INTRODUCTION: Medial open-wedge HTO is an alternative in the treatment of medial knee OA for the young and active patient. However this technique leaves an open gap that requires stable fixation to achieve bony healing. As a bone substitute injectable calcium-phosphate-cements could be an alternative to autograft.

MATERIAL AND METHODS: Biomechanical testings were performed on open wedge HTO to investigate load to failure and displacement after cyclic loading (viscous and/or damaged material response). A medial 10 mm open-wedge osteotomy was performed on 7 pairs of composite (Sawbone) left tibiaes, and 8 pairs of preserved cadaver tibiaes. Osteosynthesis where performed with the Dynafix system. In half of the bones the gap was filled with 15 g of Calcibon®. The composite tibiaes were loaded at a ramp speed of 20 mm/min and failures of the constructs were recorded visually. On the cadaver tibiaes, cyclical loading were performed with a maximum load of 2250 N.

RESULTS: Filling of the gap with Calcibon® resulted in significant different load-to-failure patterns with failure at 10.2 kN compared to 2.7 kN in the group without Calcibon®. Displacement at the end of cyclical loading was 1.2 mm in the group with Calcibon® and 2.7 mm in the group without Calcibon®. This difference also was significant.

CONCLUSION: The injectable calcium-phosphate-cement Calcibon® enhances primary stability during load to failure and during cyclical loading in open wedge osteotomies on proximal tibia. Clinical studies are performed to investigate whether Calcibon® has any clinical advantage on wedge healing and stability.

Background: The next generation of biomaterial surfaces for use in orthopaedic surgery will be functionalised to promote osteogenesis. This will be achieved in part by the stable addition of functional bioactive molecules onto the biomaterial surface. Heparan sulphate is a complex glycosaminoglycan (GAG) that displays cell and tissue specific differences in size and levels of sulphation. It is this heterogeneity that underlies the numerous biological roles of heparan sulphate, including binding of growth factors and proteases. Findings by others have shown that the addition of heparan sulphate proteoglycans stimulate osteoblast differentiation in vitro.

Aims: To characterise heparan sulphate structures that support and enhance osteogenesis and have the potential for tissue engineering.

Experiment 1. In order to further investigate the role of heparan-sulphate proteoglycans (HSPGs) in osteogenesis we supplemented cultures of differentiating rat osteo-blasts with sodium chlorate (an inhibitor of the enzyme that sulphates GAG chains) or 4-methylumbelliferyl-b-D-xyloside, BDX (an artificial acceptor of GAG chain synthesis). Interestingly the addition of chlorate to our culture system significantly stimulated alkaline phosphatase levels and increased the area of Von Kossa stained bone-like nodules. Whereas, when BDX was added to differentiating rat osteoblasts there was no increase in alkaline phosphatase activity or nodule area.

Experiment 2. Further characterisation of the HSPGs in chlorate treated osteoblasts showed that whilst they were less sulphated than untreated cells (as shown by low salt elution from an anion exchange chromatography column) they were much more abundant. These observations led us to hypothesise that less sulphated forms of heparan sulphate may well stimulate osteo-blast differentiation.

Experiment 3. To test this hypothesis we took the fully sulphated form of heparan sulphate, heparin and selectively desulphated it using DMSO/methanol (9:1) at 97°C and specifically N-resulphated or N-acetylated. These partially desulphated heparins were then added to osteoblasts cultured under osteogenic conditions. Quantification of bone nodule formation showed that specifically desulphated heparin significantly increased mineralised areas compared to controls whilst the addition of heparin inhibited osteogenesis. How these modified heparan sulphates exert their effect on bone cells is unknown, but a well characterised role of heparan sulphate is the support of FGF signalling. In preliminary studies we have shown through the activation of p42/44 MAPK and proliferation assays that the modified heparan sulphates are able to support FGF signalling in bone cells.

Experiment 4. Currently were are attaching our desulphated heparin to biomaterial scaffolds and examining osteoblast attachment and migration/ingrowth in cell culture.

Conclusion: We have isolated heparan sulphate chains that demonstrate osteogenic properties and have the potential for enhancing biological interactions of orthopaedic implant materials.

Enzymes that breakdown components of the extracellular matrix (ECM) are of fundamental importance, not only in normal bone physiology but also in pathological processes. For instance the temporal and spatial distribution of proteoglycans is not only critical for the mineralisation of bone but is also believed to be responsible for dictating the local bioavailability of glycosaminoglycan-binding growth factors. A sub-family of the ADAMs (a disintegrin and metalloproteinase) has been identified, that contains thrombospondin-like motifs (ADAMTS), and ADAMTS1, 4 and 5 have recently been shown to cleave the major proteoglycan of cartilage, aggrecan. We propose that ADAMTS family members play a novel role in regulating osteoblast function by determining the distribution of proteoglycan in bone.

RT-PCR and Northern blotting experiments have shown expression of ADAMTS1, 3, 4 and 5 in primary rat osteoblasts and in the osteosarcoma cell lines, MG63, TE85 and SaOS-2. ADAMTS1 transcript levels increased with time in primary rat osteoblasts driven by dexamethasone, beta-glycerophosphate and ascorbic acid phosphate to produce bone-like nodules in vitro. Whereas levels of ADAMTS4 that were initially raised in this culture system then became undetectable as mineralisation proceeded.

Since we are interested in the relationship between the osteoblast and matrix molecules, we plated TE85 cells onto an ECM synthesised by MG63 cells and isolated RNA at 1, 24 and 48 hours. Northern analysis showed a transient upregulation of mRNA for both ADAMTS1 and 5 at 1h that was reduced to control levels at 24 and 48h. Transcripts for ADAMTS1 and 3 were also upregulated in primary rat osteoblasts when seeded on ECM molecules like fibronectin and type I collagen for 48 hours. There was however no change in the expression levels of ADAMTS4 when plated on to any of the substrates at any of the time points tested.

These data suggests that cells of the osteoblast lineage express ADAMTS1, 3, 4 and 5 and that individual transcript levels can be regulated by ECM components. The focalised production of ADAMTS family members in response to matrix-derived and other cues may be an important part of bone formation and may have important implications for the way that cells of the osteoblast lineage interact with implant and other biomaterials.