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Orthopaedic Proceedings
Vol. 100-B, Issue SUPP_15 | Pages 17 - 17
1 Nov 2018
Dalgarno K Benning M Partridge S Tulah A Ahmed S Dickinson A Genever P Pearson R Feichtinger G Loughlin J Ferreira-Duarte A
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This paper reports on a proof of concept project funded by the UK National Council for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs), with the aim of developing an in vitro model to recapitulate the human osteoarthritic joint, based on a multiple human cell type co-culture system, for research and drug development in OA. The targets were: (i) the development of a cell culture platform that could produce a mixed stable cell culture of cell types that represent the key components of the human joint: synoviocytes – type I and type II; osteoblasts; osteoclasts; chondrocytes/cartilage or cartilage-like matrix; adipocytes; and immune cells. (ii) demonstration of cell phenotype stability and viability for at least 72 hours. In order to establish the cell culture platform we have developed an eight-channel cell printer, capable of accurately and reliably printing the required cell types to create osteochondral and synovial cell types within a transwell system. Two different sets of cells have been developed and processed using the cell printer: a set based on using an immortalised hTERT MSC line to create osteoblasts, chondrocytes and adipocytes, with commercial cells lines providing the other cell types, and a set obtained from tissue excised during orthopaedic surgery. This gives both a repeatable set of cells with which to undertake mode of action studies, and a bank of cell sets which will be representative of different stages of osteoarthritis. The co-cultures have been immunohistochemically assessed in order to demonstrate maintenance of phenotype.


Orthopaedic Proceedings
Vol. 100-B, Issue SUPP_14 | Pages 115 - 115
1 Nov 2018
Müller S Nicholson L Jone E Dickinson A Dalgarno K Wang X
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Mesenchymal stromal cells (MSCs) are widely used in clinical trials for the treatment of many bone defects. Apatite-wollastonite glass ceramic (A-W) is an osteoconductive biomaterial shown to be compatible with MSCs. This is the first study comparing the osteogenic potential of two MSC populations, heterogeneous plastic adherence MSCs (PA-MSCs) and CD271-enriched MSCs (CD271-MSCs), when cultured on A-W 3D scaffold. The paired MSC populations were assessed for their attachment, growth kinetics and ALP activity using confocal or scanning electron microscopy and the quantifications of DNA contents and p-nitrophenyl (pNP) production. While the PA-MSCs and CD271-MSCs had similar expansion and tri-lineage differentiation capacity during standard 2D culture, they showed different proliferation kinetics when seeded on the A-W scaffolds. PA-MSCs displayed a well-spread attachment with more elongated morphology compared to CD271-MSCs, signifying a different level of interaction between the cell populations and the scaffold surface. PA-MSCs also fully integrated into the scaffold surface and showed a stronger propensity for osteogenic differentiation on the A-W scaffold as indicated by higher ALP activity than CD271-MSCs. Furthermore, A-W scaffold seeded uncultured bone marrow mononuclear cells (BM-MNCs) demonstrated a higher proliferation rate and greater ALP activity compared to freshly isolated CD271-enriched BM-MNCs. Our findings suggest that enrichment of CD271-positive population is not beneficial for osteogenesis when the cells are seeded on A-W scaffold. Furthermore, unselected heterogeneous MSCs or BM-MNCs are more promising for A-W scaffold-based bone regeneration, providing novel insight with potential clinical implications in regenerative medicine for bone defects using an innovative tissue engineering approach.


Orthopaedic Proceedings
Vol. 95-B, Issue SUPP_13 | Pages 23 - 23
1 Mar 2013
Wright K Morrison R Dalgarno K Birch M McCaskie A
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hMSC cultures were prepared from osteoarthritic patients. Silicone elastomer (PDMS) culture surfaces of varying degrees of stiffness (1:10, 1:30 and 1:50 PDMS, tissue culture plastic and glass) were investigated in isolation and in combination with differentiation media. CD marker expressions of ‘stemness’ were investigated. RNA expression changes in OA-hMSCs and non-OA-hMSCs were also investigated for a panel of genes (inclusive of ‘stemness-’ and osteogenic-linked genes, FKBP5 and osteomodulin).