Preparation and characterization of vancomycin releasing PHBV coated 45S5 Bioglass®-based glass–ceramic scaffolds for bone tissue engineering

Porous 45S5 Bioglass^®-based glass–ceramic scaffolds with high porosity (96%) and interconnected pore structure (average pore size 300 μm) were prepared by foam replication method. In order to improve the mechanical properties and to incorporate a drug release function, the scaffolds were coated with a drug loaded solution, consisting of PHBV and vancomycin. The mechanical properties of the scaffolds were significantly improved by the PHBV coating. The bioactivity of scaffolds upon immersion in SBF was maintained in PHBV coated scaffolds although the formation of hydroxyapatite was slightly retarded by the presence of the coating. The encapsulated drug in coated scaffolds was released in a sustained manner (99.9% in 6 days) as compared to the rapid release (99.5% in 3 days) of drug directly adsorbed on the uncoated scaffolds. The obtained drug loaded and bioactive composite scaffolds represent promising candidates for bone tissue engineering applications.     

Sintering and crystallisation of 45S5 Bioglass® powder

The sintering process of 45S5 Bioglass^® powder (mean particle size < 5 μm) was investigated by using different thermal analysis methods. Heating microscopy and conventional dilatometry techniques showed that bioactive glass sinters in two major steps: a short stage in the temperature range 500–600 °C and a longer stage in the range 850–1100 °C. The optimal sintering temperature and time were found to be 1050 °C and 140 min, respectively. Differential thermal analysis (DTA) showed that Bioglass^® crystallises at temperatures between 600 and 750 °C. The characteristic crystalline phases were identified by Fourier Transformed Infrared Spectroscopy (FTIR), Transmission Electron Microscopy (TEM) and X-Ray Diffraction (XRD). The crystallisation kinetics was studied by DTA, using a non-isothermal method. The Kissinger plot for Bioglass^® powder heated at different heating rates between 5 and 30 °C/min yielded an activation energy of 316 kJ/mol. The average value of Avrami parameter determined using the Augis–Bennett method was 0.95 ± 0.10, confirming a surface crystallisation mechanism. After sintering at 1050 °C for 140 min, the main crystalline phase was found to be Na₂Ca₂Si₃O₉. The results of this work are useful for the design of the sintering/crystallisation heat treatment of Bioglass^® powder which is used for fabricating tissue engineering scaffolds with varying degree of bioactivity.     

Macroporous Bioglass-derived scaffolds for bone tissue regeneration

Since it was introduced at the end of the ‘60s, the 45S5 Bioglass^® has played a fundamental role among the materials for orthopedic applications because of its ability to build a stable bond with the surrounding bone. The recent development of bone tissue engineering has led the interest of many scientists in the design of Bioglass^®-based scaffolds, i.e. porous systems able to drive and foster the bone tissue regrowth. Among the available techniques to realize scaffolds, the polymer burning out method, which employs organic particles as pore generating agents in a ceramic matrix, combines versatility and low cost. In spite of the advantages of the polymer burning out method, this technique has been rarely applied to 45S5 Bioglass^® and a systematic feasibility study has not been carried out on this issue yet. In order to fill this gap, in the present contribution the polymer burning out method was employed to design macroporous scaffolds based on 45S5 Bioglass^®. Different amounts of organic phase were used to obtain samples with different porosity. The samples were characterized from a microstructural point of view, in order to evaluate the pore morphology, dimension and degree of interconnectivity. Such findings proved that a proper setting of the processing parameters made it possible to achieve very high porosity values, among the best ones obtained in the literature with the same technique, together with an appreciable mechanical behaviour, according to compression tests. Finally, the scaffolds bioactivity was assessed by means of in vitro tests in a simulated body fluid (SBF) solution. Moreover, in the view of a potential application for bone tissue engineering, a preliminary biological evaluation of the obtained scaffolds to sustain cell proliferation was carried out.     

Electrophoretic deposition of carbon nanotubes and bioactive glass particles for bioactive composite coatings

The production of bioactive coatings consisting of 45S5 Bioglass^® and mutli-walled carbon nanotubes (CNTs) by electrophoretic deposition (EPD) was investigated. In addition to pure Bioglass^® coatings, the co-deposition and sequential deposition of Bioglass^® particles (size <5 μm) and CNTs on stainless steel substrates were carried out in order to fabricate bioactive, nanostructured composite layers. The optimal experimental conditions were determined using well-dispersed suspensions by means of a trial-and-error approach by varying the relevant EPD parameters: applied voltage and deposition time. SEM images demonstrated the successful fabrication of Bioglass^®/CNT composites by revealing their morphology and topography. The co-deposition of Bioglass^® particles and CNTs resulted in homogenous and dense coatings exhibiting the presence of well-dispersed CNTs placed in-between micron-sized Bioglass^® particles. This network of high-strength CNTs embedded in the glass layer could act as reinforcing element leading to higher mechanical stability of the coatings. The coatings obtained by sequential deposition offered a two-dimensional nanostructured fibrous mesh of CNTs covering the Bioglass^® layer thus providing a controlled (ordered) nano-topographical surface. This surface nanostructure has the potential to promote the attachment and growth of osteoblast cells and to benefit the formation of bone-like nanosized hydroxyapaptite crystals in contact with body fluids.     

Pressureless spark plasma–sintered Bioglass® 45S5 with enhanced mechanical properties and stress–induced new phase formation

Commercial Bioglass^® 45S5 powder was sintered using spark plasma sintering (SPS) technique without the assistance of mechanical pressure with heating and cooling rate of 100 °C/min, dwell temperature of 1050 °C and dwell time of 30 min. Such route enabled the production of samples exhibiting superior mechanical properties in comparison with Bioglass^® sintered in furnace. In particular, flexural strength and fracture toughness reached values close to those of apatite-wollastonite bioceramics, already widely used in clinical applications. The residual stresses implemented by indentation promoted the formation of a new phase in samples sintered by SPS. Complementary use of Raman and energy dispersive spectroscopy (EDS) indicated the phase as sodium carbide and a formation mechanism was proposed.     

Fabrication and characterization of electrospun poly‐DL‐lactide (PDLLA) fibrous coatings on 45S5 Bioglass® substrates for bone tissue engineering applications

BACKGROUND: This work focuses on combining electrospun biodegradable poly‐DL‐lactide (PDLLA) fibres and 45S5 Bioglass^® for tissue engineering applications. RESULTS: A variety of fibrous structures were produced upon application of an electric field to a flowing solution of PDLLA (5 wt/v%) in di‐methyl carbonate (DMC). Electrospinning was achieved at an applied voltage of 8.5 kV for a fixed flow rate of 5 µL min^?1. Scanning electron microscopy images of PDLLA fibres deposited on 45S5 Bioglass^® sintered pellets revealed that the fibres had diameters in the range 100–200 nm, leading to increased surface roughness, as assessed by white light interferometry. Bioactivity studies on PDLLA fibre coated Bioglass^® substrates were carried out in simulated body fluid (SBF) for 7, 14 and 28 days. It was found that mineralization of PDLLA fibres on 45S5 Bioglass^® substrate (formation of hydroxyapatite) occurred after 7 days of immersion in SBF and full coverage of PDLLA fibres with HA nanocrystals was achieved after 14 days in SBF. CONCLUSION: The approach investigated represents a convenient method to develop a controlled mineralized fibrous topography on bioactive glass substrates for improved cell attachment, which can be exploited in bone tissue engineering applications. Copyright © 2009 Society of Chemical Industry     

A new potassium-based bioactive glass: Sintering behaviour and possible applications for bioceramic scaffolds

Providing structural support while maintaining bioactivity is one of the most important goals for bioceramic scaffolds, i.e. artificial templates which guide cells to grow in a 3D pattern, facilitating the formation of functional tissues. In the last few years, 45S5 Bioglass^® has been widely investigated as scaffolding material, mainly for its ability to bond to both hard and soft tissues. However, thermal treatments to improve the relatively poor mechanical properties of 45S5 Bioglass^® turn it into a glass-ceramic, decreasing its bioactivity. Therefore, the investigation of new materials as candidates for scaffold applications is necessary. Here a novel glass composition, recently obtained by substituting the sodium oxide with potassium oxide in the 45S5 Bioglass^® formulation, is employed in a feasibility study as scaffolding material. The new glass, named BioK, has the peculiarity to sinter at a relatively low temperature and shows a reduced tendency to crystallize. In this work, BioK has been employed to realize two types of scaffolds. The obtained samples have been fully characterized from a microstructural point of view and compared to each other. Additionally, their excellent bioactivity has been established by means of in vitro tests.     

Direct ink writing of highly bioactive glasses

Direct ink writing (DIW), or Robocasting, is an additive manufacturing technique that offers the opportunity to create patient specific bioactive glass scaffolds and high strength scaffolds for bone repair. The original 45S5 Bioglass^® composition crystallises during sintering and until now, robocast glass scaffolds contained at least 51.9 mol% SiO₂ or B₂O₃ to maintain their amorphous structure. Here, ICIE16 and PSrBG compositions, containing <50 mol% SiO₂, giving silicate network connectivity close to that of 45S5, were robocast and compared to 13–93 composition. Results showed Pluronic F-127 can be used as a universal binder regardless of glass reactivity and that particle size distribution affected the ink “printability”. Scaffolds with interconnects of 150 μm (41–43% porosity) had compressive strengths of 32–48 MPa, depending on the glass composition. Robocast scaffolds from these highly reactive bioactive glasses promise greatly improved bone regeneration rates compared with existing bioactive glass scaffolds.     

Porous hydroxyapatite‐bioactive glass hybrid scaffolds fabricated via ceramic honeycomb extrusion

The successful fabrication of hydroxyapatite‐bioactive glass scaffolds using honeycomb extrusion is presented herein. Hydroxyapatite was combined with either 10 wt% stoichiometric Bioglass^® (BG1), calcium‐excess Bioglass^® (BG2) or canasite (CAN). For all composite materials, glass‐induced partial phase transformation of the HA into the mechanically weaker β‐tricalcium phosphate (TCP) occurred but XRD data demonstrated that BG2 exhibited a lower volume fraction of TCP than BG1. Consequently, the maximum compressive strength observed for BG1 and BG2 were 30.3 ± 3.9 and 56.7 ± 6.9 MPa, respectively, for specimens sintered at 1300°C. CAN scaffolds, in contrast, collapsed when handled when sintered below 1300°C, and thus failed. The microstructure illustrated a morphology similar to that of BG1 sintered at 1200°C, and hence a comparable compressive strength (11.4 ± 3.1 MPa). The results highlight the great potential offered by honeycomb extrusion for fabricating high‐strength porous scaffolds. The compressive strengths exceed that of commercial scaffolds, and biological tests revealed an increase in cell viability over 7 days for all hybrid scaffolds. Thus it is expected that the incorporation of 10 wt% bioactive glass will provide the added advantage of enhanced bioactivity in concert with improved mechanical stability.     

Biphasic calcium phosphate scaffolds fabricated by direct write assembly: Mechanical,anti-microbial and osteoblastic properties

The present work reports on the fabrication of 3-D porous calcium phosphate scaffolds by robocasting from biphasic (HA/β-TCP ≈ 1.5) powders, undoped and co-doped with Sr and Ag. Scaffolds with different pore sizes and rod diameter of 410 μm were fabricated and sintered at 1100 °C. The size and morphology of the powder particles, and the concentrations of the processing additives, were shown to play major roles in the robocasting process. For all pore sizes tested, the compressive strength of scaffolds was comparable to or even higher than that of cancellous bone, and mechanical data could be systematically correlated with the porosity fraction. Co-doping the starting powders with Sr and Ag enhanced the mechanical strength of scaffolds, conferred good antimicrobial activity against Staphylococcus aureus and Escherichia coli, and did not induce any cytotoxic effects on human MG-63 cells. Furthermore, the co-doped powder was more effective in inducing pre-osteoblastic proliferation.     

Effect of Niobium Oxide on the Structure and Properties of Melt‐Derived Bioactive Glasses

The effects of adding Nb₂O₅ on the physical properties and glass structure of two glass series derived from the 45S5 Bioglass^® have been studied. The multinuclear ²⁹Si, ³¹P, and ²³Na solid‐state MAS NMR spectra of the glasses, Raman spectroscopy and the determination of some physical properties have generated insight into the structure of the glasses. The ²⁹Si MAS NMR spectra suggest that Nb⁵⁺ ions create cross‐links between several oxygen sites, breaking Si–O–Si bonds to form a range of polyhedra [Nb(OM)_6?y(OSi)_y], where 1 ≤ y ≤ 5 and M = Na, Ca, or P. The Raman spectra show that the Nb–O–P bonds would occur in the terminal sites. Adding Nb₂O₅ significantly increases the density and the stability against devitrification, as indicated by ΔT(T_x ? T_g). Bioglass particle dispersions prepared by incorporating up to 1.3 mol% Nb₂O₅ by replacing P₂O₅ or up to 1.0 mol% Nb₂O₅ by replacing SiO₂ in 45S5 Bioglass^® using deionized water or solutions buffered with HEPES showed a significant increase in the pH during the early steps of the reaction, compared using the rate and magnitude during the earliest stages of BG45S5 dissolution.     

Microstructural characterization and in vitro bioactivity of porous glass-ceramic scaffolds for bone regeneration by synchrotron radiation X-ray microtomography

One of the key purposes of bone tissue engineering is the development of new biomaterials that can stimulate the body's own regenerative mechanism for patient's anatomical and functional recovery. Bioactive glasses, due to their versatile properties, are excellent candidates to fabricate porous 3-D architectures for bone replacement. In this work, morphological and structural investigations are carried out on Bioglass^®- and CEL2-derived scaffolds produced by sponge replication (CEL2 is an experimental glass developed at Politecnico di Torino). Synchrotron radiation X-ray microtomography is used to study the samples 3-D architecture, pores size, shape, distribution and interconnectivity, as well as the growth kinetics on scaffolds struts of a newly formed apatitic phase during in vitro treatment in simulated body fluid, in order to describe from a quantitative viewpoint the bioactive potential of the analyzed biomaterials. An accurate comparison between architectural features and bioactive behaviour of Bioglass^®- and CEL2-derived scaffolds is presented and discussed.     

Reinforcement with reduced graphene oxide of bioactive glass scaffolds fabricated by robocasting

45S5 bioactive glass composite scaffolds reinforced with reduced graphene oxide (rGO) were fabricated for the first time by robocasting (direct-writing) technique. Composite scaffolds with 0–3 vol.% content of rGO platelets were printed, and then consolidated by pressureless sintering at 550 or 1000 °C in Ar atmosphere. It was found that the addition of rGO platelets up to 1.5 vol.% content enhanced the mechanical performance of the 45S5 bioactive glass scaffolds in terms of strength and toughness. Best performance was obtained for 1 vol.% rGO, which yielded an enhancement of the fracture toughness of ∼850 and 380% for sintering temperatures of 550 and 1000 °C, respectively, while the compressive strength increased by ∼290 and 75%. rGO addition thus emerges as a promising approach for the fabrication of novel bioglass scaffolds with improved mechanical performance without deterioration of their bioactivity, which may then find use in load-bearing bone tissue engineering applications.     

Bioactivity modulation of Bioglass® powder by thermal treatment

A discussion of the effects of Bioglass^® powder crystallisation on the in vitro bioactivity in simulated body fluid (SBF) is presented.Starting from Bioglass^® powder, different glass–ceramics were obtained by thermal treatments between 580 °C and 800 °C, with variable crystallisation content (from 10 to 92 wt%). All samples (glass and glass–ceramics) showed apatite formation at their surface when immersed in SBF. In case of the glass and the samples with lowest crystallinity, the first step of apatite formation involved a homogenous dissolution followed by an amorphous calcium phosphate (CaP) layer precipitation. For the samples with a high crystallisation content, heterogeneous dissolution occurred. For the first time, the Stevels number of the amorphous phase is used to explain the possible dissolution of the crystalline phase present in materials with a similar chemical composition of the Bioglass^®. All samples presented at 21 days of immersion in SBF B-type hydroxycarbonate apatite crystals.     

Suitability of Biosilicate® glass-ceramic powder for additive manufacturing of highly porous scaffolds

The Biosilicate® glass-ceramic is one of the most promising alternatives to the 45S5 Bioglass® in terms of bioactivity, osteoconductivity, osteoinductivity, non-cytotoxicity, and antibacterial properties, with significant advantages in the manufacturing of specific components of complex shapes for bone tissue application. Unlike in 45S5 glass, the crystallization does not lead to a degradation of bioactivity. In the present paper, we explored the suitability of Biosilicate® for the manufacturing of highly porous scaffolds (porosity of 50–80 vol%) by using modern additive manufacturing technologies, such as direct ink writing (DIW) and digital light processing (DLP). Both techniques could be easily applied to fine powders of Biosilicate® mixed with fugitive binders. Significant densification of the struts, despite the limited powder packing, could be achieved using liquid-assisted sintering, in turn, triggered by the phosphate-enriched residual glass phase, already at 1000 °C. The strength-to-density ratio could be variously tuned (from 1.5 to 9.5 MPa cm³/g), especially with DLP-derived samples, by adjusting both the firing temperature and the scaffold topology.     

Response of 45S5 Bioglass foams to tensile loading

The tensile strength test of highly porous ceramic foams has been developed and first results have been obtained on bioactive glass foams. The tested material was a 45S5 Bioglass^® derived foam-like scaffold intended for use in bone tissue engineering which was manufactured by Bioglass^® slurry coating of polyurethane foam and subsequent sintering. The Bioglass^® foam structure was investigated in two states: uncoated (as fabricated) and with a PDLLA polymer coating. The tensile testing procedure is based on fixation of the foam into aluminium pots by a suitable adhesive. Tensile test samples having cross-section of 10×10 mm² and a length of 30 mm were used for the experiments. Basic fractographic analysis was applied to get relevant information about specimens' behaviour during tensile loading. In Bioglass^® based scaffolds, the presence of PDLLA coating led to a significant increase of the fracture strength, which is attributed to the interaction of the polymer phase with propagating cracks, e.g. enabling a crack bridging mechanism to take place.     

Effect of uniaxial load on the sintering behaviour of 45S5 Bioglass powder compacts

The effect of a uniaxial compressive load on the sintering behaviour of 45S5 Bioglass^® powder compacts was investigated by means of sinter-forging. In comparison to free sintering, densification kinetics was enhanced and the degree of crystallization was reduced. Significantly lower sintering temperatures, i.e. 610 °C instead of 1050 °C, can be employed to obtain dense Bioglass^® parts when sintering is performed under uniaxial load. The effect of mechanical loading on microstructure (pore density, shape and orientation) is discussed. The results of the investigation are relevant in connection with the development of sintered Bioglass^® substrates for bone replacement devices, where both porosity and crystallinity of the part require careful control and low densification temperatures are sought.     

Enhancing In Vitro Bioactivity of Melt‐Derived 45S5 Bioglass® by Comminution in a Stirred Media Mill

45S5 Bioglass^® (45S5 BG) is a frequently applied Type A bioactive material, capable of forming an inherent bond to bone and soft tissue. Currently, applied melt‐derived bioactive glass powders (BG) exhibit particle sizes between a few to several hundred micrometers. Recent studies on nanometer‐sized bioactive glasses (nBGs), produced by bottom‐up methods like sol–gel processing or flame spray pyrolysis, have indicated their great potential for several biomedical applications. In this study, the feasibility of top‐down processing starting from bulk 45S5 BG by wet comminution in a stirred media mill was investigated. The products were assessed by in vitro hydroxycarbonate apatite (HCAp) formation in simulated body fluid, which is a marker for bioactive behavior. The study reveals the paramount influence of the used solvent for a successful top‐down processing: In comparison with the as‐received material bioactivity is lost for powders processed in water, preserved for comminution in ethanol and increased for powders processed using the alcohols n‐butanol, n‐pentanol, and n‐hexanol. It was also found that only for the latter solvents, the chemical composition of the glass is maintained during comminution. Flake‐like, slightly porous particles with specific surface areas of ~25–30 m²/g are obtained. Thus, the presented comminution approach offers a convenient technique to process 45S5 BG with enhanced bioactivity.     

Preparation and characterization of Bioglass®-based scaffolds reinforced by poly-vinyl alcohol/microfibrillated cellulose composite coating

The present work deals with the preparation and mechanical characterization of Bioglass^®-based porous scaffolds reinforced by a composite coating constituted by polyvinyl alcohol (PVA) and microfibrillated cellulose (MFC). Samples were produced by foam replication process, using a novel ethanol-based Bioglass^® slurry. The addition of PVA/MFC coating led to a 10 fold increase of compressive strength and a 20 fold increase of tensile strength in comparison with non-coated scaffolds. SEM observations of broken struts surfaces proved the reinforcing and toughening effect of the composite coating which were ascribed to crack bridging and fracture of cellulose fibrils. The mechanical properties of the coatings were investigated by tensile testing of PVA/MFC composite stripes. The stirring time of the PVA/MFC solution came out as a crucial parameter in order to achieve a more homogeneous dispersion of the fibers and therefore enhanced strength and stiffness.     

A study of the electrophoretic deposition of Bioglass® suspensions using the Taguchi experimental design approach

This paper presents a study of the Taguchi design method to optimise the rate of electrophoretic deposition (EPD) of Bioglass^® particles from aqueous suspensions. The effect of Bioglass^® concentration, pH and electric field was investigated. An orthogonal array of L₁₆ type with mixed levels of the control factors was utilized. Multivariate analysis of variance (MANOVA) and regression analysis based on the partial least-square method were used to identify the significant factors affecting the deposition rate and its stability during constant-voltage EPD. It was found that the pH of the suspension significantly influences the deposition rate whereas the applied electric field has the smallest effect on the deposition rate. In addition, the interaction between the Bioglass^® concentration and pH, which are key processing parameters influencing the deposition rate, was found to be significant. Although a high deposition rate was obtained at low electric field (5 V/cm) and pH 7, the instability of the suspension in particular at high Bioglass^® concentrations resulted in an increase in the variability of the deposition rate. The optimal EPD condition predicted was verified by experiments and good qualitative agreement was obtained. The experimental results and statistical analyses are discussed based on the current knowledge of the EPD of ceramic materials.