The in vitro human fracture hematoma model – a tool for preclinical drug testing

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Moritz Pfeiffenberger
Paula Hoff
Christa Thöne-Reineke
Frank Buttgereit
Annemarie Lang
Timo Gaber


The aim of the study was to establish an in vitro fracture hematoma (FH) model that mimics the in vivo situation of the human fracture gap in order to assess drug efficacy and effectiveness for the treatment of fracture healing disorders. Human peripheral blood and mesenchymal stromal cells (MSCs) were coagulated to produce in vitro FH models, which were incubated in osteogenic medium under normoxia/hypoxia and analyzed for cell composition, gene expression and cytokine/chemokine secretion. To evaluate the model, we studied the impact of dexamethasone (impairing fracture healing) and deferoxamine (promoting fracture healing). Under hypoxic conditions, MSCs represented the predominant cell population, while the frequencies of leukocyte populations decreased. Marker gene expression of osteogenesis, angiogenesis, inflammation, migration and hypoxic adaptation increased significantly over time and compared to normoxia, while cytokine/chemokine secretion remained unchanged. Dexamethasone favored the frequency of immune cells compared to MSCs, suppressed osteogenic and pro-angiogenic gene expression, and enhanced the secretion of inflammatory cytokines. Conversely, deferoxamine favored the frequency of MSCs over that of immune cells and enhanced the expression of the osteogenic marker RUNX2 and markers of hypoxic adaptation. In summary, we demonstrate that hypoxia is an important factor for modeling the initial phase of fracture healing in vitro and that both fracture-healing disrupting and promoting substances can influence the in vitro model comparable to the in vivo situation. Therefore, we conclude that our model is able to mimic in part the human FH and could reduce the number of animal experiments in early preclinical studies.

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How to Cite
Pfeiffenberger, M., Hoff, P., Thöne-Reineke, C., Buttgereit, F., Lang, A. and Gaber, T. (2020) “The in vitro human fracture hematoma model – a tool for preclinical drug testing”, ALTEX - Alternatives to animal experimentation, 37(4), pp. 561–578. doi: 10.14573/altex.1910211.

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