The ability of cells to follow gradients of extracellular matrix stiffness—durotaxis—has been implicated in development, fibrosis, and cancer. Here, we found multicellular clusters that exhibited ...durotaxis even if isolated constituent cells did not. This emergent mode of directed collective cell migration applied to a variety of epithelial cell types, required the action of myosin motors, and originated from supracellular transmission of contractile physical forces. To explain the observed phenomenology, we developed a generalized clutch model in which local stick-slip dynamics of cell-matrix adhesions was integrated to the tissue level through cell-cell junctions. Collective durotaxis is far more efficient than single-cell durotaxis; it thus emerges as a robust mechanism to direct cell migration during development, wound healing, and collective cancer cell invasion.
The migration of cancer cells is highly regulated by the biomechanical properties of their local microenvironment. Using 3D scaffolds of simple composition, several aspects of cancer cell ...mechanosensing (signal transduction, EMC remodeling, traction forces) have been separately analyzed in the context of cell migration. However, a combined study of these factors in 3D scaffolds that more closely resemble the complex microenvironment of the cancer ECM is still missing. Here, we present a comprehensive, quantitative analysis of the role of cell-ECM interactions in cancer cell migration within a highly physiological environment consisting of mixed Matrigel-collagen hydrogel scaffolds of increasing complexity that mimic the tumor microenvironment at the leading edge of cancer invasion. We quantitatively show that the presence of Matrigel increases hydrogel stiffness, which promotes β1 integrin expression and metalloproteinase activity in H1299 lung cancer cells. Then, we show that ECM remodeling activity causes matrix alignment and compaction that favors higher tractions exerted by the cells. However, these traction forces do not linearly translate into increased motility due to a biphasic role of cell adhesions in cell migration: at low concentration Matrigel promotes migration-effective tractions exerted through a high number of small sized focal adhesions. However, at high Matrigel concentration, traction forces are exerted through fewer, but larger focal adhesions that favor attachment yielding lower cell motility.
Cells can sense the density and distribution of extracellular matrix (ECM) molecules by means of individual integrin proteins and larger, integrin-containing adhesion complexes within the cell ...membrane. This spatial sensing drives cellular activity in a variety of normal and pathological contexts. Previous studies of cells on rigid glass surfaces have shown that spatial sensing of ECM ligands takes place at the nanometre scale, with integrin clustering and subsequent formation of focal adhesions impaired when single integrin-ligand bonds are separated by more than a few tens of nanometres. It has thus been suggested that a crosslinking 'adaptor' protein of this size might connect integrins to the actin cytoskeleton, acting as a molecular ruler that senses ligand spacing directly. Here, we develop gels whose rigidity and nanometre-scale distribution of ECM ligands can be controlled and altered. We find that increasing the spacing between ligands promotes the growth of focal adhesions on low-rigidity substrates, but leads to adhesion collapse on more-rigid substrates. Furthermore, disordering the ligand distribution drastically increases adhesion growth, but reduces the rigidity threshold for adhesion collapse. The growth and collapse of focal adhesions are mirrored by, respectively, the nuclear or cytosolic localization of the transcriptional regulator protein YAP. We explain these findings not through direct sensing of ligand spacing, but by using an expanded computational molecular-clutch model, in which individual integrin-ECM bonds-the molecular clutches-respond to force loading by recruiting extra integrins, up to a maximum value. This generates more clutches, redistributing the overall force among them, and reducing the force loading per clutch. At high rigidity and high ligand spacing, maximum recruitment is reached, preventing further force redistribution and leading to adhesion collapse. Measurements of cellular traction forces and actin flow speeds support our model. Our results provide a general framework for how cells sense spatial and physical information at the nanoscale, precisely tuning the range of conditions at which they form adhesions and activate transcriptional regulation.
Summary
Some bacteria have coevolved to establish symbiotic or pathogenic relationships with plants, animals or humans. With human association, the bacteria can cause a variety of diseases. Thus, ...understanding bacterial phenotypes at the single‐cell level is essential to develop beneficial applications. Traditional microbiological techniques have provided great knowledge about these organisms; however, they have also shown limitations, such as difficulties in culturing some bacteria, the heterogeneity of bacterial populations or difficulties in recreating some physical or biological conditions. Microfluidics is an emerging technique that complements current biological assays. Since microfluidics works with micrometric volumes, it allows fine‐tuning control of the test conditions. Moreover, it allows the recruitment of three‐dimensional (3D) conditions, in which several processes can be integrated and gradients can be generated, thus imitating physiological 3D environments. Here, we review some key microfluidic‐based studies describing the effects of different microenvironmental conditions on bacterial response, biofilm formation and antimicrobial resistance. For this aim, we present different studies classified into six groups according to the design of the microfluidic device: (i) linear channels, (ii) mixing channels, (iii) multiple floors, (iv) porous devices, (v) topographic devices and (vi) droplet microfluidics. Hence, we highlight the potential and possibilities of using microfluidic‐based technology to study bacterial phenotypes in comparison with traditional methodologies.
Microfluidics is an emerging technique that can complements current biological assays. Here, we review some key microfluidic‐based studies describing the effects of different microenvironmental conditions on bacterial phenotypes. We also highlight the potential and possibilities of using microfluidic‐based technology to study bacterial phenotypes in comparison with traditional methodologies.
Microfluidic devices are becoming mainstream tools to recapitulate in vitro the behavior of cells and tissues. In this study, we use microfluidic devices filled with hydrogels of mixed ...collagen-Matrigel composition to study the migration of lung cancer cells under different cancer invasion microenvironments. We present the design of the microfluidic device, characterize the hydrogels morphologically and mechanically and use quantitative image analysis to measure the migration of H1299 lung adenocarcinoma cancer cells in different experimental conditions. Our results show the plasticity of lung cancer cell migration, which turns from mesenchymal in collagen only matrices, to lobopodial in collagen-Matrigel matrices that approximate the interface between a disrupted basement membrane and the underlying connective tissue. Our quantification of migration speed confirms a biphasic role of Matrigel. At low concentration, Matrigel facilitates migration, most probably by providing a supportive and growth factor retaining environment. At high concentration, Matrigel slows down migration, possibly due excessive attachment. Finally, we show that antibody-based integrin blockade promotes a change in migration phenotype from mesenchymal or lobopodial to amoeboid and analyze the effect of this change in migration dynamics, in regards to the structure of the matrix. In summary, we describe and characterize a robust microfluidic platform and a set of software tools that can be used to study lung cancer cell migration under different microenvironments and experimental conditions. This platform could be used in future studies, thus benefitting from the advantages introduced by microfluidic devices: precise control of the environment, excellent optical properties, parallelization for high throughput studies and efficient use of therapeutic drugs.
Bone is a living tissue whose main mechanical function is to provide stiffness, strength and protection to the body. Both stiffness and strength depend on the mineralization of the organic matrix, ...which is constantly being remodelled by the coordinated action of the bone multicellular units (BMUs). Due to the dynamics of both remodelling and mineralization, each sample of bone is composed of structural units (osteons in cortical and packets in cancellous bone) created at different times, therefore presenting different levels of mineral content. In this work, a computational model is used to understand the feedback between the remodelling and the mineralization processes under different load conditions and bone porosities. This model considers that osteoclasts primarily resorb those parts of bone closer to the surface, which are younger and less mineralized than older inner ones. Under equilibrium loads, results show that bone volumes with both the highest and the lowest levels of porosity (cancellous and cortical respectively) tend to develop higher levels of mineral content compared to volumes with intermediate porosity, thus presenting higher material densities. In good agreement with recent experimental measurements, a boomerang-like pattern emerges when plotting apparent density at the tissue level versus material density at the bone material level. Overload and disuse states are studied too, resulting in a translation of the apparent-material density curve. Numerical results are discussed pointing to potential clinical applications.
The healing of bone defects is a challenge for both tissue engineering and modern orthopaedics. This problem has been addressed through the study of scaffold constructs combined with ...mechanoregulatory theories, disregarding the influence of chemical factors and their respective delivery devices. Of the chemical factors involved in the bone healing process, bone morphogenetic protein-2 (BMP-2) has been identified as one of the most powerful osteoinductive proteins. The aim of this work is to develop and validate a mechano-chemical regulatory model to study the effect of BMP-2 on the healing of large bone defects in silico. We first collected a range of quantitative experimental data from the literature concerning the effects of BMP-2 on cellular activity, specifically proliferation, migration, differentiation, maturation and extracellular matrix production. These data were then used to define a model governed by mechano-chemical stimuli to simulate the healing of large bone defects under the following conditions: natural healing, an empty hydrogel implanted in the defect and a hydrogel soaked with BMP-2 implanted in the defect. For the latter condition, successful defect healing was predicted, in agreement with previous in vivo experiments. Further in vivo comparisons showed the potential of the model, which accurately predicted bone tissue formation during healing, bone tissue distribution across the defect and the quantity of bone inside the defect. The proposed mechano-chemical model also estimated the effect of BMP-2 on cells and the evolution of healing in large bone defects. This novel in silico tool provides valuable insight for bone tissue regeneration strategies.
Dose calculation plays a critical role in radiotherapy (RT) treatment planning, and there is a growing need to develop accurate dose deposition models that incorporate heterogeneous tumour ...properties. Deterministic models have demonstrated their capability in this regard, making them the focus of recent treatment planning studies as they serve as a basis for simplified models in RT treatment planning. In this study, we present a simplified deterministic model for photon transport based on the Boltzmann transport equation (BTE) as a proof‐of‐concept to illustrate the impact of heterogeneous tumour properties on RT treatment planning. We employ the finite element method (FEM) to simulate the photon flux and dose deposition in real cases of diffuse intrinsic pontine glioma (DIPG) and neuroblastoma (NB) tumours. Importantly, in light of the availability of pipelines capable of extracting tumour properties from magnetic resonance imaging (MRI) data, we highlight the significance of such data. Specifically, we utilise cellularity data extracted from DIPG and NB MRI images to demonstrate the importance of heterogeneity in dose calculation. Our model simplifies the process of simulating a RT treatment system and can serve as a useful starting point for further research. To simulate a full RT treatment system, one would need a comprehensive model that couples the transport of electrons and photons.
We present a simplified deterministic model integrating heterogeneous tumour properties, based on the Boltzmann transport equation for photon transport in radiotherapy (RT) treatment planning. Using the finite element method, the model predicts a correlation between dose deposition in real tumour cases with cellularity. The study highlights the significance of incorporating MRI‐extracted tumour properties, like cellularity, for accurate radiation dose calculation. This model serves as a valuable starting point for further research on comprehensive RT treatment systems.
The extracellular matrix (ECM) plays an important regulatory role in the development and progression of tumoral tissue. Its functions and properties are crucial in determining tumor cell behavior ...such as invasion, migration, and malignancy development. Our study explores the role of collagen type I in cancer development and spread using engineered tumor models like multicellular spheroids grown in collagen-based hydrogels to simulate early tumor formation. We employ microfluidic techniques to test the hypothesis that (i) adding Laponite nanoclay to collagen hydrogels modifies mechanical and rheological properties and (ii) changing the stiffness of the collagen microenvironment affects tumor spheroid growth. Our findings support our theories and suggest the use of ECM components and engineered tumor models in cancer research, offering a biocompatible and biomimetic method to tailor the mechanical properties of conventional collagen hydrogels.
Wound healing is a process driven by cells. The ability of cells to sense mechanical stimuli from the extracellular matrix that surrounds them is used to regulate the forces that cells exert on the ...tissue. Stresses exerted by cells play a central role in wound contraction and have been broadly modelled. Traditionally, these stresses are assumed to be dependent on variables such as the extracellular matrix and cell or collagen densities. However, we postulate that cells are able to regulate the healing process through a mechanosensing mechanism regulated by the contraction that they exert. We propose that cells adjust the contraction level to determine the tissue functions regulating all main activities, such as proliferation, differentiation and matrix production. Hence, a closed-regulatory feedback loop is proposed between contraction and tissue formation. The model consists of a system of partial differential equations that simulates the evolution of fibroblasts, myofibroblasts, collagen and a generic growth factor, as well as the deformation of the extracellular matrix. This model is able to predict the wound healing outcome without requiring the addition of phenomenological laws to describe the time-dependent contraction evolution. We have reproduced two in vivo experiments to evaluate the predictive capacity of the model, and we conclude that there is feedback between the level of cell contraction and the tissue regenerated in the wound.
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