Gebäude 21, 1. OG
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The aim of the experiment was to establish a "Mouse Bone Induction Assay" and to compare this in vivo system with a cell culture method.The in vivo model is based on the information that implantation of bone matrix (demineralised bone) leads to the formation of new bone at sites other than bone (ectopic ossification) under specific conditions.The in vitro model used mouse osteoblasts in culture.As a consequence of the first experiments a permanent osteogenic cell line (MC3T3) had to be used in order to reach an experimental definition.The models were tested using a protein isolated from mouse embryonic tissue and a number of commercially available cytokines thought to influence the formation of bone as well as three extracts from bone matrix.In the "Mouse Bone Induction Assay" the following parameters were investigated: 45Caincorporation (Szintillationmeasurement of the explant and assessment on the basis of histological slides (Autoradiography and demonstration of mineralization collagen- and cartilage formation and presence of alkaline phosphatase by staining-techniques).The cell-culture assessment was based on the measurement of 45Ca-incorporation (autoradiographically) production of alkaline phosphatase in osteoblasts and in the culture medium,collagen production and staining for the occurrence of mineralization and collagen. The results of the investigation show a good correlation between animal experiments and cell culture experiments."False negative" results were not seen in the culture system.The number of "false positive" results was low.In both cases the embryonic protein resulted in the highest levels of mineralization, collagen formation and alkaline phosphatase activity.In the cytokine group FGF, EGF and PDGF effected low levels, IGF2,IL1a,IL1ß and 1L6 led tohigh levels in the measured parameters.As the result of the good correlation between in vivo and in vitro results the use of cell cultures can be recommended as the first step in a test protocol.