207 research outputs found
Control of crystal polymorph in microfluidics using molluscan 28 kDa Ca2+-binding protein
No full textBiominerals produced by biological systems in physiologically relevant environments possess extraordinary properties that are often difficult to replicate under laboratory conditions. Understanding the mechanism that underlies the process of biomineralisation can lead to novel strategies in the development of advanced materials. Using microfluidics, we have demonstrated for the first time, that an extrapallial (EP) 28 kDa protein, located in the extrapallial compartment between mantle and shell of Mytilus edulis, can influence, at both micro- and nanoscopic levels, the morphology, structure and polymorph that is laid down in the shell ultrastructure. Crucially, this influence is predominantly dependent on the existence of an EP protein concentration gradient and its consecutive interaction with Ca2+ ions. Novel lemon-shaped hollow vaterite structures with a clearly defined nanogranular assembly occur only where particular EP protein and Ca2+ gradients co-exist. Computational fluid dynamics enabled the progress of the reaction to be mapped and the influence of concentration gradients across the device to be calculated. Importantly, these findings could not have been observed using conventional bulk mixing methods. Our findings not only provide direct experimental evidence of the potential influence of EP proteins in crystal formation, but also offer a new biomimetic strategy to develop functional biomaterials for applications such as encapsulation and drug delivery
A genomics approach in determining nanotopographical effects on MSC phenotype
No full textTopography and its effects on cell adhesion, morphology, growth and differentiation are well documented. Thus, current advances with the use of nanotopographies offer promising results in the field of regenerative medicine. Studies have also shown nanotopographies to have strong effects on stem cell self-renewal and differentiation. What is less clear however is what mechanotransductive mechanisms are employed by the cells to facilitate such changes. In fastidious cell types, it has been suggested that direct mechanotransduction producing morphological changes in the nucleus, nucleoskeleton and chromosomes themselves may be central to cell responses to topography. In this report we move these studies into human skeletal or mesenchymal stem cells and propose that direct (mechanical) signalling is important in the early stages of tuning stem cell fate to nanotopography. Using fluorescence in situ hybridization (FISH) and Affymetrix arrays we have evidence that nanotopography stimulates changes in nuclear organisation that can be linked to spatially regulated genes expression with a particular focus on phenotypical genes. For example, chromosome 1 was seen to display the largest numbers of gene deregulations and also a concomitant change in nuclear positioning in response to nanotopography. Plotting of deregulated genes in reference to band positioning showed that topographically related changes tend to happen towards the telomeric ends of the chromosomes, where bone related genes are generally clustered. Such an approach offers a better understanding of cell-surface interaction and, critically, provides new insights of how to control stem cell differentiation with future applications in areas including regenerative medicine
Nanotopographical control of human osteoprogenitor differentiation
No full textCurrent load-bearing orthopaedic implants are produced in 'bio-inert' materials such as titanium alloys. When inserted into the reamed bone during hip or knee replacement surgery the implants interact with mesenchymal populations including the bone marrow. Bio-inert materials are shielded from the body by differentiation of the cells along the fibroblastic lineage producing scar tissue and inferior healing. This is exacerbated by implant micromotion, which can lead to capsule formation. Thus, next-generation implant materials will have to elicit influence over osteoprogenitor differentiation and mesenchymal populations in order to recruit osteoblastic cells and produce direct bone apposition onto the implant. A powerful method of delivering cues to cells is via topography. Micro-scale topography has been shown to affect cell adhesion, migration, cytoskeleton, proliferation and differentiation of a large range of cell types (thus far all cell types tested have been shown to be responsive to topographical cues). More recent research with nanotopography has also shown a broad range of cell response, with fibroblastic cells sensing down to 10 nm in height. Initial studies with human mesenchymal populations and osteoprogenitor populations have again shown strong cell responses to nanofeatures with increased levels of osteocalcin and osteopontin production from the cells on certain topographies. This is indicative of increased osteoblastic activity on the nanotextured materials. Looking at preliminary data, it is tempting to speculate that progenitor cells are, in fact, more responsive to topography than more mature cell types and that they are actively seeking cues from their environment. This review will investigate the range of nanotopographies available to researchers and our present understanding of mechanisms of progenitor cell response. Finally, it will make some speculations of the future of nanomaterials and progenitor cells in tissue engineering
Biodegradable microgrooved polymeric surfaces obtained by photolithography for skeletal muscle cell orientation and myotube development
No full textDuring 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, 100 mu m) and depths (0.5, 1, 2.5, 5 mu m), 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 1 mu m depth, presenting, in particular, a groove width of 50 and 25 mu m
Microcontact printing of fibronectin on a biodegradable polymeric surface for skeletal muscle cell orientation
No full textBACKGROUND AND OBJECTIVES:</b> Micropatterning and microfabrication techniques have been widely used to control cell adhesion and proliferation along a preferential direction according to contact guidance theory. One of these techniques is microcontact printing, a soft lithographic technique based on the transfer of a "molecular ink" from an elastomeric stamp to a surface. This method allows the useful attachment of biomolecules in a few seconds on a variety of surfaces with sub-micrometer resolution and control, without modifying the biomolecule properties. The aim of this study is to develop an easy and versatile technique for in vitro production of arrays of skeletal muscle myofibers using microcontact printing technique on biodegradable substrata.
METHODS: Microcontact printing of fibronectin stripes (10, 25, 50 μm in width) was performed onto biodegradable L-lactide/trimethylene carbonate copolymer (PLLA-TMC) films. C2C12, a murine myoblast cell line, was used for the production of parallel myofibers.
RESULTS: This approach proved to be simple, reliable and effective in obtaining a stable pattern of fibronectin on the PLLA-TMC surface as observed by fluorescence microscopy. C2C12 cells were well aligned along the pattern 24 hours after seeding, especially on fibronectin stripes 10 and 25 μm in width. Seven days after confluence cells fused and formed aligned multinucleated cells expressing a-actinin.
CONCLUSIONS: Fibronectin patterning seems to be a useful method to induce cell alignment and to improve myotube formation. Further studies will be focused on the possibility of applying external stimuli to these structures to obtain healthy myotubes and to induce myofiber development
Progress towards tubes with regular nanopatterned inner surfaces
No full textThe repair of vascular tubing is an important task in tissue engineering. The behavior of cells is strongly influenced by the topology of the surfaces, on both a micrometric and a nanometric scale, in their vicinity. Thus the authors wish to make tubes that are patterned on the inner surface. One way to do this is to use the good depth of focus capabilities of x-ray exposure to print an array of dots, 200 nm diameter and 400 nm pitch, onto a curved surface coated in resist. A die made from this structure allows nanoembossing into a biodegradable polymer. A closed vessel can then be made by adding a lid, that also has a similar nanopatterned surface. Details of the accuracy of transfer are given. It is concluded that x-ray printing is a suitable approach for the formation of internally patterned tubing
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