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ACS Applied Materials & Interfaces Feb 2024The complex interplay between cells and materials is a key focus of this research, aiming to develop optimal scaffolds for regenerative medicine. The need for tissue...
The complex interplay between cells and materials is a key focus of this research, aiming to develop optimal scaffolds for regenerative medicine. The need for tissue regeneration underscores understanding cellular behavior on scaffolds, especially cell adhesion to polymer fibers forming focal adhesions. Key proteins, paxillin and vinculin, regulate cell signaling, migration, and mechanotransduction in response to the extracellular environment. This study utilizes advanced microscopy, specifically the AiryScan technique, along with advanced image analysis employing the Density-Based Spatial Clustering of Applications with Noise (DBSCAN) cluster algorithm, to investigate protein distribution during osteoblast cell adhesion to polymer fibers and glass substrates. During cell attachment to both glass and polymer fibers, a noticeable shift in the local maxima of paxillin and vinculin signals is observed at the adhesion sites. The focal adhesion sites on polymer fibers are smaller and elliptical but exhibit higher protein density than on the typical glass surface. The characteristics of focal adhesions, influenced by paxillin and vinculin, such as size and density, can potentially reflect the strength and stability of cell adhesion. Efficient adhesion correlates with well-organized, larger focal adhesions characterized by increased accumulation of paxillin and vinculin. These findings offer promising implications for enhancing scaffold design, evaluating adhesion to various substrates, and refining cellular interactions in biomedical applications.
Topics: Paxillin; Vinculin; Focal Adhesions; Mechanotransduction, Cellular; Cell Adhesion; Polymers; Phosphoproteins; Focal Adhesion Protein-Tyrosine Kinases
PubMed: 38354103
DOI: 10.1021/acsami.3c19035 -
Medical Sciences (Basel, Switzerland) Aug 2022Vinculin and talin-1, which are cytoskeletal proteins affecting focal adhesions, were reported to be down-expressed in atherosclerotic lesions. Recently, we reported...
Vinculin and talin-1, which are cytoskeletal proteins affecting focal adhesions, were reported to be down-expressed in atherosclerotic lesions. Recently, we reported high concentrations of plasma talin-1 in patients with coronary artery disease (CAD). However, blood vinculin concentrations in CAD patients have not been clarified. Plasma vinculin concentrations as well as talin-1 were studied in 327 patients in whom coronary angiography was performed. CAD was proven in 177 patients (1-vessel, = 79; 2-vessel, = 57; 3-vessel disease, = 41). However, vinculin concentrations were not markedly different between the CAD(-) and CAD groups (median 122.5 vs. 119.6 pg/mL, = 0.325) or among patients with CAD(-), 1-, 2-, and 3-vessel diseases (122.5, 112.8, 107.9, and 137.2 pg/mL, = 0.202). In contrast, talin-1 concentrations were higher in CAD than the CAD(-) group (0.29 vs. 0.23 ng/mL, = 0.006) and increased stepwise in the number of stenotic vessels: 0.23 in CAD(-), 0.28 in 1-vessel, 0.29 in 2-vessel, and 0.33 ng/mL in 3-vessel disease ( = 0.043). No correlation was observed between vinculin and talin-1 concentrations. In multivariate analysis, vinculin concentrations were not a factor for CAD. In conclusion, plasma vinculin concentrations in patients with CAD were not high and were not associated with the presence or severity of CAD.
Topics: Coronary Angiography; Coronary Artery Disease; Focal Adhesions; Humans; Talin; Vinculin
PubMed: 36135831
DOI: 10.3390/medsci10030046 -
The Journal of International Medical... Jan 2020This study aimed to explore the expression of vinculin in non-small cell lung cancer (NSCLC) and to analyze its correlation with clinical features and prognosis of NSCLC.
OBJECTIVE
This study aimed to explore the expression of vinculin in non-small cell lung cancer (NSCLC) and to analyze its correlation with clinical features and prognosis of NSCLC.
METHODS
The expression of vinculin in cancer tissues and paracancer tissues was detected by real-time PCR, western blotting, and immunohistochemistry. Correlations between vinculin-positive expression and clinical features were analyzed by Pearson correlation analysis, and those between vinculin expression and prognosis were analyzed by Cox multivariate analysis.
RESULTS
Vinculin expression was significantly lower in cancer tissues than in paracancer tissues. Pearson correlation analysis showed that vinculin expression was significantly correlated with tumor–node–metastasis (TNM) stage and lymph node metastasis in NSCLC. Cox multivariate analysis showed that vinculin-negative expression and TNM stage were independent risk factors for NSCLC prognosis. Kaplan–Meier analysis showed that the 5-year overall survival (OS) rate was 20.51% for all NSCLC patients, and was significantly higher for vinculin-positive patients with NSCLC than vinculin-negative patients.
CONCLUSIONS
Vinculin gene transcription is inhibited in NSCLC, and low vinculin expression promotes malignancy in NSCLC. Therefore, vinculin could be used as a prognostic marker for NSCLC and a potential target for its treatment.
Topics: Carcinoma, Non-Small-Cell Lung; Female; Gene Expression Regulation, Neoplastic; Humans; Lung Neoplasms; Male; Middle Aged; Multivariate Analysis; Neoplasm Staging; Prognosis; Proportional Hazards Models; RNA, Messenger; Regression Analysis; Survival Analysis; Vinculin
PubMed: 30947597
DOI: 10.1177/0300060519839523 -
Journal of Biomaterials Applications Jan 2021Keratin-based biomaterials represent an attractive opportunity in the fields of wound healing and tissue regeneration, not only for their chemical and physical...
Keratin-based biomaterials represent an attractive opportunity in the fields of wound healing and tissue regeneration, not only for their chemical and physical properties, but also for their ability to act as a delivery system for a variety of payloads. Importantly, keratins are the only natural biomaterial that is not targeted by specific tissue turnover-related enzymes, giving it potential stability advantages and greater control over degradation after implantation. However, in-situ polymerization chemistry in some keratin systems are not compatible with cells, and incorporation within constructs such as hydrogels may lead to hypoxia and cell death. To address these challenges, we envisioned a pre-formed keratin microparticle on which cells could be seeded, while other payloads (e.g. drugs, growth factors or other biologic compounds) could be contained within, although studies investigating the potential partitioning between phases during emulsion polymerization would need to be conducted. This study employs well-established water-in-oil emulsion procedures as well as a suspension culture method to load keratin-based microparticles with bone marrow-derived mesenchymal stem cells. Fabricated microparticles were characterized for size, porosity and surface structure and further analyzed to investigate their ability to form gels upon hydration. The suspension culture technique was validated based on the ability for loaded cells to maintain their viability and express actin and vinculin proteins, which are key indicators of cell attachment and growth. Maintenance of expression of markers associated with cell plasticity was also investigated. As a comparative model, a collagen-coated microparticle (Sigma) of similar size was used. Results showed that an oxidized form of keratin ("keratose" or "KOS") formed unique microparticle structures of various size that appeared to contain a fibrous sub-structure. Cell adhesion and viability was greater on keratin microparticles compared to collagen-coated microparticles, while marker expression was retained on both.
Topics: Actins; Cell Adhesion; Coated Materials, Biocompatible; Collagen; Humans; Hydrogels; Keratins; Mesenchymal Stem Cells; Microspheres; Porosity; Surface Properties; Tissue Engineering; Tissue Scaffolds; Vinculin; Wound Healing
PubMed: 32847463
DOI: 10.1177/0885328220951892 -
The Journal of Cell Biology Jan 2024Vinculin is an actin-binding protein (ABP) that strengthens the connection between the actin cytoskeleton and adhesion complexes. It binds to β-catenin/N-cadherin...
Vinculin is an actin-binding protein (ABP) that strengthens the connection between the actin cytoskeleton and adhesion complexes. It binds to β-catenin/N-cadherin complexes in apical adherens junctions (AJs), which maintain cell-to-cell adhesions, and to talin/integrins in the focal adhesions (FAs) that attach cells to the basal membrane. Here, we demonstrate that β-catenin targets vinculin to the apical AJs and the centrosome in the embryonic neural tube (NT). Suppression of vinculin slows down the basal-to-apical part of interkinetic nuclear migration (BAINM), arrests neural stem cells (NSCs) in the G2 phase of the cell cycle, and ultimately dismantles the apical actin cytoskeleton. In the NSCs, mitosis initiates when an internalized centrosome gathers with the nucleus during BAINM. Notably, our results show that the first centrosome to be internalized is the daughter centrosome, where β-catenin and vinculin accumulate, and that vinculin suppression prevents centrosome internalization. Thus, we propose that vinculin links AJs, the centrosome, and the actin cytoskeleton where actomyosin contraction forces are required.
Topics: Actin Cytoskeleton; Actins; beta Catenin; Cell Adhesion; Cell Cycle; Focal Adhesions; Vinculin; Neural Stem Cells; Centrosome; Adherens Junctions
PubMed: 37889294
DOI: 10.1083/jcb.202106169 -
Cell Biology International Aug 2018The focal adhesion protein vinculin has been implicated in associating with soluble and membranous phospholipids. Detailed investigations over the past ten years...
The focal adhesion protein vinculin has been implicated in associating with soluble and membranous phospholipids. Detailed investigations over the past ten years describe the intermolecular interactions of the vinculin tail domain with soluble and membrane phospholipids. Previous studies have implied that the tail's unstructured C-terminal region affects the mechanical behavior of cells and that the same region, at the molecular level, has bi-stable behavior sensitive to different protonation states. The aim of this short communication is to discuss whether the C-terminal vinculin tail (Vt) domain interacts favorably with membrane-embedded phospholipids such as PIP and that the region is also an anchor for lipid membranes.
Topics: Animals; Circular Dichroism; Lipid Bilayers; Molecular Dynamics Simulation; Phosphatidylinositol 4,5-Diphosphate; Phospholipids; Protein Binding; Protein Domains; Vinculin
PubMed: 29696730
DOI: 10.1002/cbin.10978 -
Current Biology : CB Oct 2022Filopodia are narrow cell extensions involved in various physiological processes. Integrins mediate filopodia adhesion and likely transmit adhesive force to regulate...
Filopodia are narrow cell extensions involved in various physiological processes. Integrins mediate filopodia adhesion and likely transmit adhesive force to regulate filopodia formation and functions, but the force is extremely weak to study and remains poorly understood. Using integrative tension sensor (ITS), we imaged filopodia adhesive force at the single molecular tension level and investigated the force dynamics and sources. Results show that filopodia integrin tension (FIT) is generated in discrete foci (force nodes) along single filopodia with a spacing of ∼1 μm. Inhibitions of actin polymerization or myosin II activity markedly reduced FIT signals in force nodes at filopodia tips and at filopodia bases, respectively, suggesting differential force sources of FIT in the distal force nodes and proximal ones in filopodia. Using two ITS constructs with different force thresholds for activation, we showed that the molecular force level of FIT is greater at filopodia bases than that at filopodia tips. We also tested the role of vinculin and myosin X in the FIT transmission. With vinculin knockout in cells, filopodia and associated force nodes were still formed normally, suggesting that vinculin is dispensable for the formation of filopodia and force nodes. However, vinculin is indeed required for the transmission of strong FIT (capable of rupturing DNA in a shear conformation), as the strong FIT vanished in filopodia with vinculin knockout. Co-imaging of FIT and myosin X shows no apparent co-localization, demonstrating that myosin X is not directly responsible for generating FIT, despite its prominent role in filopodium elongation.
Topics: Pseudopodia; Vinculin; Integrins; Actins; Adhesives; Myosins
PubMed: 36084647
DOI: 10.1016/j.cub.2022.08.040 -
Biophysical Journal Jan 2023Focal adhesions (FAs) transmit force and mediate mechanotransduction between cells and the matrix. Previous studies revealed that integrin-transmitted force is critical...
Focal adhesions (FAs) transmit force and mediate mechanotransduction between cells and the matrix. Previous studies revealed that integrin-transmitted force is critical to regulate FA formation. As vinculin is a prominent FA protein implicated in integrin tension transmission, this work studies the relation among integrin tensions (force), vinculin (protein), and FA formation (structure) by integrin tension manipulation, force visualization and vinculin knockout (KO). Two DNA-based integrin tension tools are adopted: tension gauge tether (TGT) and integrative tension sensor (ITS), with TGT restricting integrin tensions under a designed T (tension tolerance) value and ITS visualizing integrin tensions above the T value by fluorescence. Results show that large FAs (area >1 μm) were formed on the TGT surface with T of 54 pN but not on those with lower T values. Time-series analysis of FA formation shows that focal complexes (area <0.5 μm) appeared on all TGT surfaces 20 min after cell plating, but only matured to large FAs on TGT with T of 54 pN. Next, we tested FA formation in vinculin KO cells on TGT surfaces. Surprisingly, the T value of TGT required for large FA formation is drastically decreased to 23 pN. To explore the cause, we visualized integrin tensions in both wild-type and vinculin KO cells using ITS. The results showed that integrin tensions in FAs of wild-type cells frequently activate ITS with T of 54 pN. With vinculin KO, however, integrin tensions in FAs became lower and unable to activate 54 pN ITS. Force signal intensities of integrin tensions reported by 33 and 43 pN ITS were also significantly reduced with vinculin KO, suggesting that vinculin is essential to transmit high-level integrin tensions and involved in transmitting intermediate-level integrin tensions in FAs. However, the high-level integrin tensions transmitted by vinculin are not required by FA formation.
Topics: Integrins; Focal Adhesions; Cell Adhesion; Vinculin; Mechanotransduction, Cellular
PubMed: 36352785
DOI: 10.1016/j.bpj.2022.11.013 -
Frontiers in Oncology 2022The onset and progression of cancer are strongly associated with the dissipation of adhesion forces between cancer cells, thus facilitating their incessant attachment...
The onset and progression of cancer are strongly associated with the dissipation of adhesion forces between cancer cells, thus facilitating their incessant attachment and detachment from the extracellular matrix (ECM) to move toward metastasis. During this process, cancer cells undergo mechanical stresses and respond to these stresses with membrane deformation while inducing protrusions to invade the surrounding tissues. Cellular response to mechanical forces is inherently related to the reorganization of the cytoskeleton, the dissipation of cell-cell junctions, and the adhesion to the surrounding ECM. Moreover, the role of focal adhesion proteins, and particularly the role of vinculin in cell attachment and detachment during migration, is critical, indicating the tight cell-ECM junctions, which favor or inhibit the metastatic cascade. The biomechanical analysis of these sequences of events may elucidate the tumor progression and the potential of cancer cells for migration and metastasis. In this work, we focused on the evaluation of the spreading rate and the estimation of the adhesion strength between breast cancer cells and ECM prior to and post-treatment with anti-tumor agents. Specifically, different tamoxifen concentrations were used for ER breast cancer cells, while even concentrations of trastuzumab and pertuzumab were used for HER2 cells. Analysis of cell stiffness indicated an increased elastic Young's modulus post-treatment in both MCF-7 and SKBR-3 cells. The results showed that the post-treatment spreading rate was significantly decreased in both types of breast cancer, suggesting a lower metastatic potential. Additionally, treated cells required greater adhesion forces to detach from the ECM, thus preventing detachment events of cancer cells from the ECM, and therefore, the probability of cell motility, migration, and metastasis was confined. Furthermore, post-detachment and post-treatment vinculin levels were increased, indicating tighter cell-ECM junctions, hence limiting the probability of cell detachment and, therefore, cell motility and migration.
PubMed: 36052248
DOI: 10.3389/fonc.2022.811508 -
Molecular Biology of the Cell Sep 2022Vinculin is a protein found in both focal adhesions (FAs) and adherens junctions (AJs) which regulates actin connectivity to these structures. Many studies have...
Vinculin is a protein found in both focal adhesions (FAs) and adherens junctions (AJs) which regulates actin connectivity to these structures. Many studies have demonstrated that mechanical perturbations of cells result in enhanced recruitment of vinculin to FAs and/or AJs. Likewise, many other studies have shown "cross-talk" between FAs and AJs. Vinculin itself has been suggested to be a probable regulator of this adhesion cross-talk. In this study we used MDCK as a model system of epithelia, developing cell lines in which vinculin recruitment was reduced or enhanced at AJs. Careful analysis of these cells revealed that perturbing vinculin recruitment to AJs resulted in a reduction of detectable FAs. Interestingly the cross-talk between these two structures was not due to a limited pool of vinculin, as increasing expression of vinculin did not rescue FA formation. Instead, we demonstrate that vinculin translocation between AJs and FAs is necessary for actin cytoskeleton rearrangements that occur during cell migration, which is necessary for large, well-formed FAs. Last, we show using a wound assay that collective cell migration is similarly hindered when vinculin recruitment is reduced or enhanced at AJs, highlighting that vinculin translocation between each compartment is necessary for efficient collective migration.
Topics: Adherens Junctions; Catenins; Cell Adhesion; Focal Adhesions; Vinculin; alpha Catenin
PubMed: 35921161
DOI: 10.1091/mbc.E22-02-0071