Cell differentiation through tissue elasticity-coupled, myosin-driven remodeling

https://doi.org/10.1016/j.ceb.2008.09.006Get rights and content

Cells may lack eyes to see and ears to hear, but cells do seem to have a sense of ‘touch’ that allows them to feel their microenvironment. This is achieved in part through contractility coupled adhesion to physically flexible ‘soft’ tissue. Here we summarize some of the known variations in elasticity of solid tissue and review some of the long-term effects of cells ‘feeling’ this elasticity, focusing on differentiation processes of both committed cell types and stem cells. We then highlight what is known of molecular remodeling in cells under stress on short time scales. Key roles for forces generated by ubiquitous and essential myosin-II motors in feedback remodeling are emphasized throughout.

Introduction

Solid tissues are soft, with the exception of bone, and they possess an elasticity E that not only varies between many different tissues (Figure 1a) but also exerts a considerable influence on how resident tissue cells behave. Indeed, cells that are dissociated from a solid tissue are generally not viable in a fluid suspension  they must adhere to a ‘solid’ which, by definition, recovers its shape after pushing and pulling, even at the scale of a cell. Solid tissues such as skin, muscle, and brain, are all relatively elastic, with the macroscale elasticity evident in their recovery of shape within seconds after mild poking and pinching or even after sustained compression, such as sitting. This is in contrast to fluid tissues such as blood and lymph which flow readily on a similar time scale and contain distinct cells, such as red and white blood cells, that are functional without sustained attachment. The impact of solid tissue elasticity on adherent cells is the focus here, with recent insights from stem cells and structural proteomics adding to past reviews (e.g. [1]) of findings that indicate tissue elasticity E is felt by cells, affecting cell structure and function.

Both matrix composition and cell activity contribute to tissue elasticity or stiffness at a scale that cells can actively probe and sense. With collagen as an example: collagen type, amount, diameter of fibers, crosslinking (e.g. cellular lysyl oxidase activity), plus noncovalent interactions with other matrix proteins will all contribute to the matrix elasticity. Recent measurements of the elasticity of zebrafish embryos [2••] that were treated with the nonmuscle myosin-II (NMM II)-specific inhibitor blebbistatin also document a dramatic decrease in the effective elasticity, illustrating the contribution of myosin-derived tension (like tension in a guitar string) to the elasticity of the entire organism. While adherent tissue cells and extracellular matrix contribute to a characteristic if not strictly tissue-specific elastic microenvironment, cells generally anchor and pull on their surroundings through myosin-II-based contractility and transcellular adhesions of integrins plus other adhesion molecules [3]. The resistance felt by a given cell derives from tissue matrix, an adjacent cell, or perhaps  in culture  a synthetic substrate intended to model soft tissue (Figure 1b). Disease can bring significant changes in tissue elasticity: indeed, ‘sclerosis’  as in atherosclerosis, otosclerosis, scleroderma, and more  is greek for hardening of tissue.

Contractile forces generated by ubiquitous crossbridging interactions of actin and myosin-II filaments in stress fibers are transmitted to the substrate as ‘traction’ forces that cause visible wrinkles in a thin film or lateral displacements of markers at the surface of a soft gel [4, 5, 6, 7]. On gels with collagen-I covalently attached, epithelial cells and fibroblasts [8••] were the first cells reported to detect and respond distinctly to soft versus stiff substrates; differences were suggested to depend on myosin-II as they were inhibited by BDM (2,3-butanedione monoxime)  although this drug is now known to have multiple effects beyond myosin inhibition. Since then, neurons [9, 10], muscle cells of various types [11, 12, 13], mesenchymal stem cells (MSCs) [14••], plus many other tissue cell types [15, 16, 17, 18] have been shown to sense substrate stiffness, and at least some of the results have confirmed the importance of nonmuscle myosin-II through the inhibition of elasticity-dependent behavior changes with blebbistatin. Most cell types are found to respond to the elasticity E of the substrate within hours by spreading and assembling both adhesions and cytoskeleton in proportion to E up to some saturating value beyond which changes in E exert no influence. Given that an isoform of myosin-II is also responsible for the work done by skeletal muscle, an analogy to lifting weights and exercise seems appropriate: to your bicep, a load of 1 kg undoubtedly feels very different from a load of 10 kg, whereas pushing or pulling on an immovable object like the handle of a locked door is a very distinct isometric exercise. Similar sensitivity to E seems to apply to most anchored cell types with similar implications for growth and remodeling within individual cells.

Section snippets

Soft tissue E measurements and model systems

The intrinsic resistance of a solid to a stress, regardless of topography and thickness (e.g. basement membrane), is measured by the solid’s elastic modulus E, which is most simply obtained by applying a force  such as poking with an atomic force microscope (AFM) [19]  to a section of tissue or other substrate and then measuring the relative displacement. Tissues with small E show larger indentations or displacements under a given force.

E appears to adequately characterize many tissues not only

Stem cells are particularly E-sensitive

MSCs appear especially sensitive to tissue elasticity. These cells reside in the bone marrow and are believed to enter the circulation and contribute to tissue regeneration and repair after injury, such as a muscle tear. Bone marrow aspirates are either fluid, with an ‘intercellular substance’ measured decades ago to be about 100-fold more viscous than water [29], or else have a very small E [30], and the rare MSCs in marrow are generally separated from the many other marrow cell types by their

Matrix-coupled, myosin-driven remodeling

Molecular mechanisms of elasticity sensing by cells seem likely to be collective and dependent on many interacting components of the cyto-adhesion apparatus. Cell tension is expected to be important, and differentiation of MSCs was indeed blocked by myosin-II inhibition with blebbistatin [14••]. In addition, lineage specification was associated with significant changes in the levels NMM IIB and C, with considerable downregulation on soft matrices (0.1–1 kPa) and modest upregulation on stiffer

Cys shotgun and other methods begin clarifying molecular dynamics

Cysteine is a reactive but relatively hydrophobic amino acid that is often buried within tertiary or quaternary structures. Reactivity of cysteine’s thiol group had been exploited in solution to a limited extent to probe protein interactions [43] and folding [44], and in situ Cys labeling of membrane proteins such as GPCRs had yielded insights into accessibility and ligand-induced changes in individual proteins [45]. Proteomic-scale Cys shotgun labeling of intact cells has now been shown to be

References and recommended reading

Papers of particular interest, published within the period of review, have been highlighted as:

  • • of special interest

  • •• of outstanding interest

Acknowledgements

We are grateful for grant support from the NSF, MDA, NIH (NHLBI, NIBIB, NIDDK), and NIH TG support (AZ).

References (48)

  • N. Bhasin et al.

    Chemistry on a single protein, vascular cell adhesion molecule-1, during forced unfolding

    J Biol Chem

    (2004)
  • S.D. Ward et al.

    Use of an in situ disulfide cross-linking strategy to study the dynamic properties of the cytoplasmic end of transmembrane domain VI of the M3 muscarinic acetylcholine receptor

    Biochemistry

    (2006)
  • C.P. Johnson et al.

    Forced unfolding of proteins within cells

    Science

    (2007)
  • Y. Sawada et al.

    Force sensing by mechanical extension of the Src family kinase substrate p130Cas

    Cell

    (2006)
  • D.E. Discher et al.

    Tissue cells feel and respond to the stiffness of their substrate

    Science

    (2005)
  • M. Krieg et al.

    Tensile forces govern germ-layer organization in zebrafish

    Nat Cell Biol

    (2008)
  • M.A. Schwartz et al.

    Cell adhesion receptors in mechanotransduction

    Curr Opin Cell Biol

    (2008)
  • A.K. Harris et al.

    Silicone rubber substrata: a new wrinkle in the study of cell locomotion

    Science

    (1980)
  • W.A. Marganski et al.

    Measurements of cell-generated deformations on flexible substrata using correlation-based optical flow

    Methods Enzymol

    (2003)
  • R.J. Pelham et al.

    Cell locomotion and focal adhesions are regulated by substrate flexibility

    Proc Natl Acad Sci U S A

    (1997)
  • L.A. Flanagan et al.

    Neurite branching on deformable substrates

    Neuroreport

    (2002)
  • P.C. Georges et al.

    Matrices with compliance comparable to that of brain tissue select neuronal over glial growth in mixed cortical cultures

    Biophys J

    (2006)
  • A. Engler et al.

    Substrate compliance versus ligand density in cell on gel responses

    Biophys J

    (2004)
  • A.J. Engler et al.

    Matrix elasticity directs stem cell lineage specification

    Cell

    (2006)
  • Cited by (84)

    • Engineering scaffolds for tissue engineering and regenerative medicine

      2022, Advances in Biomedical Polymers and Composites: Materials and Applications
    • Interactions at scaffold interfaces: Effect of surface chemistry, structural attributes and bioaffinity

      2019, Materials Science and Engineering C
      Citation Excerpt :

      The force application to bound integrins encourages focal adhesion assembly and causes cytoskeletal contraction and actin filament polymerization. Contractile forces are transmitted as the traction forces over to the substrate and these allow the cells to sense their surrounding matrix and judge changes in matrix elasticity [74]. The extracellular portion of integrins binds to various ECM proteins- collagen, laminin, fibronectin, and biomaterial scaffolds to mimic these ECM functions.

    • Differentiation of stem cells from the apical papilla into osteoblasts by the elastic modulus of porous silk fibroin scaffolds

      2019, Biologicals
      Citation Excerpt :

      Studies have shown that behaviour of differentiation in mesenchymal stem cell can be directed to different lineages by substrate modulus. It was reported that hBMSCs cultured on tow dimensional (2D) matrix with low (<1 kPa), intermediate (8 kPa-15 kPa), and high (25 kPa-40 kPa) modulus increase expression of neuronal, myogenic and osteogenic markers, respectively [8–12]. In addition, seeding of MSCs on three-dimensional (3-D) polyethylene glycol dimethacrylate nanofiber hydrogel matrix with elasticity range of 2 -15 kPa exhibited endothelial marker and smooth muscle marker expression in the soft and stiff area, respectively [7].

    • Poly (butylene adipate-co-terephthalate)/hydroxyapatite composite structures for bone tissue recovery

      2015, Polymer Degradation and Stability
      Citation Excerpt :

      It can be observed that the addition of nHA considerably improved the mechanical properties of the composites structures, by increasing 12.7% the elastic modulus, 25.7% the stress at break and 52% the elongation at break. Different studies have indicated that the elasticity of the substrate is an important regulator for cellular function [25–27]; for example, mesenchymal stem cells (MSCs) appear particularly sensitive to tissue elasticity [28]. The pre-calcified to rigid compact bone has an elasticity that varies from 100 kPa to 20 GPa [28]; thus, probably a composite material with an elastic modulus within this range should guide the hASC differentiation towards osteogenic lineages.

    View all citing articles on Scopus
    View full text