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Cells in the human body come across many types of information, which they respond to. Both material chemistry and topography of the surface where they adhere have an effect on cell shape, proliferation, migration, and gene expression. It is possible to create surfaces with topography at the nanometric scale to allow observation of cell-topography i...
Citations
... Note that only some of the above adhesion strength assays permit the acquisition of cell-level data to obtain the characteristics of individual cells in the assay. Parallel plate assay configurations [29][30][31][32][33] typically involve chamber heights >200 μm (Ref. 19) and can produce wall shear stress of up to a couple of tens of Pa. ...
Cell adhesion is of fundamental importance in cell and tissue organization and for designing cell-laden constructs for tissue engineering. Prior methods to assess cell adhesion strength for strongly adherent cells using hydrodynamic shear flow either involved the use of specialized flow devices to generate high shear stress or used simpler implementations like larger height parallel plate chambers that enable multihour cell culture but generate low wall shear stress and are, hence, more applicable for weakly adherent cells. Here, we propose a shear flow assay for adhesion strength assessment of strongly adherent cells that employs off-the-shelf parallel plate chambers for shear flow as well as simultaneous trypsin treatment to tune down the adhesion strength of cells. We implement the assay with a strongly adherent cell type and show that wall shear stress in the 0.07–7 Pa range is sufficient to dislodge the cells with simultaneous trypsin treatment. Imaging of cells over a square centimeter area allows cell morphological analysis of hundreds of cells. We show that the cell area of cells that are dislodged, on average, does not monotonically increase with wall shear stress at the higher end of wall shear stresses used and suggest that this can be explained by the likely higher resistance of high circularity cells to trypsin digestion. The adhesion strength assay proposed can be used to assess the adhesion strength of both weakly and strongly adherent cell types and has the potential to be adapted for substrate stiffness-dependent adhesion strength assessment in mechanobiology studies.
... Parallel plate assay configurations [29][30][31][32][33] typically involve chamber heights >200 µm [19] and can produce shear stress of up to a couple of tens of Pa. Microfluidic shear flow chambers are also effectively parallel-plate chambers but typically involve chamber heights <100 µm [19] and can produce up to 100s of Pa shear stress. ...
Cell adhesion is of fundamental importance in cell and tissue organization, and for designing cell-laden constructs for tissue engineering. Prior methods to assess cell adhesion strength for strongly adherent cells using hydrodynamic shear flow either involved the use of specialized flow devices to generate high shear stress or used simpler implementations like larger height parallel plate chambers that enable multi-hour cell culture but generate low shear stress and are hence more applicable for weakly adherent cells. Here, we propose a shear flow assay for adhesion strength assessment of strongly adherent cells that employs off-the-shelf parallel plate chambers for shear flow as well as simultaneous trypsin treatment to tune down the adhesion strength of cells. We implement the assay with a strongly adherent cell type and show that shear stress in the 0.07 to 7 Pa range is sufficient to dislodge the cells with simultaneous trypsin treatment. Imaging of cells over a square centimeter area allows cell morphological analysis of hundreds of cells. We show that the cell area of cells that are dislodged, on average, does not monotonically increase with shear stress at the higher end of shear stresses used and suggest that this can be explained by the likely higher resistance of high circularity cells to trypsin digestion. The adhesion strength assay proposed can be easily adapted by labs to assess the adhesion strength of both weakly and strongly adherent cell types and has the potential to be adapted for substrate stiffness-dependent adhesion strength assessment in mechanobiology studies.
... Similarly, centrifuge based methods have been developed to detach a fraction of the particles, depending on the adhesion strength between them and the surface [13,14]. Although this method can characterize the overall effect of surface coatings in some cases [15,16], its widespread application is hindered by certain drawbacks, most importantly the limited force range and inaccurate force characterization [17,18]. Furthermore, both shear flow and centrifuge-based methods measure the population average of the adhesion force of particles without the capability to target individual beads or cells. ...
Characterization of the binding of functionalized microparticles to surfaces with a specific chemistry sheds light on molecular scale interactions. Polymer or protein adsorption are often monitored by colloid particle deposition. Force measurements on microbeads by atomic force microscopy (AFM) or optical tweezers are standard methods in molecular biophysics, but typically have low throughput. Washing and centrifuge assays with (bio)chemically decorated microbeads provide better statistics, but only qualitative results without a calibrated binding force or energy value. In the present work we demonstrate that a computer controlled micropipette (CCMP) is a straightforward and high-throughput alternative to quantify the surface adhesion of functionalized microparticles. However, being an indirect force measurement technique, its in-depth comparison with a direct force measurement is a prerequisite of applications requiring calibrated adhesion force values. To this end, we attached polystyrene microbeads to a solid support by the avidin-biotin linkage. We measured the adhesion strength of the microbeads with both a specialized robotic fluid force microscope (FluidFM BOT) and CCMP. Furthermore, the bead-support contact zone was directly characterized on an optical waveguide biosensor to determine the density of avidin molecules. Distribution of the detachment force recorded on ∼50 individual beads by FluidFM BOT was compared to the adhesion distribution obtained from CCMP measurements on hundreds of individual beads. We found that both methods provide unimodal histograms. We conclude that FluidFM BOT can directly measure the detachment force curve of 50 microbeads in 150 min. CCMP can provide calibrated binding/adhesion force values of 120 microbeads in an hour.
... It is well known that particularly cells respond to certain surface topographies: groove-like structures on the micron scale, for example, encourage eukaryotic cells to migrate and to grow in elongated shape alongside these grooved patterns (Elter et al., 2009;Scheideler et al., 2003), which may have advantageous effects on the directed secretion of extracellular matrix and hence the reduced formation of scar tissue during the healing process of an implant (Wang et al., 2000). However, the topographic influence of surfaces on cell adhesion is not limited to dimensions which are characteristic for the morphology of the cell body; even nano-scaled surface structuring can have a significant influence on cell orientation (Monsees et al., 2005) as well as on specific cellular functions, like extracellular matrix (ECM) secretion, proliferation, and cell metabolism (Martines et al., 2004;Dalby et al., 2004). A more complicated situation arises, when micro-and nanostructure are superimposed; nevertheless, particularly this form of surface topography represents a most promising approach in order to mimic the morphology of naturally mineralized tissue (Tan and Saltzman, 2004). ...
Surface modification techniques can significantly improve the long-term in vivo performance
of biomedical implants as well as provide them with certain biological functions.
The judgement of the suitability of a certain treatment has to consider various
aspects, namely the specific properties of the chosen substrate material, the biological
environment of the respective application site, that is, the biological hard or soft tissue
adjacent to the implant surface as well as the composition of the involved body fluids,
and the particular medical situation of the patient. This chapter places the emphasis on
physical and chemical routes for surface treatment. Physical methods comprised the
fabrication of surface topographies with different scales (macro, micro, and nano),
surface treatment with PVD techniques, and the fabrication of nanotube arrays for
the application as drug delivery systems, while the subchapter about chemical modification
deals with methods to change the chemical surface properties as well as
chemical routes for the deposition of functional coatings on metallic substrates.
... Seeding concentration: 10 4 -10 6 cells /ml resulting in 100-1,000 cells/mm 2 [88], [91], [92]. ...
... In this chamber the motion of a sphere can be easily calculated. Cell trajectories, speed and adhesion process can be monitored [91]. It is a frequently applied design because of its simplicity. ...
... Endothelial cell adherence onto polymer surfaces with different hydrophilicity [90]. Investigation of nanopatterned surfaces [91]. Development of flow circuits, in which the behavior of cells can be continuously monitored on a microscope [88]. ...
Cell cell and cell matrix adhesions are fundamental in all multicellular organisms. They play a key role in cellular growth, differentiation, pattern formation and migration. Cell-cell adhesion is substantial in the immune response, pathogen host interactions, and tumor development. The success of tissue engineering and stem cell implantations strongly depends on the fine control of live cell adhesion on the surface of natural or biomimetic scaffolds. Therefore, the quantitative and precise measurement of the adhesion strength of living cells is critical, not only in basic research but in modern technologies, too. Several techniques have been developed or are under development to quantify cell adhesion. All of them have their pros and cons, which has to be carefully considered before the experiments and interpretation of the recorded data. Current review provides a guide to choose the appropriate technique to answer a specific biological question or to complete a biomedical test by measuring cell adhesion.
... It is well known that particularly cells respond to certain surface topographies: groove-like structures on the micron scale, for example, encourage eukaryotic cells to migrate and to grow in elongated shape alongside these grooved patterns (Elter et al., 2009;Scheideler et al., 2003), which may have advantageous effects on the directed secretion of extracellular matrix and hence the reduced formation of scar tissue during the healing process of an implant . However, the topographic influence of surfaces on cell adhesion is not limited to dimensions which are characteristic for the morphology of the cell body; even nano-scaled surface structuring can have a significant influence on cell orientation (Monsees et al., 2005) as well as on specific cellular functions, like extracellular matrix (ECM) secretion, proliferation, and cell metabolism (Martines et al., 2004;Dalby et al., 2004). A more complicated situation arises, when micro-and nanostructure are superimposed; nevertheless, particularly this form of surface topography represents a most promising approach in order to mimic the morphology of naturally mineralized tissue (Tan and Saltzman, 2004). ...
... Seeding concentration: 10 4 -10 6 cells /ml resulting in 100-1,000 cells/mm 2 [88], [91], [92]. ...
... In this chamber the motion of a sphere can be easily calculated. Cell trajectories, speed and adhesion process can be monitored [91]. It is a frequently applied design because of its simplicity. ...
... Endothelial cell adherence onto polymer surfaces with different hydrophilicity [90]. Investigation of nanopatterned surfaces [91]. Development of flow circuits, in which the behavior of cells can be continuously monitored on a microscope [88]. ...
Cell-cell and cell-matrix adhesions are fundamental in all multicellular organisms. They play a key role in cellular growth, differentiation, pattern formation and migration. Cell-cell adhesion is substantial in the immune response, pathogen-host interactions, and tumor development. The success of tissue engineering and stem cell implantations strongly depends on the fine control of live cell adhesion on the surface of natural or biomimetic scaffolds. Therefore, the quantitative and precise measurement of the adhesion strength of living cells is critical, not only in basic research but in modern technologies, too. Several techniques have been developed or are under development to quantify cell adhesion. All of them have their pros and cons, which has to be carefully considered before the experiments and interpretation of the recorded data. Current review provides a guide to choose the appropriate technique to answer a specific biological question or to complete a biomedical test by measuring cell adhesion.
Copyright © 2019 Elsevier B.V. All rights reserved.
... Curvature is another important consideration of ECM, as cells contain curvature sensing proteins, and, for example, nanopitpatterned surfaces decrease cell adhesion compared to flat substrates (Martines et al. 2004). The BAR domain comprises a curved protein domain that self-assembles and can sense curvature or induce curvature in membrane surfaces (Chen et al. 2012). ...
Mechanosensing is increasingly recognised as important for tumour progression. Tumours become stiff and the forces that normally balance in the healthy organism break down and become imbalanced, leading to increases in migration, invasion and metastatic dissemination. Here, we review recent advances in our understanding of how extracellular matrix properties, such as stiffness, viscoelasticity and architecture control cell behaviour. In addition, we discuss how the tumour microenvironment can be modelled in vitro, capturing these mechanical aspects, to better understand and develop therapies against tumour spread. We argue that by gaining a better understanding of the microenvironment and the mechanical forces that govern tumour dynamics, we can make advances in combatting cancer dormancy, recurrence and metastasis.
Electronic supplementary material
The online version of this article (10.1007/s12551-018-0466-8) contains supplementary material, which is available to authorized users.
... It uses a combination of a parallel flow chamber device and digital image correlation (DIC) techniques to study the deformation of biological cells in fluidic environments that are comparable to cell physiological environments. In particular, the parallel flow chamber device has been used extensively to study cell adhesion (Vankooten et al., 1992;Martines et al., 2004;Cao et al., 1997;Giavazzi et al., 1993;Hetrick and Schoenfisch, 2007), drug effects (Yellen et al., 2005;Sugiura et al., 2008;Geng et al., 2007) and cell physiological changes (Reinhart-King et al., 2008;Vaughan et al., 2013). However, there is only one prior paper (from our group) (Cao et al., 2006) on the use of the shear assay technique for the measurement of the cell viscoelasticity of human osteosarcoma cells (Cao et al., 2006). ...
An improved understanding of the evolution of cell structure and viscoelasticity with cancer malignancy could enable the development of a new generation of biomarkers and methods for cancer diagnosis. Hence, in this study, we present the viscoelastic properties (moduli and viscosities) and the actin cytoskeletal structures of triple negative breast cancer (TNBC) cells with different metastatic potential. These include: MCF-10A normal breast cells (studied as a control); MDA-MB-468 cells (less metastatic TNBC cells), and MDA-MB-231 cells (highly metastatic TNBC cells). A combination of shear assay and digital imaging correlation (DIC) techniques is used to measure the local viscoelastic properties of live breast cells subjected to constant shear stress. The local moduli and viscosities of the nuclei and cytoplasm are characterized using a generalized Maxwell model, which is used to determine the time-dependent creep responses of cells. The nuclei are shown to be stiffer and more viscous than the cytoplasms of the normal breast cells and TNBC cells. The MCF-10A normal breast cells are found to be twice as stiff as the less metastatic MDA-MB-468 breast cancer cells and over ten times stiffer than the highly metastatic MDA-MB-231 breast cancer cells. Similar trends are also observed in the viscosities of the nuclei and the cytoplasms. The measured differences in cell viscoelastic properties are also associated with significant changes in the cell cytoskeletal structure, which is studied using confocal fluorescence microscopy. This reveals significant differences in the levels of actin expression and organization in TNBC cells as they become highly metastatic. Our results suggest that the shear assay measurements of cell viscoelastic properties may be used as effective biomarkers for TNBC diagnosis and screening.
... Traditionally, parallel plate flow chamber systems have been used to quantify bacterial adhesion and detachment (Martines et al. 2004;Busscher & Van der Mei 2006;Simões et al. 2008). Recently, an interproximal (IP) space mimicked microfluidic funnel device has been used to investigate the shear tolerance of S. mutans aggregates (Shumi et al. 2013). ...
Well-established biofilms formed by Streptococcus mutans via exopolysaccharide matrix synthesis are firmly attached to tooth surfaces. Enhanced understanding of the physical properties of mature biofilms may lead to improved approaches to detaching or disassembling these highly organized and adhesive structures. Here, the mechanical stability of S. mutans biofilms was investigated by determining their ability to withstand measured applications of shear stress using a custom-built device. The data show that the initial biofilm bulk (~ 50% biomass) was removed after exposure to 0.184 and 0.449 N m−2 for 67 and 115 h old biofilms. However, removal of the remaining biofilm close to the surface was significantly reduced (vs initial bulk removal) even when shear forces were increased 10-fold. Treatment of biofilms with exopolysaccharide-digesting dextranase substantially compromised their mechanical stability and rigidity, resulting in bulk removal at a shear stress as low as 0.027 N m−2 and > a two-fold reduction in the storage modulus (G′). The data reveal how incremental increases in shear stress cause distinctive patterns of biofilm detachment, while demonstrating that the exopolysaccharide matrix modulates the resistance of biofilms to mechanical clearance.
