20 results found
Lachowski D, Matellan C, Gopal S, et al., 2022, Substrate stiffness-driven membrane tension modulates vesicular trafficking via caveolin-1., ACS Nano, Vol: 16, Pages: 4322-4337, ISSN: 1936-0851
Liver fibrosis, a condition characterized by extensive deposition and cross-linking of extracellular matrix (ECM) proteins, is idiosyncratic in cases of chronic liver injury. The dysregulation of ECM remodeling by hepatic stellate cells (HSCs), the main mediators of fibrosis, results in an elevated ECM stiffness that drives the development of chronic liver disease such as cirrhosis and hepatocellular carcinoma. Tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) is a key element in the regulation of ECM remodeling, which modulates the degradation and turnover of ECM components. We have previously reported that a rigid, fibrotic-like substrate can impact TIMP-1 expression at the protein level in HSCs without altering its mRNA expression. While HSCs are known to be highly susceptible to mechanical stimuli, the mechanisms through which mechanical cues regulate TIMP-1 at the post-translational level remain unclear. Here, we show a mechanism of regulation of plasma membrane tension by matrix stiffness. We found that this effect is orchestrated by the β1 integrin/RhoA axis and results in elevated exocytosis and secretion of TIMP-1 in a caveolin-1- and dynamin-2-dependent manner. We then show that TIMP-1 and caveolin-1 expression increases in cirrhosis and hepatocellular carcinoma. These conditions are associated with fibrosis, and this effect can be recapitulated in 3D fibrosis models consisting of hepatic stellate cells encapsulated in a self-assembling polypeptide hydrogel. This work positions stiffness-dependent membrane tension as a key regulator of enzyme secretion and function and a potential target for therapeutic strategies that aim at modulating ECM remodeling in chronic liver disease.
Lachowski D, Cortes Lopez J, Matellan C, et al., 2020, G protein-coupled estrogen receptor regulates actin cytoskeleton dynamics to impair cell polarization, Frontiers in Cell and Developmental Biology, Vol: 8, ISSN: 2296-634X
Mechanical forces regulate cell functions through multiple pathways. G protein-coupled estrogen receptor (GPER) is a seven-transmembrane receptor that is ubiquitously expressed across tissues and mediates the acute cellular response to estrogens. Here, we demonstrate an unidentified role of GPER as a cellular mechanoregulator. G protein-coupled estrogen receptor signaling controls the assembly of stress fibers, the dynamics of the associated focal adhesions, and cell polarization via RhoA GTPase (RhoA). G protein-coupled estrogen receptor activation inhibits F-actin polymerization and subsequently triggers a negative feedback that transcriptionally suppresses the expression of monomeric G-actin. Given the broad expression of GPER and the range of cytoskeletal changes modulated by this receptor, our findings position GPER as a key player in mechanotransduction.
Chronopoulos A, Thorpe SD, Cortes E, et al., 2020, Syndecan-4 tunes cell mechanics by activating the kindlin-integrin-RhoA pathway, Nature Materials, Vol: 19, Pages: 669-678, ISSN: 1476-1122
Extensive research over the past decades has identified integrins to be the primary transmembrane receptors that enable cells to respond to external mechanical cues. We reveal here a mechanism whereby syndecan-4 tunes cell mechanics in response to localized tension via a coordinated mechanochemical signalling response that involves activation of two other receptors: epidermal growth factor receptor and β1 integrin. Tension on syndecan-4 induces cell-wide activation of the kindlin-2/β1 integrin/RhoA axis in a PI3K-dependent manner. Furthermore, syndecan-4-mediated tension at the cell–extracellular matrix interface is required for yes-associated protein activation. Extracellular tension on syndecan-4 triggers a conformational change in the cytoplasmic domain, the variable region of which is indispensable for the mechanical adaptation to force, facilitating the assembly of a syndecan-4/α-actinin/F-actin molecular scaffold at the bead adhesion. This mechanotransduction pathway for syndecan-4 should have immediate implications for the broader field of mechanobiology.
Rice A, Cortes Lopez JE, Lachowski D, et al., 2020, GPER activation inhibits cancer cell mechanotransduction and basement membrane invasion via RhoA, Cancers, Vol: 12, ISSN: 2072-6694
The invasive properties of cancer cells are intimately linked to their mechanical phenotype, which can be regulated by intracellular biochemical signalling. Cell contractility, induced by mechanotransduction of a stiff fibrotic matrix, and the epithelial–mesenchymal transition (EMT) promote invasion. Metastasis involves cells pushing through the basement membrane into the stroma—both of which are altered in composition with cancer progression. Agonists of the G protein-coupled oestrogen receptor (GPER), such as tamoxifen, have been largely used in the clinic, and interest in GPER, which is abundantly expressed in tissues, has greatly increased despite a lack of understanding regarding the mechanisms which promote its multiple effects. Here, we show that specific activation of GPER inhibits EMT, mechanotransduction and cell contractility in cancer cells via the GTPase Ras homolog family member A (RhoA). We further show that GPER activation inhibits invasion through an in vitro basement membrane mimic, similar in structure to the pancreatic basement membrane that we reveal as an asymmetric bilayer, which differs in composition between healthy and cancer patients.Keywords: cancer biomechanics; metastasis; G protein-coupled receptors; tumour microenvironment
Ghose R, Rice AJ, Cortes E, et al., 2020, Implementation of a basement membrane invasion assay using mesenteric tissue, CELL-DERIVED MATRICES, PT B, Editors: Caballero, Kundu, Reis, Publisher: ACADEMIC PRESS LTD-ELSEVIER SCIENCE LTD, Pages: 99-122
Lachowski D, Cortes E, Rice A, et al., 2019, Matrix stiffness modulates the activity of MMP-9 and TIMP-1 in hepatic stellate cells to perpetuate fibrosis, Scientific Reports, Vol: 9, ISSN: 2045-2322
Liver fibrosis is characterised by a dense and highly cross-linked extracellular matrix (ECM) which promotes progression of diseases such as hepatocellular carcinoma. The fibrotic microenvironment is characterised by an increased stiffness, with rigidity associated with disease progression. External stiffness is known to promote hepatic stellate cell (HSC) activation through mechanotransduction, leading to increased secretion of ECM components. HSCs are key effector cells which maintain the composition of the ECM in health and disease, not only by regulating secretion of ECM proteins such as collagen, but also ECM-degrading enzymes called matrix metalloproteinases (MMPs) and their inhibitors (TIMPs). Uninhibited MMPs degrade ECM proteins to reduce external rigidity. Using fibronectin-coated polyacrylamide gels to alter substrate rigidity without altering ligand density, we show that fibrotic rigidities downregulate MMP-9 expression and secretion, and also upregulate secretion of TIMP-1, though not its expression. Using tissue immunofluorescence studies, we also report that the expression of MMP-9 is significantly decreased in activated HSCs in fibrotic tissues associated with hepatocellular carcinoma. This suggests the presence of a mechanical network that allows HSCs to maintain a fibrotic ECM, with external rigidity providing feedback which affects MMP-9 and TIMP-1 secretion, which may become dysregulated in fibrosis.
Cortes E, Lachowski D, Rice A, et al., 2019, Retinoic acid receptor-β is downregulated in hepatocellular carcinoma and cirrhosis and its expression inhibits myosin-driven activation and durotaxis in hepatic stellate cells, Hepatology, Vol: 69, Pages: 785-802, ISSN: 0270-9139
Hepatic stellate cells (HSCs) are essential perisinusoidal cells in the healthy and diseased liver. HSCs modulate extracellular matrix (ECM) homeostasis when quiescent, but in liver fibrosis, HSCs become activated and promote excess deposition of ECM molecules and tissue stiffening via force generation and mechanosensing. In hepatocellular carcinoma (HCC), activated HSCs infiltrate the stroma and migrate to the tumor core to facilitate paracrine signalling with cancer cells. Since the function of HSCs is known to be modulated by retinoids, we investigated the expression profile of retinoic acid receptor beta (RAR-β) in cirrhotic and HCC patients, as well as the effects of RAR-β activation in HSCs. We found that RAR-β expression is significantly reduced in cirrhotic and HCC tissues. Using a comprehensive set of biophysical methods combined with cellular and molecular biology, we have elucidated the biomechanical mechanism by which all trans-retinoic acid (ATRA) promotes HSC deactivation via RAR-β-dependent transcriptional downregulation of myosin light chain 2 (MLC-2) expression. Furthermore, this also abrogated mechanically driven migration towards stiffer substrates. CONCLUSION: Targeting mechanotransduction in HSCs at the transcriptional level may offer new therapeutic options for a range of liver diseases. This article is protected by copyright. All rights reserved.
Cortes E, Sarper M, Robinson B, et al., 2019, GPER is a mechanoregulator of pancreatic stellate cells and the tumor microenvironment, EMBO Reports, Vol: 20, ISSN: 1469-221X
The mechanical properties of the tumor microenvironment are emerging as attractive targets for the development of therapies. Tamoxifen, an agonist of the G protein-coupled estrogen receptor (GPER), is widely used to treat estrogen-positive breast cancer. Here, we show that tamoxifen mechanically reprograms the tumor microenvironment through a newly identified GPER-mediated mechanism. Tamoxifen inhibits the myofibroblastic differentiation of pancreatic stellate cells (PSCs) in the tumor microenvironment of pancreatic cancer in an acto-myosin-dependent manner via RhoA-mediated contractility, YAP deactivation, and GPER signaling. This hampers the ability of PSCs to remodel the extracellular matrix and to promote cancer cell invasion. Tamoxifen also reduces the recruitment and polarization to the M2 phenotype of tumor-associated macrophages. Our results highlight GPER as a mechanical regulator of the tumor microenvironment that targets the three hallmarks of pancreatic cancer: desmoplasia, inflammation, and immune suppression. The well-established safety of tamoxifen in clinics may offer the possibility to redirect the singular focus of tamoxifen on the cancer cells to the greater tumor microenvironment and lead a new strategy of drug repurposing.
Cortes E, Lachowski D, Robinson B, et al., 2019, Tamoxifen mechanically reprograms the tumor microenvironment via HIF-1A and reduces cancer cell survival, EMBO Reports, Vol: 20, ISSN: 1469-221X
The tumor microenvironment is fundamental to cancer progression, and the influence of its mechanical properties is increasingly being appreciated. Tamoxifen has been used for many years to treat estrogen-positive breast cancer. Here we report that tamoxifen regulates the level and activity of collagen cross-linking and degradative enzymes, and hence the organization of the extracellular matrix, via a mechanism involving both the G protein-coupled estrogen receptor (GPER) and hypoxia-inducible factor-1 alpha (HIF-1A). We show that tamoxifen reduces HIF-1A levels by suppressing myosin-dependent contractility and matrix stiffness mechanosensing. Tamoxifen also downregulates hypoxia-regulated genes and increases vascularization in PDAC tissues. Our findings implicate the GPER/HIF-1A axis as a master regulator of peri-tumoral stromal remodeling and the fibrovascular tumor microenvironment and offer a paradigm shift for tamoxifen from a well-established drug in breast cancer hormonal therapy to an alternative candidate for stromal targeting strategies in PDAC and possibly other cancers.
Cortes E, Lachowski D, Rice A, et al., 2018, Tamoxifen mechanically deactivates hepatic stellate cells via the G protein-coupled estrogen receptor, Oncogene, Vol: 38, Pages: 2910-2922, ISSN: 0950-9232
Tamoxifen has been used for many years to target estrogen receptor signalling in breast cancer cells. Tamoxifen is also an agonist of the G protein-coupled estrogen receptor (GPER), a GPCR ubiquitously expressed in tissues that mediates the acute response to estrogens. Here we report that tamoxifen promotes mechanical quiescence in hepatic stellate cells (HSCs), stromal fibroblast-like cells whose activation triggers and perpetuates liver fibrosis in hepatocellular carcinomas. This mechanical deactivation is mediated by the GPER/RhoA/myosin axis and induces YAP deactivation. We report that tamoxifen decreases the levels of hypoxia-inducible factor-1 alpha (HIF-1α) and the synthesis of extracellular matrix proteins through a mechanical mechanism that involves actomyosin-dependent contractility and mechanosensing of tissue stiffness. Our results implicate GPER-mediated estrogen signalling in the mechanosensory-driven activation of HSCs and put forward estrogenic signalling as an option for mechanical reprogramming of myofibroblast-like cells in the tumour microenvironment. Tamoxifen, with half a century of safe clinical use, might lead this strategy of drug repositioning.
Haining AWM, Rahikainen R, Cortes E, et al., 2018, Mechanotransduction in talin through the interaction of the R8 domain with DLC1, PLoS Biology, Vol: 16, Pages: 1-20, ISSN: 1544-9173
The mechanical unfolding of proteins is a cellular mechanism for force transduction with potentially broad implications in cell fate. Despite this, the mechanism by which protein unfolding elicits differential downstream signalling pathways remains poorly understood. Here, we used protein engineering, atomic force microscopy, and biophysical tools to delineate how protein unfolding controls cell mechanics. Deleted in liver cancer 1 (DLC1) is a negative regulator of Ras homolog family member A (RhoA) and cell contractility that regulates cell behaviour when localised to focal adhesions bound to folded talin. Using a talin mutant resistant to force-induced unfolding of R8 domain, we show that talin unfolding determines DLC1 downstream signalling and, consequently, cell mechanics. We propose that this new mechanism of mechanotransduction may have implications for a wide variety of associated cellular processes.
Lachowski D, Cortes E, Robinson B, et al., 2018, FAK controls the mechanical activation of YAP, a transcriptional regulator required for durotaxis, FASEB JOURNAL, Vol: 32, Pages: 1099-1107, ISSN: 0892-6638
Rice AJ, Cortes E, Lachowski D, et al., 2017, Matrix stiffness induces epithelial-mesenchymal transition and promotes chemoresistance in pancreatic cancer cells, Oncogenesis, Vol: 6, Pages: e352-e352, ISSN: 2157-9024
Increased matrix rigidity associated with the fibrotic reaction is documented to stimulate intracellular signalling pathways that promote cancer cell survival and tumour growth. Pancreatic cancer is one of the stiffest of all human solid carcinomas and is characterised by a remarkable desmoplastic reaction. Here we use mouse models, genetically engineered to recapitulate human pancreatic cancer, and several pancreatic cancer cell lines as a model to investigate the effect of matrix stiffness in epithelial-mesenchymal transition (EMT) and resistance to chemotherapeutics. We found that recapitulation of the fibrotic rigidities found in pancreatic cancer tissues promote elements of EMT, including increases in vimentin expression, decreases in E-cadherin expression, nuclear localisation of β-catenin, YAP and TAZ and changes in cell shape towards a mesenchymal phenotype. We also report that stiffness induces chemoresistance to paclitaxel, but not to gemcitabine, both commonly used therapeutics, suggesting that environmental rigidity underlies an aspect of chemoresistance.
Lachowski D, Cortes E, Pink D, et al., 2017, Substrate rigidity controls activation and durotaxis in pancreatic stellate cells, Scientific Reports, Vol: 7, ISSN: 2045-2322
Pancreatic Ductal Adenocarcinoma (PDAC) is an aggressive malignancy characterised by the presence of extensive desmoplasia, thought to be responsible for the poor response of patients to systemic therapies. Pancreatic stellate cells (PSCs) are key mediators in the production of this fibrotic stroma, upon activation transitioning to a myofibroblast-like, high matrix secreting phenotype. Given their importance in disease progression, characterisation of PSC activation has been extensive, however one aspect that has been overlooked is the mechano-sensing properties of the cell. Here, through the use of a physiomimetic system that recapitulates the mechanical microenvironment found within healthy and fibrotic pancreas, we demonstrate that matrix stiffness regulates activation and mechanotaxis in PSCs. We show the ability of PSCs to undergo phenotypic transition solely as a result of changes in extracellular matrix stiffness, whilst observing the ability of PSCs to durotactically respond to stiffness variations within their local environment. Our findings implicate the mechanical microenvironment as a potent contributor to PDAC progression and survival via induction of PSC activation and fibrosis, suggesting that direct mechanical reprogramming of PSCs may be a viable alternative in the treatment of this lethal disease.
Lachowski D, Cortes E, Robinson BK, et al., 2016, Assaying the rigidity guided migration of human tumour stromal myofibroblasts (TSMs) on polyacrylamide substrates mimicking the healthy and fibrotic tissue transition boundary, Convergent Science Physical Oncology, Vol: 2, Pages: 044502-044502
Attwood S, Cortes E, Haining, et al., 2016, Adhesive ligand tether length affects the size and length of focal adhesions and influences cell spreading and attachment, Scientific Reports, Vol: 6, ISSN: 2045-2322
Cells are known to respond to physical cues from their microenvironment such as matrix rigidity.Discrete adhesive ligands within flexible strands of fibronectin connect cell surface integrins to thebroader extracellular matrix and are thought to mediate mechanosensing through the cytoskeletonintegrin-ECMlinkage. We set out to determine if adhesive ligand tether length is another physical cuethat cells can sense. Substrates were covalently modified with adhesive arginylglycylaspartic acid (RGD)ligands coupled with short (9.5nm), medium (38.2nm) and long (318nm) length inert polyethyleneglycol tethers. The size and length of focal adhesions of human foreskin fibroblasts gradually decreasedfrom short to long tethers. Furthermore, we found cell adhesion varies in a linker length dependentmanner with a remarkable 75% reduction in the density of cells on the surface and a 50% reduction incell area between the shortest and longest linkers. We also report the interplay between RGD ligandconcentration and tether length in determining cellular spread area. Our findings show that withoutvarying substrate rigidity or ligand density, tether length alone can modulate cellular behaviour.
Lachowski D, Cortes E, Robinson B, et al., 2016, SAT-402 - Elucidating the Biomechanical Response of Human Hepatic Stellate Cells on Substrates Mimicking Healthy and Fibrotic Matrix Rigidity, EASL International Liver Congress, Publisher: Elsevier, Pages: S706-S706, ISSN: 0168-8278
Chronopoulos A, Robinson B, Sarper M, et al., 2016, ATRA mechanically reprograms pancreatic stellate cells to suppress matrix remodelling and inhibit cancer cell invasion, Nature Communications, Vol: 7, Pages: 1-12, ISSN: 2041-1723
Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive malignancy with a dismal survival rate. Persistent activation of pancreatic stellate cells (PSCs) can perturb the biomechanical homoeostasis of the tumour microenvironment to favour cancer cell invasion. Here we report that ATRA, an active metabolite of vitamin A, restores mechanical quiescence in PSCs via a mechanism involving a retinoic acid receptor beta (RAR-β)-dependent downregulation of actomyosin (MLC-2) contractility. We show that ATRA reduces the ability of PSCs to generate high traction forces and adapt to extracellular mechanical cues (mechanosensing), as well as suppresses force-mediated extracellular matrix remodelling to inhibit local cancer cell invasion in 3D organotypic models. Our findings implicate a RAR-β/MLC-2 pathway in peritumoural stromal remodelling and mechanosensory-driven activation of PSCs, and further suggest that mechanical reprogramming of PSCs with retinoic acid derivatives might be a viable alternative to stromal ablation strategies for the treatment of PDAC.
Sarper M, Cortes E, Lieberthal T, et al., 2016, ATRA modulates mechanical activation of TGF-β by pancreatic stellate cells, Scientific Reports, Vol: 6, ISSN: 2045-2322
The hallmark of pancreatic ductal adenocarcinoma (PDAC) is abundant desmoplasia, which is orchestrated by pancreatic stellate cells (PSCs) and accounts for the majority of the stroma surrounding the tumour. Healthy PSCs are quiescent, but upon activation during disease progression, they adopt a myofibroblast-contractile phenotype and secrete and concomitantly reorganise the stiff extracellular matrix (ECM). Transforming growth factor β (TGF-β) is a potent activator of PSCs, and its activation requires spatiotemporal organisation of cellular and extracellular cues to liberate it from an inactive complex with latent TGF-β binding protein (LTBP). Here we study the mechanical activation of TGF-β by PSCs in vitro by investigating LTBP-1 organisation with fibrillar fibronectin and show that all trans-retinoic acid (ATRA), which induces PSC quiescence, down-regulates the ability of PSCs to mechanically organise LTBP-1 and activate TGF-β through a mechanism involving myosin II dependent contractility. Therefore, ATRA inhibits the ability of PSCs to mechanically release active TGF-β, which might otherwise act in an autocrine manner to sustain PSCs in an active state and a tumour-favouring stiff microenvironment.
Robinson BK, Cortes E, Rice AJ, et al., 2016, Quantitative analysis of 3D extracellular matrix remodelling bypancreatic stellate cells, Biology Open, Vol: 5, Pages: 875-882, ISSN: 2046-6390
Extracellular matrix (ECM) remodelling is integral to numerousphysiological and pathological processes in biology, such asembryogenesis, wound healing, fibrosis and cancer. Until recently,most cellular studies have been conducted on 2D environmentswhere mechanical cues significantly differ from physiologicallyrelevant 3D environments, impacting cellular behaviour andmasking the interpretation of cellular function in health and disease.We present an integrated methodology where cell-ECM interactionscan be investigated in 3D environments via ECM remodelling.Monitoring and quantification of collagen-I structure in remodelledmatrices, through designated algorithms, show that 3D matrices canbe used to correlate remodelling with increased ECM stiffnessobserved in fibrosis. Pancreatic stellate cells (PSCs) are the keyeffectors of the stromal fibrosis associated to pancreatic cancer. Weuse PSCs to implement our methodology and demonstrate that PSCmatrix remodelling capabilities depend on their contractile machineryand β1 integrin-mediated cell-ECM attachment.
This data is extracted from the Web of Science and reproduced under a licence from Thomson Reuters. You may not copy or re-distribute this data in whole or in part without the written consent of the Science business of Thomson Reuters.