Article
Modelling delayed bone healing in a mouse-osteotomy model to evaluate therapeutic strategies for affected patients with RA
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Authors
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Annemarie Lang
- Charité-Universitätsmedizin Berlin, Medizinische Klinik mit Schwerpunkt Rheumatologie und klinische Immunologie, Berlin
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Sarah Fügener
- Charité-Universitätsmedizin Berlin, Medizinische Klinik mit Schwerpunkt Rheumatologie und klinische Immunologie, Berlin
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Paula Hoff
- Charité-Universitätsmedizin Berlin, Medizinische Klinik mit Schwerpunkt Rheumatologie und klinische Immunologie, Berlin
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Timo Gaber
- Charité-Universitätsmedizin Berlin, Medizinische Klinik mit Schwerpunkt Rheumatologie und klinische Immunologie, Berlin
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Frank Buttgereit
- Charité-Universitätsmedizin Berlin, Medizinische Klinik mit Schwerpunkt Rheumatologie und klinische Immunologie, Berlin
| Published: | September 4, 2017 |
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Outline
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Background: Anti-inflammatory treatment of rheumatoid arthritis (RA) using non- steroidal anti-inflammatory drugs (NSAID) or glucocorticoids (GC) as well as the disease itself, are supposed to negatively influence bone metabolism and healing. However, in vivo models allowing to evaluate therapeutic strategies for patients suffering from delayed bone healing are scarce and mainly created by critical size defects that are not representative for comorbity-induced disorders. In addition, there are no adequate rodent models allowing the analyses of the influence on the bone metabolism by the complete dysregulation of the immune system in RA as well as long-term medications with NSAID or GC.
Methods: Female C57BL aged 12 weeks underwent osteotomy of the femur (fracture gap 0,7 mm) that was fixated with an external fixator (RIS-System). Lyostypt® (based on bovine Col I; mimicking extracellular bone matrix) was applied in the fracture gap and analysis was performed 2 and 3 weeks after surgery. The ratio of bone volume (BV) per total volume (TV) in the fracture gap was evaluated using in vitro µCT. In addition, Movat’s pentachrome staining was performed to analyze the cellular and tissue composition within the fracture gap. To investigate the number of cells as well as the vessel formation, we used immunofluorescence to stain for Endomucin, PECAM-1 and DAPI. Quantitative analysis of histological staining was conducted by using ImageJ.
Results: We developed an in vivo mouse-osteotomy model showing delayed bone healing by applying Lyostypt® in the fracture gap. The BV, TV as well as the ratio was significantly higher in the controls (empty gap = normal healing; n=8 per timepoint) as compared to the Lyostypt® group(n=8 per timepoint) 2 weeks after osteotomy and slightly higher after 3 weeks. Histological investigation showed the clear presence of the scaffold on both timepoints without briding cartilaginous tissue. The cell number as well as the vessel formation was significantly reduced within the fracture gap.
Conclusion: The results obtained so far support the hypothesis that we were able to develop a delayed healing or even non-union osteotomy model in mice by avoiding to create a critical size defect. Therefore, this approach represents a promising alternative animal model to evaluate therapeutic strategies to overcome bone healing complications in RA patients and perhaps other immunologically restricted patients.
