An ex vivo mouse mandible culture model to study inflammatory bone disease

Project Background

Periodontal diseases are a group of chronic inflammatory diseases of the supporting tissues of the teeth. Chronic periodontitis is the major cause of tooth loss in the developed world with approximately 54% of the UK adult population are affected by the condition. The consequences of the disease extend beyond oral health as bacterial-induced periodontal inflammation represents a risk factor for heart attacks, bacterial pneumonia and stroke. Destructive forms of periodontal disease result in degradation of the alveolar bone, thought to result from host mediated reactions to pathogenic factors secreted by the bacteria, such as lipopolysaccharide (LPS). Current systems for understanding periodontal disease require animal models to ensure the inclusion of cell/cell and host/bacterial interactions are present.

Why we funded it

This Project Grant aims to reduce the number of animals needed to study periodontal tissue and disease by developing an ex vivo prototype rodent mandibular organ culture system.

In vivo studies are currently used to investigate pathogenesis of periodontal diseases and for testing novel therapeutic treatments. An in vivo study of periodontal disease with five time points uses approximately five animals per time point and a control, thus significant numbers of animals are required to obtain statistically robust data. In vitro studies using rodent cells can also be used but rodent mandibles contain only a small amount of tissue. For enough viable cells to be obtained to create reproducible assays a study may require between 20 and 25 animals. The use of immortalised cell lines overcomes this, but these cells may not behave in the same way as primary, unmodified cells. In addition, cells may behave differently when cultured in 3D in tissue-like environments rather than in dishes as this is more representative of their natural environment. The prototype ex vivo culture system in this Project Grant aims to obtain multiple slices of whole tissue from each mandible, which would be cultured as separate entities. Obtaining multiple slices from each mandible has the potential to reduce the number of animals required by up to 80% per experiment whilst still providing a representative tissue environment.

Research Methods

This prototype ex vivo mandible model aims to maintain the cellular activity, tissue architecture and physiology of the periodontal ligament and surrounding alveolar bone, whilst supporting the wide population of cells present in the tissue. Initial work will seek to compare tissue culture methods to assess the best method for maintaining cell activity and tissue architecture. The cultures will be assessed for viability, with expression of relevant markers and differentiation factors analysed to identify whether relevant cell types are present in the cultures. To ensure the model mimics the degradation of bone prevalent in destructive periodontal disease, osteoclasts and innate immune cells will be transplanted into the ex vivo model. Cultures will be maintained and activity monitored before the tissue is exposed to pro-inflammatory cytokines to investigate the pathways leading to the pathogenesis of periodontal disease.   

    Roberts JL, Maillard JY, Waddington RJ, Denyer SP, Lynch CD, Sloan AJ (2013) Development of an ex vivo coculture system to model pulpal infection by Streptococcus anginosus group bacteria. Journal of Endodontics 39(1): 49-56 doi:10.1016/j.joen.2012.09.005

    Sloan AJ, Taylor SY, Smith EL, Roberts JL, Chen L, Wei XQ, Waddington RJ (2013) A novel ex vivo culture model for inflammatory bone destruction. Journal of Dental Research 92(8): 728-34 doi:10.1177/0022034513495240

    Sloan AJ, Taylor SY, Smith EL (2012). Organotypic mandibular cultures for the study of inflammatory bone pathology. In: Replacing animal models: a practical guide to creating and using biomimetic alternatives. Ed. J Davies Wiley Blackwell, London doi:10.1002/9781119940685.ch15

    Sloan AJ, Lynch CD (2012) Dental tissue repair: novel models for tissue regeneration strategies. Open Dentistry Journal 6: 214-219 doi:10.2174/1874210601206010214 

    Sloan AJ, Smith EL, Locke M, Waddington RJ (2010) An ex vivo rodent mandible culture model for bone repair. Tissue Engineering 16: 1287-1296 doi:10.1089/ten.TEC.2009.0698 

    Sloan AJ, Singhrao SK, Smith EL, Archer CW (2010) Technical advances in the sectioning of dental tissue and of on-section cross-linked collagen detection in mineralised teeth. Microscopy Research and Technique 73 (8): 741-5 doi:10.1002/jemt.20815

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    Project grant

    Status:

    Closed

    Principal investigator

    Professor Alastair Sloan

    Institution

    Cardiff University

    Co-Investigator

    Professor Rachel Waddington
    Dr Xiao-Qing Wei

    Grant reference number

    G0600349/1

    Award date:

    Feb 2007 - Jul 2009

    Grant amount

    £193,675