65 research outputs found

    Quantifying adhesive interactions between cells and extracellular matrix by single-cell force spectroscopy

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    Interactions of cells with their environment regulate important cellular functions and are required for the organization of cells into tissues and complex organisms. These interactions involve different types of adhesion receptors. Interactions with extracellular matrix (ECM) proteins are mainly mediated by the integrin family of adhesion molecules. Situations in which integrin-ECM interactions are deregulated cause diseases and play a crucial role in cancer cell invasion. Thus, the mechanisms underlying integrin-binding and regulation are of high interest, particularly at the molecular level. How can cell-ECM interactions be studied? While there are several methods to analyze cell adhesion, few provide quantitative data on adhesion forces. One group, single-cell force spectroscopy (SCFS), quantifies adhesion at the single-cell level and can therefore differentiate the adhesive properties of individual cells. One implementation of SCFS is based on atomic force microscopy (AFM); this technique has been employed in the presented work. Advantageously AFM-SCFS combines high temporal and spatial cell manipulation, the ability to measure a large range of adhesion forces and sufficiently high-force resolution to allow the study of single-molecule binding events in the context of a living cell. Since individual adhesion receptors can be analyzed within their physiological environment, AFM-SCFS is a powerful tool to study the mechanisms underlying integrin-regulation. The presented work is split into six chapters. Chapter one gives background information about cell-ECM interactions. In chapter two, different adhesion assays are compared and contrasted. The theoretical Bell-Evans model which is used to interpret integrin-mediated cell adhesion is discussed in chapter three. Thereafter, the three projects that form the core of the thesis are detailed in chapters four through six. In the first project (chapter 4), α2ÎČ1-integrin mediated cell adhesion to collagen type I, the most abundant structural protein in vertebrates, was quantified using CHO cells. Firstly, α2ÎČ1-collagen interactions were investigated at the single-molecule level. Dynamic force spectroscopy permitted calculation of bond specific parameters, such as the bond dissociation rate koff (1.3 ± 1.3 sec-1) and the barrier width xu (2.3 ± 0.3 Å). Next, α2ÎČ1-integrin mediated cell adhesion to collagen type I was monitored over contact times between 0 and 600 sec. Thereby the kinetics of α2ÎČ1-integrin mediated interactions was explored and insights into the underlying binding mechanisms were gained. In the second project (chapter five), effects of cryptic integrin binding sites within collagen type I exerted on pre-osteoblasts were investigated. Collagen type I matrices were thermally denatured which lead to exposure of cryptic RGD (Arg-Gly-Asp)-motifs. As a consequence pre-osteoblasts enhanced their adhesion to denatured collagen. Compared to native collagen type I, adhesion to denatured collagen was mediated by a different set of integrins, including αv- and α5ÎČ1-integrins. Cells grown on denatured collagen showed enhanced spreading and motility, which correlated with increased focal adhesion kinase phosphorylation levels. Moreover, osteogenic differentiation kinetics and differentiation potential were increased on denatured collagen. The findings of this project open new perspectives for optimization of tissue engineering substrates. In the third part (chapter six), the effect of the fusion protein BCR/ABL, a hallmark of chronic myeloid leukemia, on adhesion of myeloid progenitor cells was studied. Adhesion between BCR/ABL transformed progenitor cells to bone marrow derived stromal cells and to different ECM proteins was quantitatively compared to that of control cells. The tyrosine kinase activity of BCR/ABL enhanced cell adhesion, which was blocked by imatinib mesylate, a drug interfering with BCR/ABL activity. BCR/ABL-enhanced adhesion correlated with increased ÎČ1-integrin cell surface concentrations. Since adhesion of leukemic cells to the bone marrow compartment is critical for the development of drug resistance, the reported results may provide a basis for optimized target therapies. In the three described projects AFM-based SCFS was applied to investigate early steps of integrin-mediated adhesion at the molecular level. Taken together, the results demonstrate that AFM-SCFS is a versatile tool that permits monitoring of cell adhesion from single-molecule interactions to the formation of more complex adhesion sites at the force level.Interaktionen zwischen Zellen und ihrer Umgebung sind maßgeblich an der Regulierung zellulĂ€rer Funktionen beteiligt und daher notwendig fĂŒr die Organisation von Zellen in Geweben und komplexen Organismen. Zellinteraktionen mit der extrazellulĂ€ren Matrix (EZM) werden hauptsĂ€chlich durch Integrine vermittelt. Situationen, in denen Integrin- EZM Interaktionen verĂ€ndert sind, können Krankheiten verursachen und spielen zudem eine wichtige Rolle bei der Invasion von Krebszellen. Daher besteht ein großes Interesse darin, die molekularen Mechanismen, die Integrin-EZM Interaktionen regulieren, besser zu verstehen. Wie können Zell-EZM Interaktionen untersucht werden? Obwohl es mehrere Methoden gibt, mit denen ZelladhĂ€sion untersucht werden kann, sind die wenigsten dazu geeignet, ZelladhĂ€sionskrĂ€fte zu quantifizieren. Einzelzellspektroskopie erfasst die AdhĂ€sionskrĂ€fte einzelner Zellen quantitativ und ermöglicht dadurch eine differenzierte Betrachtung der AdhĂ€sion individueller Zellen. Eine Variante der Einzelzellspektroskopie basiert auf der Rasterkraftmikroskopie (AFM); diese Technik wurde in der vorliegenden Arbeit verwendet. Ein Vorteil von AFM- Einzelzellspektroskopie besteht darin, dass Zellen mit hoher zeitlicher und rĂ€umlicher PrĂ€zision manipuliert werden können. ZelladhĂ€sionskrĂ€fte können zudem ĂŒber einen großen Kraftbereich hinweg untersucht werden. Dabei ermöglicht es die hohe Kraftauflösung, einzelne Integrin-Ligandenbindungen in lebenden Zellen zu untersuchen. Die vorliegende Arbeit gliedert sich in sechs Kapitel. Kapitel eins gibt Hintergrundinformationen ĂŒber Zell-EZM Wechselwirkungen. In Kapitel zwei werden verschiedene AdhĂ€sionsassays einander gegenĂŒber gestellt. Das theoretische Bell-Evans Modell, mit dessen Hilfe die gewonnenen Daten interpretiert wurden, wird in Kapitel drei diskutiert. Im Anschluss werden drei Projekte, welche das HerzstĂŒck dieser Doktorarbeit bilden, in Kapiteln vier bis sechs nĂ€her ausgefĂŒhrt. Im ersten Projekt (Kapitel vier) wurde die AdhĂ€sion von α2ÎČ1-Integrin exprimierenden CHO Zellen zu Kollagen I, dem hĂ€ufigsten strukturellen Protein in Wirbeltieren, quantitativ untersucht. ZunĂ€chst wurden α2ÎČ1-Kollagen-Interaktionen auf EinzelmolekĂŒlebene analysiert. Mithilfe der dynamischen Kraftspektroskopie wurden fĂŒr diese Bindung Dissoziationsrate koff (1.3 ± 1.3 sec-1) und Potentialbarrierenbreite xu (2.3 ± 0.3 Å) bestimmt. Daraufhin wurde die α2ÎČ1-vermittelte AdhĂ€sion ĂŒber einen Zeitraum von zehn Minuten untersucht. Dadurch konnten Einblicke in die Kinetik von α2ÎČ1-integrin vermittelter ZelladhĂ€sion sowie in die zugrunde liegenden Regulationsmechanismen gewonnen werden. Im zweiten Projekt (Kapitel fĂŒnf) wurde die Rolle von kryptischen Integrin-Bindungsstellen in Kollagen I untersucht. Die zuvor verwendeten KollagenoberflĂ€chen wurden thermisch denaturiert, wodurch versteckte RGD (Arg-Gly-Asp)-Sequenzen freigelegt wurden. Die partielle Denaturierung hatte- verglichen mit nativem Kollagen I- eine erhöhte AdhĂ€sion von PrĂ€osteoblasten (MC3T3-E1) zur Folge, was auf das Binden zusĂ€tzlicher Integrine zurĂŒckgefĂŒhrt wurde. Im Unterschied zu nativem Kollagen wurde die ZelladhĂ€sion zu denaturiertem Kollagen I u.a. durch αv- and α5ÎČ1-Integrine vermittelt. PrĂ€osteoblasten zeigten verstĂ€rktes Zellspreiten sowie höhere MotilitĂ€t auf denaturiertem Kollagen I; zudem wurde ein erhöhtes Differenzierungpotential der PrĂ€osteoblasten festgestellt. Die in diesem Projekt erhaltenen Einblicke bilden eine hilfreiche Basis fĂŒr die Entwicklung optimierter OberflĂ€chen fĂŒr diverse Zell- und Gewebekulturanwendungen. Im dritten Projekt (Kapitel sechs) wurde der Einfluss des Fusionproteins BCR/ABL, charakteristisch fĂŒr chronische myeloische LeukĂ€mie, auf die AdhĂ€sion von myeloischen VorlĂ€uferzellen untersucht. Dazu wurde die AdhĂ€sion von BCR/ABL transformierten VorlĂ€uferzellen (32D Zellen) bzw. Kontrollzellen zu Stromazellen (M2-10B4) sowie verschiedenen EZM Proteinen untersucht. BCR/ABL erhöhte die ZelladhĂ€sion der myeloischen VorlĂ€uferzellen signifikant. Dieser Effekt wurde durch die Zugabe von Imatinib, welches die TyrosinkinaseaktivitĂ€t von BCR/ABL inhibiert, aufgehoben. Die BCR/ABL-verstĂ€rkte ZelladhĂ€sion korrelierte mit erhöhten ÎČ1-Integrin-konzentrationen. Da die AdhĂ€sion von LeukĂ€miezellen im Knockenmark bekanntermaßen kritisch fĂŒr die Entwicklung von Resistenzen gegenĂŒber verschiedenen Wirkstoffen ist, könnten die Ergebnisse dieser Studie eine Grundlage fĂŒr die Entwicklung optimierter Target-Therapien sein. In den drei beschriebenen Projekten wurde AFM Einzelzellspektroskopie verwendet, um Integrin- vermittelte AdhĂ€sion auf molekularer Ebene zu untersuchen. Die Ergebnisse zeigen, dass AFM-Einzelzellspektroskopie ein vielseitiges Werkzeug darstellt, das ĂŒberaus geeignet dazu ist, ZelladhĂ€sion- ausgehend von EinzelmolekĂŒlinteraktionen bis hin zur Entstehung komplexerer AdhĂ€sionsstellen- auf der Kraftebene zu verfolgen

    Adipose cells and tissues soften with lipid accumulation while in diabetes adipose tissue stiffens

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    Adipose tissue expansion involves both differentiation of new precursors and size increase of mature adipocytes. While the two processes are well balanced in healthy tissues, obesity and diabetes type II are associated with abnormally enlarged adipocytes and excess lipid accumulation. Previous studies suggested a link between cell stiffness, volume and stem cell differentiation, although in the context of preadipocytes, there have been contradictory results regarding stiffness changes with differentiation. Thus, we set out to quantitatively monitor adipocyte shape and size changes with differentiation and lipid accumulation. We quantified by optical diffraction tomography that differentiating preadipocytes increased their volumes drastically. Atomic force microscopy (AFM)-indentation and -microrheology revealed that during the early phase of differentiation, human preadipocytes became more compliant and more fluid-like, concomitant with ROCK-mediated F-actin remodelling. Adipocytes that had accumulated large lipid droplets were more compliant, and further promoting lipid accumulation led to an even more compliant phenotype. In line with that, high fat diet-induced obesity was associated with more compliant adipose tissue compared to lean animals, both for drosophila fat bodies and murine gonadal adipose tissue. In contrast, adipose tissue of diabetic mice became significantly stiffer as shown not only by AFM but also magnetic resonance elastography. Altogether, we dissect relative contributions of the cytoskeleton and lipid droplets to cell and tissue mechanical changes across different functional states, such as differentiation, nutritional state and disease. Our work therefore sets the basis for future explorations on how tissue mechanical changes influence the behaviour of mechanosensitive tissue-resident cells in metabolic disorders

    Actin stress fiber organization promotes cell stiffening and proliferation of pre-invasive breast cancer cells

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    This deposit is composed by the main article and supplementary files of the publication.Studies of the role of actin in tumour progression have highlighted its key contribution in cell softening associated with cell invasion. Here, using a human breast cell line with conditional Src induction, we demonstrate that cells undergo a stiffening state prior to acquiring malignant features. This state is characterized by the transient accumulation of stress fibres and upregulation of Ena/VASP-like (EVL). EVL, in turn, organizes stress fibres leading to transient cell stiffening, ERK-dependent cell proliferation, as well as enhancement of Src activation and progression towards a fully transformed state. Accordingly, EVL accumulates predominantly in premalignant breast lesions and is required for Src-induced epithelial overgrowth in Drosophila. While cell softening allows for cancer cell invasion, our work reveals that stress fibre-mediated cell stiffening could drive tumour growth during premalignant stages. A careful consideration of the mechanical properties of tumour cells could therefore offer new avenues of exploration when designing cancer-targeting therapies.Bloomington Drosophila Stock Centre; Vienna Drosophila Research Center (VDRC); Developmental Studies Hybridoma Bank (DSHB); Fundação para a CiĂȘncia e Tecnologia (FCT) grant: (IF/01031/2012); Laço Grant in breast cancer 2015; Alexander von Humboldt Foundation grant: (Alexander von Humboldt Professorship); Liga Portuguesa contra o Cancro/Pfizer.info:eu-repo/semantics/publishedVersio

    In vitro microenvironments to study breast cancer bone colonisation

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    Bone metastasis occurs frequently in patients with advanced breast cancer and is a major cause of morbidity and mortality in these patients. In order to advance current therapies, the mechanisms leading to the formation of bone metastases and their pathophysiology have to be better understood. Several in vitro models have been developed for systematic studies of interactions between breast cancer cells and the bone microenvironment. Such models can provide insights into the molecular basis of bone metastatic colonisation and also may provide a useful platform to design more physiologically relevant drug testing assays. This review describes different in vitro approaches and discusses their advantages and disadvantages

    Quantifying adhesive interactions between cells and extracellular matrix by single-cell force spectroscopy

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    Interactions of cells with their environment regulate important cellular functions and are required for the organization of cells into tissues and complex organisms. These interactions involve different types of adhesion receptors. Interactions with extracellular matrix (ECM) proteins are mainly mediated by the integrin family of adhesion molecules. Situations in which integrin-ECM interactions are deregulated cause diseases and play a crucial role in cancer cell invasion. Thus, the mechanisms underlying integrin-binding and regulation are of high interest, particularly at the molecular level. How can cell-ECM interactions be studied? While there are several methods to analyze cell adhesion, few provide quantitative data on adhesion forces. One group, single-cell force spectroscopy (SCFS), quantifies adhesion at the single-cell level and can therefore differentiate the adhesive properties of individual cells. One implementation of SCFS is based on atomic force microscopy (AFM); this technique has been employed in the presented work. Advantageously AFM-SCFS combines high temporal and spatial cell manipulation, the ability to measure a large range of adhesion forces and sufficiently high-force resolution to allow the study of single-molecule binding events in the context of a living cell. Since individual adhesion receptors can be analyzed within their physiological environment, AFM-SCFS is a powerful tool to study the mechanisms underlying integrin-regulation. The presented work is split into six chapters. Chapter one gives background information about cell-ECM interactions. In chapter two, different adhesion assays are compared and contrasted. The theoretical Bell-Evans model which is used to interpret integrin-mediated cell adhesion is discussed in chapter three. Thereafter, the three projects that form the core of the thesis are detailed in chapters four through six. In the first project (chapter 4), α2ÎČ1-integrin mediated cell adhesion to collagen type I, the most abundant structural protein in vertebrates, was quantified using CHO cells. Firstly, α2ÎČ1-collagen interactions were investigated at the single-molecule level. Dynamic force spectroscopy permitted calculation of bond specific parameters, such as the bond dissociation rate koff (1.3 ± 1.3 sec-1) and the barrier width xu (2.3 ± 0.3 Å). Next, α2ÎČ1-integrin mediated cell adhesion to collagen type I was monitored over contact times between 0 and 600 sec. Thereby the kinetics of α2ÎČ1-integrin mediated interactions was explored and insights into the underlying binding mechanisms were gained. In the second project (chapter five), effects of cryptic integrin binding sites within collagen type I exerted on pre-osteoblasts were investigated. Collagen type I matrices were thermally denatured which lead to exposure of cryptic RGD (Arg-Gly-Asp)-motifs. As a consequence pre-osteoblasts enhanced their adhesion to denatured collagen. Compared to native collagen type I, adhesion to denatured collagen was mediated by a different set of integrins, including αv- and α5ÎČ1-integrins. Cells grown on denatured collagen showed enhanced spreading and motility, which correlated with increased focal adhesion kinase phosphorylation levels. Moreover, osteogenic differentiation kinetics and differentiation potential were increased on denatured collagen. The findings of this project open new perspectives for optimization of tissue engineering substrates. In the third part (chapter six), the effect of the fusion protein BCR/ABL, a hallmark of chronic myeloid leukemia, on adhesion of myeloid progenitor cells was studied. Adhesion between BCR/ABL transformed progenitor cells to bone marrow derived stromal cells and to different ECM proteins was quantitatively compared to that of control cells. The tyrosine kinase activity of BCR/ABL enhanced cell adhesion, which was blocked by imatinib mesylate, a drug interfering with BCR/ABL activity. BCR/ABL-enhanced adhesion correlated with increased ÎČ1-integrin cell surface concentrations. Since adhesion of leukemic cells to the bone marrow compartment is critical for the development of drug resistance, the reported results may provide a basis for optimized target therapies. In the three described projects AFM-based SCFS was applied to investigate early steps of integrin-mediated adhesion at the molecular level. Taken together, the results demonstrate that AFM-SCFS is a versatile tool that permits monitoring of cell adhesion from single-molecule interactions to the formation of more complex adhesion sites at the force level.Interaktionen zwischen Zellen und ihrer Umgebung sind maßgeblich an der Regulierung zellulĂ€rer Funktionen beteiligt und daher notwendig fĂŒr die Organisation von Zellen in Geweben und komplexen Organismen. Zellinteraktionen mit der extrazellulĂ€ren Matrix (EZM) werden hauptsĂ€chlich durch Integrine vermittelt. Situationen, in denen Integrin- EZM Interaktionen verĂ€ndert sind, können Krankheiten verursachen und spielen zudem eine wichtige Rolle bei der Invasion von Krebszellen. Daher besteht ein großes Interesse darin, die molekularen Mechanismen, die Integrin-EZM Interaktionen regulieren, besser zu verstehen. Wie können Zell-EZM Interaktionen untersucht werden? Obwohl es mehrere Methoden gibt, mit denen ZelladhĂ€sion untersucht werden kann, sind die wenigsten dazu geeignet, ZelladhĂ€sionskrĂ€fte zu quantifizieren. Einzelzellspektroskopie erfasst die AdhĂ€sionskrĂ€fte einzelner Zellen quantitativ und ermöglicht dadurch eine differenzierte Betrachtung der AdhĂ€sion individueller Zellen. Eine Variante der Einzelzellspektroskopie basiert auf der Rasterkraftmikroskopie (AFM); diese Technik wurde in der vorliegenden Arbeit verwendet. Ein Vorteil von AFM- Einzelzellspektroskopie besteht darin, dass Zellen mit hoher zeitlicher und rĂ€umlicher PrĂ€zision manipuliert werden können. ZelladhĂ€sionskrĂ€fte können zudem ĂŒber einen großen Kraftbereich hinweg untersucht werden. Dabei ermöglicht es die hohe Kraftauflösung, einzelne Integrin-Ligandenbindungen in lebenden Zellen zu untersuchen. Die vorliegende Arbeit gliedert sich in sechs Kapitel. Kapitel eins gibt Hintergrundinformationen ĂŒber Zell-EZM Wechselwirkungen. In Kapitel zwei werden verschiedene AdhĂ€sionsassays einander gegenĂŒber gestellt. Das theoretische Bell-Evans Modell, mit dessen Hilfe die gewonnenen Daten interpretiert wurden, wird in Kapitel drei diskutiert. Im Anschluss werden drei Projekte, welche das HerzstĂŒck dieser Doktorarbeit bilden, in Kapiteln vier bis sechs nĂ€her ausgefĂŒhrt. Im ersten Projekt (Kapitel vier) wurde die AdhĂ€sion von α2ÎČ1-Integrin exprimierenden CHO Zellen zu Kollagen I, dem hĂ€ufigsten strukturellen Protein in Wirbeltieren, quantitativ untersucht. ZunĂ€chst wurden α2ÎČ1-Kollagen-Interaktionen auf EinzelmolekĂŒlebene analysiert. Mithilfe der dynamischen Kraftspektroskopie wurden fĂŒr diese Bindung Dissoziationsrate koff (1.3 ± 1.3 sec-1) und Potentialbarrierenbreite xu (2.3 ± 0.3 Å) bestimmt. Daraufhin wurde die α2ÎČ1-vermittelte AdhĂ€sion ĂŒber einen Zeitraum von zehn Minuten untersucht. Dadurch konnten Einblicke in die Kinetik von α2ÎČ1-integrin vermittelter ZelladhĂ€sion sowie in die zugrunde liegenden Regulationsmechanismen gewonnen werden. Im zweiten Projekt (Kapitel fĂŒnf) wurde die Rolle von kryptischen Integrin-Bindungsstellen in Kollagen I untersucht. Die zuvor verwendeten KollagenoberflĂ€chen wurden thermisch denaturiert, wodurch versteckte RGD (Arg-Gly-Asp)-Sequenzen freigelegt wurden. Die partielle Denaturierung hatte- verglichen mit nativem Kollagen I- eine erhöhte AdhĂ€sion von PrĂ€osteoblasten (MC3T3-E1) zur Folge, was auf das Binden zusĂ€tzlicher Integrine zurĂŒckgefĂŒhrt wurde. Im Unterschied zu nativem Kollagen wurde die ZelladhĂ€sion zu denaturiertem Kollagen I u.a. durch αv- and α5ÎČ1-Integrine vermittelt. PrĂ€osteoblasten zeigten verstĂ€rktes Zellspreiten sowie höhere MotilitĂ€t auf denaturiertem Kollagen I; zudem wurde ein erhöhtes Differenzierungpotential der PrĂ€osteoblasten festgestellt. Die in diesem Projekt erhaltenen Einblicke bilden eine hilfreiche Basis fĂŒr die Entwicklung optimierter OberflĂ€chen fĂŒr diverse Zell- und Gewebekulturanwendungen. Im dritten Projekt (Kapitel sechs) wurde der Einfluss des Fusionproteins BCR/ABL, charakteristisch fĂŒr chronische myeloische LeukĂ€mie, auf die AdhĂ€sion von myeloischen VorlĂ€uferzellen untersucht. Dazu wurde die AdhĂ€sion von BCR/ABL transformierten VorlĂ€uferzellen (32D Zellen) bzw. Kontrollzellen zu Stromazellen (M2-10B4) sowie verschiedenen EZM Proteinen untersucht. BCR/ABL erhöhte die ZelladhĂ€sion der myeloischen VorlĂ€uferzellen signifikant. Dieser Effekt wurde durch die Zugabe von Imatinib, welches die TyrosinkinaseaktivitĂ€t von BCR/ABL inhibiert, aufgehoben. Die BCR/ABL-verstĂ€rkte ZelladhĂ€sion korrelierte mit erhöhten ÎČ1-Integrin-konzentrationen. Da die AdhĂ€sion von LeukĂ€miezellen im Knockenmark bekanntermaßen kritisch fĂŒr die Entwicklung von Resistenzen gegenĂŒber verschiedenen Wirkstoffen ist, könnten die Ergebnisse dieser Studie eine Grundlage fĂŒr die Entwicklung optimierter Target-Therapien sein. In den drei beschriebenen Projekten wurde AFM Einzelzellspektroskopie verwendet, um Integrin- vermittelte AdhĂ€sion auf molekularer Ebene zu untersuchen. Die Ergebnisse zeigen, dass AFM-Einzelzellspektroskopie ein vielseitiges Werkzeug darstellt, das ĂŒberaus geeignet dazu ist, ZelladhĂ€sion- ausgehend von EinzelmolekĂŒlinteraktionen bis hin zur Entstehung komplexerer AdhĂ€sionsstellen- auf der Kraftebene zu verfolgen

    Single-cell force spectroscopy, an emerging tool to quantify cell adhesion to biomaterials

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    Cell adhesion receptors play a central role in sensing and integrating signals provided by the cellular environment. Thus, understanding adhesive interactions at the cell-biomaterial interface is essential to improve the design of implants that should emulate certain characteristics of the cell's natural environment. Numerous cell adhesion assays have been developed; among these, atomic force microscopy-based single-cell force spectroscopy (AFM-SCFS) provides a versatile tool to quantify cell adhesion at physiological conditions. Here we discuss how AFM-SCFS can be used to quantify the adhesion of living cells to biomaterials and give examples of using AFM-SCFS in tissue engineering and regenerative medicine. We anticipate that in the near future, AFM-SCFS will be established in the biomaterial field as an important technique to quantify cell-biomaterial interactions and thereby will contribute to the optimization of implants, scaffolds, and medical devices

    EMT‐Induced Cell‐Mechanical Changes Enhance Mitotic Rounding Strength

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    To undergo mitosis successfully, most animal cells need to acquire a round shape to provide space for the mitotic spindle. This mitotic rounding relies on mechanical deformation of surrounding tissue and is driven by forces emanating from actomyosin contractility. Cancer cells are able to maintain successful mitosis in mechanically challenging environments such as the increasingly crowded environment of a growing tumor, thus, suggesting an enhanced ability of mitotic rounding in cancer. Here, it is shown that the epithelial–mesenchymal transition (EMT), a hallmark of cancer progression and metastasis, gives rise to cell-mechanical changes in breast epithelial cells. These changes are opposite in interphase and mitosis and correspond to an enhanced mitotic rounding strength. Furthermore, it is shown that cell-mechanical changes correlate with a strong EMT-induced change in the activity of Rho GTPases RhoA and Rac1. Accordingly, it is found that Rac1 inhibition rescues the EMT-induced cortex-mechanical phenotype. The findings hint at a new role of EMT in successful mitotic rounding and division in mechanically confined environments such as a growing tumor
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