Autogenous bone grafting limitations have motivated the development of Tissue-Engineered (TE) biomaterials that offer an alternative as bone void fillers. However, the lack of a blood supply within implanted constructs may result in avascular necrosis and construct failure. 1. The aim of this project was to investigate the potential of novel TE constructs to promote vascularisation and bone defect repair using two distinct approaches. In Study 1, we investigated the potential of a mesenchymal stem cell (MSC) and endothelial cell (EC) co-culture to stimulate pre-vascularisation of biomaterials prior to in vivo implantation. 2. In Study 2, we investigated the potential of TE hypertrophic cartilage to promote the release of angiogenic factors such as VEGF, vascular invasion and subsequent endochondral bone formation in an in vivo model. Collagen-only (Coll), collagen-glycosaminoglycan (CG) and collagen-hydroxyapatite (CHA) scaffolds were fabricated by freeze-drying. 3. , seeded with cells and implanted into critical-sized calvarial and femoral defects in immunocompetent rats. In Study 1, Coll and CG scaffolds were initially seeded with ECs, allowed to form capillary-like networks before the delayed addition of MSCs and continued culture prior to calvarial implantation. In Study 2, CG and CHA scaffolds were seeded with MSCs and cultured under chondrogenic and subsequent hypertrophic conditions to form a cartilage pre-cursor prior to calvarial and femoral implantation in vivo. MicroCT and histomorphometry quantification demonstrated the ability of both systems to support increased bone formation compared to controls. Moreover, the greatest levels of bone formation were observed in the CG groups, notably in those containing cartilage tissue (Study 2). Assessment of the immune response suggests the addition of MSCs promotes the polarisation of macrophages away from inflammation (M1) towards a pro-remodelling phenotype (M2). We have developed distinct collagen-based systems that promote vascularisation and ultimately enhance bone formation, confirming their potential as advanced strategies for bone repair applications

Introduction. Autologous fat grafting has favourable potential as a regenerative strategy and is the current gold-standard to repair large contour defects, as needed in breast reconstruction after mastectomy and traumatic soft tissue reconstruction. Clinically, there is a limit on the volume of lipoaspirate which can be utilised to repair a soft-tissue defect. Surgical complications are the result of poor structural fidelity of lipoaspirate and graft resorption as a filling material and are hindered further by poor graft vascularisation. This study aims to develop injectable lipoaspirate-derived adipose tissue grafts with enhanced biologically and clinically-admissible structural and functional properties adopting light photocrosslinking of unmodified lipoaspirate. Methods. Patient-derived lipoaspirate was harvested and crosslinked using novel photoinitiator and exposure to visible light (wavelength 450nm) in surgery, establishing bonds between extracellular matrix (ECM) proteins within the material. The degree of crosslinking was tuned (photoinitiator concentration, light exposure, light intensity) and covalent bond formation measured using mass spectrometry. To predict patient response, SWATH-MS was used to identify differences in patient ECM and crosslinked grafts were implanted in vivo using a subcutaneous mouse model. Functional vessel formation and resorption were quantified using micro-CT and tissue-remodelling was assessed via histology. Results. There was an increase in the relative abundance of covalent bonds present with increasing degree of crosslinking. When injected, crosslinked lipoaspirate had better shape fidelity compared with native lipoaspirate – demonstrated by a smaller fibre diameter. Crosslinked lipoaspirate remained viable over long term culture and resulted in more predictable resorption profiles when implanted in vivo. Conclusions. The crosslinking approach described here is tunable and functional across different patient samples. Improving the structural properties of lipoaspirate through minimal manipulation has clinical utility for the delivery of grafts with higher shape fidelity and therefore increased graft survival when implanted

Infection is a common complication of severe open fractures and compromises bone healing. The present standard of care is a two-stage approach comprising of initial placement of antibiotic-impregnated PMMA beads to control infection followed later by bone grafting. Although the systemic antibiotics and PMMA/antibiotic beads control the infection initially, there are often residual bacteria within the wound. After grafting and definitive closure, the implanted graft is placed in an avascular defect and could function as a nidus for infection. Bioactive porous polyurethane (PUR) scaffolds have been shown to improve bone healing by delivering recombinant human bone morphogenetic protein-2 (BMP-2) and reduce infection by delivering antibiotics. The release kinetics of the BMP-2 were an initial burst to recruit cells and sustained release to induce the migrating cells. The Vancomycin (Vanc) release kinetics were designed to protect the graft from contamination until vascularisation by having an initial burst and then remaining over the MIC for Staph. aueus for two months. In this study, PUR+BMP-2+Vanc scaffolds were first tested in a non-infected critical size rat femoral segmental defect and was found to perform comparably to PUR+BMP-2, thus indicating that Vanc did not hinder bone healing. PUR+BMP-2+Vanc scaffolds were subsequently evaluated in an infected critical size rat femoral segmental defect. The dual delivery approach resulted in significantly more new bone formation and infection control than both PUR+BMP-2 and the collagen+BMP-2 treatments. These data indicate that the dual-delivery strategy effectively protects the graft from infection during wound healing and regenerates more bone in contaminated defects. This moderately osteoconductive bone graft is capable of being injected, provides a more sustained release of BMP-2 than the collagen sponge, and can release antibiotics for over 8 weeks. The dual-delivery approach may improve patient outcomes of open fractures by protecting the osteoinductive graft from colonization until vascularization occurs

Autologous cell therapy using stem cells and progenitor cells is considered to be a popular approach in regenerative medicine for the repair and regeneration of tissue and organs. In orthopaedic practice, autologous cell therapy has become a major focus, particularly, as a feasible treatment for tendon injury. Tendons are dense connective tissue that bridge bone to muscle and transmit forces between muscle and bone to maintain mechanical movement. Tendons are poorly vascularised and have very little capacity to self-regenerate. Degeneration of tendon is often caused by injury. The pathogenesis of tendon injury, commonly known as tendinosis, is not an inflammatory condition but is secondary to degenerative changes, including disruption of the collagen matrix, calcification, vascularisation and adipogenesis. The aetiology of tendinosis is considered to be multifactorial and the pathogenesis is still unclear. Intrinsic factors such as a lack of blood and nutrition supply and extrinsic factors such as acute trauma and overuse injury caused by repetitive strain, have been implicated as contributors to the pathogenesis of tendinosis. More recent studies suggest that programmed tendon cell death (tenocyte apoptosis) may play a major role in the development of tendinosis. Such cellular abnormalities may influence the capacity of tendon to maintain its integrity. Traditional treatments such as anti-inflammatory drugs, steroid injections and physiotherapy are aimed at symptom relief and do not address the underlying pathological changes of degeneration. Here, we propose that autologous cell therapy may be an innovative and promising treatment for tendon injury. We will present evidence that suggest that autologous tendon cell therapy may be feasible to repair and regenerate tendon. We will also present data summarising the preclinical evaluation of autologous tendon cell therapy in animal models and the safety and tolerability of autologous tendon cell therapy in humans in studies, which are currently conducted at the Centre for Orthopaedic Research at the University of Western Australia

Currently, there is no animal model in which to evaluate the underlying physiological processes leading to the heterotopic ossification (HO) which forms in most combat-related and blast wounds. We sought to reproduce the ossification that forms under these circumstances in a rat by emulating patterns of injury seen in patients with severe injuries resulting from blasts. We investigated whether exposure to blast overpressure increased the prevalence of HO after transfemoral amputation performed within the zone of injury. We exposed rats to a blast overpressure alone (BOP-CTL), crush injury and femoral fracture followed by amputation through the zone of injury (AMP-CTL) or a combination of these (BOP-AMP). The presence of HO was evaluated using radiographs, micro-CT and histology. HO developed in none of nine BOP-CTL, six of nine AMP-CTL, and in all 20 BOP-AMP rats. Exposure to blast overpressure increased the prevalence of HO.

This model may thus be used to elucidate cellular and molecular pathways of HO, the effect of varying intensities of blast overpressure, and to evaluate new means of prophylaxis and treatment of heterotopic ossification.

Cite this article: Bone Joint J 2015;97-B:572–6