ECTS Abstracts (2015) 1 P173

Osteoblast Differentiation of Mesenchymal Stem Cells in Different 3D Culture Models: Issues in the Study of Bone Tumours

Anne-Laure Gamblin, Meadhbh Brennan, Jérôme Amiaud, Kevin Biteau, Dominique Heymann, Françoise Rédini, Valérie Trichet & Pierre Layrolle

INSERM UMR U957, Equipe Ligue 2012, Université de Nantes. Laboratoire de Physiopathologie de la Résorption Osseuse et Thérapie des Tumeurs Osseuses Primitives, Nantes, France.

Background: The antitumour potential of new molecules is tested first on tumour cell lines cultured on plastic (2D) and in different animal models. However, less than 10% of the molecules validated by preclinical tests are effective in clinical tests. To reduce this failure rate, an intermediate stage of testing molecules could be achieved in vitro 3D culture systems including the stroma on which cancer cells develop. In order to test new molecules targeting bone tumour cells, we develop 3D scaffold presenting some characteristics of the bone environment.

Methods: Mesenchymal stem cells (MSCs) derived from human bone marrow, were cultured either on a biphasic calcium phosphate ceramic (BCP) or nanofibers of polycaprolactone in osteogenic medium (100 nM dexamethasone, 250 μM ascorbic acid, 10 mM β-glycerophosphate). Cell viability studies, electron microscopy, immunohistochemistry (alkaline phosphatase, type I collagen and bone sialo protein), quantification of mineralisation (Alizarin Red) and collagen (Sirius Red) and studies gene expression (COL1A1, CBFA1, ALP, SOST, BSP, BMP2, OC) were performed.

Results: Viability of MSCs was observed on long culture times (30-90 days) and their differentiation to the osteoblast phenotype was confirmed on both 3D culture systems. The abundant presence of type I collagen and the presence of non-collagenous bone proteins were observed, indicating the synthesis of an osteoid extracellular matrix.

Conclusion: These culture systems have not resulted in a lamellar bone tissue in vitro, but an osteoid matrix was obtained and used to culture osteosarcoma or Ewing’s sarcoma cells.

Disclosure: The authors declared no competing interests. This work was supported by a grant (HEALTH-2009-1.4.2-241879) from the 7th Framework Programme of the European Commission REBORNE (Regenerating Bone Defects Using New Biomedical Engineering Approaches). A.L.G. is the recipient of a PhD fellowship from the regional council of “Pays de la Loire”.

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