Impaction allografting is a bone reconstruction technique currently used in lower limb revision arthroplasty. Demineralisation and addition of osteogenic protein-1 (OP-1) can improve the osteoinductivity of the allograft however recent reports indicate significant allograft resorption when it is combined with OP-1 during impaction. Our hypothesis was that hydroxyapatite (HA) and OP-1 could effectively replace demineralised allograft. The objective was to evaluate human mesenchymal stem cell (h-MSC) proliferation (tritiated thymidine incorporation, total DNA Hoechst 33258 and scanning electron microscopy) and osteogenic differentiation (alkaline phosphatase activity) in human demineralised bone matrix (h-DBM) and HA, with or without OP-1. Cell proliferation on HA+OP-1 was significantly higher compared to HA at all time points (p< 0.05) and to DBM alone (day 1, p=0.042; day 14, p< 0.001). Cell proliferation was higher in DBM+OP-1, at all time points compared to HA+OP-1 but only in absolute values. Cell differentiation was significantly higher in HA+OP-1 compared to HA (p< 0.05) but comparable to DBM alone. Differentiation was significantly higher on DBM+OP-1 at all time points compared to HA (p< 0.05) and to HA+OP-1 (p< 0.05). HA is a potential graft expander in impaction allografting. When combined with OP-1 is comparable to DBM alone and being non absorbable may support the impacted graft in the early stages after the administration of OP-1.

Impaction allografting is a bone tissue engineering technique currently used in lower limb reconstruction orthopaedic surgery. Our hypothesis was that biological optimisation can be achieved by demineralisation and addition of osteogenic protein-1(OP-1) to the allograft. The objective of our in vitro study was to evaluate human mesenchymal stem cell (MSC) proliferation (Alamar Blue assay, titrated thymidine assay, total DNA Hoechst 33258 and scanning electron microscopy) and osteogenic differentiation (alkaline phosphatase assay) in two types of impacted carrier, namely demineralised bone matrix (DBM) and insoluble collagenous bone matrix (ICBM), with or without OP-1. The objective in vivo was to compare the osteogenic potential of impacted DBM with or without OP-1, with that of impacted fresh frozen allograft (FFA), again with or without OP-1. DBM+OP-1 optimized osteoinduction and significantly improved (p< 0.05) proliferation and differentiation in comparison to the majority of all other graft preparation in vitro. In addition DBM+OP-1 was significantly superior, with regard to osteogenesis, compared to the impacted FFA alone (p< 0.001), FFA+OP-1 (p=0.01) and DBM alone (p=0.02) in vivo. We propose that partial demineralisation and addition of OP-1 provides a good method for improving the osteoinductive properties of fresh allograft currently used in the impaction grafting technique.

Aim: Mesenchymal stem cells (MSCñs) attach to hydroxyapaptite surfaces (HA) surfaces and given appropriate stimuli from human Bone Morphogenetic Protein 7 (OP -1), will differentiate into osteogenic cells. Our hypothesis is that combining HA/MSC/BMP-7 will provide a superior osseoinductive property compared to HA alone. Methods: Porous hydroxyapatite (74.6% porosity, and 0.65% closed porosity) loaded with MSCñs (2 x 105) were compared to samples loaded with rhBMP-7 (400 ng/0.1g HA,) of the same MSC concentration over a fourteen day period. Quantitative analysis (Cell proliferation, measured by total DNA and the Alamar blue assay and Cell differentiation- alkaline phosphatase activity) and qualitative (Light and Scanning Electron Microscopy) were performed. The Students T-test was performed. Ethical approval for the use of human tissue was obtained prior to experimentation Results: Cell proliferation as indicated by total DNA, and Alamar blue was signiþcantly enhanced (P< 0.05) in the BMP-7 loaded composite at all time points. ALP production and release was enhanced in loaded samples. ALP production per unit DNA was also enhanced in the loaded samples and was signiþ-cant at day fourteen. Conclusion: Results indicate that the loaded composites showed enhanced cell proliferation, and ALP production and release. SEM analysis also demonstrated enhanced cell attachment and an increase number of proliferative cells. Thus the HA/MSC/BMP-7 composite displayed superior osseo-inductive properties in comparison to the HA/MSC composite.