JoVE Visualize What is visualize?
Stop Reading. Start Watching.
Advanced Search
Stop Reading. Start Watching.
Regular Search
Find video protocols related to scientific articles indexed in Pubmed.
Nanotopographical Induction of Osteogenesis through Adhesion, Bone Morphogenic Protein Cosignaling, and Regulation of MicroRNAs.
ACS Nano
PUBLISHED: 09-23-2014
Show Abstract
Hide Abstract
It is emerging that nanotopographical information can be used to induce osteogenesis from mesenchymal stromal cells from the bone marrow, and it is hoped that this nanoscale bioactivity can be utilized to engineer next generation implants. However, the osteogenic mechanism of surfaces is currently poorly understood. In this report, we investigate mechanism and implicate bone morphogenic protein (BMP) in up-regulation of RUNX2 and show that RUNX2 and its regulatory miRNAs are BMP sensitive. Our data demonstrate that osteogenic nanotopography promotes colocalization of integrins and BMP2 receptors in order to enhance osteogenic activity and that vitronectin is important in this interface. This provides insight that topographical regulation of adhesion can have effects on signaling cascades outside of cytoskeletal signaling and that adhesions can have roles in augmenting BMP signaling.
Related JoVE Video
Nanotopographical effects on mesenchymal stem cell morphology and phenotype.
J. Cell. Biochem.
PUBLISHED: 07-24-2014
Show Abstract
Hide Abstract
There is a rapidly growing body of literature on the effects of topography and critically, nanotopography on cell adhesion, apoptosis and differentiation. Understanding the effects of nanotopography on cell adhesion and morphology and the consequences of cell shape changes in the nucleus, and consequently, gene expression offers new approaches to the elucidation and potential control of stem cell differentiation. In the current study we have used molecular approaches in combination with immunohistology and transcript analysis to understand the role of nanotopography on mesenchymal stem cell morphology and phenotype. Results demonstrate large changes in cell adhesion, nucleus and lamin morphologies in response to the different nanotopographies. Furthermore, these changes relate to alterations in packing of chromosome territories within the interphase nucleus. This, in turn, leads to changes in transcription factor activity and functional (phenotypical) signalling including cell metabolism. Nanotopography provides a useful, non-invasive tool for studying cellular mechanotransduction, gene and protein expression patterns, through effects on cell morphology. The different nanotopographies examined, result in different morphological changes in the cyto- and nucleo-skeleton. We propose that both indirect (biochemical) and direct (mechanical) signalling are important in these early stages of regulating stem cell fate as a consequence of altered metabolic changes and altered phenotype. The current studies provide new insight on cell-surface interactions and enhance our understanding of the modulation of stem cell differentiation with significant potential application in regenerative medicine.
Related JoVE Video
Design and fabrication of memory devices based on nanoscale polyoxometalate clusters.
Nature
PUBLISHED: 06-26-2014
Show Abstract
Hide Abstract
Flash memory devices-that is, non-volatile computer storage media that can be electrically erased and reprogrammed-are vital for portable electronics, but the scaling down of metal-oxide-semiconductor (MOS) flash memory to sizes of below ten nanometres per data cell presents challenges. Molecules have been proposed to replace MOS flash memory, but they suffer from low electrical conductivity, high resistance, low device yield, and finite thermal stability, limiting their integration into current MOS technologies. Although great advances have been made in the pursuit of molecule-based flash memory, there are a number of significant barriers to the realization of devices using conventional MOS technologies. Here we show that core-shell polyoxometalate (POM) molecules can act as candidate storage nodes for MOS flash memory. Realistic, industry-standard device simulations validate our approach at the nanometre scale, where the device performance is determined mainly by the number of molecules in the storage media and not by their position. To exploit the nature of the core-shell POM clusters, we show, at both the molecular and device level, that embedding [(Se(iv)O3)2](4-) as an oxidizable dopant in the cluster core allows the oxidation of the molecule to a [Se(v)2O6](2-) moiety containing a {Se(v)-Se(v)} bond (where curly brackets indicate a moiety, not a molecule) and reveals a new 5+ oxidation state for selenium. This new oxidation state can be observed at the device level, resulting in a new type of memory, which we call 'write-once-erase'. Taken together, these results show that POMs have the potential to be used as a realistic nanoscale flash memory. Also, the configuration of the doped POM core may lead to new types of electrical behaviour. This work suggests a route to the practical integration of configurable molecules in MOS technologies as the lithographic scales approach the molecular limit.
Related JoVE Video
Harnessing nanotopography and integrin-matrix interactions to influence stem cell fate.
Nat Mater
PUBLISHED: 04-09-2014
Show Abstract
Hide Abstract
Stem cells respond to nanoscale surface features, with changes in cell growth and differentiation mediated by alterations in cell adhesion. The interaction of nanotopographical features with integrin receptors in the cells' focal adhesions alters how the cells adhere to materials surfaces, and defines cell fate through changes in both cell biochemistry and cell morphology. In this Review, we discuss how cell adhesions interact with nanotopography, and we provide insight as to how materials scientists can exploit these interactions to direct stem cell fate and to understand how the behaviour of stem cells in their niche can be controlled. We expect knowledge gained from the study of cell-nanotopography interactions to accelerate the development of next-generation stem cell culture materials and implant interfaces, and to fuel discovery of stem cell therapeutics to support regenerative therapies.
Related JoVE Video
Development of a Novel 3D Culture System for Screening Features of a Complex Implantable Device for CNS Repair.
Mol. Pharm.
PUBLISHED: 12-06-2013
Show Abstract
Hide Abstract
Tubular scaffolds which incorporate a variety of micro- and nanotopographies have a wide application potential in tissue engineering especially for the repair of spinal cord injury (SCI). We aim to produce metabolically active differentiated tissues within such tubes, as it is crucially important to evaluate the biological performance of the three-dimensional (3D) scaffold and optimize the bioprocesses for tissue culture. Because of the complex 3D configuration and the presence of various topographies, it is rarely possible to observe and analyze cells within such scaffolds in situ. Thus, we aim to develop scaled down mini-chambers as simplified in vitro simulation systems, to bridge the gap between two-dimensional (2D) cell cultures on structured substrates and three-dimensional (3D) tissue culture. The mini-chambers were manipulated to systematically simulate and evaluate the influences of gravity, topography, fluid flow, and scaffold dimension on three exemplary cell models that play a role in CNS repair (i.e., cortical astrocytes, fibroblasts, and myelinating cultures) within a tubular scaffold created by rolling up a microstructured membrane. Since we use CNS myelinating cultures, we can confirm that the scaffold does not affect neural cell differentiation. It was found that heterogeneous cell distribution within the tubular constructs was caused by a combination of gravity, fluid flow, topography, and scaffold configuration, while cell survival was influenced by scaffold length, porosity, and thickness. This research demonstrates that the mini-chambers represent a viable, novel, scale down approach for the evaluation of complex 3D scaffolds as well as providing a microbioprocessing strategy for tissue engineering and the potential repair of SCI.
Related JoVE Video
The origin of off-resonance non-linear optical activity of a gold chiral nanomaterial.
Nanoscale
PUBLISHED: 11-05-2013
Show Abstract
Hide Abstract
We demonstrate that engineered artificial gold chiral nanostructures display significant levels of non-linear optical activity even without plasmonic enhancement. Our work suggests that although plasmonic excitation enhances the intensity of second harmonic emission it is not a prerequisite for significant non-linear (second harmonic) optical activity. It is also shown that the non-linear optical activities of both the chiral nanostructures and simple chiral molecules on surfaces have a common origin, namely pure electric dipole excitation. This is a surprising observation given the significant difference in length scales, three orders of magnitude, between the nanostructures and simple chiral molecules. Intuitively, given that the dimensions of the nanostructures are comparable to the wavelength of visible light, one would expect non-localised higher multipole excitation (e.g. electric quadrupole and magnetic dipole) to make the dominant contribution to non-linear optical activity. This study provides experimental evidence that the electric dipole origin of non-linear optical activity is a generic phenomenon which is not limited to sub-wavelength molecules and assemblies. Our work suggests that viewing non-plasmonic nanostructures as "meta-molecules" could be useful for rationally designing substrates for optimal non-linear optical activity.
Related JoVE Video
Nanotopographical cues augment mesenchymal differentiation of human embryonic stem cells.
Small
PUBLISHED: 01-30-2013
Show Abstract
Hide Abstract
The production of bone-forming osteogenic cells for research purposes or transplantation therapies remains a significant challenge. Using planar polycarbonate substrates lacking in topographical cues and substrates displaying a nanotopographical pattern, mesenchymal differentiation of human embryonic stem cells is directed in the absence of chemical factors and without induction of differentiation by embryoid body formation. Cells incubated on nanotopographical substrates show enhanced expression of mesenchymal or stromal markers and expression of early osteogenic progenitors at levels above those detected in cells on planar substrates in the same basal media. Evidence of epithelial-to-mesenchymal transition during substrate differentiation and DNA methylation changes akin to chemical induction are also observed. These studies provide a suitable approach to overcome regenerative medical challenges and describe a defined, reproducible platform for human embryonic stem cell differentiation.
Related JoVE Video
A genomics approach in determining nanotopographical effects on MSC phenotype.
Biomaterials
PUBLISHED: 01-09-2013
Show Abstract
Hide Abstract
Topography 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.
Related JoVE Video
Label-free segmentation of Co-cultured cells on a nanotopographical gradient.
Nano Lett.
PUBLISHED: 01-02-2013
Show Abstract
Hide Abstract
The function and fate of cells is influenced by many different factors, one of which is surface topography of the support culture substrate. Systematic studies of nanotopography and cell response have typically been limited to single cell types and a small set of topographical variations. Here, we show a radical expansion of experimental throughput using automated detection, measurement, and classification of co-cultured cells on a nanopillar array where feature height changes continuously from planar to 250 nm over 9 mm. Individual cells are identified and characterized by more than 200 descriptors, which are used to construct a set of rules for label-free segmentation into individual cell types. Using this approach we can achieve label-free segmentation with 84% confidence across large image data sets and suggest optimized surface parameters for nanostructuring of implant devices such as vascular stents.
Related JoVE Video
Nanoscale surfaces for the long-term maintenance of mesenchymal stem cell phenotype and multipotency.
Nat Mater
PUBLISHED: 05-31-2011
Show Abstract
Hide Abstract
There is currently an unmet need for the supply of autologous, patient-specific stem cells for regenerative therapies in the clinic. Mesenchymal stem cell differentiation can be driven by the material/cell interface suggesting a unique strategy to manipulate stem cells in the absence of complex soluble chemistries or cellular reprogramming. However, so far the derivation and identification of surfaces that allow retention of multipotency of this key regenerative cell type have remained elusive. Adult stem cells spontaneously differentiate in culture, resulting in a rapid diminution of the multipotent cell population and their regenerative capacity. Here we identify a nanostructured surface that retains stem-cell phenotype and maintains stem-cell growth over eight weeks. Furthermore, the study implicates a role for small RNAs in repressing key cell signalling and metabolomic pathways, demonstrating the potential of surfaces as non-invasive tools with which to address the stem cell niche.
Related JoVE Video
Protein Expression of STRO-1 Cells in Response to Different Topographic Features.
J Tissue Eng
PUBLISHED: 04-09-2011
Show Abstract
Hide Abstract
Human skeletal stem cells (STRO-1 positive) display distinct responses to different topographical features. On a flat surface, skeletal cells spread, and in vitro, they typically display a polarized, fibroblast-like morphology. However, on microgrooved surfaces, these cells prefer to stretch along the grooves forming a similar morphology to in vivo, bipolarized fibroblasts. In contrast, on nanopits, these cells display a polygonal and osteoblastic phenotype. We have examined mechanotransduction events of STRO-1 positive in response to fibroblastic, microgrooved and osteogenic, controlled disorder nanopit, topographies using proteomics after 3 days in culture. Protein expression profiles were analyzed by difference gel electrophoresis to identify proteins that showed modulation of expression in response to different topographic features to assess early decision events in these cells on these discrete topographies. After only 72 hours in culture, STRO-1 positive displayed differential regulations of families of proteins involved in cell migration and proliferation. The current study indicated that osteogenic decision specific events had already occurred. Runx2 was localized in nuclei of the skeletal stem cells on the osteogenic nanopits; however, few signaling pathway changes were observed. This study demonstrated that micro- and nanotopographies activated skeletal stem cells at different times and with distinct mechanotransduction profiles.
Related JoVE Video
A biodegradable and biocompatible regular nanopattern for large-scale selective cell growth.
Small
PUBLISHED: 11-12-2010
Show Abstract
Hide Abstract
A biodegradable substrate with a regular array of nanopillars fabricated by electron-beam lithography and hot embossing is used to address the mechanisms of nanotopographical control of cell behavior. Two different cell lines cultured on the nanopillars show striking differences in cell coverage. These changes are topography- and cell-dependent, and are not mediated by air bubbles trapped on the nanopattern. For the first time, a strong cell-selective effect of the same nanotopography has been clearly demonstrated on a large area; while fibroblast proliferation is inhibited, endothelial cell spreading is visibly enhanced. The reduced fibroblast proliferation indicates that a reduction of available surface area induced by nanotopography might be the main factor affecting cell growth on nanopatterns. The results presented herein pave the way towards the development of permanent vascular replacements, where non-adhesive, inert, surfaces will induce rapid in situ endothelialization to reduce thrombosis and occlusion.
Related JoVE Video
The interactions of astrocytes and fibroblasts with defined pore structures in static and perfusion cultures.
Biomaterials
PUBLISHED: 10-22-2010
Show Abstract
Hide Abstract
Open pores to maintain nutrient diffusion and waste removal after cell colonization are crucial for the successful application of constructs based on assembled membranes, in our case tubular scaffolds made of ?-polycaprolactone (PCL), for use in tissue engineering. Due to the complex three-dimensional structure and large size of such scaffolds needed for transplantable tissues, it is difficult to investigate the cell-pore interactions in situ. Therefore miniaturized bioreactors inside Petri dishes (30 mm in diameter), containing porous PCL or poly-dimethylsiloxane (PDMS) membranes, were developed to allow the interactions of different cells with defined pores to be investigated in situ during both static and perfusion cultures. Investigation of two different cell types (fibroblasts and cortical astrocytes) and how they interact with a range of pores (100-350 ?m in diameter) for up to 50 days indicated that the cells either covered or bridged the pores. Three distinct behaviors were observed in the way cortical astrocytes interacted with pores, while fibroblasts were able to quickly bridge the pores based on consistent "joint efforts". Our studies demonstrate that the distinct pore sealing behaviors of both cell types were influenced by pore size, initial cell density and culture period, but not by medium perfusion within the range of shear forces investigated. These findings form important basic data about the usability of pores within scaffolds that could inform the design and fabrication of suitable scaffolds for various applications in tissue engineering.
Related JoVE Video
Control of crystal polymorph in microfluidics using molluscan 28 kDa Ca²(+)-binding protein.
Integr Biol (Camb)
PUBLISHED: 08-27-2010
Show Abstract
Hide Abstract
Biominerals 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.
Related JoVE Video
Microcontact printing of fibronectin on a biodegradable polymeric surface for skeletal muscle cell orientation.
Int J Artif Organs
PUBLISHED: 07-05-2010
Show Abstract
Hide Abstract
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.
Related JoVE Video
Preparation of Caco-2 cell sheets using plasma polymerised acrylic acid as a weak boundary layer.
Biomaterials
PUBLISHED: 03-24-2010
Show Abstract
Hide Abstract
The use of cell sheets for tissue engineering applications has considerable advantages over single cell seeding techniques. So far, only thermoresponsive surfaces have been used to manufacture cell sheets without chemically disrupting the cell-surface interactions. Here, we present a new and facile technique to prepare sheets of epithelial cells using plasma polymerised acrylic acid films. The cell sheets are harvested by gentle agitation of the media without the need of any additional external stimulus. We demonstrate that the plasma polymer deposition conditions affect the viability and metabolic activity of the cells in the sheet and relate these effects to the different surface properties of the plasma polymerised acrylic acid films. Based on surface analysis data, a first attempt is made to explain the mechanism behind the cell sheet formation. The advantage of the epithelial cell sheets generated here over single cell suspensions to seed a PLGA scaffold is presented. The scaffold itself, prepared using a mould fabricated via photolithography, exhibits a unique architecture that mimics closely the dimensions of the native tissue (mouse intestine).
Related JoVE Video
Interactions with nanoscale topography: adhesion quantification and signal transduction in cells of osteogenic and multipotent lineage.
J Biomed Mater Res A
PUBLISHED: 11-03-2009
Show Abstract
Hide Abstract
Polymeric medical devices widely used in orthopedic surgery play key roles in fracture fixation and orthopedic implant design. Topographical modification and surface micro-roughness of these devices regulate cellular adhesion, a process fundamental in the initiation of osteoinduction and osteogenesis. Advances in fabrication techniques have evolved the field of surface modification; in particular, nanotechnology has allowed the development of nanoscale substrates for the investigation into cell-nanofeature interactions. In this study human osteoblasts (HOBs) were cultured on ordered nanoscale pits and random nano "craters" and "islands". Adhesion subtypes were quantified by immunofluorescent microscopy and cell-substrate interactions investigated via immuno-scanning electron microscopy. To investigate the effects of these substrates on cellular function 1.7 k microarray analysis was used to establish gene profiles of enriched STRO-1+ progenitor cell populations cultured on these nanotopographies. Nanotopographies affected the formation of adhesions on experimental substrates. Adhesion formation was prominent on planar control substrates and reduced on nanocrater and nanoisland topographies; nanopits, however, were shown to inhibit directly the formation of large adhesions. STRO-1+ progenitor cells cultured on experimental substrates revealed significant changes in genetic expression. This study implicates nanotopographical modification as a significant modulator of osteoblast adhesion and cellular function in mesenchymal populations.
Related JoVE Video
Use of neutron reflectivity to measure the dynamics of solvation and structural changes in polyvinylferrocene films during electrochemically controlled redox cycling.
Langmuir
PUBLISHED: 08-29-2009
Show Abstract
Hide Abstract
Time-resolved specular neutron reflectivity measurements are presented and interpreted for electroactive polyvinylferrocene (PVF) films subject to potentiodynamic electrochemical control. New data acquisition methodology allows an effective measurement time scale on the order of seconds, which is an improvement over conventional methodology by 2 to 3 orders of magnitude. Reflectivity profiles were obtained for PVF films exposed to aqueous 0.1 M NaClO4 in which PVF films are thermodynamically permselective, with contrast variation via H2O and D2O. Irrespective of any model, the raw profiles show chemically reversible film "breathing" due to redox-driven solvent entry and exit during polymer oxidation and reduction, respectively. Modeling reveals three compositionally distinct regions within the polymer film: interfacial regions at the electrode and solution interfaces and a "bulk" interior. The new methodology, supported by simultaneous in situ visible transmission spectroscopy, reveals an unprecedented level of insight into the temporal and spatial mechanistic details of film solvation changes, including a two-stage (de)solvation mechanism for redox switching, differences in interior (in)homogeneity for reduced and oxidized films, and permselectivity failure under dynamic electrochemical conditions for the reduced (but not oxidized) state, in contrast to static conditions that allow permselectivity for both states.
Related JoVE Video
Fixation and drying protocols for the preparation of cell samples for time-of-flight secondary ion mass spectrometry analysis.
Anal. Chem.
PUBLISHED: 07-31-2009
Show Abstract
Hide Abstract
Time-of-flight secondary ion mass spectrometry (TOF-SIMS) is a promising tool for subcellular chemical analysis of biological cells. However, to obtain relevant information, the method used for sample preparation is critical. In this work, we have used TOF-SIMS, scanning electron microscopy (SEM), and interference reflection microscopy (IRM) to study the effects of different fixation and drying methods on the morphology and chemical structure of human fibroblast cells (hTERT) adhered to a silicon surface. Specifically, two fixation techniques (chemical fixation with glutaraldehyde and cryofixation by plunge freezing) and two drying techniques (freeze drying and alcohol substitution drying) were investigated. Cryofixation followed by freeze drying was determined to produce dried cells with preserved cell morphology, intact cell membranes, and retained sodium/potassium ion concentration gradients across the plasma membrane. By washing samples in an aqueous solution of ammonium formate (AF) before cryofixation, the accumulation of salts on the sample surface during drying could be suppressed. IRM measurements showed that the cell morphology was preserved during washing with ammonium formate, although some swelling occurred. Compared with cryofixation, cells fixed with glutaraldehyde showed finer structures on the cell surface in SEM and similar lipid distributions in TOF-SIMS, but the sodium/potassium ion gradients were not retained. Alcohol drying was determined to remove cell membrane phospholipids significantly, although the use of osmium tetroxide as a post-fixative was shown to decrease this effect.
Related JoVE Video
Proteomic analysis of human osteoprogenitor response to disordered nanotopography.
J R Soc Interface
PUBLISHED: 06-25-2009
Show Abstract
Hide Abstract
Previous studies have shown that microgroove-initiated contact guidance can induce bone formation in osteoprogenitor cells (OPGs) and produce changes in the cell proteome. For proteomic analysis, differential in-gel electrophoresis (DIGE) can be used as a powerful diagnostic method to provide comparable data between the proteomic profiles of cells cultured in different conditions. This study focuses on the response of OPGs to a novel nanoscale pit topography with osteoinductive properties compared with planar controls. Disordered near-square nanopits with 120 nm diameter and 100 nm depth with an average 300 nm centre-to-centre spacing (300 nm spaced pits in square pattern, but with +/-50 nm disorder) were fabricated on 1x1 cm2 polycaprolactone sheets. Human OPGs were seeded onto the test materials. DIGE analysis revealed changes in the expression of a number of distinct proteins, including upregulation of actin isoforms, beta-galectin1, vimentin and procollagen-proline, 2-oxoglutarate 4-dioxygenase and prolyl 4-hydroxylase. Downregulation of enolase, caldesmon, zyxin, GRASP55, Hsp70 (BiP/GRP78), RNH1, cathepsin D and Hsp27 was also observed. The differences in cell morphology and mineralization are also reported using histochemical techniques.
Related JoVE Video
Influence of the plasma sheath on plasma polymer deposition in advance of a mask and down pores.
J Phys Chem B
PUBLISHED: 06-03-2009
Show Abstract
Hide Abstract
Plasma species that form plasma polymer deposits readily penetrate through small openings and are therefore well suited to coat the interior of porous objects. Here, we show how the size of the cross section of square channels influences the penetration of active species from a hexane plasma and how it affects the formation of surface chemical gradients in the interior of these model pores. WCA mapping and ToF-SIMS imaging are used to visualize the plasma polymer deposit in the interior of the model pores and demonstrate that a strong dependence of the wettability gradient profile only exists up to a channel cross section of about 1 mm. XPS data allow us to calculate a deposition rate of plasma polymerized hexane (ppHex) at discrete positions on the surface and show that the deposition rate of ppHex is reduced by the presence of the mask up to a distance of 16 mm in advance of the channel opening. A strong dependence of the ppHex deposition rate on the cross-section of the channels is found within the first 2 mm in front of the pore opening. An estimation of the sheath thickness suggests that this effect can be attributed to the plasma sheath that perturbs the plasma in front of the pores. Plasma mass spectrometry allows us to identify the nature of the plasma species penetrating from the plasma through the pores and shows that no negatively charged ions are able to penetrate through the small channels. Neutral and positively charged species penetrate several millimeters down the channels and both species are therefore likely to contribute to the formation of the deposit on the sample. In addition, the formation of positively charged higher molecular mass hexane fragments is observed in the gas phase, demonstrating the likelihood of neutral-positive reactions in the plasma.
Related JoVE Video
Effect of sessile drop volume on the wetting anisotropy observed on grooved surfaces.
Langmuir
PUBLISHED: 05-14-2009
Show Abstract
Hide Abstract
This study reports experimental measurements of the water contact angle (WCA) measured on surfaces with grooves of different widths using drop volumes ranging from 400 pL to 4.5 microL. These measurements were carried out on both relatively hydrophobic and hydrophilic surface chemistry formed using a conformal plasma polymer coating of topographically embossed poly(methyl methacrylate) (PMMA). Anisotropic wetting of the grooved surfaces was found to be more marked for larger drops on both the hydrophilic and hydrophobic surfaces. Above a certain drop base diameter to groove width ratio, topography had no effect on the measured WCA; this ratio was found to be dependent on the water drop volume. The WCA measured from the direction perpendicular to the grooves using submicroliter water drops is found to be a good indicator of the WCA on the flat surface with equivalent wettabilities. To the best of our knowledge, this is the first study on the phenomenon of anisotropic wetting using picoliter water drops.
Related JoVE Video
The use of nanoscale topography to modulate the dynamics of adhesion formation in primary osteoblasts and ERK/MAPK signalling in STRO-1+ enriched skeletal stem cells.
Biomaterials
PUBLISHED: 04-28-2009
Show Abstract
Hide Abstract
The physiochemical characteristics of a material with in vivo applications are critical for the clinical success of the implant and regulate both cellular adhesion and differentiated cellular function. Topographical modification of an orthopaedic implant may be a viable method to guide tissue integration and has been shown in vitro to dramatically influence osteogenesis, inhibit bone resorption and regulate integrin mediated cell adhesion. Integrins function as force dependant mechanotransducers, acting via the actin cytoskeleton to translate tension applied at the tissue level to changes in cellular function via intricate signalling pathways. In particular the ERK/MAPK signalling cascade is a known regulator of osteospecific differentiation and function. Here we investigate the effects of nanoscale pits and grooves on focal adhesion formation in human osteoblasts (HOBs) and the ERK/MAPK signalling pathway in mesenchymal populations. Nanopit arrays disrupted adhesion formation and cellular spreading in HOBs and impaired osteospecific differentiation in skeletal stem cells. HOBs cultured on 10 microm wide groove/ridge arrays formed significantly less focal adhesions than cells cultured on planar substrates and displayed negligible differentiation along the osteospecific lineage, undergoing up-regulations in the expression of adipospecific genes. Conversely, osteospecific function was correlated to increased integrin mediated adhesion formation and cellular spreading as noted in HOBS cultured on 100 microm wide groove arrays. Here osteospecific differentiation and function was linked to focal adhesion growth and FAK mediated activation of the ERK/MAPK signalling pathway in mesenchymal populations.
Related JoVE Video
Whole proteome analysis of osteoprogenitor differentiation induced by disordered nanotopography and mediated by ERK signalling.
Biomaterials
PUBLISHED: 03-03-2009
Show Abstract
Hide Abstract
Topographic features can modulate cell behaviours such as proliferation, migration, differentiation and apoptosis. Biochemical mechanotransduction implies the conversion of mechanical forces (e.g. changes in cell spreading and morphology from changing surface topography) into biochemical signal via biomolecules. Still, little is known concerning which pathways may be directly involved in cell response to changes in the material surface. A number of pathways have been implicated using focused studies of selected biomolecules rather than a global analysis of signal pathways. This study used a controlled disorder nanopit topography (NSQ50, fabricated by electron beam lithography) to direct osteoblast differentiation of progenitor cells. This topography is unique as it represents a middle route (from absolute order or random roughness) that allows osteoconversion with similar efficiency as dexamethasone and ascorbate treatment. Two direct-comparison proteomics techniques, firstly gel-based and then chromatography-based, were used to analyse progenitor proteome changes in response to the nanotopography. Many of the changed proteins form part of the Extracellular Signal-regulated Kinase (ERK1/2) pathway.
Related JoVE Video
Micropatterned surfaces with covalently grafted unsymmetrical polyoxometalate-hybrid clusters lead to selective cell adhesion.
J. Am. Chem. Soc.
PUBLISHED: 01-14-2009
Show Abstract
Hide Abstract
Mn-Anderson based polyoxometalate clusters with different terminal groups have been patterned successfully onto self-assembled monolayer (SAM) using microcontact printing. Studies of the interactions between the designed SAMs and human fibroblast (hTERT-BJ1) cells have been reported, and it was observed that cells attach and spread efficiently for monolayer presenting a terminal aromatic pyrene platform with a polyoxometalate Mn-Anderson cluster as linker, demonstrating the crucial role played by the polyoxometalate metal oxide cluster as an intermediary in cell adhesion to the surface.
Related JoVE Video
Screening of biomineralization using microfluidics.
Anal. Chem.
PUBLISHED: 01-02-2009
Show Abstract
Hide Abstract
Biomineralization is the process where biological systems produce well-defined composite structures such as shell, teeth, and bones. Currently, there is substantial momentum to investigate the processes implicated in biomineralization and to unravel the complex roles of proteins in the control of polymorph switching. An understanding of these processes may have wide-ranging significance in health care applications and in the development of advanced materials. We have demonstrated a microfluidic approach toward these challenges. A reversibly sealed T-junction microfluidic device was fabricated to investigate the influence of extrapallial (EP) fluid proteins in polymorph control of crystal formation in mollusk shells. A range of conditions were investigated on chip, allowing fast screening of various combinations of ion, pH, and protein concentrations. The dynamic formation of crystals was monitored on chip and combined with in situ Raman to reveal the polymorph in real time. To this end, we have demonstrated the unique advantages of this integrated approach in understanding the processes involved in biomineralization and revealing information that is impossible to obtain using traditional methods.
Related JoVE Video
Using nanotopography and metabolomics to identify biochemical effectors of multipotency.
ACS Nano
Show Abstract
Hide Abstract
It is emerging that mesenchymal stem cell (MSC) metabolic activity may be a key regulator of multipotency. The metabolome represents a "snapshot" of the stem cell phenotype, and therefore metabolic profiling could, through a systems biology approach, offer and highlight critical biochemical pathways for investigation. To date, however, it has remained difficult to undertake unbiased experiments to study MSC multipotency in the absence of strategies to retain multipotency without recourse to soluble factors that can add artifact to experiments. Here we apply a nanotopographical systems approach linked to metabolomics to regulate plasticity and demonstrate rapid metabolite reorganization, allowing rational selection of key biochemical targets of self-renewal (ERK1/2, LDL, and Jnk). We then show that these signaling effectors regulate functional multipotency.
Related JoVE Video
The development of a ?-polycaprolactone scaffold for central nervous system repair.
Tissue Eng Part A
Show Abstract
Hide Abstract
Potential treatment strategies for the repair of spinal cord injury (SCI) currently favor a combinatorial approach incorporating several factors, including exogenous cell transplantation and biocompatible scaffolds. The use of scaffolds for bridging the gap at the injury site is very appealing although there has been little investigation into the central nervous system neural cell interaction and survival on such scaffolds before implantation. Previously, we demonstrated that aligned microgrooves 12.5-25 ?m wide on ?-polycaprolactone (PCL) promoted aligned neurite orientation and supported myelination. In this study, we identify the appropriate substrate and its topographical features required for the design of a three-dimensional scaffold intended for transplantation in SCI. Using an established myelinating culture system of dissociated spinal cord cells, recapitulating many of the features of the intact spinal cord, we demonstrate that astrocytes plated on the topography secrete soluble factors(s) that delay oligodendrocyte differentiation, but do not prevent myelination. However, as myelination does occur after a further 10-12 days in culture, this does not prevent the use of PCL as a scaffold material as part of a combined strategy for the repair of SCI.
Related JoVE Video
A dual gradient assay for the parametric analysis of cell-surface interactions.
Small
Show Abstract
Hide Abstract
Cellular response to microgrooves is addressed using a new assay format, comprising orthogonal gradients of continuously varied groove pitch and depth. Dual layer etch masks are created using a combination of micropatterning and plasma polymer deposition. A silicon substrate with a constant groove width of 8 ?m and with ridge width increasing from 8 ?m in 0.5 ?m steps across 10 mm is fabricated by photolithography. A plasma-polymerized hexane film which is 120 nm thick at one end of these grooves, and 10 nm at the other, is deposited under a diffusion mask. Reactive etching of the patterned sample transfers a gradient of groove pitch and groove depth into the silicon substrate. A silicon master with a gradient of groove depth spanning more than two orders of magnitude (less than 10 nm to over 1000 nm) is used to create an injection molding inlay for mass replication of the screening topography. Polycarbonate replicas are molded for use in cell culture studies, and the functionality of the topography as a high-throughput screening platform is investigated. The response of MDCK, h-TERT fibroblasts, and LE2 endothelial cells is examined, in terms of attachment and morphological response to the variation in topographical cues, with the aim of pinpointing the optimal combination of groove pitch and depth to elicit a tailored response from each cell type. When the range of topographical features screened on a single substrate is considered, this new assay represents a significant step forward in the parametric design and analysis of topographical cues at the biomaterial interface.
Related JoVE Video
Enhanced light extraction in ITO-free OLEDs using double-sided printed electrodes.
Nanoscale
Show Abstract
Hide Abstract
We show how nanoimprint lithographic techniques are particularly suited for the realization of OLED device structures. We tested them to realize nanopatterned metallic electrodes containing photonic crystals to couple the light out and plasmonic crystals showing extraordinary transmission. At similar current densities, a two-fold electroluminescence is achieved with devices having double-sided structured metallic electrodes as compared to a control OLED with an ITO anode. The use of combined nanoimprint lithography processes has the potential to expand the performance range of various organic optoelectronic devices.
Related JoVE Video
Induced chirality through electromagnetic coupling between chiral molecular layers and plasmonic nanostructures.
Nano Lett.
Show Abstract
Hide Abstract
We report a new approach for creating chiral plasmonic nanomaterials. A previously unconsidered, far-field mechanism is utilized which enables chirality to be conveyed from a surrounding chiral molecular material to a plasmonic resonance of an achiral metallic nanostructure. Our observations break a currently held preconception that optical properties of plasmonic particles can most effectively be manipulated by molecular materials through near-field effects. We show that far-field electromagnetic coupling between a localized plasmon of a nonchiral nanostructure and a surrounding chiral molecular layer can induce plasmonic chirality much more effectively (by a factor of 10(3)) than previously reported near-field phenomena. We gain insight into the mechanism by comparing our experimental results to a simple electromagnetic model which incorporates a plasmonic object coupled with a chiral molecular medium. Our work offers a new direction for the creation of hybrid molecular plasmonic nanomaterials that display significant chiroptical properties in the visible spectral region.
Related JoVE Video

What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.