Enamel derived matrix (EMD) increases plasma resistin in vivo and resistin secretion in cells of mesenchymal origin in vitro

S162

Abstracts / Bone 48 (2011) S161–S172

matched WTs. Thus increased bone remodelling in CD59−/− may be at the expense of bone quality. Our findings signify an important homeostatic role for CD59 in bone. Our findings could have important implications in understanding pathological mechanisms involved in osteoporosis, bone fractures and the early metastatic spread of breast cancer. This article is part of a Special Issue entitled ECTS 2011. Disclosure of interest: None declared.

doi:10.1016/j.bone.2011.03.357

PP198-T Enamel derived matrix (EMD) increases plasma resistin in vivo and resistin secretion in cells of mesenchymal origin in vitro M.V. Obregon-Whittle a, A.K. Stunes b, S. Almqvist c, G. Kvalheim d, U. Syversen b,e, J. Reseland a a Department of Biomaterials, Faculty of Dentistry, University of Oslo, Oslo, Norway b Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway c Department of Biomaterials, Sahlgrenska Academy at University of Gothenburg, Göteborg, Sweden d Department of Cellular Therapy, Radiumhospitalet, Oslo University Hospital, Oslo, Norway e Department of Endocrinology, St. Olavs University Hospital, Trondheim, Norway Abstract: Enamel matrix derivative (purified extract of proteins from pig enamel matrix, EMD) is reported to promote proliferation, migration, adhesion and differentiation of cells associated with healing periodontal tissues in vivo. EMD induces local growth factor and cytokine expression in the treated tissues and is used to induce the regenerative process that mimics odontogenesis. Several studies also show a positive influence of EMD on wound healing and bone formation, suggesting new applications. In this study we wanted to identify the effect of EMD on adipokine levels (resistin, adiponectin and leptin), as these adipokines are reported to directly influence the metabolism in various tissues, including bone. Rats were injected with either 2.0 mg EMD or saline (vehicle) intramuscularly every 3rd day for 14 days. Blood samples were taken before and after injections, and the level of resistin in rat plasma was measured by a resistin ELISA (B-Bridge International). Primary human cells of mesenchymal (osteoblasts (NHO) and mesenchymal stem cells (MSC)) and hematopoietic and hematopoietic (monocytes from human peripheral blood mononuclear cell (PBMC)) origin, were incubated with EMD. The levels of adipokines in cell culture medium were quantified by Lincoplex human adipocyte panel (leptin, adiponectin, resistin, IL-6 and TNFalpha) (Luminex). EMD injections significantly enhanced the circulating levels of resistin in rats. The secretion of resistin from MSC increased dramatically upon EMD stimulation for 1–7 days, and this effect was confirmed in NHO. Even though the main source of circulating resistin is believed to be monocytes/macrophages, we observed, no significant changes in the resistin levels in monocytes from PBMC treated with EMD. Our results indicate an important role for the adipokine resistin in generating the biological effect of EMD in mesenchymal tissues. This article is part of a Special Issue entitled ECTS 2011. Disclosure of interest: None declared.

doi:10.1016/j.bone.2011.03.358

PP199-S The efficacy of mechanical stimulation in acceleration the tibial rabbit fracture repair A. Shadmehr, A. Esteki, G.R. Oliaie Physical Therapy, Tehran University of Medical Sciences, Tehran, Islamic Republic of Iran Abstract: Introduction: It is a widespread understanding that the mechanical stimulation to the tissue within the fracture gap may accelerate bone healing. The aim of this study was to examine the influence of a well defined mechanical stimulation, externally applied to bone fixator, on acceleration of bone healing process. Materials/methods: A randomised, prospective in vivo animal study was designed. Eight skeletally mature white rabbits (2–2.5 kg) were randomly assigned to two groups. A four-pin, double bar external fixator was fixed to the right tibia. The mid shaft of the tibia was then osteotomized, leaving a 2 mm gap. External stimulation was performed in four rabbits with the use of a motor-driven actuator unit. The actuator could provide a well controlled axial cyclic compressive movement to the osteotomized gap with frequency of 1 Hz, amplitude of 0.5 mm and

duration of 15 minutes per day. The rabbits in experimental group received mechanical stimulations for 14 days and then were sacrificed. The animals in control group received no mechanical stimulation and were sacrificed on postoperative day 21.The mechanical properties of healed bone were determined using a standard three-point bending test. Results: Statistical significance was evaluated using nonparametric Mann–Whitney U test. Callus stiffness and maximum force up to failure were significantly superior on the stimulated group compared to the control (P < 0.05) after 14 and 21 days postoperative respectively. Conclusions: Current study demonstrates the effect of direct low frequency, low amplitude mechanical stimulation on acceleration of bone healing. This article is part of a Special Issue entitled ECTS 2011. Disclosure of interest: None declared. doi:10.1016/j.bone.2011.03.359

PP200-M Extension of the sheep sequence space using composite transcriptome assembly of RNA-Seq data in a sheep model for delayed bone healing C.E. Ott a, M. Jäger a,b, J. Grünhagen a, J. Hecht b,c, H. Schell b,d, S. Mundlos a,b,c, G.N. Duda b,d, P.N. Robinson a,b,c, J. Lienau b,d a Institute for Medical and Human Genetics, Charité-Universitätsmedizin Berlin, Germany b Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité-Universitätsmedizin Berlin, Germany c Max Planck Institute for Molecular Genetics, MPG, Charité-Universitätsmedizin Berlin, Germany d Julius Wolff Institute and Center for Musculoskeletal Surgery, Charité-Universitätsmedizin Berlin, Berlin, Germany Abstract: Background: The sheep is an important model organism for many types of medically relevant research, but molecular genetic experiments in the sheep have been limited by the lack of knowledge about ovine gene sequences. Results: Prior to our study, mRNA sequences for only 1,556 partial or complete ovine genes were publicly available. Therefore, we developed a composite de novo transcriptome assembly method for nextgeneration sequence data to combine known ovine mRNA and EST sequences, mRNA sequences from mouse and cow, and sequences assembled de novo from short read RNA-Seq data into a composite reference transcriptome, and identified transcripts from over 12 thousand previously undescribed ovine genes. Gene expression analysis based on these data revealed substantially different expression profiles in standard versus delayed bone healing in an ovine tibial osteotomy model. Hundreds of transcripts were differentially expressed between standard and delayed healing and between the time points of the standard and delayed healing groups. We used the sheep sequences to design quantitative RT-PCR assays with which we validated the differential expression of 25 genes that had been identified by RNA-seq analysis. A number of clusters of characteristic expression profiles could be identified, some of which showed striking differences between the standard and delayed healing groups. Gene Ontology (GO) analysis showed that the differentially expressed genes were enriched in terms including extracellular matrix, cartilage development, contractile fiber, and chemokine activity. Conclusions: Our results provide a first atlas of gene expression profiles and differentially expressed genes in standard and delayed bone healing in a largeanimal model and provide a number of clues as to the shifts in gene expression that underlie delayed bone healing. In the course of our study, we identified transcripts of 13,987 ovine genes, including 12,431 genes for which no sequence information was previously available. This information will provide a basis for future molecular research involving the sheep as a model organism. This article is part of a Special Issue entitled ECTS 2011. Disclosure of interest: None declared. doi:10.1016/j.bone.2011.03.360

PP201-T The effect of a single zoledronic acid 5 mg on periprosthetic bone loss after non cemented primary total hip arthroplasty A quantitative analysis using DXA E.J. Nieto a, J.E. Uzcategui b, D.J. Saavedra b, J.R. Salinas c and Grupo de Investigacion del Metabolismo Oseo (GIMO) a Ortogeriatria, Instituto Clinico Medico Quirurgico, Bolivarian Republic of Venezuela b Ortopedia y Traumatologia, Universidad de Los Andes, Bolivarian Republic of Venezuela c Medicina Nuclear, Instituto de Osteoporosis y Densitometria Osea, Merida, Bolivarian Republic of Venezuela