China
Africa
Explore chapters and articles related to this topic
Organoids as an Emerging Tool for Nano-Pharmaceuticals
Published in Harishkumar Madhyastha, Durgesh Nandini Chauhan, Nanopharmaceuticals in Regenerative Medicine, 2022
At a cellular level, after sensing the cues, these cues may be processed as external stimuli by modulating the microenvironment and inducing specific responses through activation of genetic programme, known as the mechanotransduction process (Chan et al. 2017, Davidson 2017). These biophysical signals are interpreted by a set of specialised proteins and their dysregulation, in turn, leads to various pathologies. Vinculin is an important mechanotransducter, serving as a linker for integrins to cytoskeleton, and pathogenic variations in this protein in humans are reported in various pathologies like cancer, cardiomyopathies, and neural defects (Olson et al. 2002, Vasile et al. 2006, Liang et al. 2014, Chinthalapudi et al. 2016).
Cell and Extracellular Matrix Interactions in a Dynamic Biomechanical Environment:
Published in Michel R. Labrosse, Cardiovascular Mechanics, 2018
External mechanical forces are detected by cells through a number of mechanosensors, proteins that are directly modified by mechanical force to change conformation and/or enzymatic activity, which can then modulate intracellular signaling pathways to regulate cell behavior and gene transcription. A number of mechanosensors play key roles in the activation and assembly of focal adhesions, including integrins (Friedland et al. 2009, Chen et al. 2012), talin (del Rio et al. 2009), FAK (Zhou et al. 2015), and p130Crk-associated substrate (p130CAS) (Sawada et al. 2006). Integrin conformational change between inactive and active states is assisted by intracellular force through actin–myosin contraction (Friedland et al. 2009) as well as by extracellular force applied by the attached ECM ligand (Chen et al. 2012). Talin links integrins to actin and contains helical bundles in its structure. These bundles unfold when mechanically stretched to expose vinculin-binding domains, allowing vinculin recruitment to focal adhesions (del Rio et al. 2009). Vinculin mediates actin cytoskeleton reorganization and focal adhesion stability and also recruits other focal adhesion proteins such as Arp2/3 and paxillin in a force-dependent manner (Carisey and Ballestrem 2011, Carisey et al. 2013). Full activation of FAK requires recruitment and phosphorylation by Src, a key tyrosine kinase that phosphorylates many focal adhesion-associated proteins, at tyrosines 576 and 577 (Calalb et al. 1995). Initial recruitment of FAK to integrin and talin allows autophosphorylation of tyrosin 397, which opens a binding site for Src (Schaller et al. 1994) and is a process that in silico models predict to be mechanically assisted by pulling forces between FAK’s attachment to the cell membrane and actin cytoskeleton (Zhou et al. 2015). Similar to talin, p130CAS also extends under applied force to expose its tyrosine phosphorylation site (Sawada et al. 2006). p130CAS is a substrate phosphorylated by Src (Ruest et al. 2001) and can enhance Src activity in reciprocity (Burnham et al. 2000). It is also an important scaffold for other proteins (Defilippi et al. 2006) and controls the activation of the Rac1 GTPase (Sharma and Mayer 2008).
Responsiveness of voltage-gated calcium channels in SH-SY5Y human neuroblastoma cells on micropillar substrates
Published in Journal of Biomaterials Science, Polymer Edition, 2018
Wenxu Wang, Donghuo Zhong, Yu Lin, Rong Fan, Zhengjun Hou, Xiumei Cao, Yubin Ren
For the immunofluorescence staining of F-actin, vinculin and N-type VGCC, the samples were washed with PBS twice, fixed with 4% paraformaldehyde (Electron Microscopy Science) in PBS for 20 min and permeabilized with 0.1% Triton-X100 (Sigma-Aldrich, U.S.A.) in PBS at room temperature for 15 min. Then, the cells were treated with 10% goat serum for 1 h to avoid unspecific labeling. Actin filaments were detected with FITC-conjugated Phalloidin (1:200, Sigma-Aldrich, U.S.A.) overnight at 4 °C. Vinculin were labeled with a mouse anti-vinculin monoclonal antibody (1:200, Santa, U.S.A.) overnight at 4 °C and followed by incubation with a secondary antibody coupled to Cy3 (1:200, Sigma, U.S.A.) for 1 h at 37 °C after washed with PBS twice. N-type VGCC were labeled with a rabbit anti-α1 subunit for Cav2.2 monoclonal antibody (1:200, Rabbit Anti-CaV2.2 antibody, Alomone Lab, Israel) overnight at 4 °C and followed by incubation with a secondary antibody coupled to Cy3 (1:200, Sigma, U.S.A.) for 1 h at 37 °C. The samples were washed with PBS twice and then stained by 1 μg/ml DAPI (Invitrogen, U.S.A.) for 10 min. Afterwards, samples were washed 3 times with PBS, mounted by glycerol and stored at 4 °C for observation.
Astral microtubules determine the final division axis of cells confined on anisotropic surface topography
Published in Journal of Experimental Nanoscience, 2020
Kyunghee Lee, Yen Ling Koon, Jaewon Kim, Keng-Hwee Chiam, Sungsu Park
Cortical cues have been shown to dictate spindle orientation of cells cultured on 2D FN coated surfaces [12]. To further investigate whether cortical cues can also affect spindle orientation of cells grown on microgratings, CD was treated to cells to perturb focal adhesion contacts and abolish cortical cues. Low concentrations of CD (0.2 μM) have been found to be sufficient to prevent membrane ruffling and disrupt treadmilling [28]. We immunostained RPE-1 and HeLa cells with vinculin both in the absence and presence of 1 µM CD. Vinculin is a membrane-cytoskeleton protein that is a constituent of focal adhesion plaques, and is involved in the linkage to actin. For control cells grown on 1 µm grating, vinculin accumulated along the ridges of the grating, with a preference for the ridge edge (Figure 2(A) top row). With the addition of CD, fluorescence intensities of both actin and vinculin decreased suggesting that focal adhesion cues were inhibited at these CD concentrations (Figure 2(A) bottom row). When cells are treated with 1 μM CD, aspect ratios for RPE-1 and HeLa increased by approximately 50% and 30%, respectively (Figure 2(B,C), top row and Table 4). In accordance to Hertwig’s rule, spindle angles of both cell lines decreased after CD treatment (Figure 2(B,C) bottom row and Table 4). To ensure consistency in our results, we also treated RPE-1 cells with another actin polymerisation inhibitor, Latrunculin A (Lat A). Lat A at low concentrations is known to induce the loss of stress fibers in fibroblasts [29,30]. Fluorescence intensities of both actin and vinculin disappeared in RPE-1 cells on the 1 μm grating when cells are treated with 100 nM Lat A for 12 h (Supplementary Figure 1(A)). In agreement with the CD results, aspect ratios of RPE-1 cells increased while spindle angles decreased in the presence of Lat A (Supplementary Figure 1(B,C)).
Regulation of stem cell fate and function by using bioactive materials with nanoarchitectonics for regenerative medicine
Published in Science and Technology of Advanced Materials, 2022
Wei Hu, Jiaming Shi, Wenyan Lv, Xiaofang Jia, Katsuhiko Ariga
The topographical features are one of the biophysical cues that cells in vivo encounter ranging from protein folding to collage nanofibrils [132]. Understanding the cell–nanotopography interaction enables us to a better design of implant materials and stem cell therapeutics. For example, Dalby et al. fabricated five nanotopographies with varying degrees of disorder using electron beam lithography. Nanoscale disorder is found to promote MSCs osteogenic differentiation and production of bone minerals. In addition, the osteogenic differentiation profile of MSCs topographically stimulated and dexamethasone treated is distinct. Following this study, they hypothesized that absolute square lattice symmetry can retain MSC multipotency and phenotype. On this nanostructured surface, MSCs show minimum metabolism and repress differentiation-related canonical signalling, which may be regulated by extracellular signal-regulated kinase (ERK) and small RNAs. It can maintain the self-renewal and differentiation capacity of MSCs over 8 weeks. As shown in Figure 4, Han et al. investigate how hMSC fates are temporally modulated by different lateral nanospacings (from 30 to 60 nm) [133]. Smaller spacings recruit more activated focal adhesion kinase (FAK) and Src proteins, along with more myosin IIA. It promotes vinculin recruitment to more mature and stable focal adhesions and transduction of higher tension forces (by >5 pN/FA). Consequently, nuclear YAP/TAZ localization is increased with further increasing RUNX2 and decreasing β-catenin. They explain the underlying molecular mechanisms by which smaller nanoscale spacings bias the osteogenic commitment of hMSCs. Werner et al. constructed 3D macrotopographic cell culture chips to investigate the impact of surface curvature on hMSCs behaviours. hMSCs show distinct migration regimes on convex and concave structures with the latter obviously faster. It can be interpreted by different cytoskeletal forces. In addition, cytoskeletal forces of convex surfaces lead to substantial nuclear deformation and increasing lamin-A levels, which promote osteogenic differentiation on convex spherical structures.