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Biosensors for studies on adhesion-mediated cellular responses to their microenvironment

Cells interact with their microenvironment by constantly sensing mechanical and chemical cues converting them into biochemical signals. These processes allow cells to respond and adapt to changes in their environment, and are crucial for most cellular functions. Understanding the mechanism underlying this complex interplay at the cell-matrix interface is of fundamental value to decipher key biochemical and mechanical factors regulating cell fate. The combination of material science and surface chemistry aided in the creation of controllable environments to study cell mechanosensing and mechanotransduction. Biologically inspired materials tailored with specific bioactive molecules, desired physical properties and tunable topography have emerged as suitable tools to study cell behavior. Among these materials, synthetic cell interfaces with built-in sensing capabilities are highly advantageous to measure biophysical and biochemical interaction between cells and their environment. In this review, we discuss the design of micro and nanostructured biomaterials engineered not only to mimic the structure, properties, and function of the cellular microenvironment, but also to obtain quantitative information on how cells sense and probe specific adhesive cues from the extracellular domain. This type of responsive biointerfaces provides a readout of mechanics, biochemistry, and electrical activity in real time allowing observation of cellular processes with molecular specificity. Specifically designed sensors based on advanced optical and electrochemical readout are discussed. We further provide an insight into the emerging role of multifunctional micro and nanosensors to control and monitor cell functions by means of material design.Fil: Saffioti, Nicolas Andres. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de San Martin. Instituto de Nanosistemas; ArgentinaFil: Cavalcanti Adam, Elisabetta Ada. Max Planck Institute for Medical Research. Department Of Cellular Biophysics; AlemaniaFil: Pallarola, Diego Andres. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de San Martin. Instituto de Nanosistemas; Argentin

Amoeboid-mesenchymal migration plasticity promotes invasion only in complex heterogeneous microenvironments

During tissue invasion individual tumor cells exhibit two interconvertible migration modes, namely mesenchymal and amoeboid migration. The cellular microenvironment triggers the switch between both modes, thereby allowing adaptation to dynamic conditions. It is, however, unclear if this amoeboid-mesenchymal migration plasticity contributes to a more effective tumor invasion. We address this question with a mathematical model, where the amoeboid-mesenchymal migration plasticity is regulated in response to local extracellular matrix resistance. Our numerical analysis reveals that extracellular matrix structure and presence of a chemotactic gradient are key determinants of the model behavior. Only in complex microenvironments, if the extracellular matrix is highly heterogeneous and a chemotactic gradient directs migration, the amoeboid-mesenchymal migration plasticity allows a more widespread invasion compared to the non-switching amoeboid and mesenchymal modes. Importantly, these specific conditions are characteristic for in vivo tumor invasion. Thus, our study suggests that in vitro systems aiming at unraveling the underlying molecular mechanisms of tumor invasion should take into account the complexity of the microenvironment by considering the combined effects of structural heterogeneities and chemical gradients on cell migration

Copresentation of BMP-6 and RGD ligands enhances cell adhesion and BMP-mediated signaling

We report on the covalent immobilization of bone morphogenetic protein 6 (BMP-6) and its co-presentation with integrin ligands on a nanopatterned platform to study cell adhesion and signaling responses which regulate the transdifferentiation of myoblasts into osteogenic cells. To immobilize BMP-6, the heterobifunctional linker MU-NHS is coupled to amine residues of the growth factor; this prevents its internalization while ensuring that its biological activity is maintained. Additionally, to allow cells to adhere to such platform and study signaling events arising from the contact to the surface, we used click-chemistry to immobilize cyclic-RGD carrying an azido group reacting with PEG-alkyne spacers via copper-catalyzed 1,3-dipolar cycloaddition. We show that the copresentation of BMP-6 and RGD favors focal adhesion formation and promotes Smad 1/5/8 phosphorylation. When presented in low amounts, BMP-6 added to culture media of cells adhering to the RGD ligands is less effective than BMP-6 immobilized on the surfaces in inducing Smad complex activation and in inhibiting myotube formation. Our results suggest that a local control of ligand density and cell signaling is crucial for modulating cell response

Adherent cells avoid polarization gradients on periodically poled LiTaO\u3csub\u3e3\u3c/sub\u3e ferroelectrics

The response of fibroblast cells to periodically poled LiTaO3 ferroelectric crystals has been studied. While fibroblast cells do not show morphological differences on the two polarization directions, they show a tendency to avoid the field gradients that occur between polarization domains of the ferroelectric. The response to the field gradients is fully established after one hour, a time at which fibroblasts form their first focal contacts. If suspension cells, with a lower tendency to establish strong surface contacts are used, no influence of the field gradients is observed

Adherent cells avoid polarization gradients on periodically poled LiTaO\u3csub\u3e3\u3c/sub\u3e ferroelectrics

The response of fibroblast cells to periodically poled LiTaO3 ferroelectric crystals has been studied. While fibroblast cells do not show morphological differences on the two polarization directions, they show a tendency to avoid the field gradients that occur between polarization domains of the ferroelectric. The response to the field gradients is fully established after one hour, a time at which fibroblasts form their first focal contacts. If suspension cells, with a lower tendency to establish strong surface contacts are used, no influence of the field gradients is observed

Mimicking bone extracellular matrix: from BMP-2-derived sequences to osteogenic-multifunctional coatings

Cell–material interactions are regulated by mimicking bone extracellular matrix on the surface of biomaterials. In this regard, reproducing the extracellular conditions that promote integrin and growth factor (GF) signaling is a major goal to trigger bone regeneration. Thus, the use of synthetic osteogenic domains derived from bone morphogenetic protein 2 (BMP-2) is gaining increasing attention, as this strategy is devoid of the clinical risks associated with this molecule. In this work, the wrist and knuckle epitopes of BMP-2 are screened to identify peptides with potential osteogenic properties. The most active sequences (the DWIVA motif and its cyclic version) are combined with the cell adhesive RGD peptide (linear and cyclic variants), to produce tailor-made biomimetic peptides presenting the bioactive cues in a chemically and geometrically defined manner. Such multifunctional peptides are next used to functionalize titanium surfaces. Biological characterization with mesenchymal stem cells demonstrates the ability of the biointerfaces to synergistically enhance cell adhesion and osteogenic differentiation. Furthermore, in vivo studies in rat calvarial defects prove the capacity of the biomimetic coatings to improve new bone formation and reduce fibrous tissue thickness. These results highlight the potential of mimicking integrin-GF signaling with synthetic peptides, without the need for exogenous GFs.Peer ReviewedPostprint (published version

A comprehensive evaluation of the activity and selectivity profile of ligands for RGD-binding integrins

Integrins, a diverse class of heterodimeric cell surface receptors, are key regulators of cell structure and behaviour, affecting cell morphology, proliferation, survival and differentiation. Consequently, mutations in specific integrins, or their deregulated expression, are associated with a variety of diseases. In the last decades, many integrin-specific ligands have been developed and used for modulation of integrin function in medical as well as biophysical studies. The IC50-values reported for these ligands strongly vary and are measured using different cell-based and cell-free systems. A systematic comparison of these values is of high importance for selecting the optimal ligands for given applications. In this study, we evaluate a wide range of ligands for their binding affinity towards the RGD-binding integrins avß3, avß5, avß6, avß8, a5ß1, aIIbß3, using homogenous ELISA-like solid phase binding assay.Postprint (published version

Epithelial cell cluster size affects force distribution in response to EGF-induced collective contractility

Several factors present in the extracellular environment regulate epithelial cell adhesion and dynamics. Among them, growth factors such as EGF, upon binding to their receptors at the cell surface, get internalized and directly activate the acto-myosin machinery. In this study we present the effects of EGF on the contractility of epithelial cancer cell colonies in confined geometry of different sizes. We show that the extent to which EGF triggers contractility scales with the cluster size and thus the number of cells. Moreover, the collective contractility results in a radial distribution of traction forces, which are dependent on integrin β1 peripheral adhesions and transmitted to neighboring cells through adherens junctions. Taken together, EGF-induced contractility acts on the mechanical crosstalk and linkage between the cell-cell and cell-matrix compartments, regulating collective responses

Functionalizing avß3- or a5ß1-selective integrin antagonists for surface coating: A new tool to discriminate integrin subtypes in vitro

Thumbnail image of graphical abstract Stuck with the right choice: avß3- or a5ß1-selective integrin ligands were functionalized for surface coating without losing activity and selectivity. The coating of nanostructured gold surfaces with these compounds stimulated subtype-selective cell adhesion of genetically modified avß3- or a5ß1-expressing fibroblasts in vitro.Postprint (published version