Rapid fibroblast adhesion to 27nm high polymer demixed nano-topography.

It is well known that many cell types react strongly to micro-topography. It is rapidly becoming clear than cells will also react to nano-topography. Polymer demixing is a rapid and low-cost chemical method of producing nano-topography. This manuscript investigates human fibroblast response to 27nm high nano-islands produced by polymer demixing. Cell spreading, cytoskeleton, focal adhesion and Rac localisation were studied. The results showed that an initial rapid adhesion and cytoskeletal formation on the islands at 4 days of culture gave way to poorly formed contacts and vimentin cytoskeleton at 30 days of culture.

Cells react to nanoscale order and symmetry in their surroundings.

Mammalian cells react to microstructured surfaces, but there is little information on the reactions to nanostructured surfaces, and such as have been tested are poorly ordered or random in their structure. We now report that ordered surface arrays (orthogonal or hexagonal) of nanopits in polycaprolactone or polymethylmethacrylate have marked effects in reducing cell adhesion compared with less regular arrays or planar surfaces. The pits had diameters of 35, 75, and 120 nm, respectively, with pitch between the pits of 100, 200, and 300 nm, respectively. The cells appear to be able to distinguish between different symmetries of array. We suggest that interfacial forces may be organized by the nanostructures to affect the cells in the same way as they affect liquid crystal orientations.

Polyelectrolyte multilayers generated in a microfluidic device with pH gradients direct adhesion and movement of cells.

In this study, multilayers from polyethylene imine, heparin and chitosan are prepared at three different pH values of 5, 7 and 9. Water contact angle and quartz microbalance measurements show that resulting multilayers differ in terms of wetting behaviour, layer mass and mechanical properties. The multilayer is then formed within a gradient generation microfluidic (µFL) device. Polyethylene imine or heparin solutions of pH 5 are introduced into one inlet and the same solutions but at pH 9 into another inlet of the µFL device. The pH gradient established during the multilayer formation can be visualized inside the microchamber by pH sensitive fluorophores and confocal laser scanning microscopy. From this setup it is expected that properties of multilayers displayed at distinct pH values can be realised in a gradient manner inside the µFL device. Behaviour of the osteoblast cell line MG-63 seeded and cultured on top of multilayers created inside the µFL device support this hypothesis. It is observed that more cells adhere and spread on multilayers build-up at the basic side of the µFL channel, while those cells on top of multilayers built at pH 5 are fewer and smaller. These results are consistent with the behaviour of MG-63 cells seeded on multilayers formed at discrete pH values. It is particularly interesting to see that cells start to migrate from multilayers built at pH 5 to those built at pH 9 during 6 h of culture. Overall, the presented multilayer formation setup applying pH gradients leads to surfaces that promote migration of cells.

Biodegradable microgrooved polymeric surfaces obtained by photolithography for skeletal muscle cell orientation and myotube development.

During tissue formation, skeletal muscle precursor cells fuse together to form multinucleated myotubes. To understand this mechanism, in vitro systems promoting cell alignment need to be developed; for this purpose, micrometer-scale features obtained on substrate surfaces by photolithography can be used to control and affect cell behaviour. This work was aimed at investigating how differently microgrooved polymeric surfaces can affect myoblast alignment, fusion and myotube formation in vitro. Microgrooved polymeric films were obtained by solvent casting of a biodegradable poly-l-lactide/trimethylene carbonate copolymer (PLLA-TMC) onto microgrooved silicon wafers with different groove widths (5, 10, 25, 50, 100microm) and depths (0.5, 1, 2.5, 5microm), obtained by a standard photolithographic technique. The surface topography of wafers and films was evaluated by scanning electron microscopy. Cell assays were performed using C2C12 cells and myotube formation was analysed by immunofluorescence assays. Cell alignment and circularity were also evaluated using ImageJ software. The obtained results confirm the ability of microgrooved surfaces to influence myotube formation and alignment; in addition, they represent a novel further improvement to the comprehension of best features to be used. The most encouraging results were observed in the case of microstructured PLLA-TMC films with grooves of 2.5 and 1microm depth, presenting, in particular, a groove width of 50 and 25microm.

The effects of nanoscale pits on primary human osteoblast adhesion formation and cellular spreading.

Current understanding of the mechanisms involved in ossesoinegration following implantation of a biomaterial has led to an emphasis being placed on the modification of material topography to control interface reactions. Recent studies have inferred nanoscale topography as an important mediator of cell adhesion and differentiation. Biomimetic strategies in orthopaedic research aim to exploit these influences to regulate cellular adhesion and subsequent bony tissue formation. Here experimental topographies of nanoscale pits demonstrating varying order have been fabricated by electron-beam lithography in (poly)carbonate. Osteoblast adhesion to these nanotopographies was ascertained by quantification of the relation between adhesion complex formation and total cell area. This study is specifically concerned with the effects these nanotopographies have on adhesion formation in S-phase osteoblasts as identified by BrdU incorporation. Nanopits were found to reduce cellular spreading and adhesion formation.

3D polymer scaffolds for tissue engineering.

This review discusses some of the most common polymer scaffold fabrication techniques used for tissue engineering applications. Although the field of scaffold fabrication is now well established and advancing at a fast rate, more progress remains to be made, especially in engineering small diameter blood vessels and providing scaffolds that can support deep tissue structures. With this in mind, we introduce two new lithographic methods that we expect to go some way to addressing this problem.