V. Švorčík

Modulation of cell adhesion, proliferation and differentiation on materials designed for body implants.

The interaction of cells and tissues with artificial materials designed for applications in biotechnologies and in medicine is governed by the physical and chemical properties of the material surface. There is optimal cell adhesion to moderately hydrophilic and positively charged substrates, due to the adsorption of cell adhesion-mediating molecules (e.g. vitronectin, fibronectin) in an advantageous geometrical conformation, which makes specific sites on these molecules (e.g. specific amino acid sequences) accessible to cell adhesion receptors (e.g. integrins). Highly hydrophilic surfaces prevent the adsorption of proteins, or these molecules are bound very weakly. On highly hydrophobic materials, however, proteins are adsorbed in rigid and denatured forms, hampering cell adhesion. The wettability of the material surface, particularly in synthetic polymers, can be effectively regulated by physical treatments, e.g. by irradiation with ions, plasma or UV light. The irradiation-activated material surface can be functionalized by various biomolecules and nanoparticles, and this further enhances its attractiveness for cells and its effectiveness in regulating cell functions. Another important factor for cell-material interaction is surface roughness and surface topography. Nanostructured substrates (i.e. substrates with irregularities smaller than 100nm), are generally considered to be beneficial for cell adhesion and growth, while microstructured substrates behave more controversially (e.g. they can hamper cell spreading and proliferation but they enhance cell differentiation, particularly in osteogenic cells). A factor which has been relatively less investigated, but which is essential for cell-material interaction, is material deformability. Highly soft and deformable substrates cannot resist the tractional forces generated by cells during cell adhesion, and cells are not able to attach, spread and survive on such materials. Local variation in the physical and chemical properties of the material surface can be advantageously used for constructing patterned surfaces. Micropatterned surfaces enable regionally selective cell adhesion and directed growth, which can be utilized in tissue engineering, in constructing microarrays and in biosensorics. Nanopatterned surfaces are an effective tool for manipulating the type, number, spacing and distribution of ligands for cell adhesion receptors on the material surface. As a consequence, these surfaces are able to control the size, shape, distribution and maturity of focal adhesion plaques on cells, and thus cell adhesion, proliferation, differentiation and other cell functions.

Properties of gold nanostructures sputtered on glass

We studied the electrical and optical properties, density, and crystalline structure of Au nanostructures prepared by direct current sputtering on glass. We measured temperature dependence of sheet resistance and current-voltage characteristics and also performed scanning electron microscopy [SEM] analysis of gold nanolayers. It was shown that within the wide range of temperatures, gold nanolayers (<10 nm) exhibit both metal and semiconducting-like type of conductivity. UV/Vis analysis proved the semiconducting characteristic of intrinsic Au clusters. SEM analysis showed the initiatory stadium of gold layer formation to be running over isolated islands. Gold density calculated from the weight and effective thickness of the layers is an increasing function of the layer thickness up to approximately 100 nm. In thin layers deposited on solid surface, a lattice expansion is observed, which is manifested in the increase of the lattice parameter and the decrease of metal density. With increasing layer thickness, the lattice parameter and the density approach the bulk values.

Adhesion and proliferation of human endothelial cells on photochemically modified polytetrafluoroethylene

We studied the adhesion and proliferation of human endothelial cells on photochemically modified polytetrafluoroethylene samples. The polymer surfaces were modified by exposure to the ultraviolet light of a Xe(2)(*)-excimer lamp at a wavelength of 172 nm in an ammonia atmosphere. Treatment times were between 10 and 20 min. The endothelial cell density was determined 1, 3 and 8 days after seeding by image analysis. Surface modification of the samples resulted in a significant increase in the number of adhering cells and in the formation of a confluent cell layer after 3-8 days. The results were comparable than those obtained on polystyrene Petri dishes, which are used as standard substrates in cell cultivation. Thus modified PTFE appears to be a promising material for the fabrication of artificial vascular prostheses coated with endothelial cells.

Fluorine ion-implanted polystyrene improves growth and viability of vascular smooth muscle cells in culture.

Vascular smooth muscle cells derived from the rat aorta were cultured on unmodified or F(+) ion-implanted polystyrene (5 x 10(12) or 5 x 10(14) ions/cm(2), energy 150 keV). In 1-day-old cultures, the cells adhered to the modified polystyrene in higher numbers and over larger contact areas. Increased resistance of the cells to trypsin-mediated detachment from the growth support indicated an improved adhesion of cells to the modified polymer at later culture intervals. The cells cultured on ion-modified polymers also were larger and had a higher total protein content. By use of immunocytochemistry, several specific protein species were increased, including the cytoskeletal alpha-actin and vimentin and the plasma membrane-associated vinculin, talin, alpha-v integrins, ICAM-1, and VCAM-1, which account for stronger cell-cell and cell-extracellular matrix adhesion. The lower number of cells found floating in the medium suggests that the spontaneous detachment of cells from the modified polystyrene was lower and that the viability of the adhered cell population was higher. As was shown by the two-parameter flow-cytometric measurements of BrdU incorporation and DNA content, as well as by (3)H-thymidine autoradiography, the cell proliferation on samples modified by the dose of 5 x 10(12) ions/cm(2) was similar to that in controls; and at the dose of 5 x 10(14) ions/cm(2), it tended to be even lower. The cells grown on the polymer implanted with the dose of 5 x 10(12) ions/cm(2) responded to a new artificially created cell-free area in a confluent cell layer by more intense migration whereas at the dose of 5 x 10(14) ions/cm(2), the migration ability of cells was similar to that on the unmodified polymer. The data revealed a higher biocompatibility of ion-implanted polystyrene with vascular smooth muscle cells in culture. There was better adhesion, differentiation, and survival, and there was neither excessive migration nor proliferation.

Molecular mechanisms of improved adhesion and growth of an endothelial cell line cultured on polystyrene implanted with fluorine ions

Endothelial cells derived from the bovine pulmonary artery (line CPAE, CCL 209, American Tissue Culture Collection, Rockville, MD, USA) were cultured on pristine or fluorine ion-irradiated polystyrene (5 x 10(12) or 5 x 10(14) F ions/cm2, 150 keV). At 24-h post-seeding interval, the number of cells which adhered to the ion-modified polystyrene was significantly higher than on the unmodified material (+20 and +58% in cultures with the polystyrene irradiated by lower and higher ion doses, respectively). On day 7, the populations cultured on the irradiated substrates grew to higher densities, exceeding the controls at the lower and higher ion doses by 69 and 180%, respectively. The cells on ion-implanted samples were also larger (+70-95% and +90-99% at the lower and higher ion doses, respectively) and contained more protein (+16% at both ion doses). As was shown by ELISA, the polystyrene irradiated by the higher ion dose enhanced the expression of a cytoskeletal protein, vimentin (+65%) and protein of focal adhesion plaques, talin (+15%). The content of integrin alpha5beta1 (VLA-5), receptor for fibronectin, was increased at both lower and higher ion doses (+22 and +57%). In contrast to this, the content of ICAM-1 and vinculin was similar in cells grown on both pristine and ion-irradiated growth substrates. Moreover, the expression of VCAM-1 and ELAM-1 was lower by 11-14% in both ion dose groups. The present study has shown that ion implantation of polymers improves the adhesion and growth of endothelial cells without elevating the expression of immunoglobulin and selectin types of adhesion molecules. This surface modification should promote colonization of an artificial vascular prosthesis by endothelial cells and make it less vulnerable by immune system cells of the recipient.

Effect of plasma treatment on cellulose fiber

Cotton cellulose fibers were modified in inert plasma. Surface morphology of the modified fibers was studied by SEM and changes in the surface composition by XPS and FTIR. Standard goniometry was used for determination of contact angle as a function of modified fiber aging. Absorptivity of modified fibers was determined by gravimetry and fiber width in physiological solution, simulating body liquids, by confocal microscopy. Antibacterial effect of pristine and plasma treated samples was examined by following growth of Escherichia coli. Plasma treatment led to surface ablation, changes in surface morphology and fiber width. Surface of the plasma modified fibers was oxidized and their water absorptivity was reduced. The plasma modification did not affect E. coli growth substantially.

AFM surface morphology investigation of ion beam modified polyimide

Abstract Polyimide Upilex R was irradiated with 90 keV N + ions to the fluences of 1 × 10 14 –2 × 10 17 cm −2 . The surface morphology and the structure of the ion beam modified PI were examined using atomic force microscopy and X-ray difraction. Sheet resistance as a function of the ion fluence and the sample temperature was measured by standard two point technique. Significant changes of the surface morphology and production of graphitic phase in the sample surface layer modified by the ion irradiation were observed. Strong decrease of the sheet resistance (by 11 orders of magnitude) in the ion beam modified samples is connected with progressive carbonization and graphitization of the degraded polymer. Electrical charge transport is mediated by variable-range hopping mechanism. Drastic structural changes initiated by the ion irradiation to high fluences are similar to those observed in polymer pyrolysis.

Noble metal nanostructures influence of structure and environment on their optical properties

Optical properties of nanostructuredmaterials, isolated nanoparticles, and structures composed of both metals and semiconductors are broadly discussed. Fundamentals of the origin of surface plasmons as well as the surface plasmon resonance sensing are described and documented on a number of examples. Localized plasmon sensing and surface-enhanced Raman spectroscopy are subjected to special interest since those techniques are inherently associated with the direct application of plasmonic structures. The possibility of tailoring the optical properties of ultra-thin metal layers via controlling their shape and morphology by postdeposition annealing is documented. Special attention is paid to the contribution of bimetallic particles and layers as well as metal structures encapsulated in semiconductors and dielectrics to the optical response. The opportunity to tune the properties of materials over a large scale of values opens up entirely new application possibilities of optical active structures. The nature of surface plasmons predetermines noble metal nanostructures to be promising great materials for development of modern label-free sensing methods based on plasmon resonance--SPR and LSPR sensing.

Ablation and water etching of plasma-treated polymers

Ablation and water etching of low-density polyethylene (LDPE), polytetrafluorethylene, polystyrene, and polyethyleneterephtalate (PET) modified by Ar plasma was studied. The amount of ablated and etched material was determined by gravimetry. The changes in polymer surface morphology were observed using atomic force microscopy, UV-Vis and FTIR spectroscopy. This was used for the determination of the chemical structure of modified polymers. Wettability of polymers modified in plasma and water etched was determined from contact angle measurement. It was found that under the plasma discharge the polymers are ablated and their surface morphology and roughness are changed dramatically. The plasma treatment leads to an increase in the concentration of conjugated double bonds. Dissolution of a thin surface layer of plasma-modified PET and LDPE during water etching was observed and preferential liberation of low-molecular weight segments was proved.

Silver release and antimicrobial properties of PMMA films doped with silver ions, nano-particles and complexes

Materials prepared on the base of bioactive silver compounds have become more and more popular due to low microbial resistance to silver. In the present work, the efficiency of polymethylmethacrylate (PMMA) thin films doped with silver ions, nanoparticles and silver-imidazole polymer complex was studied by a combination of AAS, XPS and AFM techniques. The biological activities of the proposed materials were discussed in view of the rate of silver releasing from the polymer matrix. Concentrations of Ag active form were estimated by its ability to interact with l-cysteine using electronic circular dichroism spectroscopy. Rates of the released silver were compared with the biological activity in dependence on the form of embedded silver. Antimicrobial properties of doped polymer films were studied using two bacterial strains: Staphylococcus epidermidis and Escherichia coli. It was found that PMMA films doped with Ag(+) had greater activity than those doped with nanoparticles and silver-imidazole polymeric complexes. However, the antimicrobial efficiency of Ag(+) doped films was only short-term. Contrary, the antimicrobial activity of silver-imidazole/PMMA films increased in time of sample soaking.