Versatile Biodegradable Poly(acrylic acid)-Based Hydrogels Infiltrated in Porous Titanium Implants to Improve the Biofunctional Performance.

This research work proposes a synergistic approach to improve implants' performance through the use of porous Ti substrates to reduce the mismatch between Young's modulus of Ti (around 110 GPa) and the cortical bone (20-25 GPa), and the application of a biodegradable, acrylic acid-based polymeric coating to reduce bacterial adhesion and proliferation, and to enhance osseointegration. First, porous commercially pure Ti substrates with different porosities and pore size distributions were fabricated by using space-holder techniques to obtain substrates with improved tribomechanical behavior. On the other hand, a new diacrylate cross-linker containing a reduction-sensitive disulfide bond was synthesized to prepare biodegradable poly(acrylic acid)-based hydrogels with 1, 2, and 4% cross-linker. Finally, after the required characterization, both strategies were implemented, and the combination of 4% cross-linked poly(acrylic acid)-based hydrogel infiltrated in 30 vol % porosity, 100-200 µm average pore size, was revealed as an outstanding choice for enhancing implant performance.

Nanotextured porous titanium scaffolds by argon ion irradiation: Toward conformal nanopatterning and improved implant osseointegration.

Stress shielding and osseointegration are two main challenges in bone regeneration, which have been targeted successfully by chemical and physical surface modification methods. Direct irradiation synthesis (DIS) is an energetic ion irradiation method that generates self-organized nanopatterns conformal to the surface of materials with complex geometries (e.g., pores on a material surface). This work exposes porous titanium samples to energetic argon ions generating nanopatterning between and inside pores. The unique porous architected titanium (Ti) structure is achieved by mixing Ti powder with given amounts of spacer NaCl particles (vol % equal to 30%, 40%, 50%, 60%, and 70%), compacted and sintered, and combined with DIS to generate a porous Ti with bone-like mechanical properties and hierarchical topography to enhance Ti osseointegration. The porosity percentages range between 25% and 30% using 30 vol % NaCl space-holder (SH) volume percentages to porosity rates of 63%-68% with SH volume of 70 vol % NaCl. Stable and reproducible nanopatterning on the flat surface between pores, inside pits, and along the internal pore walls are achieved, for the first time on any porous biomaterial. Nanoscale features were observed in the form of nanowalls and nanopeaks of lengths between 100 and 500 nm, thicknesses of 35-nm and heights between 100 and 200 nm on average. Bulk mechanical properties that mimic bone-like structures were observed along with increased wettability (by reducing contact values). Nano features were cell biocompatible and enhanced in vitro pre-osteoblast differentiation and mineralization. Higher alkaline phosphatase levels and increased calcium deposits were observed on irradiated 50 vol % NaCl samples at 7 and 14 days. After 24 h, nanopatterned porous samples decreased the number of attached macrophages and the formation of foreign body giant cells, confirming nanoscale tunability of M1-M2 immuno-activation with enhanced osseointegration.

Antimicrobial and Antibiofilm Effect of 4,4'-Dihydroxy-azobenzene against Clinically Resistant Staphylococci.

The spread of antibiotic resistance among human and animal pathogens is one of the more significant public health concerns. Moreover, the restrictions on the use of particular antibiotics can limit the options for the treatment of infections in veterinary clinical practice. In this context, searching for alternative antimicrobial substances is crucial nowadays. In this study, 4,4'-dihydroxy-azobenzene (DHAB) was tested for its potential in vitro as an antimicrobial agent against two relevant human and animal pathogens, namely Staphylococcus aureus and Staphylococcus pseudintermedius. The values of minimal inhibitory concentration (MIC) were 64 and 32 mg/L respectively, and they comparable to other azo compounds of probed antimicrobial activity. In addition, the minimal bactericidal concentrations (MCB) were 256 and 64 mg/L. The mechanism by which DHAB produces toxicity in staphylococci has been investigated. DHAB caused membrane damage as revealed by the increase in thiobarbituric acid reactive substances (TBARS) such as malondialdehyde. Furthermore, differential induction of the enzymes peroxidases and superoxide dismutase in S. aureus and S. pseudintermedius suggested their prevalent role in ROS-scavenging due to the oxidative burst induced by this compound in either species. In addition, this substance was able to inhibit the formation of biofilms by both bacteria as observed by colorimetric tests and scanning electron microscopy. In order to assess the relevance of DHAB against clinical strains of MRSA, 10 clinical isolates resistant to either methicillin or daptomycin were assayed; 80% of them gave values of CMI and CMB similar to those of the control S. aureus strain. Finally, cutaneous plasters containing a composite formed by an agar base supplemented with DHAB were designed. These plasters were able to inhibit in vitro the growth of S. aureus and S. pseudintermedius, particularly the later, and this suggests that this substance could be a promising candidate as an alternative to antibiotics in the treatment of animal skin infections, as it has been proven that the toxicity of this substance is very low particularly at a dermal level.

Approach to the Fatigue and Cellular Behavior of Superficially Modified Porous Titanium Dental Implants.

In this work, the fatigue and cellular performance of novel superficially treated porous titanium dental implants made up using conventional powder metallurgy and space-holder techniques (30 vol.% and 50 vol.%, both with a spacer size range of 100-200 µm) are evaluated. Before the sintering stage, a specific stage of CNC milling of the screw thread of the implant is used. After the consolidation processing, different surface modifications are performed: chemical etching and bioactive coatings (BG 45S5 and BG 1393). The results are discussed in terms of the effect of the porosity, as well as the surface roughness, chemical composition, and adherence of the coatings on the fatigue resistance and the osteoblast cells' behavior for the proposed implants. Macro-pores are preferential sites of the nucleation of cracks and bone cell adhesion, and they increase the cellular activity of the implants, but decrease the fatigue life. In conclusion, SH 30 vol.% dental implant chemical etching presents the best bio-functional (in vitro osseointegration) and bio-mechanical (stiffness, yield strength and fatigue life) balance, which could ensure the required characteristics of cortical bone tissue.

Novel Utilization of Therapeutic Coatings Based on Infiltrated Encapsulated Rose Bengal Microspheres in Porous Titanium for Implant Applications.

Despite the increasing progress achieved in the last 20 years in both the fabrication of porous dental implants and the development of new biopolymers for targeting drug therapy, there are important issues such as bone resorption, poor osseointegration, and bacterial infections that remain as critical challenges to avoid clinical failure problems. In this work, we present a novel microtechnology based on polycaprolactone microspheres that can adhere to porous titanium implant models obtained by the spacer holder technique to allow a custom biomechanical and biofunctional balance. For this purpose, a double emulsion solvent evaporation technique was successfully employed for the fabrication of the microparticles properly loaded with the antibacterial therapeutic agent, rose bengal. The resulting microspheres were infiltrated into porous titanium substrate and sintered at 60 °C for 1 h, obtaining a convenient prophylactic network. In fact, the sintered polymeric microparticles were demonstrated to be key to controlling the drug dissolution rate and favoring the early healing process as consequence of a better wettability of the porous titanium substrate to promote calcium phosphate nucleation. Thus, this joint technology proposes a suitable prophylactic tool to prevent both early-stage infection and late-stage osseointegration problems.

Influence of Femtosecond Laser Modification on Biomechanical and Biofunctional Behavior of Porous Titanium Substrates.

Bone resorption and inadequate osseointegration are considered the main problems of titanium implants. In this investigation, the texture and surface roughness of porous titanium samples obtained by the space holder technique were modified with a femtosecond Yb-doped fiber laser. Different percentages of porosity (30, 40, 50, and 60 vol.%) and particle range size (100-200 and 355-500 µm) were compared with fully-dense samples obtained by conventional powder metallurgy. After femtosecond laser treatment the formation of a rough surface with micro-columns and micro-holes occurred for all the studied substrates. The surface was covered by ripples over the micro-metric structures. This work evaluates both the influence of the macro-pores inherent to the spacer particles, as well as the micro-columns and the texture generated with the laser, on the wettability of the surface, the cell behavior (adhesion and proliferation of osteoblasts), micro-hardness (instrumented micro-indentation test, P-h curves) and scratch resistance. The titanium sample with 30 vol.% and a pore range size of 100-200 µm was the best candidate for the replacement of small damaged cortical bone tissues, based on its better biomechanical (stiffness and yield strength) and biofunctional balance (bone in-growth and in vitro osseointegration).

Fabrication and Characterization of Bioactive Gelatin-Alginate-Bioactive Glass Composite Coatings on Porous Titanium Substrates.

In this research work, the fabrication of biphasic composite implants has been investigated. Porous, commercially available pure Ti (50 vol % porosity and pore distributions of 100-200, 250-355, and 355-500 µm) has been used as a cortical bone replacement, while different composites based on a polymer blend (gelatin and alginate) and bioactive glass (BG) 45S5 have been applied as a soft layer for cartilage tissues. The microstructure, degradation rates, biofunctionality, and wear behavior of the different composites were analyzed to find the best possible coating. Experiments demonstrated the best micromechanical balance for the substrate containing 200-355 µm size range distribution. In addition, although the coating prepared from alginate presented a lower mass loss, the composite containing 50% alginate and 50% gelatin showed a higher elastic recovery, which entails that this type of coating could replicate the functions of the soft tissue in areas of the joints. Therefore, results revealed that the combinations of porous commercially pure Ti and composites prepared from alginate/gelatin/45S5 BG are candidates for the fabrication of biphasic implants not only for the treatment of osteochondral defects but also potentially for any other diseases affecting simultaneously hard and soft tissues.

Electrical Impedance of Surface Modified Porous Titanium Implants with Femtosecond Laser.

The chemical composition and surface topography of titanium implants are essential to improve implant osseointegration. The present work studies a non-invasive alternative of electrical impedance spectroscopy for the characterization of the macroporosity inherent to the manufacturing process and the effect of the surface treatment with femtosecond laser of titanium discs. Osteoblasts cell culture growths on the titanium surfaces of the laser-treated discs were also studied with this method. The measurements obtained showed that the femtosecond laser treatment of the samples and cell culture produced a significant increase (around 50%) in the absolute value of the electrical impedance module, which could be characterized in a wide range of frequencies (being more relevant at 500 MHz). Results have revealed the potential of this measurement technique, in terms of advantages, in comparison to tiresome and expensive techniques, allowing semi-quantitatively relating impedance measurements to porosity content, as well as detecting the effect of surface modification, generated by laser treatment and cell culture.

Improved Corrosion Behavior and Biocompatibility of Porous Titanium Samples Coated with Bioactive Chitosan-Based Nanocomposites.

Porous titanium implants can be a good solution to solve the stress shielding phenomenon. However, the presence of pores compromises mechanical and corrosion resistance. In this work, porous titanium samples obtained using a space-holder technique are coated with Chitosan, Chitosan/AgNPs and Chitosan/Hydroxyapatite using only one step and an economic electrodeposition method. The coatings' topography, homogeneity and chemical composition were analyzed. A study of the effect of the porosity and type of coating on corrosion resistance and cellular behavior was carried out. The electrochemical studies reveal that porous samples show high current densities and an unstable oxide film; therefore, there is a need for surface treatments to improve corrosion resistance. The Chitosan coatings provide a significant improvement in the corrosion resistance, but the Chitosan/AgNPs and Chitosan/HA coatings showed the highest protection efficiency, especially for the more porous samples. Furthermore, these coatings have better adherence than the chitosan coatings, and the higher surface roughness obtained favors cell adhesion and proliferation. Finally, a combination of coating and porous substrate material with the best biomechanical balance and biofunctional behavior is proposed as a potential candidate for the replacement of small, damaged bone tissues.

Effect of the Processing Parameters on the Porosity and Mechanical Behavior of Titanium Samples with Bimodal Microstructure Produced via Hot Pressing.

Commercially pure (c.p.) titanium grade IV with a bimodal microstructure is a promising material for biomedical implants. The influence of the processing parameters on the physical, microstructural, and mechanical properties was investigated. The bimodal microstructure was achieved from the blends of powder particles with different sizes, while the porous structure was obtained using the space-holder technique (50 vol.% of ammonium bicarbonate). Mechanically milled powders (10 and 20 h) were mixed in 50 wt.% or 75 wt.% with c.p. titanium. Four different mixtures of powders were precompacted via uniaxial cold pressing at 400 MPa. Then, the specimens were sintered at 750 °C via hot pressing in an argon gas atmosphere. The presence of a bimodal microstructure, comprised of small-grain regions separated by coarse-grain ones, was confirmed by optical and scanning electron microscopies. The samples with a bimodal microstructure exhibited an increase in the porosity compared with the commercially available pure Ti. In addition, the hardness was increased while the Young's modulus was decreased in the specimens with 75 wt.% of the milled powders (20 h).

Characterization and Monitoring of Titanium Bone Implants with Impedance Spectroscopy.

Porous titanium is a metallic biomaterial with good properties for the clinical repair of cortical bone tissue, although the presence of pores can compromise its mechanical behavior and clinical use. It is therefore necessary to characterize the implant pore size and distribution in a suitable way. In this work, we explore the new use of electrical impedance spectroscopy for the characterization and monitoring of titanium bone implants. Electrical impedance spectroscopy has been used as a non-invasive route to characterize the volumetric porosity percentage (30%, 40%, 50% and 60%) and the range of pore size (100-200 and 355-500 mm) of porous titanium samples obtained with the space-holder technique. Impedance spectroscopy is proved to be an appropriate technique to characterize the level of porosity of the titanium samples and pore size, in an affordable and non-invasive way. The technique could also be used in smart implants to detect changes in the service life of the material, such as the appearance of fractures, the adhesion of osteoblasts and bacteria, or the formation of bone tissue.

Villari Effect at Low Strain in Magnetoactive Materials.

Magnetic composites of soft magnetic FeGa particles embedded in a silicone matrix have been synthesized. The Villari effect has been studied depending on the size and concentration of the particles and on the magnetic state of the composite. The results indicate a decrease in the Villari effect when the concentration of the magnetic particles increases. These results suggest a relationship between the Villari effect and the mechanical properties of the composites. The Young's modulus of the composites has been obtained by microindentation and their values related to the intensity and slope of the Villari signals. The results are explained on the basis that the reduction in the cross section of the composite when submitted to stress is the main origin of the variation of the magnetic flux in the Villari effect in this kind of composite. It has also been obtained that the magnetic state of the composite plays an important role in the Villari signal. When the magnetization of the composite is greater, the magnetic flux across the composite is greater too and, so, the same reduction in the cross section originates a greater Villari signal.

Biofunctional and Tribomechanical Behavior of Porous Titanium Substrates Coated with a Bioactive Glass Bilayer (45S5-1393).

The porous substrates of commercially pure titanium have been coated with a novel bilayer of bioactive glasses (BGs), 45S5 and 1393, to improve the osseointegration and solve the stress-shielding phenomenon of titanium partial implants. The porosity of the substrates and the scratch resistance and bioactivity of the coating have been evaluated. Results are discussed in terms of stiffness and yield strength of the substrates, as well as the chemical composition, thickness, and design of the bioglass coating (monolithic vs bilayer). The role of the pores was a crucial issue in the anchoring of the coating, both in porosity percentage (30 and 60 vol %) and in pore range size (100-200 and 355-500 µm). The study was focused on the adhesion and infiltration of a 1393 bioglass layer (in contact with a porous titanium substrate), in combination with the biofunctionality of the 45S5 bioglass layer (surrounded by the host bone tissue), as 1393 bioglass enhances the adherence, while 45S5 bioglass promotes higher bioactivity. This bioactivity of the raw powder was initially estimated by nuclear magnetic resonance, through the evaluation of the chemical environments, and confirmed by the formation of hydroxyapatite when immersed in a simulated body fluid. The results revealed that the substrate with 30 vol % of porosity and a range of 355-500 µm pore size, coated with this novel BG bilayer, presented the best combination in terms of mechanical and biofunctional properties.

Balancing biofunctional and biomechanical properties using porous titanium reinforced by carbon nanotubes.

Despite the well-known advantages of the titanium-based implant systems, they still lack an optimal balance between biofunctionality and mechanical strength, especially regarding the modulation of cellular response and a desired implant osseointegration. In this work, we fabricated a nanocomposite based on porous commercially pure grade 4 titanium (c.p. Ti) reinforced with carbon nanotubes (CNT) at 5% and 10% w/w, with the aim of obtaining a nanocomposite with lower stiffness compared to traditional titanium-based implants and with the mechanical strength and bioactivity owed by the CNT. Results obtained by scanning electron microscopy, X-ray photoelectron spectroscopy, and atomic force microscopy characterization showed that the CNT dispersed and incorporated into the porous c.p. Ti matrix. Interestingly, CNT were associated with a higher twining within neighbor Ti grains, which was indeed consistent with an increased in nano-hardness. Biological evaluation by MTT and Comet assay revealed that the nanocomposites did not induce genotoxicity and cytotoxicity on two different cells lines despite the presence of nickel at the surface. Accordingly, a purification step would be required before these CNT can be used for biomedical applications. Our results indicate that incorporation of CNT into porous c.p. Ti is a promising avenue to achieve an adequate balance between biofunctionality and mechanical strength in Ti-based scaffolds for tissue replacement. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 719-731, 2019.

Development of porous titanium for biomedical applications: A comparison between loose sintering and space-holder techniques.

One of the most important concerns in long-term prostheses is bone resorption as a result of the stress shielding due to stiffness mismatch between bone and implant. The aim of this study was to obtain porous titanium with stiffness values similar to that exhibited by cortical bone. Porous samples of commercial pure titanium grade-4 were obtained by following both loose-sintering processing and space-holder technique with NaCl between 40 and 70% in volume fraction. Both mechanical properties and porosity morphology were assessed. Young's modulus was measured using uniaxial compression testing, as well as ultrasound methodology. Complete characterization and mechanical testing results allowed us to determine some important findings: (i) optimal parameters for both processing routes; (ii) better mechanical response was obtained by using space-holder technique; (iii) pore geometry of loose sintering samples becomes more regular with increasing sintering temperature; in the case of the space-holder technique that trend was observed for decreasing volume fraction; (iv) most reliable Young's modulus measurements were achieved by ultrasound technique.