenesis, the process of new blood vessel formation essential for both health and disease. Endothelial cells are strongly regulated by the juxtacrine signaling pathway Notch. Recent studies have shown that both Notch and angiogenesis are influenced by extracellular matrix stiffness; however, the underlying mechanisms are poorly understood. Here, we addressed this challenge by combining computational models of Notch signaling and YAP/TAZ, stiffness- and cytoskeleton-regulated mechanotransducers whose activity inhibits both Dll4 (Notch ligand) and LFng (Notch-Dll4 binding modulator). Our simulations successfully mimicked previous experiments, indicating that this YAP/TAZ-Notch crosstalk elucidates the Notch and angiogenesis mechanoresponse to stiffness. Additional simulations also identified possible strategies to control Notch activity and sprouting angiogenesis via cytoskeletal manipulations or spatial patterns of alternating stiffnesses. Our study thus inspires new experimental avenues and provides a promising modeling framework for further investigations into the role of Notch, YAP/TAZ, and mechanics in determining endothelial cell behavior during angiogenesis and similar processes.

Calciprotein particles induce arterial stiffening ex vivo and impair vascular cell function

Calciprotein particles (CPPs) are an endogenous buffering system, clearing excessive amounts of Ca2+ and PO43- from the circulation and thereby preventing ectopic mineralization. CPPs circulate as primary CPPs (CPP1), which are small spherical colloidal particles, and can aggregate to form large, crystalline, secondary CPPs (CPP2). Even though it has been reported that CPPs are toxic to vascular smooth muscle cells (VSMC) in vitro, their effect(s) on the vasculature remain unclear. Here we have shown that CPP1, but not CPP2, increased arterial stiffness ex vivo. Interestingly, the effects were more pronounced in the abdominal infrarenal aorta compared to the thoracic descending aorta. Further, we demonstrated that CPP1 affected both endothelial and VSMC function, impairing vasorelaxation and contraction respectively. Concomitantly, arterial glycosaminoglycan accumulation was observed as well, which is indicative of an increased extracellular matrix stiffness. However, these effects were not observed in vivo. Hence, we concluded that CPP1 can induce vascular dysfunction.

PIEZO1 mediates matrix stiffness-induced tumor progression in kidney renal clear cell carcinoma by activating the Ca2+/Calpain/YAP pathway

ObjectiveThe significance of physical factors in the onset and progression of tumors has been increasingly substantiated by a multitude of studies. The extracellular matrix, a pivotal component of the tumor microenvironment, has been the subject of extensive investigation in connection with the advancement of KIRC (Kidney Renal Clear Cell Carcinoma) in recent years. PIEZO1, a mechanosensitive ion channel, has been recognized as a modulator of diverse physiological processes. Nonetheless, the precise function of PIEZO1 as a transducer of mechanical stimuli in KIRC remains poorly elucidated. MethodsA bioinformatics analysis was conducted using data from The Cancer Genome Atlas (TCGA) and the Clinical Proteomic Tumor Analysis Consortium (CPTAC) to explore the correlation between matrix stiffness indicators, such as COL1A1 and LOX mRNA levels, and KIRC prognosis. Expression patterns of mechanosensitive ion channels, particularly PIEZO1, were examined. Collagen-coated polyacrylamide hydrogel models were utilized to simulate varying stiffness environments and study their effects on KIRC cell behavior in vitro. Functional experiments, including PIEZO1 knockdown and overexpression, were performed to investigate the molecular mechanisms underlying matrix stiffness-induced cellular changes. Interventions in the Ca2+/Calpain/YAP Pathway were conducted to evaluate their effects on cell growth, EMT, and stemness characteristics. ResultsOur findings indicate a significant correlation between matrix stiffness and the prognosis of KIRC patients. It is observed that higher mechanical stiffness can facilitate the growth and metastasis of KIRC cells. Notably, we have also observed that the deficiency of PIEZO1 hinders the proliferation, EMT, and stemness characteristics of KIRC cells induced by a stiff matrix. Our study suggests that PIEZO1 plays a crucial role in mediating KIRC growth and metastasis through the activation of the Ca2+/Calpain/YAP Pathway. ConclusionThis study elucidates a novel mechanism through which the activation of PIEZO1 leads to calcium influx, subsequent calpain activation, and YAP nuclear translocation, thereby contributing to the progression of KIRC driven by matrix stiffness.

Matrix Stiffness of GelMA Hydrogels Regulates Lymphatic Endothelial Cells toward Enhanced Lymphangiogenesis.

Lymphatic vessel regeneration is crucial for various tissue engineering strategies, particularly in resolving inflammation and restoring tissue homeostasis. In our study, we focused on investigating how hydrogel matrix stiffness influences lymphatic endothelial cells (LECs) in promoting lymphatic vessel regeneration. Gelatin methacrylate (GelMA) was chosen as our biomaterial due to its versatility in tissue engineering and biofabrication. We fabricated GelMA hydrogels at concentrations of 5, 7.5, and 15% (w/v) with corresponding Young's modulus values of 1.55 kPa (soft matrix), 12.02 kPa (medium matrix), and 48.50 kPa (stiff matrix). Among these, the 7.5% GelMA hydrogel exhibited optimal stiffness for promoting lymphangiogenesis. LECs seeded either on the hydrogel surface or within spontaneously formed a more stable lymphatic capillary network compared with other GelMA formulations. Furthermore, we investigated the enhancement of lymphangiogenesis by incorporating VEGF-C into the GelMA hydrogel, leveraging the synergistic effects of mechanical and chemical cues. Our results underscored the critical role of FAK-phosphorylation in this process; treatment with an FAK-specific inhibitor prevented the formation of tube-like structures by LECs and attenuated the expression of lymphatic markers. Overall, our findings highlight how the mechanical and chemical cues provided by GelMA hydrogels can effectively regulate LEC behavior toward enhanced lymphangiogenesis via the integrin/FAK mechanotransduction pathway. This study proposes a promising strategy for developing hydrogel-based scaffolds or bioinks tailored to promote lymphatic vessel regeneration in therapeutic applications.

The biomechanics of turgor pressure

If you ever forget to water your houseplant, you may find its leaves getting soft and droopy - if you water it again in time, the leaves may stiffen, spring back up, and resist gravity. During this recovery, plant cells absorb water and build up an intracellular pressure, called turgor pressure, similar to inflating a balloon. Turgor pressure is an intrinsic component of plant physiology, and its biomechanical role as the 'hydroskeleton' is generally appreciated either statically in structural stability, like leaves resisting gravity, or dynamically in rapid motions, like Venus flytrap snapping, Mimosa closing, or stomatal opening. Slow but non-static processes like plant cell expansion also rely on turgor pressure, and it has been increasingly realized that turgor pressure and water fluxes in plant tissues play active roles in plant growth and morphogenesis. But where does turgor pressure come from, and how does it interact with other biomechanical properties of a plant cell? This primer aims to answer these questions by taking a brief tour from osmosis, to pressure vessel theory, then plant cell rheology. Although this primer is centered around intracellular pressure in plant cells, the biomechanical concepts are generally applicable to other organisms like bacteria and fungi, and even animal cells, which do not have cell walls, but are either embedded in stiff extracellular matrix (ECM) or are contracting (see the following section). To explain some of these concepts, we included a few equations, most of which are not hard to derive at all. We encourage readers to check the included examples, try for themselves, and look up derivations in suggested further reading materials.

Geometric constraint of mechanosensing by modification of hydrogel thickness prevents stiffness-induced differentiation in bone marrow stromal cells.

Extracellular matrix (ECM) stiffness is fundamental in cell division, movement and differentiation. The stiffness that cells sense is determined not only by the elastic modulus of the ECM material but also by ECM geometry and cell density. We hypothesized that these factors would influence cell traction-induced matrix deformations and cellular differentiation in bone marrow stromal cells (BMSCs). To achieve this, we cultivated BMSCs on polyacrylamide hydrogels that varied in elastic modulus and geometry and measured cell spreading, cell-imparted matrix deformations and differentiation. At low cell density BMSCs spread to a greater extent on stiff compared with soft hydrogels, or on thin compared with thick hydrogels. Cell-imparted matrix deformations were greater on soft compared with stiff hydrogels or thick compared with thin hydrogels. There were no significant differences in osteogenic differentiation relative to hydrogel elastic modulus and thickness. However, increased cell density and/or prolonged culture significantly reduced matrix deformations on soft hydrogels to levels similar to those on stiff substrates. This suggests that at high cell densities cell traction-induced matrix displacements are reduced by both neighbouring cells and the constraint imposed by an underlying stiff support. This may explain observations of the lack of difference in osteogenic differentiation as a function of stiffness.

Matrix stiffening from collagen fibril density and alignment modulates YAP-mediated T-cell immune suppression

T-cells are essential components of the immune system, adapting their behavior in response to the mechanical environments they encounter within the body. In pathological conditions like cancer, the extracellular matrix (ECM) often becomes stiffer due to increased density and alignment of collagen fibrils, which can have a significant impact on T-cell function. In this study, we explored how these ECM properties—density and fibrillar alignment—affect T-cell behavior using three-dimensional (3D) collagen matrices that mimic these conditions. Our results show that increased matrix stiffness, whether due to higher density or alignment, significantly suppresses T-cell activation, reduces cytokine production, and limits proliferation, largely through enhanced YAP signaling. Individually, matrix alignment appears to lower actin levels in activated T-cells and changes migration behavior in both resting and activated T-cells, an effect not observed in matrices with randomly oriented fibrils. Notably, inhibiting YAP signaling was able to restore T-cell activation and improve immune responses, suggesting a potential strategy to boost the effectiveness of immunotherapy in stiff ECM environments. Overall, this study provides new insights into how ECM characteristics influence T-cell function, offering potential avenues for overcoming ECM-induced immunosuppression in diseases such as cancer.