Biomaterials and scaffolds for tissue engineering

Every day thousands of surgical procedures are performed to replace or repair tissue that has been damaged through disease or trauma. The developing field of tissue engineering (TE) aims to regenerate damaged tissues by combining cells from the body with highly porous scaffold biomaterials, which act as templates for tissue regeneration, to guide the growth of new tissue. This article describes the functional requirements, and types, of materials used in developing state of the art of scaffolds for tissue engineering applications. Furthermore, it describes the challenges and where future research and direction is required in this rapidly advancing field.</p

Visualising feasible operating ranges within tissue engineering systems using a "windows of operation" approach: A perfusion-scaffold bioreactor case study.

Tissue engineering approaches to developing functional substitutes are often highly complex, multivariate systems where many aspects of the biomaterials, bio-regulatory factors or cell sources may be controlled in an effort to enhance tissue formation. Furthermore, success is based on multiple performance criteria reflecting both the quantity and quality of the tissue produced. Managing the trade-offs between different performance criteria is a challenge. A "windows of operation" tool that graphically represents feasible operating spaces to achieve user-defined levels of performance has previously been described by researchers in the bio-processing industry. This paper demonstrates the value of "windows of operation" to the tissue engineering field using a perfusion-scaffold bioreactor system as a case study. In our laboratory, perfusion bioreactor systems are utilised in the context of bone tissue engineering to enhance the osteogenic differentiation of cell-seeded scaffolds. A key challenge of such perfusion bioreactor systems is to maximise the induction of osteogenesis but minimise cell detachment from the scaffold. Two key operating variables that influence these performance criteria are the mean scaffold pore size and flow-rate. Using cyclooxygenase-2 and osteopontin gene expression levels as surrogate indicators of osteogenesis, we employed the "windows of operation" methodology to rapidly identify feasible operating ranges for the mean scaffold pore size and flow-rate that achieved user-defined levels of performance for cell detachment and differentiation. Incorporation of such tools into the tissue engineer's armoury will hopefully yield a greater understanding of the highly complex systems used and help aid decision making in future translation of products from the bench top to the market place. Biotechnol. Bioeng. © 2012 Wiley Periodicals, Inc.</p

Influence of shear stress in perfusion bioreactor cultures for the development of three-dimensional bone tissue constructs: a review.

Bone tissue engineering aims to generate clinically applicable bone graft substitutes in an effort to ease the demands and reduce the potential risks associated with traditional autograft and allograft bone replacement procedures. Biomechanical stimuli play an important role under physiologically relevant conditions in the normal formation, development, and homeostasis of bone tissue--predominantly, strain (predicted levels in vivo for humans με) caused by physical deformation, and fluid shear stress (0.8-3 Pa), generated by interstitial fluid movement through lacunae caused by compression and tension under loading. Therefore, in vitro bone tissue cultivation strategies seek to incorporate biochemical stimuli in an effort to create more physiologically relevant constructs for grafting. This review is focused on collating information pertaining to the relationship between fluid shear stress, cellular deformation, and osteogenic differentiation, providing further insight into the optimal culture conditions for the creation of bone tissue substitutes.</p

Visualising feasible operating ranges within tissue engineering systems using a "windows of operation" approach: A perfusion-scaffold bioreactor case study.

Tissue engineering approaches to developing functional substitutes are often highly complex, multivariate systems where many aspects of the biomaterials, bio-regulatory factors or cell sources may be controlled in an effort to enhance tissue formation. Furthermore, success is based on multiple performance criteria reflecting both the quantity and quality of the tissue produced. Managing the trade-offs between different performance criteria is a challenge. A "windows of operation" tool that graphically represents feasible operating spaces to achieve user-defined levels of performance has previously been described by researchers in the bio-processing industry. This paper demonstrates the value of "windows of operation" to the tissue engineering field using a perfusion-scaffold bioreactor system as a case study. In our laboratory, perfusion bioreactor systems are utilised in the context of bone tissue engineering to enhance the osteogenic differentiation of cell-seeded scaffolds. A key challenge of such perfusion bioreactor systems is to maximise the induction of osteogenesis but minimise cell detachment from the scaffold. Two key operating variables that influence these performance criteria are the mean scaffold pore size and flow-rate. Using cyclooxygenase-2 and osteopontin gene expression levels as surrogate indicators of osteogenesis, we employed the "windows of operation" methodology to rapidly identify feasible operating ranges for the mean scaffold pore size and flow-rate that achieved user-defined levels of performance for cell detachment and differentiation. Incorporation of such tools into the tissue engineer's armoury will hopefully yield a greater understanding of the highly complex systems used and help aid decision making in future translation of products from the bench top to the market place. Biotechnol. Bioeng. © 2012 Wiley Periodicals, Inc.</p

Influence of a novel calcium-phosphate coating on the mechanical properties of highly porous collagen scaffolds for bone repair.

Lyophilised collagen scaffolds have shown enormous potential in tissue engineering in a number of areas due to their excellent biological performance. However, they are limited for use in bone tissue engineering due to poor mechanical properties. This paper discusses the development of a calcium-phosphate coating for collagen scaffolds in order to improve their mechanical properties for bone tissue engineering. Pure collagen scaffolds produced in a lyophilization process were coated by immersing them in sodium ammonium hydrogen phosphate (NaNH(4)HPO(4)) followed by calcium chloride (CaCl(2)). The optimal immersing sequence, duration, as well as the optimal solution concentration which facilitated improved mechanical properties of the scaffolds was investigated. The influence of the coating on composition, structural and material properties was analysed. This investigation successfully developed a novel collagen/calcium-phosphate composite scaffold. An increase in the mechanical properties of the scaffolds from 0.3 kPa to up to 90 kPa was found relative to a pure collagen scaffold, while the porosity was maintained as high as 92%, indicating the potential of the scaffold for bone tissue engineering or as a bone graft substitute

The effect of bone microstructure on the initiation and growth of microcracks.

Osteonal bone is often compared to a composite material and to metals as discontinuities within the material may provide sites of stress concentration for crack initiation and serve as barriers to crack growth. However, little experimental data exist to back up these hypotheses. Fluorescent chelating agents were applied at specific intervals to bone specimens fatigue tested in cyclic compression at a stress range of 80 MPa. The failed specimens were sectioned and labelled microcracks identified using UV epifluorescence microscopy. Microcrack lengths were measured and their relationship to cement lines surrounding secondary osteons recorded. Microcrack length at the time of encountering a cement line was also measured. Microcracks of less than 100mum stopped growing when they encountered a cement line. Microcracks of greater than 100mum in length continued to grow after encountering a cement line surrounding an osteon. Only microcracks greater than 300mum in length were capable of penetrating osteons and these microcracks were the only ones which were observed to cause failure in the specimen. These experimental data support the hypothesis that secondary osteons act as barriers to crack propagation in compact bone. However, it shows that this microstructural barrier effect is dependent on the crack length at the time of encountering an osteon. For the vast majority of cracks, osteons act as barriers to growth but for the minority of cracks that are long enough and do break through the cement line, an osteon may actually act as a weakness in the bone and facilitate crack propagation

Staphylococcus aureus protein A causes osteoblasts to hyper-mineralise in a 3D extra-cellular matrix environment.

Osteomyelitis is an inflammatory bone infection that is caused most commonly by the opportunistic pathogen Staphylococcus aureus. Research into staphylococcal induced bone infection is typically conducted using traditional 2D in vitro culture settings, which is not fully representative of the dynamic in vivo environment. In this study we utilised a collagen glycosaminoglycan scaffold, previously developed for bone tissue engineering, as a representative 3D model of infection. The scaffold resisted degradation and retained its pore structure, which is important for cellular function and survival, when seeded with both cells and bacteria. Using this model, we showed that in the presence of S. aureus, osteoblast proliferation was reduced over 21 days. Interestingly however these cells were more metabolically active compared to the uninfected cells and demonstrated increased mineralisation. Protein A (SpA) is a virulence factor found on the surface of S. aureus and has been shown to interact with osteoblasts. When SpA was removed from the surface of S. aureus, the osteoblasts show comparable activity with the uninfected cells-demonstrating the importance of SpA in the interaction between bone cells and S. aureus. Our results suggest that infected osteoblasts are capable of over-compensating for bone loss and bone destruction by increasing mineralisation in a 3D environment, key elements required for ensuring bone strength. It also reinforces our previously established result that S. aureus SpA is a critical mediator in osteomyelitis and might be a potential novel drug target to treat osteomyelitis by preventing the interaction between S. aureus and osteoblasts.</p

Bone as a composite material: the role of osteons as barriers to crack growth in compact bone.

This article summarises a number of studies in the area of bone microdamage which were carried out in our laboratory over the past 5 years. A technique was developed to label microcracks during mechanical testing. Fluorescent chelating agents were applied at intervals to bone specimens fatigue tested in cyclic compression until failure occurred. Microcrack densities were measured and microcrack length at the time of encountering the cement line surrounding an osteon was also recorded. Microcracks were shown to develop rapidly during the first stage of testing but then further accumulation of cracks did not occur until the period just before failure. The majority of microcracks were found in interstitial bone and did not penetrate cement lines. Only microcracks greater than 300 μm in length were found to be capable of penetrating osteons. This work provides experimental data to support the hypothesis that secondary osteons act as barriers to crack propagation in compact bone

An improved labelling technique for monitoring microcrack growth in compact bone.

Fatigue-induced damage plays an important role in bone remodelling and in the formation of stress and fragility fractures. Recently, a technique has been developed (Lee, T.C. et al., Sequential labelling of microdamage in bone using chelating agents. Journal of Orthopedic Research, 18 (2000) 322-325) which allows microcrack growth in trabecular bone to be monitored by the application of a series of chelating fluorochromes, however, some limitations were identified with the process. The aims of this study were to refine the method of detection using these agents in order to determine the optimal sequence of application and the optimal concentrations which allowed all the agents to fluoresce equally brightly using UV epifluorescence. A chemical analysis process, ion chromatography, followed by validation tests on bone samples showed that the optimal sequence of application and concentration of each agent was alizarin complexone (0.0005 M) followed by xylenol orange (0.0005 M), calcein (0.0005 M) and calcein blue (0.0001 M). A fifth agent, oxytetracycline was excluded from the study after recurring problems were found with its ability to chelate exposed calcium when applied in sequence with the other agents. This work has developed a sequential labelling technique, which allows for microcrack propagation during fatigue testing of bone specimens to be monitored without the problem of chelating agent substitution occurring

Articulation inspired by nature: a review of biomimetic and biologically active 3D printed scaffolds for cartilage tissue engineering

In the human body, articular cartilage facilitates the frictionless movement of synovial joints. However, due to its avascular and aneural nature, it has a limited ability to self-repair when damaged due to injury or wear and tear over time. Current surgical treatment options for cartilage defects often lead to the formation of fibrous, non-durable tissue and thus a new solution is required. Nature is the best innovator and so recent advances in the field of tissue engineering have aimed to recreate the microenvironment of native articular cartilage using biomaterial scaffolds. However, the inability to mirror the complexity of native tissue has hindered the clinical translation of many products thus far. Fortunately, the advent of 3D printing has provided a potential solution. 3D printed scaffolds, fabricated using biomimetic biomaterials, can be designed to mimic the complex zonal architecture and composition of articular cartilage. The bioinks used to fabricate these scaffolds can also be further functionalised with cells and/or bioactive factors or gene therapeutics to mirror the cellular composition of the native tissue. Thus, this review investigates how the architecture and composition of native articular cartilage is inspiring the design of biomimetic bioinks for 3D printing of scaffolds for cartilage repair. Subsequently, we discuss how these 3D printed scaffolds can be further functionalised with cells and bioactive factors, as well as looking at future prospects in this field