Resorbable composite scaffolds for craniofacial bone tissue engineering

Aim: Bone loss associated with trauma, osteo-degenerative diseases and tumors has tremendous socioeconomic impact related to personal and occupation disability and health care costs. In the present climate of increasing life expectancy with an ensuing increase in bone-related injuries, orthopaedic surgery is undergoing a paradigm shift from bone-grafting to bone engineering, where a scaffold is implanted to provide adequate load bearing and enhance tissue regeneration. We aim to develop composite scaffolds for bone tissue engineering applications to replace the current gold standard of autografting. ---------- Methods: Medical grade polycaprolactone-tricalcium phosphate (mPCL/TCP) scaffolds (80/20 wt%) were custom made using fused deposition modelling to produce 1x1.5x2 cm sized implants for critical-sized pig cranial implantations, empty defects were used as a control. Autologous bone marrow stromal cells (BMSCs) were extracted and precultured for 2 weeks, dispersed within fibrin glue and injected during scaffold implantation. After 2 years, microcomputed tomography and histology were used to assess bone regenerative capabilities of cell versus cell-free scaffolds. ---------- Results: Extensive bone regeneration was evident throughout the entire scaffold. Clear osteocytes embedded within mineralised matrix and active osteoblasts present around scaffold struts were observed. Cell groups performed better than cell-free scaffolds. ---------- Conclusions: Bone regeneration within defects which cannot heal unassisted can be achieved using mPCL/TCP scaffolds. This is improved by the inclusion of autogenous BMSCs. Further work will include the inclusion of growth factors including BMP-2, VEGF and PDGF to provide multifunctional scaffolds, where the three-dimensional (3D) template itself acts as a biomimetic, programmable and multi-drug delivery device.

Ovine bone and marrow derived progenitor cells : isolation, characterization, and osteogenic potential

Recently, research has focused on bone marrow derived multipotent mesenchymal precursor cells (MPC) for their potential clinical use in bone engineering. Prior to clinical application, MPC-based treatment concepts need to be evaluated in preclinical, immunocompetent, large animal models. Sheep in particular are considered a valid model for orthopaedic and trauma related research. However, ovine MPC and their osteogenic potential remain poorly characterized. In the present study, ex vivo expanded MPC isolated from ovine bone marrow proliferated at a higher rate than osteoblasts (OB) derived from tibial compact bone as assessed using standard 2D culture. MPC expressed the respective phenotypic profile typical for different mesenchymal cell populations (CD14-/CD31-/CD45- /CD29+/CD44+/CD166+) and showed a multilineage differentiation potential. When compared to OB, MPC had a higher mineralization potential under standard osteogenic culture conditions and expressed typical markers such as osteocalcin, osteonectin and type I collagen at the mRNA and protein level. After 4 weeks in 3D culture, MPC constructs demonstrated higher cell density and mineralization, whilst cell viability on the scaffolds was assessed >90%. Cells displayed a spindle-like morphology and formed an interconnected network. Implanted subcutaneously into NOD/SCID mice on type I collagen coated polycaprolactone-tricalciumphosphate (mPCL-TCP) scaffolds, MPC presented a higher developmental potential than osteoblasts. In summary, this study provides a detailed in vitro characterisation of ovine MPC from a bone engineering perspective and suggests that MPC provide promising means for future bone disease related treatment applications.

Resorbable composite scaffolds for bone tissue engineering

Bone loss associated with trauma osteo-degenerative diseases and tumors has tremendous socioeconomic impact related to personal and occupation disability and health care costs. Bone grafting is often critical to surgical therapies. Autogenous bone is presently the preferred grafting material; however, this holds several disadvantages such as donor site morbidity. In the present climate of increasing life expectancy with an ensuing increase in bone-related injuries, orthopaedic surgery is undergoing a paradigm shift from bone-grafting to bone engineering, where a scaffold is implanted to provide adequate load bearing and enhance tissue regeneration. Our group at Queensland University of Technology (QUT) have developed, characterised and tested polycaprolactone/ tricalcium phosphate (PCL/TCP) composite scaffolds for low load-bearing bone defects. These scaffolds are being further developed for application in higher load bearing sites. Our approach emphasizes the importance of the biomaterials’ structural design, the scaffold architecture and structural and nutritional requirements for cell culture. These first-generation scaffolds made from medical grade PCL (mPCL) have been studied for more than 5 years within a clinical setting 1. This paper describes the application of second-generation scaffolds in small and large animal bone defect models and the ensuing bone regeneration as shown by histology and µCT.

Apatite-Polymer Composites for the Controlled Dual Delivery of BMP-2 and BMP-6 for Bone Tissue Engineering

The release of growth factors from tissue engineering scaffolds provides signals that influence the migration, differentiation, and proliferation of cells. The incorporation of a drug delivery platform that is capable of tunable release will give tissue engineers greater versatility in the direction of tissue regeneration. We have prepared a novel composite of two biomaterials with proven track records - apatite and poly(lactic-co-glycolic acid) (PLGA) – as a drug delivery platform with promising controlled release properties. These composites have been tested in the delivery of a model protein, bovine serum albumin (BSA), as well as therapeutic proteins, recombinant human bone morphogenetic protein-2 (rhBMP-2) and rhBMP-6. The controlled release strategy is based on the use of a polymer with acidic degradation products to control the dissolution of the basic apatitic component, resulting in protein release. Therefore, any parameter that affects either polymer degradation or apatite dissolution can be used to control protein release. We have modified the protein release profile systematically by varying the polymer molecular weight, polymer hydrophobicity, apatite loading, apatite particle size, and other material and processing parameters. Biologically active rhBMP-2 was released from these composite microparticles over 100 days, in contrast to conventional collagen sponge carriers, which were depleted in approximately 2 weeks. The released rhBMP-2 was able to induce elevated alkaline phosphatase and osteocalcin expression in pluripotent murine embryonic fibroblasts. To augment tissue engineering scaffolds with tunable and sustained protein release capabilities, these composite microparticles can be dispersed in the scaffolds in different combinations to obtain a superposition of the release profiles. We have loaded rhBMP-2 into composite microparticles with a fast release profile, and rhBMP-6 into slow-releasing composite microparticles. An equi-mixture of these two sets of composite particles was then injected into a collagen sponge, allowing for dual release of the proteins from the collagenous scaffold. The ability of these BMP-loaded scaffolds to induce osteoblastic differentiation in vitro and ectopic bone formation in a rat model is being investigated. We anticipate that these apatite-polymer composite microparticles can be extended to the delivery of other signalling molecules, and can be incorporated into other types of tissue engineering scaffolds.

Preparation of porous calcium phosphate ceramic scaffold in bone tissue engineering

Calcium Phosphate ceramic has been widely used in bone tissue engineering due to its excellent biocompatibility and biodegradability. However, low mechanical properties and biodegradability limit their potential applications. In this project, hydroxyapatite (HA) and calcium phosphate bioglass were used to produce porous tri-calcium phosphate (TCP) bio-ceramic scaffolds. It was found that porous TCP bioceramic could be obtained when 20wt percent bioglass addition and sintered in 1400 degrees celsius for 3 h. Significantly higher compressive strength (9.98 MPa) was achieved in the scaffolds as compared to those produced from tCP power (<3 MPa). The biocompatibility of the scaffold was also estimated.

Preparation of mesoporous bioglass coated zirconia scaffold for bone tissue engineering

Porous yttria-stabilized zirconia (YSZ) has been regarded as a potential candidate for bone substitute due to its high mechanical strength. However, porous YSZ is biologically inert to bone tissue. It is therefore necessary to introduce bioactive coatings onto the walls of the porous structures to enhance its bioactivity. In this study, porous YSZ scaffolds were prepared using a replication technique and then coated with mesoporous bioglass due to its excellent bioactivity. The microstructures were examined using scanning electron microscopy and the mechanical strength was evaluated via compression test. The biocompatibility and bioactivity were also evaluated using bone marrow stromal cell (BMSC) proliferation test and simulated body fluid test.

Mesoporous bioglass/silk composite scaffolds for bone tissue engineering

In the past 20 years, mesoporous materials have been attracted great attention due to their significant feature of large surface area, ordered mesoporous structure, tunable pore size and volume, and well-defined surface property. They have many potential applications, such as catalysis, adsorption/separation, biomedicine, etc. [1]. Recently, the studies of the applications of mesoporous materials have been expanded into the field of biomaterials science. A new class of bioactive glass, referred to as mesoporous bioactive glass (MBG), was first developed in 2004. This material has a highly ordered mesopore channel structure with a pore size ranging from 5–20 nm [1]. Compared to non-mesopore bioactive glass (BG), MBG possesses a more optimal surface area, pore volume and improved in vitro apatite mineralization in simulated body fluids [1,2]. Vallet-Regí et al. has systematically investigated the in vitro apatite formation of different types of mesoporous materials, and they demonstrated that an apatite-like layer can be formed on the surfaces of Mobil Composition of Matters (MCM)-48, hexagonal mesoporous silica (SBA-15), phosphorous-doped MCM-41, bioglass-containing MCM-41 and ordered mesoporous MBG, allowing their use in biomedical engineering for tissue regeneration [2-4]. Chang et al. has found that MBG particles can be used for a bioactive drug-delivery system [5,6]. Our study has shown that MBG powders, when incorporated into a poly (lactide-co-glycolide) (PLGA) film, significantly enhance the apatite-mineralization ability and cell response of PLGA films. compared to BG [7]. These studies suggest that MBG is a very promising bioactive material with respect to bone regeneration. It is known that for bone defect repair, tissue engineering represents an optional method by creating three-dimensional (3D) porous scaffolds which will have more advantages than powders or granules as 3D scaffolds will provide an interconnected macroporous network to allow cell migration, nutrient delivery, bone ingrowth, and eventually vascularization [8]. For this reason, we try to apply MBG for bone tissue engineering by developing MBG scaffolds. However, one of the main disadvantages of MBG scaffolds is their low mechanical strength and high brittleness; the other issue is that they have very quick degradation, which leads to an unstable surface for bone cell growth limiting their applications. Silk fibroin, as a new family of native biomaterials, has been widely studied for bone and cartilage repair applications in the form of pure silk or its composite scaffolds [9-14]. Compared to traditional synthetic polymer materials, such as PLGA and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), the chief advantage of silk fibroin is its water-soluble nature, which eliminates the need for organic solvents, that tend to be highly cytotoxic in the process of scaffold preparation [15]. Other advantages of silk scaffolds are their excellent mechanical properties, controllable biodegradability and cytocompatibility [15-17]. However, for the purposes of bone tissue engineering, the osteoconductivity of pure silk scaffolds is suboptimal. It is expected that combining MBG with silk to produce MBG/silk composite scaffolds would greatly improve their physiochemical and osteogenic properties for bone tissue engineering application. Therefore, in this chapter, we will introduce the research development of MBG/silk scaffolds for bone tissue engineering.

Scaffolds for growth factor delivery as applied to bone tissue engineering

There remains a substantial shortfall in treatment of severe skeletal injuries. The current gold standard of autologous bone grafting from the same patient, has many undesirable side effects associated such as donor site morbidity. Tissue engineering seeks to offer a solution to this problem. The primary requirements for tissue engineered scaffolds have already been well established, and many materials, such as polyesters, present themselves as potential candidates for bone defects; they have comparable structural features, but they often lack the required osteoconductivity to promote adequate bone regeneration. By combining these materials with biological growth factors; which promote the infiltration of cells into the scaffold as well as the differentiation into the specific cell and tissue type, it is possible to increase the formation of new bone. However cost and potential complications associated with growth factors means controlled release is an important consideration in the design of new bone tissue engineering strategies. This review will cover recent research in the area of encapsulation and release of growth factors within a variety of different polymeric scaffolds.

Preparation of mesoporous bioglass coated zirconia scaffold for bone tissue engineering

Porous yttria-stabilized zirconia (YSZ) has been regarded as a potential candidate for bone substitute due to its high mechanical strength. However, porous YSZ is biologically inert to bone tissue. It is therefore necessary to introduce bioactive coatings onto the walls of the porous structures to enhance its bioactivity. In this study, porous YSZ scaffolds were prepared using a replication technique and then coated with mesoporous bioglass due to its excellent bioactivity. The microstructures were examined using scanning electron microscopy and the mechanical strength was evaluated via compression test. The biocompatibility and bioactivity were also evaluated using bone marrow stromal cell (BMSC) proliferation test and simulated body fluid test.