Corneal Fibroblasts Respond Rapidly to Changes in Local Mechanical Stress

Department of Ophthalmology, University of Texas Southwestern Medical Center, Dallas, Texas 75390-9057, USA.
Investigative Ophthalmology & Visual Science (Impact Factor: 3.4). 11/2004; 45(10):3466-74. DOI: 10.1167/iovs.04-0361
Source: PubMed


To investigate the response of corneal fibroblasts to local changes in extracellular matrix (ECM) tension.
Rabbit and human corneal fibroblasts were plated inside fibrillar collagen matrices. After 18 to 72 hours, a glass microneedle was inserted into the ECM and either pushed toward a cell to reduce local tension, or pulled away to increase tension. Time-lapse differential interference contrast (DIC) imaging was performed both before and after needle micromanipulation. ECM displacements were quantified, and strain maps were generated by finite element modeling. In some experiments, cells were treated with the Rho-kinase inhibitor Y-27632 either 30 minutes before, or 1 hour after they were pushed with the microneedle. Changes in focal adhesion organization were also evaluated in a subset of cells expressing green fluorescent protein (GFP)-zyxin, by simultaneous fluorescent and DIC imaging.
Pulling on the ECM resulted in initial cell elongation, followed by disengagement and retraction of pseudopodia. In contrast, pushing the ECM toward a cell induced rapid shortening (contraction), presumably since existing cellular forces were no longer counterbalanced by ECM tension. Pseudopodial extension (spreading) was then observed at both ends of the cell. The ECM was pulled inward during this secondary spreading, and rapid turnover of focal adhesions was observed along extending pseudopodia. Preincubation with Y-27632 or cytochalasin D blocked both the initial contractile and secondary spreading responses.
Overall, the data suggest that corneal fibroblasts actively respond to increases or decreases in local matrix stress in an attempt to maintain tensional homeostasis (constant tension), and that this response may be mediated by Rho and/or Rac.

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Available from: Walter Matthew Petroll, Jul 15, 2014
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    • "traction to the extracellular matrix (ECM) depends on cell shape, size and focal adhesions (Rape et al., 2011). Cells monitor their mechanical state and adjust traction forces depending on the environmental conditions to reach 'tensional homeostasis' (Brown et al., 1998; Saez et al., 2005; Ghibaudo et al., 2008; Petroll et al., 2004). Cell traction results in alignment of the collagen network (Stopak and Harris, 1982), which in turn becomes a topological trigger for cells to orient. "
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    ABSTRACT: Soft biological tissues adapt their collagen network to the mechanical environment. Collagen remodeling and cell traction are both involved in this process. The present study presents a collagen adaptation model which includes strain-dependent collagen degradation and contact-guided cell traction. Cell traction is determined by the prevailing collagen structure and is assumed to strive for tensional homeostasis. In addition, collagen is assumed to mechanically fail if it is over-strained. Care is taken to use principally measurable and physiologically meaningful relationships. This model is implemented in a fibril-reinforced biphasic finite element model for soft hydrated tissues. The versatility and limitations of the model are demonstrated by corroborating the predicted transient and equilibrium collagen adaptation under distinct mechanical constraints against experimental observations from the literature. These experiments include overloading of pericardium explants until failure, static uniaxial and biaxial loading of cell-seeded gels in vitro and shortening of periosteum explants. In addition, remodeling under hypothetical conditions is explored to demonstrate how collagen might adapt to small differences in constraints. Typical aspects of all essentially different experimental conditions are captured quantitatively or qualitatively. Differences between predictions and experiments as well as new insights that emerge from the present simulations are discussed. This model is anticipated to evolve into a mechanistic description of collagen adaptation, which may assist in developing load-regimes for functional tissue engineered constructs, or may be employed to improve our understanding of the mechanisms behind physiological and pathological collagen remodeling. Copyright © 2014 Elsevier Ltd. All rights reserved.
    Preview · Article · Dec 2014 · Journal of Biomechanics
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    • "FEM analysis showed that fibroblasts partially re-establish baseline ECM tension during this secondary spreading, presumably in an attempt to maintain tensional homeostasis [19]. Overall, while durotaxis has been shown to regulate cell alignment and migration within collagen matrices under static conditions, tensional homeostasis may modulate cell behavior in response to more transient changes in ECM tension, at both the local and global level [19] [97] [98]. Interestingly, when a needle is pushed toward the trailing edge of a migrating cell in 3-D, the same initial contraction and secondary spreading response identified at the leading edge is induced. "
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    ABSTRACT: Cellular interactions with extracellular matrices (ECM) through the application of mechanical forces mediate numerous biological processes including developmental morphogenesis, wound healing and cancer metastasis. They also play a key role in the cellular repopulation and/or remodeling of engineered tissues and organs. While 2-D studies can provide important insights into many aspects of cellular mechanobiology, cells reside within 3-D ECMs in vivo, and matrix structure and dimensionality have been shown to impact cell morphology, protein organization and mechanical behavior. Global measurements of cell-induced compaction of 3-D collagen matrices can provide important insights into the regulation of overall cell contractility by various cytokines and signaling pathways. However, to understand how the mechanics of cell spreading, migration, contraction and matrix remodeling are regulated at the molecular level, these processes must also be studied in individual cells. Here we review the evolution and application of techniques for imaging and assessing local cell-matrix mechanical interactions in 3-D culture models, tissue explants and living animals.
    Full-text · Article · Jun 2013 · Experimental Cell Research
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    • "Raghavan and co-workers negated transport limitations by fabricating microgels of the order of micrometers [20]. Local mechanical stress can be changed by pulling/pushing on gel regions adjacent to the cells [24]. Some contact guidance experiments utilized electrospun fibers or cell-generated matrices [10] [25]. "
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    ABSTRACT: Cells reside in a complex microenvironment in situ, with a number of chemical and physical parameters interacting to modulate cell phenotype and activities. To understand cell behaviors in 3D, recent studies have utilized natural or synthetic hydrogel or fibrous materials. Taking cues from the nucleation and growth characteristics of collagen fibrils in shear flow, we generate cell-laden 3D collagen hydrogels with aligned collagen fibrils using a simple microfluidic device driven by hydrostatic flow. Furthermore, by regulating collagen hydrogel thickness, the surface effective stiffness can be modulated to change the mechanical environment of the cell. Dimensionality, topography, and substrate thickness/stiffness change cell morphology and migration. Interactions amongst these parameters further influence cell behaviors. For instance, while cells responded similarly to the change in substrate thickness/stiffness on 2D random gels, dimensionality and fiber alignment both interacted with substrate thickness/stiffness to change cell morphology and motility. This economical, simple to use, and fully-biocompatible platform highlights the importance of well-controlled physical parameters in the cellular microenvironment.
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