Impaired bone healing biology secondary to soft tissue deficits and chemotherapy contribute to non-union, fracture and infection following limb salvage surgery in Osteosarcoma patients. Approved bone healing augments such as recombinant human bone morphogenetic protein-2 (rhBMP-2) have great potential to mitigate these complications. rhBMP-2 use in sarcoma surgery is limited, however, due to concerns of pro-oncogenic signalling within the tumour resection bed. To the contrary, recent pre-clinical studies demonstrate that BMP-2 may induce Osteosarcoma differentiation and limit tumour growth. Further pre-clinical studies evaluating the oncologic influences of BMP-2 in Osteosarcoma are needed. The purpose of this study is to evaluate how BMP-2 signalling affects Osteosarcoma cell proliferation and metastasis in an active tumour bed. Two Osteosarcoma cell lines (143b and SaOS-2) were assessed for proliferative capacity and invasion. 143b and SaOS-2 cells were engineered to upregulate BMP-2. In vitro proliferation was assessed using a cell viability assay, motility was assessed with a scratch wound healing assay, and degree of osteoblastic differentiation was assessed using qRT-PCR of Osteoblastic markers (CTGF, ALP, Runx-2 and Osx). For in vivo evaluation, Osteosarcoma cells were injected into the intramedullary proximal tibia of immunocompromised (NOD-SCID) mice and local tumour growth and metastases were assessed using weekly bioluminescence imaging (BLI) and tumour volume measurements for 4–6 weeks. At the experimental end point we assessed radiographic tumour burden using ex-vivo micro-CT, as well as tibial and pulmonary gross and histologic pathology. SaOS-2 was more differentiated than 143b, with increased expression of Runx-2 (p = 0.009), Osx (p = 0.004) and ALP (p = 0.035). BMP-2 upregulation did not stimulate an osteoblast differentiation response in 143b, but stimulated an increase in Osx expression in SaOS-2 (p = 0.002). BMP-2 upregulation in 143b cells resulted in increased proliferation in vitro (p = 0.014), faster in vitro wound healing (p = 0.03), significantly increased tumour volume (p = 0.001) with enhanced osteolysis detected on micro-CT, but did not affect rates of lung metastasis (67% vs. 71%, BMP-2 vs. Control). BMP-2 over-expression in SaOS-2 cells reduced in vitro proliferation when grown in partial osteogenic-differentiation media (p < 0.001), had no effect on in vitro wound healing (p = 0.28), reduced in vivo SaOS-2 tumour burden at 6 weeks (photon counts, p < 0.0001), decreased tumour-associated matrix deposition as assessed by trabecular thickness (p = 0.02), and did not affect rates of lung metastasis (0% vs. 0%). Our results indicate BMP-2 signalling incites a proliferative effect on a poorly differentiated Osteosarcoma cell line (143b), but conditionally reduces proliferative capacity and induces a partial differentiation response in a moderately-differentiated Osteosarcoma cell line (SaOS-2). This dichotomous effect may be due to the inherent ability for Osteosarcoma cells to undergo BMP-2 mediated terminal differentiation. Importantly, these results do not support the clinical application of BMP-2 in Osteosarcoma limb salvage surgery due to the potential for stimulating growth of poorly differentiated Osteosarcoma cells within the tumour bed. Additional studies assessing the effects of BMP-2 in an immune-competent mouse model are ongoing

Impaired bone healing biology secondary to soft tissue deficits and chemotherapy contribute to non-union, fracture and infection following limb salvage surgery in Osteosarcoma patients. Approved bone healing augments such as recombinant human bone morphogenetic protein-2 (rhBMP-2) have great potential to mitigate these complications. rhBMP-2 use in sarcoma surgery is limited, however, due to concerns of pro-oncogenic signalling within the tumour resection bed. To the contrary, recent pre-clinical studies demonstrate that BMP-2 may induce Osteosarcoma differentiation and limit tumour growth. Further pre-clinical studies evaluating the oncologic influences of BMP-2 in Osteosarcoma are needed. The purpose of this study is to evaluate how BMP-2 signalling affects Osteosarcoma cell proliferation and metastasis in an active tumour bed. Two Osteosarcoma cell lines (143b and SaOS-2) were assessed for proliferative capacity and invasion. 143b and SaOS-2 cells were engineered to upregulate BMP-2. In vitro proliferation was assessed using a cell viability assay, motility was assessed with a scratch wound healing assay, and degree of osteoblastic differentiation was assessed using qRT-PCR of Osteoblastic markers (CTGF, ALP, Runx-2 and Osx). For in vivo evaluation, Osteosarcoma cells were injected into the intramedullary proximal tibia of immunocompromised (NOD-SCID) mice and local tumour growth and metastases were assessed using weekly bioluminescence imaging and tumour volume measurements for 4–6 weeks. At the experimental end point we assessed radiographic tumour burden using ex-vivo micro-CT, as well as tibial and pulmonary gross and histologic pathology. SaOS-2 was more differentiated than 143b, with significantly increased expression of the Osteoblast markers Osx (p = 0.004) and ALP (p = 0.035). BMP-2 upregulation did not stimulate an osteoblast differentiation response in 143b, but stimulated an increase in Osx expression in SaOS-2 (p = 0.002). BMP-2 upregulation in 143b cells resulted in increased proliferation in vitro (p = 0.014), faster in vitro wound healing (p = 0.03), significantly increased tumour volume (p = 0.001) with enhanced osteolysis detected on micro-CT, but did not affect rates of lung metastasis (67% vs. 71%, BMP-2 vs. Control). BMP-2 over-expression in SaOS-2 cells reduced in vitro proliferation when grown in osteogenic-differentiation media (p < 0.001), had no effect on in vitro wound healing (p = 0.28), reduced in vivo SaOS-2 tumour burden at 6 weeks (photon counts, p < 0.0001), decreased tumour-associated matrix deposition as assessed by trabecular thickness (p = 0.02), but did not affect rates of lung metastasis (0% vs. 0%). Our results indicate BMP-2 signalling incites a proliferative effect on a poorly differentiated Osteosarcoma cell line (143b), but conditionally reduces proliferative capacity and induces a partial differentiation response in a moderately-differentiated Osteosarcoma cell line (SaOS-2). This dichotomous effect may be due to the inherent ability for Osteosarcoma cells to undergo BMP-2 mediated terminal differentiation. Importantly, these results do not support the clinical application of BMP-2 in Osteosarcoma limb salvage surgery due to the potential for stimulating growth of poorly differentiated Osteosarcoma cells within the tumour bed. Additional studies assessing the effects of BMP-2 in an immune-competent mouse model are ongoing

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

Treatment of segmental bone loss remains a major challenge in orthopaedic surgery. This study evaluated the healing potential of a series of highly porous tissue engineering scaffolds with the current clinical gold standard. We compare healing of collagen-glycosaminoglycan (CG) and collagen micro-hydroxyapatite (CHA) scaffolds, with and without recombinant bone morphogenetic protein-2 (BMP2), with autogenous bone graft (ABG) in the healing of a 15mm rabbit radius defect, which were filled with either CG scaffold, CHA scaffold, CG-BMP2, CHA-BMP2 or ABG. Serial radiographs and micro-computed tomography (µCT) at six week radiographs demonstrated complete defect bridging with callus using CHA and CG-BMP2 while the CHA-BMP2 was already in an advanced state of healing with cortical remodeling. By sixteen weeks CHA, CG-BMP2 and ABG all had advanced healing with cortical remodeling while CHA-BMP2 had complete anatomic healing. Quantitative histomorphometry values demonstrated similarly high healing levels of healing in CHA, CG-BMP2 and ABG with highest overall values in the CHA-BMP2 group. Thus, treatment of a critical sized, weight bearing, rabbit radius defect with a CHA scaffold can result in full cortical bridging with medullary cavity development. In addition, a CHA-BMP2 combination can result in fully mature, anatomic healing. The use of an off-the-shelf CHA scaffold for direct surgical placement into a defect site may be an effective bone graft substitute in the treatment of skeletal defects. The ease of manufacture, storage and peri-operative preparation may offer an alternative to traditional strategies, as well as to more recent BMP2 devices. This study provides clear evidence that CHA scaffolds can perform as well as autogenous bone grafts and supports their use as a viable alternative. Where the use of BMP2 may be desirable, these materials provide an ideal delivery mechanism and using a very low (near physiological) dose, healing superior to autogenous graft may be achieved

Implant-associated infection is a major source of morbidity in orthopaedic surgery. There has been extensive research into the development of materials that prevent biofilm formation, and hence, reduce the risk of infection. Silver nanoparticle technology is receiving much interest in the field of orthopaedics for its antimicrobial properties, and the results of studies to date are encouraging. Antimicrobial effects have been seen when silver nanoparticles are used in trauma implants, tumour prostheses, bone cement, and also when combined with hydroxyapatite coatings. Although there are promising results with in vitro and in vivo studies, the number of clinical studies remains small. Future studies will be required to explore further the possible side effects associated with silver nanoparticles, to ensure their use in an effective and biocompatible manner. Here we present a review of the current literature relating to the production of nanosilver for medical use, and its orthopaedic applications.

Cite this article: Bone Joint J 2015; 97-B:582–9.