Cell migration on material-driven fibronectin microenvironments

Cell migration is a fundamental process involved in a wide range of biological phenomena. However, how the underlying mechanisms that control migration are orchestrated is not fully understood. In this work, we explore the migratory characteristics of human fibroblasts using different organisations of fibronectin (FN) triggered by two chemically similar surfaces, poly(ethyl acrylate) (PEA) and poly(methyl acrylate) (PMA); cell migration is mediated via an intermediate layer of fibronectin (FN). FN is organised into nanonetworks upon simple adsorption on PEA whereas a globular conformation is observed on PMA. We studied cell speed over the course of 24 h and the morphology of focal adhesions in terms of area and length. Additionally, we analysed the amount of cell-secreted FN as well as FN remodelling. Velocity of human fibroblasts was found to exhibit a biphasic behaviour on PEA, whereas it remained fairly constant on PMA. FA analysis revealed more mature focal adhesions on PEA over time contrary to smaller FAs found on PMA. Finally, human fibroblasts seemed to remodel adsorbed FN more on PMA than on PEA. Overall, these results indicate that the cell–protein–material interface affects cell migratory behaviour. Analysis of FAs together with FN secretion and remodelling were associated with differences in cell velocity providing insights into the factors that can modulate cell motility

Electrospun fibrinogen-PLA nanofibres for vascular tissue engineering

Here we report on the development of a new type of hybrid fibrinogen–polylactic acid (FBG–PLA) nanofibres (NFs) with improved stiffness, combining the good mechanical properties of PLA with the excellent cell recognition properties of native FBG. We were particularly interested in the dorsal and ventral cell response to the nanofibres' organization (random or aligned), using human umbilical endothelial cells (HUVECs) as a model system. Upon ventral contact with random NFs, the cells developed a stellate-like morphology with multiple projections. The well-developed focal adhesion complexes suggested a successful cellular interaction. However, time-lapse analysis shows significantly lowered cell movements, resulting in the cells traversing a relatively short distance in multiple directions. Conversely, an elongated cell shape and significantly increased cell mobility were observed in aligned NFs. To follow the dorsal cell response, artificial wounds were created on confluent cell layers previously grown on glass slides and covered with either random or aligned NFs. Time-lapse analysis showed significantly faster wound coverage (within 12 h) of HUVECs on aligned samples vs. almost absent directional migration on random ones. However, nitric oxide (NO) release shows that endothelial cells possess lowered functionality on aligned NFs compared to random ones, where significantly higher NO production was found. Collectively, our studies show that randomly organized NFs could support the endothelization of implants while aligned NFs would rather direct cell locomotion for guided neovascularization

Bacteria-laden microgels as autonomous three-dimensional environments for stem cell engineering

A one-step microfluidic system is developed in this study which enables the encapsulation of stem cells and genetically engineered non-pathogenic bacteria into a so-called three-dimensional (3D) pearl lace–like microgel of alginate with high level of monodispersity and cell viability. The alginate-based microgel constitutes living materials that control stem cell differentiation in either an autonomous or heteronomous manner. The bacteria (Lactococcus lactis) encapsulated within the construct surface display adhesion fragments (III7-10 fragment of human fibronectin) for integrin binding while secreting growth factors (recombinant human bone morphogenetic protein-2) to induce osteogenic differentiation of human bone marrow–derived mesenchymal stem cells. We concentrate on interlinked pearl lace microgels that enabled us to prototype a low-cost 3D bioprinting platform with highly tunable properties

Assembly of living building blocks to engineer complex tissues

The great demand for tissue and organ grafts, compounded by an aging demographic and a shortage of available donors, has driven the development of bioengineering approaches that can generate biomimetic tissues in vitro. Despite the considerable progress in conventional scaffold‐based tissue engineering, the recreation of physiological complexity has remained a challenge. Bottom‐up tissue engineering strategies have opened up a new avenue for the modular assembly of living building blocks into customized tissue architectures. This Progress Report overviews the recent progress and trends in the fabrication and assembly of living building blocks, with a key highlight on emerging bioprinting technologies that can be used for modular assembly and complexity in tissue engineering. By summarizing the work to date, providing new classifications of different living building blocks, highlighting state‐of‐the‐art research and trends, and offering personal perspectives on future opportunities, this Progress Report aims to aid and inspire other researchers working in the field of modular tissue engineering

Molecular clutch drives cell response to surface viscosity

Cell response to matrix rigidity has been explained by the mechanical properties of the actin-talin-integrin-fibronectin clutch. Here the molecular clutch model is extended to account for cell interactions with purely viscous surfaces (i.e., without an elastic component). Supported lipid bilayers present an idealized and controllable system through which to study this concept. Using lipids of different diffusion coefficients, the mobility (i.e., surface viscosity) of the presented ligands (in this case RGD) was altered by an order of magnitude. Cell size and cytoskeletal organization were proportional to viscosity. Furthermore, there was a higher number of focal adhesions and a higher phosphorylation of FAK on less-mobile (more-viscous) surfaces. Actin retrograde flow, an indicator of the force exerted on surfaces, was also seen to be faster on more mobile surfaces. This has consequential effects on downstream molecules; the mechanosensitive YAP protein localized to the nucleus more on less-mobile (more-viscous) surfaces and differentiation of myoblast cells was enhanced on higher viscosity. This behavior was explained within the framework of the molecular clutch model, with lower viscosity leading to a low force loading rate, preventing the exposure of mechanosensitive proteins, and with a higher viscosity causing a higher force loading rate exposing these sites, activating downstream pathways. Consequently, the understanding of how viscosity (regardless of matrix stiffness) influences cell response adds a further tool to engineer materials that control cell behavior

Molecular clutch drives cell response to surface viscosity

Cell response to matrix rigidity has been explained by the mechanical properties of the actin-talin-integrin-fibronectin clutch. Here the molecular clutch model is extended to account for cell interactions with purely viscous surfaces (i.e., without an elastic component). Supported lipid bilayers present an idealized and controllable system through which to study this concept. Using lipids of different diffusion coefficients, the mobility (i.e., surface viscosity) of the presented ligands (in this case RGD) was altered by an order of magnitude. Cell size and cytoskeletal organization were proportional to viscosity. Furthermore, there was a higher number of focal adhesions and a higher phosphorylation of FAK on less-mobile (more-viscous) surfaces. Actin retrograde flow, an indicator of the force exerted on surfaces, was also seen to be faster on more mobile surfaces. This has consequential effects on downstream molecules; the mechanosensitive YAP protein localized to the nucleus more on less-mobile (more-viscous) surfaces and differentiation of myoblast cells was enhanced on higher viscosity. This behavior was explained within the framework of the molecular clutch model, with lower viscosity leading to a low force loading rate, preventing the exposure of mechanosensitive proteins, and with a higher viscosity causing a higher force loading rate exposing these sites, activating downstream pathways. Consequently, the understanding of how viscosity (regardless of matrix stiffness) influences cell response adds a further tool to engineer materials that control cell behavior

Hybrid Core-Shell Scaffolds for Bone Tissue Engineering

The tissue engineering applications of coaxial electrospinning are growing due to the potential increased functionality of the fibres compared to basic electrospinning. Previous studies of core and shell scaffolds have placed the active elements in the core, however, the surface response to a biomaterial affects the subsequent behaviour, thus here hydroxyapatite (HA) was added to the shell. Coaxial electrospun polycaprolactone (PCL)-polylactic acid (PLA)/HA (core-shell) scaffolds were produced in 2D sheets using a plate collector, or 3D tubes for bone tissue engineering using a rotating needle collector. The scaffolds include high hydroxyapatite content while retaining their structural and mechanical integrity. The effect of the collector type on fibre diameter, fibre alignment and mechanical properties have been evaluated, and the impact of HA incorporation on bioactivity, BMP-2 release, cell behaviour and mechanical properties for up to 12 weeks degradation were assessed. Fibre uniformity in coaxial electrospinning depends on the relative flow rate of the core and shell solutions. Using a rotating needle collector increased fibre alignment compared to a stationary collector, without affecting fibre diameter significantly, while HA content increased fibre non-uniformity. Coaxial PCL-PLA/HA fibres exhibited significantly higher bioactivity compared to PCL-PLA scaffolds due to the surface exposure of the HA particles. Apatite formation increased with increasing SBF immersion time. Coaxial tubular scaffolds with and without HA incorporation showed gradual reductions in their mechanical properties over 12 weeks in PBS or SBF but still retained their structural integrity. Coaxial scaffolds with and without HA exhibited gradual and sustained BMP-2 release and supported MSCs proliferation and differentiation with no significant difference between the two scaffolds types. These materials therefore show potential applications as bone tissue engineering scaffolds