Phosphate Glass Microspheres as Cell Microcarrier Substrates for Bone Tissue Engineering Applications

Phosphate glasses have demonstrated a high degree of suitability for use as biomaterials in a wide range of biomedical applications involving both hard and soft tissue regeneration. This study focused on the use of these glasses as substrate microcarrier materials for three-dimensional bone tissue formation. For this purpose, the successful production of phosphate glass microspheres in the ~10–200 μm size range was demonstrated using a simple, inexpensive and industrially scalable process. Microspheres made out of two different series of phosphate glass compositions were investigated: (a) iron phosphate glasses 0.5P2O5–0.4CaO–(0.1 – x)Na2O–xFe2O3 where x = 0.00, 0.01, 0.03 and 0.05 mole fraction and (b) titanium phosphate glasses 0.5P2O5–0.4CaO–(0.1 – x)Na2O–xTiO2 where x = 0.00, 0.01, 0.03, 0.05 and 0.07 mole fraction. Investigations of the microsphere physicochemical properties revealed the densification of the glass structure with increased metal oxide incorporation in the glass. Glass structural characterisation studies provided valuable information relating the physicochemical properties to glass structural arrangements at the glass phase and atomic levels. Cell culture studies involving culture of the microspheres with MG63 osteosarcoma cells and human mesenchymal stem cells indicated that microspheres made of glasses containing 3–7 mol% metal oxides, and particularly those containing 5 mol% TiO2, showed favourable characteristics in terms of cell attachment, viability, proliferation and release of proteins related to cell differentiation and metabolism under both static conditions in culture well plates and dynamic conditions in spinner flask bioreactors. Taken together, the results provide evidence of the potential of the investigated glass microspheres to function as effective microcarrier substrates for bone tissue engineering application

Titanium phosphate glass microcarriers induce enhanced osteogenic cell proliferation and human mesenchymal stem cell protein expression.

In this study, we have developed 50- to 100-µm-sized titanium phosphate glass microcarriers (denoted as Ti5) that show enhanced proliferation of human mesenchymal stem cells and MG63 osteosarcoma cells, as well as enhanced human mesenchymal stem cell expression of bone differentiation markers, in comparison with commercially available glass microspheres at all time points. We also demonstrate that these microcarriers provide superior human mesenchymal stem cell proliferation with conventional Dulbecco's Modified Eagle medium than with a specially developed commercial stem cell medium. The microcarrier proliferative capacity is revealed by a 24-fold increase in MG63 cell numbers in spinner flask bioreactor studies performed over a 7-day period, versus only a 6-fold increase in control microspheres under the same conditions; the corresponding values of Ti5 and control microspheres under static culture are 8-fold and 7-fold, respectively. The capability of guided osteogenic differentiation is confirmed by ELISAs for bone morphogenetic protein-2 and osteopontin, which reveal significantly greater expression of these markers, especially osteopontin, by human mesenchymal stem cells on the Ti5 microspheres than on the control. Scanning electron microscopy and confocal laser scanning microscopy images reveal favorable MG63 and human mesenchymal stem cell adhesion on the Ti5 microsphere surfaces. Thus, the results demonstrate the suitability of the developed microspheres for use as microcarriers in bone tissue engineering applications

Novel sol–gel preparation of (P2O5)0.4–(CaO)0.25–(Na2O)X–(TiO2)(0.35−X) bioresorbable glasses (X = 0.05, 0.1, and 0.15)

Quaternary phosphate-based glasses in the P2O5–CaO–Na2O–TiO2 system with a fixed P2O5 and CaO content of 40 and 25 mol% respectively have been successfully synthesised via sol–gel method and bulk, transparent samples were obtained. The structure, elemental proportion, and thermal properties of stabilised sol–gel glasses have been characterised using X-ray diffraction (XRD), energy dispersive X-ray spectroscopy (EDX), 31P nuclear magnetic resonance (31P NMR), titanium K-edge X-ray absorption near-edge structure (XANES), fourier transform infrared (FTIR) spectroscopy, and differential thermal analysis (DTA). The XRD results confirmed the amorphous nature for all stabilized sol–gel derived glasses. The EDX result shows the relatively low loss of phosphorus during the sol–gel process and Ti K-edge XANES confirmed titanium in the glass structure is in mainly six-fold coordination environment. The 31P NMR and FTIR results revealed that the glass structure consist of mainly Q1 and Q2 phosphate units and the Ti4+ cation was acting as a cross-linking between phosphate units. In addition DTA results confirmed a decrease in the glass transition and crystallisation temperature with increasing Na2O content. Ion release studies also demonstrated a decrease in degradation rates with increasing TiO2 content therefore supporting the use of these glasses for biomedical applications that require a degree of control over glass degradation. These sol–gel glasses also offer the potential to incorporate proactive molecules for drug delivery application due to the low synthesis temperature employed

In vitro bioactivity of titanium-doped bioglass

Previous studies have suggested that incorporating relatively small quantities of titanium dioxide into bioactive glasses may result in an increase in bioactivity and hydroxyapatite formation. The present work therefore investigated the in vitro bioactivity of a titanium doped bioglass and compared the results with 45S5 bioglass. Apatite formation was evaluated for bioglass and Ti-bioglass in the presence and absence of foetal calf serum. Scanning electron microscopy (SEM) images were used to evaluate the surface development and energy dispersive X-ray measurements provided information on the elemental ratios. X-ray diffraction spectra confirmed the presence of apatite formation. Cell viability was assessed for bone marrow stromal cells under direct and indirect contact conditions and cell adhesion was assessed using SEM

Comprehensive in vitro and in vivo studies of novel melt-derived Nb-substituted 45S5 bioglass reveal its enhanced bioactive properties for bone healing

The present work presents and discusses the results of a comprehensive study on the bioactive properties of Nb-substituted silicate glass derived from 45S5 bioglass. In vitro and in vivo experiments were performed. We undertook three different types of in vitro analyses: (i) investigation of the kinetics of chemical reactivity and the bioactivity of Nb-substituted glass in simulated body fluid (SBF) by 31P MASNMR spectroscopy, (ii) determination of ionic leaching profiles in buffered solution by inductively coupled plasma optical emission spectrometry (ICP-OES), and (iii) assessment of the compatibility and osteogenic differentiation of human embryonic stem cells (hESCs) treated with dissolution products of different compositions of Nb-substituted glass. The results revealed that Nb-substituted glass is not toxic to hESCs. Moreover, adding up to 1.3 mol% of Nb2O5 to 45S5 bioglass significantly enhanced its osteogenic capacity. For the in vivo experiments, trial glass rods were implanted into circular defects in rat tibia in order to evaluate their biocompatibility and bioactivity. Results showed all Nb-containing glass was biocompatible and that the addition of 1.3 mol% of Nb2O5, replacing phosphorous, increases the osteostimulation of bioglass. Therefore, these results support the assertion that Nb-substituted glass is suitable for biomedical applications

Influence of single and binary doping of strontium and lithium on in vivo biological properties of bioactive glass scaffolds

Effects of strontium and lithium ion doping on the biological properties of bioactive glass (BAG) porous scaffolds have been checked in vitro and in vivo. BAG scaffolds were prepared by conventional glass melting route and subsequently, scaffolds were produced by evaporation of fugitive pore formers. After thorough physico-chemical and in vitro cell characterization, scaffolds were used for pre-clinical study. Soft and hard tissue formation in a rabbit femoral defect model after 2 and 4 months, were assessed using different tools. Histological observations showed excellent osseous tissue formation in Sr and Li + Sr scaffolds and moderate bone regeneration in Li scaffolds. Fluorochrome labeling studies showed wide regions of new bone formation in Sr and Li + Sr doped samples as compared to Li doped samples. SEM revealed abundant collagenous network and minimal or no interfacial gap between bone and implant in Sr and Li + Sr doped samples compared to Li doped samples. Micro CT of Li + Sr samples showed highest degree of peripheral cancellous tissue formation on periphery and cortical tissues inside implanted samples and vascularity among four compositions. Our findings suggest that addition of Sr and/or Li alters physico-chemical properties of BAG and promotes early stage in vivo osseointegration and bone remodeling that may offer new insight in bone tissue engineering

Development of microspheres for biomedical applications: a review

An overview of microspheres manufactured for use in biomedical applications based on recent literature is presented in this review. Different types of glasses (i.e. silicate, borate, and phosphates), ceramics and polymer-based microspheres (both natural and synthetic) in the form of porous , non-porous and hollow structures that are either already in use or are currently being investigated within the biomedical area are discussed. The advantages of using microspheres in applications such as drug delivery, bone tissue engineering and regeneration, absorption and desorption of substances, kinetic release of the loaded drug components are also presented. This review also reports on the preparation and characterisation methodologies used for the manufacture of these microspheres. Finally, a brief summary of the existing challenges associated with processing these microspheres which requires further research and development are presented