Plasticity of the actin cytoskeleton in response to extracellular matrix nanostructure and dimensionality

2014 ◽  
Vol 42 (5) ◽  
pp. 1356-1366 ◽  
Author(s):  
Josefine Starke ◽  
Bernhard Wehrle-Haller ◽  
Peter Friedl

Mobile cells discriminate and adapt to mechanosensory input from extracellular matrix (ECM) topographies to undergo actin-based polarization, shape change and migration. We tested ‘cell-intrinsic’ and adaptive components of actin-based cell migration in response to widely used in vitro collagen-based substrates, including a continuous 2D surface, discontinuous fibril-based surfaces (2.5D) and fibril-based 3D geometries. Migrating B16F1 mouse melanoma cells expressing GFP–actin developed striking diversity and adaptation of cytoskeletal organization and migration efficacy in response to collagen organization. 2D geometry enabled keratinocyte-like cell spreading and lamellipod-driven motility, with barrier-free movement averaging the directional vectors from one or several leading edges. 3D fibrillar collagen imposed spindle-shaped polarity with a single cylindrical actin-rich leading edge and terminal filopod-like protrusions generating a single force vector. As a mixed phenotype, 2.5D environments prompted a broad but fractalized leading lamella, with multiple terminal filopod-like protrusions engaged with collagen fibrils to generate an average directional vector from multiple, often divergent, interactions. The migratory population reached >90% of the cells with high speeds for 2D, but only 10–30% of the cells and a 3-fold lower speed range for 2.5D and 3D substrates, suggesting substrate continuity as a major determinant of efficient induction and maintenance of migration. These findings implicate substrate geometry as an important input for plasticity and adaptation of the actin cytoskeleton to cope with varying ECM topography and highlight striking preference of moving cells for 2D continuous-shaped over more complex-shaped discontinuous 2.5 and 3D substrate geometries.

1997 ◽  
Vol 78 (02) ◽  
pp. 934-938 ◽  
Author(s):  
Hsiun-ing Chen ◽  
Yueh-I Wu ◽  
Yu-Lun Hsieh ◽  
Guey-Yueh Shi ◽  
Meei-Jyh Jiang ◽  
...  

SummaryTo investigate whether the endothelium-platelet interactions may be altered by plasminogen activation, cultured human umbilical vein endothelial cells (ECs) were treated with tissue-type plasminogen activator (t-PA) in the presence of plasminogen, and platelet adhesion to ECs was subsequently measured by using a tapered flow chamber. Our results demonstrated that platelets adhered more readily to t-PA treated EC monolayer than to the control monolayer at all shear stress levels tested. This phenomenon was treatment time-dependent and dose-dependent, and it could be blocked by adding plasmin inhibitors, such as e-amino caproic acid and aprotinin. Adherent platelets on t-PA treated EC monolayer underwent more severe shape change than those on the control monolayer. While the extracellular matrix directly treated with t-PA attracted less platelets than the control matrix did, platelet adhesion to the matrix that was produced by t-PA-treated ECs was unaltered. These data suggest that t-PA treatment on ECs compromised antiplatelet-adhesion capability on their apical surface without altering the reactivity of their extracellular matrix towards platelets.


2021 ◽  
Author(s):  
◽  
Sandi Grainne Dempsey

<p>Biomaterials derived from decellularised extracellular matrices have shown promise as tools in tissue regeneration and wound healing. Such materials display biocompatibility as well as inherent bioactivity, promoting constructive remodelling in healing tissues. In this study, the bioactivity of ovine forestomach matrix (a decellularised extracellular matrix biomaterial) is assessed based on its ability to affect the proliferation and migration of wound healing cells.  This material supported cell attachment and proliferation, but did not allow cell infiltration in vitro. Enzymatic digestion of the material rendered soluble components that were able to induce proliferation and migration of some cell types. Cell-mediated processing of the material generated a protein or proteins with chemotactic activity for mesenchymal stem cells in vitro. Mass spectrometry analysis indicated the bioactive component consisted of the proteoglycan decorin, or fragments thereof. Decorin has not previously been shown to induce mesenchymal stem cell motility, and these findings may add to what is known about decorin and its role in constructive remodelling. Furthermore, this cell-mediated approach for ECM breakdown could lead to the discovery of other bioactive peptides involved in ECM remodelling and wound healing.</p>


2021 ◽  
Author(s):  
◽  
Sandi Grainne Dempsey

<p>Biomaterials derived from decellularised extracellular matrices have shown promise as tools in tissue regeneration and wound healing. Such materials display biocompatibility as well as inherent bioactivity, promoting constructive remodelling in healing tissues. In this study, the bioactivity of ovine forestomach matrix (a decellularised extracellular matrix biomaterial) is assessed based on its ability to affect the proliferation and migration of wound healing cells.  This material supported cell attachment and proliferation, but did not allow cell infiltration in vitro. Enzymatic digestion of the material rendered soluble components that were able to induce proliferation and migration of some cell types. Cell-mediated processing of the material generated a protein or proteins with chemotactic activity for mesenchymal stem cells in vitro. Mass spectrometry analysis indicated the bioactive component consisted of the proteoglycan decorin, or fragments thereof. Decorin has not previously been shown to induce mesenchymal stem cell motility, and these findings may add to what is known about decorin and its role in constructive remodelling. Furthermore, this cell-mediated approach for ECM breakdown could lead to the discovery of other bioactive peptides involved in ECM remodelling and wound healing.</p>


2019 ◽  
Vol 20 (10) ◽  
pp. 2592 ◽  
Author(s):  
Wei-Hsiang Hsu ◽  
Se-Chun Liao ◽  
Yau-Jan Chyan ◽  
Kai-Wen Huang ◽  
Shih-Lan Hsu ◽  
...  

Background and Aims: Liver fibrosis is the excessive accumulation of extracellular matrix proteins, including collagen, which occurs in most types of chronic liver diseases. Advanced liver fibrosis results in cirrhosis, liver failure, and portal hypertension. Activated hepatic perivascular stellate cells, portal fibroblasts, and myofibroblasts of bone marrow origin have been identified as major collagen-producing cells in the injured liver. These cells are activated by fibrogenic cytokines, such as TGF-β1. The inhibition of TGF-β1 function or synthesis is a major target for the development of antifibrotic therapies. Our previous study showed that the water and ethanol extracts of Graptopetalum paraguayense (GP), a Chinese herbal medicine, can prevent dimethylnitrosamine (DMN)-induced hepatic inflammation and fibrosis in rats. Methods: We used rat hepatic stellate HSC-T6 cells and a diethylnitrosamine (DEN)-induced rat liver injury model to test the potential mechanism of GP extracts and its fraction, HH-F3. Results: We demonstrated that GP extracts and HH-F3 downregulated the expression levels of extracellular matrix (ECM) proteins and inhibited the proliferation and migration via suppression of the TGF-β1 pathway in rat hepatic stellate HSC-T6 cells. Moreover, the HH-F3 fraction decreased hepatic collagen content and reduced plasma AST, ALT, and γ-GT activities in a DEN-induced rat liver injury model, suggesting that GP/HH-F3 has hepatoprotective effects against DEN-induced liver fibrosis. Conclusion: These findings indicate that GP/HH-F3 may be a potential therapeutic agent for the treatment of liver fibrosis. The inhibition of TGF-β-mediated fibrogenesis may be a central mechanism by which GP/HH-F3 protects the liver from injury.


2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Emily Nguyen ◽  
Olha Koval ◽  
Isabella Grumbach

Background: Restenosis after angioplasty for coronary vascular disease remains a critical problem in cardiovascular medicine. Vascular smooth muscle cell (VSMC) migration and proliferation cause restenosis through neointima formation. Mitochondrial motility is likely necessary for cell proliferation and migration, and is inhibited in microdomains with increased Ca 2+ . The Ca 2+ /calmodulin-dependent kinase II (CaMKII) in mitochondria (mtCaMKII) is proposed to control mitochondrial matrix Ca 2+ uptake through mitochondrial Ca 2+ uniporter (MCU). Thus, we hypothesized that blocking mtCaMKII decreases VSMC migration and neointima formation by decreasing mitochondrial motility. Methods: mtCaMKII was inhibited by expression of the mitochondria-targeted CaMKII inhibitor peptide (CaMKIIN) in a novel transgenic mouse model in smooth muscle only (SM-mtCaMKIIN) or delivered by adenoviral transduction (Ad-mtCaMKIIN). Results: In our models, mtCaMKIIN was detected selectively in mitochondria of VSMC. mtCaMKIIN significantly reduced mitochondrial Ca 2+ current and Ca 2+ content compared to WT in vivo and in vitro. SM-mtCaMKIIN mice showed significantly reduced neointimal area 28 days after endothelial injury (n=8, p<0.05) and fewer proliferating neointimal cells by PCNA staining. In vitro, Ad-mtCaMKIIN mildly reduced VSMC proliferation and mitochondrial ROS production without altering maximal respiration after PDGF treatment. Ad-mtCaMKIIN abolished VSMC migration, as did mitoTEMPO and MCU inhibitor Ru360. Ad-mtCaMKIIN blocked mitochondrial mobility towards the leading edge, while relocation of mitochondria was seen in WT cells 6 h after PDGF treatment. Mitochondrial redistribution was also inhibited by Ru360, but not by mitoTEMPO or cytoplasmic CaMKII inhibition. Mitochondrial fission promotes cell migration. Accordingly, PDGF increased mitochondrial particles in WT VSMC, while mitochondria in Ad-mtCaMKIIN cells were fragmented and unresponsive to PDGF treatment. Conclusions: mtCaMKIIN prevents mitochondrial distribution to the leading edge and reduces VSMC migration and neointima formation. These data suggest mitochondrial Ca 2+ regulation plays an important role in VSMC migration by altering mitochondrial location.


Development ◽  
1973 ◽  
Vol 29 (1) ◽  
pp. 1-13
Author(s):  
Marc H. Glickman ◽  
Georgia E. Lesh-Laurie

A chemically defined in vitro system for Tubularia has been developed. At 24 h after explantation of coenosarc, digestive cells attached to the substrate and migrated from the explant. The digestive cells migrated by a gliding motion with a fan-like membrane acting as a leading edge. Within 48 h a digestive cell monolayer was formed and invasion of this area by epithelio-muscular, gland and interstitial cells occurred. Autoradiographic study of 48–72 h cultures treated with [3H]thymidine showed nuclear incorporation of the label in digestive, epithelio-muscular, interstitial, cnidoblast and gland cells. When explants were grown on collagen-coated coverslips, accelerated attachment and migration of digestive cells was observed. Explants were also grown on Millipore filters. No digestive cell attachment occurred but epithelio-muscular, gland and interstitial cell attachments to the filter were observed. From these experiments, a morphological role for the digestive cells as a substrate for other cells of the coenosarc is postulated. Hydranth extract was supplemented to the culture medium. Studies with this material were performed with coenosarc from ‘late summer’ animals in which only 10–15% of the explants normally entered culture. However, with the addition of the extract, 100% of the explants went into culture. Interstitial cell populations increased 2–3 times in extract-treated explants.


Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3374-3374
Author(s):  
Neeta Shirvaikar ◽  
Ali Jalili ◽  
Mariusz Z. Ratajczak ◽  
Anna Janowska-Wieczorek

Abstract Thrombin, an important serine protease, not only plays a pivotal role in platelet aggregation and coagulation, but also through activation of its receptor, seven transmembrane, G-protein-coupled receptor PAR-1, elicits numerous cellular responses in platelets and endothelial cells such as induction of adhesion molecules, production of chemokines, activation of matrix metalloproteinase (MMP)-2, cytoskeletal reorganization and migration. Thrombin is also one of the inflammatory molecules elevated during G-CSF mobilization of hematopoietic stem/progenitor cells (HSPC) and their collection by leukapheresis. We recently reported that components of leukapheresis products including thrombin enhance in vitro chemotaxis of CD34+ cells towards an SDF-1 gradient and in vivo homing to bone marrow (BM) niches in a murine model (Blood2005; 105:40). In this study we investigated whether thrombin enhances the homing-related responses of human HSPC (CD34+ cells) through MMPs, especially membrane-type (MT)1-MMP which is known to be localized on the leading edge of migrating cells and both activates latent proMMPs (MMP-2, -9) and itself has strong pericellular proteolytic activity. We found that stimulation of CD34+ cells with thrombin upregulates mRNA for MT1-MMP and MMP-9 as well as MT1-MMP protein expression (Western blot, flow cytometry) and proMMP-2 and proMMP-9 secretion (zymography). Thrombin was also found to (i) prime trans-Matrigel chemoinvasion of CD34+ cells towards a low SDF-1 gradient (20 ng/mL), which was inhibited by epigallocatechin-3-gallate, a potent inhibitor of MT1-MMP, and (ii) activate MMP-2 in of co-cultures of CD34+ cells with stromal cells (BM fibroblasts and HUVEC) which secrete proMMP-2. We also found that SDF-1 upregulates mRNA and protein expression of MT1-MMP. Moreover, using confocal microscopy we demonstrate for the first time that in CD34+ cells, PAR-1, like CXCR4, is localized in the GM1 fraction of lipid rafts and stimulation of these cells with thrombin as well as SDF-1 increases incorporation of MT1-MMP into membrane lipid rafts. Furthermore, disruption of lipid raft formation by the cholesterol-depleting agent methyl-b-cyclodextrin inhibits MT1-MMP incorporation into membrane lipid rafts and also trans-Matrigel chemoinvasion of CD34+ cells towards SDF-1. Thus we conclude that thrombin, through PAR-1 signalling and the SDF-1-CXCR4 axis, upregulates the incorporation of MT1-MMP into membrane lipid rafts and the interaction of these axes enhances the homing-related responses of HSPC towards SDF-1.


2013 ◽  
Vol 453 (1) ◽  
pp. 17-25 ◽  
Author(s):  
Fumihiko Nakamura

Cell migration, phagocytosis and cytokinesis are mechanically intensive cellular processes that are mediated by the dynamic assembly and contractility of the actin cytoskeleton. GAPs (GTPase-activating proteins) control activities of the Rho family proteins including Cdc42, Rac1 and RhoA, which are prominent upstream regulators of the actin cytoskeleton. The present review concerns a class of Rho GAPs, FilGAP (ARHGAP24 gene product) and its close relatives (ARHGAP22 and AHRGAP25 gene products). FilGAP is a GAP for Rac1 and a binding partner of FLNa (filamin A), a widely expressed F-actin (filamentous actin)-cross-linking protein that binds many different proteins that are important in cell regulation. Phosphorylation of FilGAP serine/threonine residues and binding to FLNa modulate FilGAP's GAP activity and, as a result, its ability to regulate cell protrusion and spreading. FLNa binds to FilGAP at F-actin-enriched sites, such as at the leading edge of the cell where Rac1 activity is controlled to inhibit actin assembly. FilGAP then dissociates from FLNa in actin networks by myosin-dependent mechanical deformation of FLNa's FilGAP-binding site to relocate at the plasma membrane by binding to polyphosphoinositides. Since actomyosin contraction is activated downstream of RhoA–ROCK (Rho-kinase), RhoA activity regulates Rac1 through FilGAP by signalling to the force-generating system. FilGAP and the ARHGAP22 gene product also act as mediators between RhoA and Rac1 pathways, which lead to amoeboid and mesenchymal modes of cell movements respectively. Therefore FilGAP and its close relatives are key regulators that promote the reciprocal inhibitory relationship between RhoA and Rac1 in cell shape changes and the mesenchymal–amoeboid transition in tumour cells.


1999 ◽  
Vol 146 (2) ◽  
pp. 517-529 ◽  
Author(s):  
Claire Legrand ◽  
Christine Gilles ◽  
Jean-Marie Zahm ◽  
Myriam Polette ◽  
Anne-Cécile Buisson ◽  
...  

Cell spreading and migration associated with the expression of the 92-kD gelatinase (matrix metalloproteinase 9 or MMP-9) are important mechanisms involved in the repair of the respiratory epithelium. We investigated the location of MMP-9 and its potential role in migrating human bronchial epithelial cells (HBEC). In vivo and in vitro, MMP-9 accumulated in migrating HBEC located at the leading edge of a wound and MMP-9 expression paralleled cell migration speed. MMP-9 accumulated through an actin-dependent pathway in the advancing lamellipodia of migrating cells and was subsequently found active in the extracellular matrix (ECM). Lamellipodia became anchored through primordial contacts established with type IV collagen. MMP-9 became amassed behind collagen IV where there were fewer cell–ECM contacts. Both collagen IV and MMP-9 were involved in cell migration because when cell–collagen IV interaction was blocked, cells spread slightly but did not migrate; and when MMP-9 activation was prevented, cells remained fixed on primordial contacts and did not advance at all. These observations suggest that MMP-9 controls the migration of repairing HBEC by remodeling the provisional ECM implicated in primordial contacts.


2010 ◽  
Vol 188 (1) ◽  
pp. 157-173 ◽  
Author(s):  
Feng Ye ◽  
Guiqing Hu ◽  
Dianne Taylor ◽  
Boris Ratnikov ◽  
Andrey A. Bobkov ◽  
...  

Increased affinity of integrins for the extracellular matrix (activation) regulates cell adhesion and migration, extracellular matrix assembly, and mechanotransduction. Major uncertainties concern the sufficiency of talin for activation, whether conformational change without clustering leads to activation, and whether mechanical force is required for molecular extension. Here, we reconstructed physiological integrin activation in vitro and used cellular, biochemical, biophysical, and ultrastructural analyses to show that talin binding is sufficient to activate integrin αIIbβ3. Furthermore, we synthesized nanodiscs, each bearing a single lipid-embedded integrin, and used them to show that talin activates unclustered integrins leading to molecular extension in the absence of force or other membrane proteins. Thus, we provide the first proof that talin binding is sufficient to activate and extend membrane-embedded integrin αIIbβ3, thereby resolving numerous controversies and enabling molecular analysis of reconstructed integrin signaling.


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