The effect of unlocking RGD-motifs in collagen I on pre-osteoblast adhesion and differentiation

Denaturation of extracellular matrix proteins exposes cryptic binding sites. It is hypothesized that binding of cell adhesion receptors to these cryptic binding sites regulates cellular behaviour during tissue repair and regeneration. To test this hypothesis, we quantify the adhesion of pre-osteoblastic cells to native (Col) and partially-denatured (pdCol) collagen I using single-cell force spectroscopy. During early stages of cell attachment (≤180 s) pre-osteoblasts (MC3T3-E1) adhered significantly stronger to pdCol compared to Col. RGD (Arg-Gly-Asp)-containing peptides suppressed this elevated cell adhesion. We show that the RGD-binding α5β1- and αv-integrins mediated pre-osteoblast adhesion to pdCol, but not to Col. On pdCol pre-osteoblasts had a higher focal adhesion kinase tyrosine-phosphorylation level that correlated with enhanced spreading and motility. Moreover, pre-osteoblasts cultured on pdCol showed a pronounced matrix mineralization activity. Our data suggest that partially-denatured collagen exposes RGD-motifs that trigger binding of α5β1- and αv-integrins. These integrins initiate cellular processes that stimulate osteoblast adhesion, spreading, motility and differentiation. Taken together, these quantitative insights reveal an approach for the development of alternative collagen I- based surfaces for tissue engineering applications.

Thin film deposition and characterization of pure and iron-doped electron-beam evaporated tungsten oxide for gas sensors

Pure Tungsten Oxide (WO3) and Iron-doped (10 at%) Tungsten Oxide (WO3:Fe) nanostructured thin films were prepared using a dual crucible Electron Beam Evaporation techniques. The films were deposited at room temperature in high vacuum condition on glass substrate and post-heat treated at 300 oC for 1 hour. From the study of X-ray diffraction and Raman the characteristics of the as-deposited WO3 and WO3:Fe films indicated non-crystalline nature. The surface roughness of all the films showed in the order of 2.5 nm as observed using Atomic Force Microscopy (AFM). X-Ray Photoelectron Spectroscopy (XPS) analysis revealed tungsten oxide films with stoichiometry close to WO3. The addition of Fe to WO3 produced a smaller particle size and lower porosity as observed using Transmission Electron Microscopy (TEM). A slight difference in optical band gap energies of 3.22 eV and 3.12 eV were found between the as-deposited WO3 and WO3:Fe films, respectively. However, the difference in the band gap energies of the annealed films were significantly higher having values of 3.12 eV and 2.61 eV for the WO3 and WO3:Fe films, respectively. The heat treated samples were investigated for gas sensing applications using noise spectroscopy and doping of Fe to WO3 reduced the sensitivity to certain gasses. Detailed study of the WO3 and WO3:Fe films gas sensing properties is the subject of another paper.

PDMS spreading morphological patterns on substrates of different hydrophilicity in air vacuum and water

In paper has been to investigate the morphological patterns and kinetics of PDMS spreading on silicon wafer using combination of techniques like ellipsometry, atomic force microscope (AFM), scanning electron microscope (SEM) and optical microscopy. A macroscopic silicone oil drops as well as PDMS water based emulsions were studied after deposition on a flat surface of silicon wafer in air, water and vacuum. our own measurements using an imaging ellipsometer, which also clearly shows the presence of a precursor film. The diffusion constant of this film, measured with a 60 000 cS PDMS sample spreading on a hydrophilic silicon wafer, is Df = 1.4  10-11 m2/s. Regardless of their size, density and method of deposition, droplets on both types of wafer (hydrophilic and hydrophobic) flatten out over a period of many hours, up to 3 days. During this process neighbouring droplets may coalesce, but there is strong evidence that some of the PDMS from the droplets migrates into a thin, continuous film that covers the surface in between droplets. The thin film appears to be ubiquitous if there has been any deposition of PDMS. However, this statement needs further verification. One question is whether the film forms immediately after forced drying, or whether in some or all cases it only forms by spreading from isolated droplets as they slowly flatten out.

AFM study of forces between silicon oil and hydrophobic - hydrophylic surfaces in aqueous solutions

An investigation has been made of the interactions between silicone oil and various solid substrates immersed in aqueous solutions. Measurements were made using an atomic force microscope (AFM) using the colloid-probe method. The silicone oil drop is simulated by coating a small silica sphere with the oil, and measuring the force as this coated sphere is brought close to contact with a flat solid surface. It is found that the silicone oil surface is negatively charged, which causes a double-layer repulsion between the oil drop and another negatively charged surface such as mica. With hydrophilic solids, this repulsion is strong enough to prevent attachment of the drop to the solid. However, with hydrophobic surfaces there is an additional attractive force which overcomes the double-layer repulsion, and the silicone oil drop attaches to the solid. A "ramp" force appears in some, but not all, of the data sets. There is circumstantial evidence that this force results from compression of the silicone oil film coated on the glass sphere.

Kaolinite platelet orientation for XRD and AFM applications

The structure-building phenomena within clay aggregates are governed by forces acting between clay particles. The nature of such forces is important to understand in order to manipulate the aggregate structure for applications such as settling and dewatering. A parallel particle orientation is required when conducting force measurements acting between the basal planes of clay mineral platelets using atomic force microscopy (AFM). In order to prepare a film of clay particles with the optimal orientation for conducting AFM measurements, the influences of particle concentration in suspension, suspension pH and particle size on the clay platelet orientation were investigated using scanning electron microscopy (SEM) and X-ray diffraction (XRD) methods. From these investigations, we conclude that high clay (dry mass) concentrations and larger particle diameters (up to 5 µm) in suspension result in random orientation of platelets on the substrate. The best possible laminar orientation in the clay dried film as represented in the XRD by the 001/020 intensity ratio of more than 150 and by SE micrograph assessments, was obtained by drying thin layers from 0.2 wt% of -5 µm clay suspensions at pH 10.5. These dried films are stable and suitable for close-approach AFM studies in solution.

Influence of hydrocarbon contamination on clay soil microstructure

Microstructural (fabric, forces and composition) changes due to hydrocarbon contamination in a clay soil were studied using Scanning Electron Microscope (micro-fabric analysis), Atomic Force Microscope (forces measurement) and sedimentation bench test (particle size measurements). The non-polluted and polluted glacial till from north-eastern Poland (area of a fuel terminal) were used for the study. Electrostatic repelling forces for the polluted sample were much lower than for the non-polluted sample. In comparison to non-polluted sample, the polluted sample exhibited lower electric charge, attractive forces on approach and strong adhesion on retrieve. The results of the sedimentation tests indicate that clay particles form larger aggregates and settle out of the suspension rapidly in diesel oil. In non-polluted soil, the fabric is strongly aggregated – densely packed, dominate the face-to-face and edge-to-edge types of contacts, clay film tightly adheres to the surface of larger grains and interparticle pores are more common. In polluted soil, the clay matrix is less aggregated – loosely packed, dominate the edge-to-face types of contacts and inter-micro-aggregate pores are more frequent. Substantial differences were observed in the morphometric and geometrical parameters of pore space. The polluted soil micro-fabric proved to be more isotropic and less oriented than in non-polluted soil. The polluted soil, in which electrostatic forces were suppressed by hydrocarbon interaction, displays more open porosity and larger voids than non-polluted soil, which is characterized by occurrence of the strong electrostatic interaction between clay particles.

Microscopic and spectroscopic investigation of poly(3-hexylthiophene) interaction with carbon nanotubes

The inclusion of carbon nanotubes in polymer matrix has been proposed to enhance the polymer’s physical and electrical properties. In this study, microscopic and spectroscopic techniques are used to investigate the interaction between poly(3-hexylthiophene) (P3HT) and nanotubes and the reciprocal modification of physical properties. The presence of P3HT-covered nanotubes dispersed in the polymer matrix has been observed by atomic force microscopy and transmission electron microscopy. Then, the modification of P3HT optical properties due to nanotube inclusion has been evidenced with spectroscopic techniques like absorption and Raman spectroscopy. The study is completed with detailed nanoscale analysis by scanning probe techniques. The ordered self assembly of polymer adhering on the nanotube is unveiled by showing an example of helical wrapping of P3HT. Scanning tunneling spectroscopy study provides information on the electronic structure of nanotube-polymer assembly, revealing the charge transfer from P3HT to the nanotube.

Reduction of functionalized graphite oxides by trioctylphosphine in non-polar organic solvents

Graphene, functionalized with oleylamine (OA) and soluble in non-polar organic solvents, was produced on a large scale with a high yield by combining the Hummers process for graphite oxidation, an amine-coupling process to make OA-functionalized graphite oxide (OA-GO), and a novel reduction process using trioctylphosphine (TOP). TOP acts as both a reducing agent and an aggregation-prevention surfactant in the reduction of OA-GO in 1,2-dichlorobenzene (DCB). The reduction of OA-GO is confirmed by X-ray photoelectron spectroscopy, Fourier-transform infrared spectroscopy, X-ray diffraction, thermogravimetric analysis, and Raman spectroscopy. The exfoliation of GO, OA GO, and OA-functionalized graphene (OA-G) is verified by atomic force microscopy. The conductivity of TOP-reduced OA G, which is deduced from the current–voltage characteristics of a vacuum-filtered thin film, shows that the reduction of functionalized GO by TOP is as effective as the reduction of GO by hydrazine.

The effect of various pH, ionic strength and temperature on papain hydrolysis of salivary film

Stimulated human whole saliva (WS) was used to study the dynamics of papain hydrolysis at defined pH, ionic strength and temperature with the view of reducing an acquired pellicle. A quartz crystal microbalance with dissipation (QCM-D) was used to monitor the changes in frequency due to enzyme hydrolysis of WS films and the hydrolytic parameters were calculated using an empirical model. The morphological and conformational changes of the salivary films before and after enzymatic hydrolysis were characterized by atomic force microscopy (AFM) imaging and grazing angle infrared spectroscopy (GA-FTIR) spectra, respectively. The characteristics of papain hydrolysis of WS films were pH-, ionic strength- and temperature-dependent. The WS films were partially removed by the action of enzyme, resulting thinner and smoother surfaces. The IR data suggested that hydrolysis-induced deformation did not occur onto the remnants salivary films. The processes of papain hydrolysis of WS films can be controlled by properly regulating pH, ionic strength and temperature.

Polycaprolactone microspheres as carriers for dry powder inhalers : effect of surface coating on aerosolization of salbutamol sulfate

This study reports the factors controlling aerosolization of salbutamol sulfate (SS) from mixtures with polycaprolactone (PCL) microspheres fabricated using an emulsion technique with polyvinyl alcohol (PVA) as stabilizer. The fine particle fraction (FPF) of SS from PCL measured by a twin-stage impinger was unexpectedly found to be zero, although scanning electron microscopy showed that the drug coated the entire microsphere. Precoating the microspheres with magnesium stearate (MgSt) excipient solutions (1%–2%) significantly increased (p < 0.05, n = 5) the FPF of SS (11.4%–15.4%), whereas precoating with leucine had a similar effect (FPF = 11.3 ± 1.1%), but was independent of the solution concentration. The force of adhesion (by atomic force microscopy) between the PCL microspheres and SS was reduced from 301.4 ± 21.7 nN to 110.9 ± 30.5 nN and 121.8 ± 24.6 nN, (p < 0.05, n = 5) for 1% and 2% MgSt solutions, respectively, and to 148.1 ± 21.0 nN when coated with leucine. The presence of PVA on the PCL microspheres (detected by X-ray photoelectron spectroscopy) affected the detachment of SS due to strong adhesion between the two, presumably due to capillary forces acting between them. Precoating the microspheres with excipients increased the FPF significantly by reducing the drug–carrier adhesion. © 2011 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 101:733–745, 2012

Dynamic covalent chemistry on surfaces employing highly reactive cyclopentadienyl moieties

Silicon substrates coated with a bromide-terminated silane are transformed into highly reactive, cyclopentadiene covered analogues. These surfaces undergo rapid cycloaddition reactions with various dienophile-capped polymers. Mild heating of the substrates causes the retro-Diels-Alder reaction to occur, thus reforming the reactive cyclopentadiene surface, generating an efficiently switchable surface.

Thermally evaporated tungsten oxide (WO3) thin films for gas sensing applications

In this thesis, the author proposed and developed gas sensors made of nanostructured WO3 thin film by a thermal evaporation technique. This technique gives control over film thickness, grain size and purity. The device fabrication, nanostructured material synthesis, characterization and gas sensing performance have been undertaken. Three different types of nanostructured thin films, namely, pure WO3 thin films, iron-doped WO3 thin films by co-evaporation and Fe-implanted WO3 thin films have been synthesized. All the thin films have a film thickness of 300 nm. The physical, chemical and electronic properties of these films have been optimized by annealing heat treatment at 300ºC and 400ºC for 2 hours in air. Various analytical techniques were employed to characterize these films. Atomic Force Microscopy and Transmission Electron Microscopy revealed a very small grain size of the order 5-10 nm in as-deposited WO3 films, and annealing at 300ºC or 400ºC did not result in any significant change in grain size. X-ray diffraction (XRD) analysis revealed a highly amorphous structure of as-deposited films. Annealing at 300ºC for 2 hours in air did not improve crystallinity in these films. However, annealing at 400ºC for 2 hours in air significantly improved the crystallinity in pure and iron-doped WO3 thin films, whereas it only slightly improved the crystallinity of iron-implanted WO3 thin film as a result of implantation. Rutherford backscattered spectroscopy revealed an iron content of 0.5 at.% and 5.5 at.% in iron-doped and iron-implanted WO3 thin films, respectively. The RBS results have been confirmed using energy dispersive x-ray spectroscopy (EDX) during analysis of the films using transmission electron microscopy (TEM). X-ray photoelectron spectroscopy (XPS) revealed significant lowering of W 4f7/2 binding energy in all films annealed at 400ºC as compared with the as-deposited and 300ºC annealed films. Lowering of W 4f7/2 is due to increase in number of oxygen vacancies in the films and is considered highly beneficial for gas sensing. Raman analysis revealed that 400ºC annealed films except the iron-implanted film are highly crystalline with significant number of O-W-O bonds, which was consistent with the XRD results. Additionally, XRD, XPS and Raman analyses showed no evidence of secondary peaks corresponding to compounds of iron due to iron doping or implantation. This provided an understanding that iron was incorporated in the host WO3 matrix rather than as a separate dispersed compound or as catalyst on the surface. WO3 thin film based gas sensors are known to operate efficiently in the temperature range 200ºC-500 ºC. In the present study, by optimizing the physical, chemical and electronic properties through heat treatment and doping, an optimum response to H2, ethanol and CO has been achieved at a low operating temperature of 150ºC. Pure WO3 thin film annealed at 400ºC showed the highest sensitivity towards H2 at 150ºC due to its very small grain size and porosity, coupled with high number of oxygen vacancies, whereas Fe-doped WO3 film annealed at 400ºC showed the highest sensitivity to ethanol at an operating temperature of 150ºC due to its crystallinity, increased number of oxygen vacancies and higher degree of crystal distortions attributed to Fe addition. Pure WO3 films are known to be insensitive to CO, but iron-doped WO3 thin film annealed at 300ºC and 400ºC showed an optimum response to CO at an operating temperature of 150ºC. This result is attributed to lattice distortions produced in WO3 host matrix as a result of iron incorporation as substitutional impurity. However, iron-implanted WO3 thin films did not show any promising response towards the tested gases as the film structure has been damaged due to implantation, and annealing at 300ºC or 400ºC was not sufficient to induce crystallinity in these films. This study has demonstrated enhanced sensing properties of WO3 thin film sensors towards CO at lower operating temperature, which was achieved by optimizing the physical, chemical and electronic properties of the WO3 film through Fe doping and annealing. This study can be further extended to systematically investigate the effects of different Fe concentrations (0.5 at.% to 10 at.%) on the sensing performance of WO3 thin film gas sensors towards CO.

Studies on the surface properties of biodegradable polymer carriers in respiratory delivery of drug from Dry Powder Inhaler formulations

Dry Powder Inhaler (DPI) technology has a significant impact in the treatment of various respiratory disorders. DPI formulations consist of a micronized drug (<5ìm) blended with an inert coarse carrier, for which lactose is widely used to date. DPIs are one of the inhalation devices which are used to target the delivery of drugs to the lungs. Drug delivery via DPI formulations is influenced by the physico-chemical characteristics of lactose particles such as size, shape, surface roughness and adhesional forces. Commercially available DPI formulations, which utilise lactose as the carrier, are not efficient in delivering drug to the lungs. The reasons for this are the surface morphology, adhesional properties and surface roughness of lactose. Despite several attempts to modify lactose, the maximum efficient drug delivery to the lungs remains limited; hence, exploring suitable alternative carriers for DPIs is of paramount importance. Therefore, the objective of the project was to study the performance of spherical polymer microparticles as drug carriers and the factors controlling their performance. This study aimed to use biodegradable polymer microspheres as alternative carriers to lactose in DPIs for achieving efficient drug delivery into the lungs. This project focused on fabricating biodegradable polymer microparticles with reproducible surface morphology and particle shape. The surface characteristics of polymeric carriers and the adhesional forces between the drug and carrier particles were investigated in order to gain a better understanding of their influence on drug dispersion. For this purpose, two biodegradable polymers- polycaprolactone (PCL) and poly (DL-lactide-co-glycolide) (PLGA) were used as the carriers to deliver the anti-asthmatic drug - Salbutamol Sulphate (SS). The first study conducted for this dissertation was the aerosolization of SS from mixtures of SS and PCL or PLGA microparticles. The microparticles were fabricated using an emulsion technique and were characterized by laser diffraction for particle size analysis, Scanning Electron Microscopy (SEM) for surface morphology and X-ray Photoelectron Spectroscopy (XPS) to obtain surface elemental composition. The dispersion of the drug from the DPI formulations was determined by using a Twin Stage Impinger (TSI). The Fine particle Fraction (FPF) of SS from powder mixtures was analyzed by High Performance Liquid Chromatography (HPLC). It was found that the drug did not detach from the surface of PCL microspheres. To overcome this, the microspheres were coated with anti-adherent agents such as magnesium stearate and leucine to improve the dispersion of the drug from the carrier surfaces. It was found that coating the PCL microspheres helped in significantly improving the FPF of SS from the PCL surface. These results were in contrast to the PLGA microspheres which readily allowed detachment of the SS from their surface. However, coating PLGA microspheres with antiadherent agents did not further improve the detachment of the drug from the surface. Thus, the first part of the study demonstrated that the surface-coated PCL microspheres and PLGA microspheres can be potential alternatives to lactose as carriers in DPI formulations; however, there was no significant improvement in the FPF of the drug. The second part of the research studied the influence of the size of the microspheres on the FPF of the drug. For this purpose, four different sizes (25 ìm, 48 ìm, 100 ìm and 150 ìm) of the PCL and PLGA microspheres were fabricated and characterized. The dispersion of the drug from microspheres of different sizes was determined. It was found that as the size of the carrier increased there was a significant increase in the FPF of SS. This study suggested that the size of the carrier plays an important role in the dispersion of the drug from the carrier surface. Subsequent experiments in the third part of the dissertation studied the surface properties of the polymeric carrier. The adhesion forces existing between the drug particle and the polymer surfaces, and the surface roughness of the carriers were quantified using Atomic Force Microscopy (AFM). A direct correlation between adhesion forces and dispersion of the drug from the carrier surface was observed suggesting that adhesion forces play an important role in determining the detachment potential of the drug from the carrier surface. However, no direct relationship between the surface roughness of the PCL or PLGA carrier and the FPF of the drug was observed. In conclusion, the body of work presented in this dissertation demonstrated the potential of coated PCL microspheres and PLGA microspheres to be used in DPI formulations as an alternative carrier to sugar based carriers. The study also emphasized the role of the size of the carrier particles and the forces of interaction prevailing between the drug and the carrier particle surface on the aerosolization performances of the drug.

Characterisation of multi wall carbon nanotube–polymer composites for strain sensing applications

Carbon nanotubes (CNTs) have excellent electrical, mechanical and electromechanical properties. When CNTs are incorporated into polymers, electrically conductive composites with high electrical conductivity at very low CNT content (often below 1% wt CNT) result. Due to the change in electrical properties under mechanical load, carbon nanotube/polymer composites have attracted significant research interest especially due to their potential for application in in-situ monitoring of stress distribution and active control of strain sensing in composite structures or as strain sensors. To sucessfully develop novel devices for such applications, some of the major challenges that need to be overcome include; in-depth understanding of structure-electrical conductivity relationships, response of the composites under changing environmental conditions and piezoresistivity of different types of carbon nanotube/polymer sensing devices. In this thesis, direct current (DC) and alternating current (AC) conductivity of CNT-epoxy composites was investigated. Details of microstructure obtained by scanning electron microscopy were used to link observed electrical properties with structure using equivalent circuit modeling. The role of polymer coatings on macro and micro level electrical conductivity was investigated using atomic force microscopy. Thermal analysis and Raman spectroscopy were used to evaluate the heat flow and deformation of carbon nanotubes embedded in the epoxy, respectively, and related to temperature induced resistivity changes. A comparative assessment of piezoresistivity was conducted using randomly mixed carbon nanotube/epoxy composites, and new concept epoxy- and polyurethane-coated carbon nanotube films. The results indicate that equivalent circuit modelling is a reliable technique for estimating values of the resistance and capacitive components in linear, low aspect ratio-epoxy composites. Using this approach, the dominant role of tunneling resistance in determining the electrical conductivity was confirmed, a result further verified using conductive-atomic force microscopy analysis. Randomly mixed CNT-epoxy composites were found to be highly sensitive to mechanical strain and temperature variation compared to polymer-coated CNT films. In the vicinity of the glass transition temperature, the CNT-epoxy composites exhibited pronounced resistivity peaks. Thermal and Raman spectroscopy analyses indicated that this phenomenon can be attributed to physical aging of the epoxy matrix phase and structural rearrangement of the conductive network induced by matrix expansion. The resistivity of polymercoated CNT composites was mainly dominated by the intrinsic resistivity of CNTs and the CNT junctions, and their linear, weakly temperature sensitive response can be described by a modified Luttinger liquid model. Piezoresistivity of the polymer coated sensors was dominated by break up of the conducting carbon nanotube network and the consequent degradation of nanotube-nanotube contacts while that of the randomly mixed CNT-epoxy composites was determined by tunnelling resistance between neighbouring CNTs. This thesis has demonstrated that it is possible to use microstructure information to develop equivalent circuit models that are capable of representing the electrical conductivity of CNT/epoxy composites accurately. New designs of carbon nanotube based sensing devices, utilising carbon nanotube films as the key functional element, can be used to overcome the high temperature sensitivity of randomly mixed CNT/polymer composites without compromising on desired high strain sensitivity. This concept can be extended to develop large area intelligent CNT based coatings and targeted weak-point specific strain sensors for use in structural health monitoring.

Graphene-like nano-sheets/36° LiTaO3 surface acoustic wave hydrogen gas sensor

Presented is the material and gas sensing properties of graphene-like nano-sheets deposited on 36° YX lithium tantalate (LiTaO3) surface acoustic wave (SAW) transducers. The graphene-like nano-sheets were characterized via scanning electron microscopy (SEM), atomic force microscopy(AFM)and X-ray photoelectron spectroscopy (XPS). The graphenelike nano-sheet/SAW sensors were exposed to different concentrations of hydrogen (H2) gas in a synthetic air at room temperature. The developed sensors exhibit good sensitivity towards low concentrations of H2 in ambient conditions, as well as excellent dynamic performance towards H2 at room temperature.