Fabrication and characterization of three dimensional composite scaffolds from 45S5 and copper doped bioactive glasses

Tissue damage and degenerative diseases such as large bone defects are among the most human health issues which lead to organ failure and death all over the world. Traditional methods such as autografts, allografts or metallic implants suffer limitations for treatment of these injuries. For instance, autografts have limited availability of tissues and it has a long time for treatment of damages. Also, allografts have a very high cost and they may cause adverse reaction in human body. Therefore, tissue engineering emerges a promising approach to overcome these problems. Bone tissue engineering is a science that aim to repair, replace or renew of damaged bones which caused by any reason such as illness, accident or trauma. Cells, biomolecules and scaffolds are three main components in tissue engineering approaches. In bone tissue engineering applications, scaffolds are mostly applied in the form of three dimensional porous structures. Three-dimensional scaffolds are prepared that mimic the structure of bone mineral and adjust with the cells to repair and refunction of bone tissue. Several fabrication methods such as sol gel process, foam replica technique, solvent casting with or without particule leaching, microsphere sintering, and solid free form fabrication, have been applied to produce these scaffolds for tissue engineering applications. Foam replica method is a simple process that provides to fabricate three dimensional porous scaffolds in a short time through these methods. Biomaterials, metals, ceramics, polymers and composite materials are used in production of these scaffolds. Among these materials, bioactive glasses which are in class of biological used materials (bioceramics), represent promising scaffolding materials for bone tissue regeneration. Bioactive glasses have very high biocompatible, bioactive, osteoconductive and osteoproductive properties that can be used for the repair and reconstruction of damaged bone tissues. They are mainly composed of silicon, sodium, calcium and phosphate component. When they are implanted to human body, they are able to bind with bone through a layer of hydroxyapatite formed on their surfaces which react with the surrounding tissue to form a mechanically strong interfacial bond between a host tissue and an implant. Bioactive glasses can be produced by conventional melting and sol-gel methods. Conventional melting method is a simple method and also it provides to fabricate bioglasses in a short time. Properties of bioactive glasses depend on their chemical structures and in order to enhance functional properties of them and improve bone production properties of these materials, different ions such as copper (Cu2+), strontium (Sr2+), magnesium (Mg2+), zinc (Zn2+) and silver (Ag1+) can be incorporated in their structures. All of these ions have essential biological effects. For instance, strontium accelerate bone healing processes and have positive effects on bone tissue repair. Among these ions, copper plays an active role in stimulating angiogenesis. Angiogenesis can be stimulated by delivery of the cells or growth factors such as endothelial growth factor (VEGF) from biodegradable scaffolds. However, these prices are very high and they have potential diverse effects in clinical applications. Copper ions are attractive for tissue engineering applications because of their low cost, high stability and potentially greater safety compared with recombinant proteins. In recent years, scientists have been working on polymer coating of bioglass based scaffolds in order to improve mechanical properties. Also, bioactivity and biodegradability behavior can be enhanced with this way. For this purpose, different natural and synthetic polymers are used. Among these materials, natural polymers such as alginate, gelatin and chitosan are remarkable materials. Especially, alginate is a special polymer which is widely used for cell immobilization and encapsulation. In this study, three dimensional scaffolds were fabricated based on 45S5 (composition in weight; %45 SiO2, %24.5 Na2O, %6 P2O5, %24.5 CaO) and copper doped (composition in weight; %45 SiO2, %24.5 Na2O, %6 P2O5, %24.5 CaO, %2 CuO) bioactive glasses. Conventional melting method was used to produce bioactive glasses. DTA analysis is applied to find glass transition and crystallization temperatures of the bioactive glass samples. Also, amorphous and crystalline phases of the glasses are determined with using XRD, and FTIR analysis was done to research chemical bonds. Crystallization temperatures of the glasses, which were found with DTA analysis, show that they were appropriate for sintering process in scaffold production. As a result of XRD and FTIR analysis, it was detected that glasses have fully amorphous structures. At the same time, it did not emerge a significant difference on the structure of glass with doping copper. Three dimensional scaffolds were fabricated from bioactive glasses by the foam replica method. Then in order to improve bioactivity and biodegradability behavior of scaffolds, they were coated with polymer. With this aim alginate which is a natural polymer was used at different percentages (1, 2, 3 % weight). Copper ions were dropped on some coated scaffolds which were produced by 45S5 bioactive glass and thus cross-linking of the copper with alginate was provided. Physical and micro structural properties of the obtained scaffolds were determined with using different characterization methods. It was observed that scaffolds have a very high porous structure and there was a good pore interconnectivity in the scaffold microstructure from the SEM images. As a result of FTIR and SEM analysis, it was detected that alginate attached on the surface of the scaffolds without clogging of the pores and also it was provided that homogeneous coating on the surface of the samples. The obtained scaffolds were immersed in simulated body fluid at different time points (1, 7, 14 and 28 day) to research of bioactivity and biodegradability behavior. As it was observed from the SEM images, it was seen that significantly hydroxyapatite layer formed on the surface of the scaffolds after immersion in SBF for 28 day. Furthermore, XRD patterns and FTIR spectra were verified this result. Characteristic chemical bonds which belongs to hydroxyapatite were obtained with FTIR and degrees of hydroxyapatite was detected from the result of XRD analysis. Copper releasing of the scaffolds in simulated body fluid in vitro were investigated with ICP analysis. As a result of this analysis the samples coated with alginate cross linked with copper, have more potential in stimulating angionesis compared with samples which were produced with based on copper doped bioactive glasses. At the same time, all copper releasing values of scaffolds were remained below the limit of cytotoxic effects of copper (58 ppm) in human body. The results showed that three dimensional composite scaffolds could be promising candidates for bone tissue engineering applications.