A qualitative method for testing the antimicrobial ability of osteosynthetic fixation material by simulating in vitro contamination by Staphylococcus aureus.

External fixators of serious fractures could be an attractive substrate on which microorganisms can accumulate. Therefore, this study aimed to develop a suitable method for enabling the simulation of a real situation when osteosynthetic fixation material is open for the potential threat of bacterial attack. Agar-based media represented human tissue, and the metallic pin characterized the screw in the fixation. Various types of agar, supplements, and contamination strategy by Staphylococcus aureus were tested. The influence of the initial bacterial concentration was also examined. Surfaces were observed by scanning electron microscopy (SEM), and all results were compared. Brain Heart Infusion Agar with the Egg Yolk Tellurite Emulsion was established in a transparent test tube as a suitable system for enabling the good interpretability of bacterial contamination in the pin's surroundings. Pin contamination has been found to be an appropriate approach for testing microbial growth, rather than agar surface contamination, which distorted obtained results. A lower initial colony forming units (CFU) provided better clarity of the test. SEM observation of the pin surface was comparable with the visual evaluations in the test tubes. Results were assembled for positive and negative control samples as well. Screening method for the most common bacteria S. aureus has been standardized and developed. This experimental setup could also be a useful tool for surface modification with antibacterial properties testing.

Mesenchymal stem cell interaction with Ti6Al4V alloy pre-exposed to simulated body fluid.

Titanium and its alloys are widely used for substitution of hard tissues, especially in orthopaedic and dental surgery. Despite the benefit of the use of titanium for such applications, there are still questions which must be sorted out. Surface properties are crucial for cell adhesion, proliferation and differentiation. Mainly, micro/nanostructured surfaces positively influence osteogenic differentiation of human mesenchymal stem cells. Ti6Al4V is a biocompatible α + ß alloy which is widely used in orthopaedics. The aim of this study was to investigate the interaction of the nanostructured and ground Ti6Al4V titanium alloys with simulated body fluid complemented by the defined precipitation of hydroxyapatite-like coating and to study the cytotoxicity and differentiation capacity of cells with such a modified titanium alloy. Nanostructures were fabricated using electrochemical oxidation. Human mesenchymal stem cells (hMSC) were used to evaluate cell adhesion, metabolic activity and proliferation on the specimens. The differentiation potential of the samples was investigated using PCR and specific staining of osteogenic markers collagen type I and osteocalcin. Our results demonstrate that both pure Ti6Al4V, nanostructured samples, and hydroxyapatite-like coating supported hMSC growth and metabolic activity. Nanostructured samples improved collagen type I synthesis after 14 days, while both nanostructured and hydroxyapatite-like coated samples enhanced collagen synthesis on day 21. Osteocalcin synthesis was the most enhanced by hydroxyapatite-like coating on the nanostructured surfaces. Our results indicate that hydroxyapatite-like coating is a useful tool guiding hMSC osteogenic differentiation.

In vitro evaluation of a novel nanostructured Ti-36Nb-6Ta alloy for orthopedic applications.

Aim: To evaluate the impact of a nanostructured surface created on ß-titanium alloy, Ti-36Nb-6Ta, on the growth and differentiation of human mesenchymal stem cells. Materials & methods: The nanotubes, with average diameters 18, 36 and 46 nm, were prepared by anodic oxidation. Morphology, hydrophilicity and mechanical properties of the nanotube layers were characterized. The biocompatibility and osteogenic potential of the nanostructured surfaces were established using various in vitro assays, scanning electron microscopy and confocal microscopy. Results: The nanotubes lowered elastic modulus close to that of bone, positively influenced cell adhesion, improved ALP activity, synthesis of type I collagen and osteocalcin expression, but diminished early cell proliferation. Conclusion: Nanostructured Ti-36Nb-6Ta with nanotube diameters 36 nm was the most promising material for bone implantation.

A two-phase gradual silver release mechanism from a nanostructured TiAlV surface as a possible antibacterial modification in implants.

Titanium biomaterials are widely used in the medical field due to their biocompatibility and excellent corrosion and mechanical resistance. However, these materials have no antibacterial properties. To obtain an antibacterial active surface, a nanostructure of Ti6Al4V alloy was created. This specific nanostructure contained nanotubes and micro-cavities and was used as a substrate for silver anchoring. The electrochemical approach to silver reduction was studied. It is a common approach for silver deposition and in this work, inhomogeneities in the nanostructure were used as a preferential area for silver localisation. The galvanostatic regimen of deposition allowed for a technically quantitative process and the required silver placement. The experimental conditions used enabled testing and silver dissolution rate evaluation within a reasonable time span. Based on the corrosion and analytical results (EDS, XPS and ICP-MS), a two-phase silver release mechanism was confirmed. The openings of the individual nanotubes were filled with silver nanoparticles, whose release was relatively fast. By contrast, the silver anchored inside the cavities allowed the silver to release gradually. Antibacterial efficiency against Staphylococcus aureus and Escherichia coli was successfully demonstrated. Cytotoxicity testing with murine fibroblasts showed cell metabolic activity far above the normative limit of 70%.