Marketa Bacakova

Surface treatment by electric discharge machining of Ti-6Al-4V alloy for potential application in orthopaedics.

This study investigated the properties of Ti-6Al-4V alloy after surface treatment by the electric discharge machining (EDM) process. The EDM process with high peak currents proved to induce surface macro-roughness and to cause chemical changes to the surface. Evaluations were made of the mechanical properties by means of tensile tests, and of surface roughness for different peak currents of the EDM process. The EDM process with peak current of 29 A was found to induce sufficient surface roughness, and to have a low adverse effect on tensile properties. The chemical changes were studied by scanning electron microscopy equipped with an energy dispersive X-ray analyser (EDX). The surface of the benchmark samples was obtained by plasma-spraying a titanium dioxide coating. An investigation of the biocompatibility of the surface-treated Ti-6Al-4V samples in cultures of human osteoblast-like MG 63 cells revealed that the samples modified by EDM provided better substrates for the adhesion, growth and viability of MG 63 cells than the TiO2 coated surface. Thus, EDM treatment can be considered as a promising surface modification to orthopaedic implants, in which good integration with the surrounding bone tissue is required.

Adhesion and Growth of Vascular Smooth Muscle Cells on Nanostructured and Biofunctionalized Polyethylene

Cell colonization of synthetic polymers can be regulated by physical and chemical modifications of the polymer surface. High-density and low-density polyethylene (HDPE and LDPE) were therefore activated with Ar+ plasma and grafted with fibronectin (Fn) or bovine serum albumin (BSA). The water drop contact angle usually decreased on the plasma-treated samples, due to the formation of oxidized groups, and this decrease was inversely related to the plasma exposure time (50–300 s). The presence of nitrogen and sulfur on the polymer surface, revealed by X-ray photoelectron spectroscopy (XPS), and also by immunofluorescence staining, showed that Fn and BSA were bound to this surface, particularly to HDPE. Plasma modification and grafting with Fn and BSA increased the nanoscale surface roughness of the polymer. This was mainly manifested on HDPE. Plasma treatment and grafting with Fn or BSA improved the adhesion and growth of vascular smooth muscle cells in a serum-supplemented medium. The final cell population densities on day 6 after seeding were on an average higher on LDPE than on HDPE. In a serum-free medium, BSA grafted to the polymer surface hampered cell adhesion. Thus, the cell behavior on polyethylene can be modulated by its type, intensity of plasma modification, grafting with biomolecules, and composition of the culture medium.

The potential applications of fibrin-coated electrospun polylactide nanofibers in skin tissue engineering

Fibrin plays an important role during wound healing and skin regeneration. It is often applied in clinical practice for treatment of skin injuries or as a component of skin substitutes. We prepared electrospun nanofibrous membranes made from poly(l-lactide) modified with a thin fibrin nanocoating. Fibrin surrounded the individual fibers in the membrane and also formed a thin fibrous mesh on several places on the membrane surface. The cell-free fibrin nanocoating remained stable in the cell culture medium for 14 days and did not change its morphology. On membranes populated with human dermal fibroblasts, the rate of fibrin degradation correlated with the degree of cell proliferation. The cell spreading, mitochondrial activity, and cell population density were significantly higher on membranes coated with fibrin than on nonmodified membranes, and this cell performance was further improved by the addition of ascorbic acid in the cell culture medium. Similarly, fibrin stimulated the expression and synthesis of collagen I in human dermal fibroblasts, and this effect was further enhanced by ascorbic acid. The expression of beta1-integrins was also improved by fibrin, and on pure polylactide membranes, it was slightly enhanced by ascorbic acid. In addition, ascorbic acid promoted deposition of collagen I in the form of a fibrous extracellular matrix. Thus, the combination of nanofibrous membranes with a fibrin nanocoating and ascorbic acid seems to be particularly advantageous for skin tissue engineering.

Adhesion, Growth, and Maturation of Vascular Smooth Muscle Cells on Low-Density Polyethylene Grafted with Bioactive Substances

The attractiveness of synthetic polymers for cell colonization can be affected by physical, chemical, and biological modification of the polymer surface. In this study, low-density polyethylene (LDPE) was treated by an Ar+ plasma discharge and then grafted with biologically active substances, namely, glycine (Gly), polyethylene glycol (PEG), bovine serum albumin (BSA), colloidal carbon particles (C), or BSA+C. All modifications increased the oxygen content, the wettability, and the surface free energy of the materials compared to the pristine LDPE, but these changes were most pronounced in LDPE with Gly or PEG, where all the three values were higher than in the only plasma-treated samples. When seeded with vascular smooth muscle cells (VSMCs), the Gly- or PEG-grafted samples increased mainly the spreading and concentration of focal adhesion proteins talin and vinculin in these cells. LDPE grafted with BSA or BSA+C showed a similar oxygen content and similar wettability, as the samples only treated with plasma, but the nano- and submicron-scale irregularities on their surface were more pronounced and of a different shape. These samples promoted predominantly the growth, the formation of a confluent layer, and phenotypic maturation of VSMC, demonstrated by higher concentrations of contractile proteins alpha-actin and SM1 and SM2 myosins. Thus, the behavior of VSMC on LDPE can be regulated by the type of bioactive substances that are grafted.

Protein nanocoatings on synthetic polymeric nanofibrous membranes designed as carriers for skin cells

Protein-coated resorbable synthetic polymeric nanofibrous membranes are promising for the fabrication of advanced skin substitutes. We fabricated electrospun polylactic acid and poly(lactide-co-glycolic acid) nanofibrous membranes and coated them with fibrin or collagen I. Fibronectin was attached to a fibrin or collagen nanocoating, in order further to enhance the cell adhesion and spreading. Fibrin regularly formed a coating around individual nanofibers in the membranes, and also formed a thin noncontinuous nanofibrous mesh on top of the membranes. Collagen also coated most of the fibers of the membrane and randomly created a soft gel on the membrane surface. Fibronectin predominantly adsorbed onto a thin fibrin mesh or a collagen gel, and formed a thin nanofibrous structure. Fibrin nanocoating greatly improved the attachment, spreading, and proliferation of human dermal fibroblasts, whereas collagen nanocoating had a positive influence on the behavior of human HaCaT keratinocytes. In addition, fibrin stimulated the fibroblasts to synthesize fibronectin and to deposit it as an extracellular matrix. Fibrin coating also showed a tendency to improve the ultimate tensile strength of the nanofibrous membranes. Fibronectin attached to fibrin or to a collagen coating further enhanced the adhesion, spreading, and proliferation of both cell types.

Effects of fiber density and plasma modification of nanofibrous membranes on the adhesion and growth of HaCaT keratinocytes.

It may be possible to regulate the cell colonization of biodegradable polymer nanofibrous membranes by plasma treatment and by the density of the fibers. To test this hypothesis, nanofibrous membranes of different fiber densities were treated by oxygen plasma with a range of plasma power and exposure times. Scanning electron microscopy and mechanical tests showed significant modification of nanofibers after plasma treatment. The intensity of the fiber modification increased with plasma power and exposure time. The exposure time seemed to have a stronger effect on modifying the fiber. The mechanical behavior of the membranes was influenced by the plasma treatment, the fiber density, and their dry or wet state. Plasma treatment increased the membrane stiffness; however, the membranes became more brittle. Wet membranes displayed significantly lower stiffness than dry membranes. X-ray photoelectron spectroscopy (XPS) analysis showed a slight increase in oxygen-containing groups on the membrane surface after plasma treatment. Plasma treatment enhanced the adhesion and growth of HaCaT keratinocytes on nanofibrous membranes. The cells adhered and grew preferentially on membranes of lower fiber densities, probably due to the larger area of void spaces between the fibers.

Nanofibrous Scaffolds as Promising Cell Carriers for Tissue Engineering

Nanofibers are promising cell carriers for tissue engineering of a variety of tissues and organs in the human organism. They have been experimentally used for reconstruction of tissues of cardiovascular, respiratory, digestive, urinary, nervous and musculoskele‐ tal systems. Nanofibers are also promising for drug and gene delivery, construction of biosensors and biostimulators, and wound dressings. Nanofibers can be created from a wide range of natural polymers or synthetic biostable and biodegradable polymers. For hard tissue engineering, polymeric nanofibers can be reinforced with various ceramic, metal-based or carbon-based nanoparticles, or created directly from hard materials. The nanofibrous scaffolds can be loaded with various bioactive molecules, such as growth, differentiation and angiogenic factors, or funcionalized with ligands for the cell adhesion receptors. This review also includes our experience in skin tissue engineering using nanofibers fabricated from polycaprolactone and its copolymer with polylactide, cellulose acetate, and particularly from polylactide nanofibers modified by plasma activation and fibrin coating. In addition, we studied the interaction of human bone-derived cells with nanofibrous scaffolds loaded with hydroxyapatite or diamond nanoparticles. We also created novel nanofibers based on diamond deposition on a SiO2 template, and tested their effects on the adhesion, viability and growth of human vascular endothelial cells.

A Virtual Reality Visualization Tool for Three-Dimensional Biomedical Nanostructures

Nanoscience is the source of many advanced techniques and applications, the areas of life sciences and materials sciences included. In the field of materials, the latest technological developments have enabled the emergence of innovative materials. There are numerous of nanomaterials in the field of healthcare (nano-, bio- electronic, tools for medical diagnostics, theragnostics, etc.). It also can be noted that the involvement of nanotechnology to problems of medicine and health care is causing a revolution in the biomedical field, including nanomedicine with the emergence of new disciplines such as nanobiomechanics and nano-biology (mechanics of cells, cancer, microbiology, virology, etc.). The paper describes our new approach to creating 3D nanostructures on the SEM frame base. The impact of our research is demonstrated on the 3D digital model of nanostructures which includes 3D coordinates of nanostructures and is ready for the next research simulation process and also for virtual reality (VR) of the research results.

Morphology of a fibrin nanocoating influences dermal fibroblast behavior

Background Our study focuses on the fabrication of appropriate scaffolds for skin wound healing. This research brings valuable insights into the molecular mechanisms of adhesion, proliferation, and control of cell behavior through the extracellular matrix represented by synthetic biodegradable nanofibrous membranes coated by biomolecules. Methods Nanofibrous polylactic acid (PLA) membranes were prepared by a needle-less electrospinning technology. These membranes were coated with fibrin according to two preparation protocols, and additionally they were coated with fibronectin in order to increase the cell affinity for colonizing the PLA membranes. The adhesion, growth, and extracellular matrix protein production of neonatal human dermal fibroblasts were evaluated on the nanofibrous membranes. Results Our results showed that fibrin-coated membranes improved the adhesion and proliferation of human dermal fibroblasts. The morphology of the fibrin nanocoating seems to be crucial for the adhesion of fibroblasts, and consequently for their phenotypic maturation. Fibrin either covered the individual fibers in the membrane (F1 nanocoating), or covered the individual fibers and also formed a fine homogeneous nanofibrous mesh on the surface of the membrane (F2 nanocoating), depending on the mode of fibrin preparation. The fibroblasts on the membranes with the F1 nanocoating remained in their typical spindle-like shape. However, the cells on the F2 nanocoating were spread mostly in a polygon-like shape, and their proliferation was significantly higher. Fibronectin formed an additional mesh attached to the surface of the fibrin mesh, and further enhanced the cell adhesion and growth. The relative gene expression and protein production of collagen I and fibronectin were higher on the F2 nanocoating than on the F1 nanocoating. Conclusion A PLA membrane coated with a homogeneous fibrin mesh seems to be promising for the construction of temporary full-thickness skin tissue substitutes.