L.E. Fratila-Apachitei

In vitro antibacterial activity of porous TiO2-Ag composite layers against methicillin-resistant Staphylococcus aureus.

The aim of this study was the synthesis of a porous TiO(2)-Ag composite coating and assessment of its in vitro bactericidal activity against methicillin-resistant Staphylococcus aureus. The coating was produced by plasma electrolytic oxidation of Ti-6Al-7Nb medical alloy in a calcium acetate/calcium glycerophosphate electrolyte bearing Ag nanoparticles. Following oxidation, the surface of the titanium substrate was converted into the corresponding oxide (TiO(2)) bearing Ca and P species from the electrolyte. In addition, Ag was detected associated with particles present in the oxide layers. The coatings revealed a porous interconnected structure with pores up to 3 microm in size, a threefold increase in roughness and improved wettability relative to the non-oxidized specimens. The composite TiO(2)-Ag coating showed complete killing of methicillin-resistant S. aureus within 24h in all culture conditions, whereas a 1000-fold increase in bacterial numbers was recorded with the ground titanium specimens and the samples oxidized in the absence of Ag nanoparticles.

An electron microscopical study on the growth of TiO2–Ag antibacterial coatings on Ti6Al7Nb biomedical alloy

This research was aimed at investigating the growth mechanism of TiO(2)-Ag antibacterial coatings during plasma electrolytic oxidation (PEO) of Ti6Al7Nb biomedical alloy in an electrolyte based on calcium acetate/calcium glycerophosphate bearing Ag nanoparticles. The focus was on the mechanism of incorporation of Ag nanoparticles, their distribution and chemical composition within the porous coatings using high resolution transmission electron microscopy (HRTEM) and scanning electron microscopy (SEM) imaging techniques combined with energy dispersive X-ray spectroscopy (EDX) for chemical analyses. The PEO coatings were grown using different oxidation times, 10, 30, 60, 90, 120, 180, 240 and 300 s. The electron microscopy results confirmed the formation of a porous coating with incorporated Ag nanoparticles from the initial stages of oxidation (i.e. 10 s), with further Ag incorporation as the PEO process was continued for longer durations. The Ag nanoparticles were embedded in the dense oxide layer, fused into the pore walls and on the surface of the coatings without any change in their morphology or chemistry as detected by HRTEM, SEM and EDX. Ag seems to be delivered to the sites of coating growth (where dielectric breakdown occurs) through different transport pathways, i.e. open pores, cracks and short-circuit channels.

In vitro cytotoxicity evaluation of porous TiO2–Ag antibacterial coatings for human fetal osteoblasts

Implant-associated infections (IAIs) may be prevented by providing antibacterial properties to the implant surface prior to implantation. Using a plasma electrolytic oxidation (PEO) technique, we produced porous TiO₂ coatings bearing various concentrations of Ag nanoparticles (Ag NPs) (designated as 0 Ag, 0.3 Ag and 3.0 Ag) on a Ti-6Al-7Nb biomedical alloy. This study investigates the cytotoxicity of these coatings using a human osteoblastic cell line (SV-HFO) and evaluates their bactericidal activity against methicillin-resistant Staphylococcus aureus (MRSA). The release of Ag and the total amount of Ag in the coatings were determined using a graphite furnace atomic absorption spectrometry technique (GF-AAS) and flame-AAS, respectively. Cytotoxicity was evaluated using the AlamarBlue assay coupled with the scanning electron microscopy (SEM) observation of seeded cells and by fluorescence microscopy examination of the actin cytoskeleton and nuclei after 48 h of incubation. Antibacterial activity was assessed quantitatively using a direct contact assay. AlamarBlue viability assay, SEM and fluorescence microscopy observation of the SV-HFO cells showed no toxicity for 0 Ag and 0.3 Ag specimens, after 2, 5 and 7 days of culture, while 3.0 Ag surfaces appeared to be extremely cytotoxic. All Ag-bearing surfaces had good antibacterial activity, whereas Ag-free coatings showed an increase in bacterial numbers. Our results show that the 0.3 Ag coatings offer conditions for optimum cell growth next to antibacterial properties, which makes them extremely useful for the development of new antibacterial dental and orthopedic implants.

Nanopattern-induced osteogenic differentiation of stem cells - A systematic review.

It is well known that biomaterials topography can influence the behavior of stem cells. Nevertheless, the fundamentals and the impact of nanoscale topography are just emerging. The main objective of this review has been to reveal the state-of-the-art on the effects of controlled nanoscale topographies (nanopatterns) on in vitro osteogenic differentiation of mesenchymal stem cells (MSCs) in the absence of osteogenic supplements. The findings indicate that nanopatterns with specific feature sizes, spatial arrangements, or shapes may induce osteogenic differentiation of MSCs. Regardless of substrate chemistry, nanopattern-induced osteogenic differentiation is associated with large focal adhesions, enhanced cell areas, and well organized cytoskeleton. These results suggest that earlier interactions between nanopattern features and cell receptors are involved, with effects on the entire cell structure and subsequent differentiation. Such events are possibly mediated by nanotopography-induced mechanotransduction pathways. The findings so far reveal that nanoscale topography has potential for directing differentiation of MSCs towards the osteogenic lineage in non-osteogenic media and should be harnessed for possible synergistic effects in bone regenerative therapies. STATEMENT OF SIGNIFICANCE The use of nanotopography to induce cellular responses represents a novel and rapidly growing area of research. Nevertheless, the findings and trends so far are difficult to identify and discuss mostly due to a non-systematic research approach. The present manuscript is providing a systematic review focused on nanopattern-induced osteogenic differentiation of mesenchymal stem cells. The coverage of the most relevant aspects including nanopatterns fabrication methods, their effects on osteogenic differentiation of mesenchymal stem cells as well as the related effects on adhesion and cell morphology has enabled an integrated discussion including the potential mechanotransduction mechanisms involved. Furthermore, a clear distinction between the studies that use only surface nanotopographies and the ones that mix nanotopographical features with osteogenic supplements has been made. This delineation is essential for revealing and understanding the role of biomaterials nanotopography per se on stem cells differentiation based on which novel osteoinductive biomaterials can be developed.

Biofunctional surfaces by plasma electrolytic oxidation on titanium biomedical alloys

The effects of substrate composition on oxide layer properties following plasma electrolytic oxidation under similar conditions have been evaluated for α-cpTi, α/β-Ti6Al7Nb, β-Ti35Zr10Nb and β-Ti45Nb alloys. All oxidised surfaces revealed enhanced wettability, surface free energy and roughness relative to the non-oxidised surfaces. Nevertheless, the resultant oxides differed with respect to average pore size, pores density, layer chemistry and phase composition. The β-titanium alloys developed oxides with a larger average pore size and lower pore density relative to the α-cpTi and α/β-Ti6Al7Nb substrates. Anatase dominated the oxide layer formed on α-cpTi and β-Ti45Nb alloys, a mixture of anatase and rutile was present on the oxidised α/β-Ti6Al7Nb surface, whereas Ti2ZrO6 was the only phase detected on the oxidised surface of the β-Ti35Zr10Nb alloy.

Synthesis and characterization of hybrid micro/nano-structured NiTi surfaces by a combination of etching and anodizing

The purpose of this study was to generate hybrid micro/nano-structures on biomedical nickel-titanium alloy (NiTi). To achieve this, NiTi surfaces were firstly electrochemically etched and then anodized in fluoride-containing electrolyte. With the etching process, the NiTi surface was micro-roughened through the formation of micropits uniformly distributed over the entire surface. Following the subsequent anodizing process, self-organized nanotube structures enriched in TiO2 could be superimposed on the etched surface under specific conditions. Furthermore, the anodizing treatment significantly reduced water contact angles and increased the surface free energy compared to the surfaces prior to anodizing. The results of this study show for the first time that it is possible to create hybrid micro/nano-structures on biomedical NiTi alloys by combining electrochemical etching and anodizing under controlled conditions. These novel structures are expected to significantly enhance the surface biofunctionality of the material when compared to conventional implant devices with either micro- or nano-structured surfaces.

Drug Release from PLGA Microspheres Attached to Solids Using Supercritical CO2

Functionalization of a porous orthopedic implant with dexamethasone, a widely used anti-inflammatory drug, encapsulated within a biodegradable polymer for controlled release could help reduce or eliminate the inflammation response by the local tissue. In this research, we investigated the possibility of using supercritical carbon dioxide (CO2) for attaching dexamethasone-loaded PLGA (polylactic-co-glycolic acid) microspheres to porous CoCrMo alloy for continuous delivery of dexamethasone. Supercritical CO2 has been shown to be effective for attachment of PLGA microspheres to glass plates and porous CoCrMo alloy. Attached microspheres showed similar dexamethasone release profiles but different magnitude of burst release. Microspheres attached to the porous alloy samples using supercritical CO2 at 10 bar and 40°C for 30 min showed a release profile similar to that of the nonattached microspheres. The microsphere morphology and the release profiles of microspheres attached to the glass plates and to the porous alloy samples suggest that dexamethasone burst release is enhanced by PLGA swelling at higher CO 2 pressures and better dispersion of microspheres. This study shows that microspheres can be incorporated into porous solids using supercritical CO2, allowing for a wide variety of drug-biodegradable polymer formulations prepared using the proven emulsion/solvent evaporation method to be tested.

Porous TiO2 surface formed on nickel‐titanium alloy by plasma electrolytic oxidation: A prospective polymer‐free reservoir for drug eluting stent applications

In this study, a porous oxide layer was formed on the surface of nickel-titanium alloy (NiTi) by plasma electrolytic oxidation (PEO) with the aim to produce a polymer-free drug carrier for drug eluting stent (DES) applications. The oxidation was performed galvanostatically in concentrated phosphoric acid electrolyte at low temperature. It was found that the response of NiTi substrate during the PEO process was different from that of bulk Ti, since the presence of large amount of Ni delayed the initial formation of a compact oxide layer that is essential for the PEO to take place. Under optimized PEO conditions, the resultant surface showed porosity, pore density and oxide layer thickness of 14.11%, 2.40 × 10⁵ pores/mm² and 0.8 μm, respectively. It was additionally noted that surface roughness after PEO did not significantly increase as compared with that of original NiTi substrate and the EDS analyses revealed a decrease in Ni/Ti ratio on the surface after PEO. The cross-section morphology showed no discontinuity between the PEO layer and the NiTi substrate. Furthermore, wettability and surface free energy of the NiTi substrate increased significantly after PEO treatment. The PEO process could be successfully translated to NiTi stent configuration proving for the first time its feasibility for such a medical device and offering potential for development of alternative, polymer-free drug carriers for NiTi DES.

Characterization of Porous TiO2 Surfaces Formed on 316L Stainless Steel by Plasma Electrolytic Oxidation for Stent Applications

In this study, a porous oxide layer was formed on the surface of 316L stainless steel (SS) by combining Ti magnetron sputtering and plasma electrolytic oxidation (PEO) with the aim to produce a polymer-free drug carrier for drug eluting stent (DES) applications. The oxidation was performed galvanostatically in Na3PO4 electrolyte. The surface porosity, average pore size and roughness varied with PEO treatment duration, and under optimum conditions, the surface showed a porosity of 7.43%, an average pore size of 0.44 µm and a roughness (Ra) of 0.34 µm. The EDS analyses revealed that the porous layer consisted of Ti, O and P. The cross-sectional morphology evidenced a double-layer structure, with a porous titania surface and an un-oxidized dense Ti film towards the interface with 316L SS. After the PEO treatment, wettability and surface free energy increased significantly. The results of the present study confirm the feasibility of forming a porous TiO2 layer on stainless steel by combining sputtering technology and PEO. Further, the resultant porous oxide layer has the potential to be used as a drug carrier for DES, thus avoiding the complications associated with the polymer based carriers.

Effects of dexamethasone-loaded PLGA microspheres on human fetal osteoblasts

Integration of a drug delivery function into implantable medical devices enables local release of specific bioactives to control cells–surface interactions. One alternative to achieve this biofunctionality for bone implants is to incorporate particulate drug delivery systems (DDSs) into the rough or porous implant surfaces. The scope of this study was to assess the effects of a model DDS consisting of poly(D,L-lactide-co-glycolide) (PLGA) microspheres loaded with an anti-inflammatory drug, dexamethasone (DXM), on the response of Simian Virus-immortalized Human Fetal Osteoblast (SV-HFO) cells. The microspheres were prepared by the oil-in-water emulsion/solvent evaporation method, whereas cells response was investigated by Alamar Blue test for viability, alkaline phosphatase (ALP) activity for differentiation, and Alizarin Red staining for matrix mineralization. Cell viability was not affected by the presence of increased concentrations of polymeric microspheres in the culture media. Furthermore, in the cultures with DXM-loaded microspheres, ALP activity was expressed at levels similar with those obtained under osteogenic conditions, indicating that DXM released from the microsphere-stimulated cell differentiation. Matrix mineralization occurred preferentially around the DXM-loaded microspheres confirming that the released DXM could act as osteogenic supplement for the cells. These in vitro findings suggest that a particulate PLGA-DXM DDS may actually provide dual, anti-inflammatory and osteogenic functions when incorporated on the surface of bone implants.