Publikationsdatenbank

HIF-stabilization to accelerate fracture healing:
evaluation of a new therapeutic strategy to treat delayed bone regeneration (2019)

Art

Hochschulschrift

Autor

Lang, Annemarie (WE 12)

Quelle

Berlin: Mensch & Buch Verlag Berlin, 2019 — ix, 150 Seiten

ISBN: 978-3-96729-026-4

Sprache

Englisch

Verweise

URL (Volltext): https://refubium.fu-berlin.de/handle/fub188/26365

Kontakt

Institut für Tierpathologie

Robert-von-Ostertag-Str. 15
14163 Berlin
+49 30 838 62450
pathologie@vetmed.fu-berlin.de

Abstract / Zusammenfassung

The hypoxic microenvironment during the initial phase of fracture healing is essential for initiating immunological process that further the regeneration and restoration of the bone. The hypoxia-inducible factor (HIF)-1α closely regulates the cellular adaption under hypoxic conditions. HIF-1α can be chemically stabilized by different factors which either inhibit the O2-sensing prolyl hydroxylase e.g. deferoxamine (DFO) or directly interfere with the downstream effects after nuclear translocation e.g. the macrophage-migration inhibitory factor (MIF). As a transcription factor, HIF-1α initiates the consecutive processes of bone regeneration and human mesenchymal stromal cell (hMSC) differentiation. Hence, the aim was to promote the cellular adaptation towards hypoxia in order to specifically accelerate fracture healing under compromised conditions (inhibited mineralization and vascularization). First, in vitro studies were performed to i) evaluate the potential of DFO and MIF in combination to counteract a glucocorticoid-induced inhibition of hMSC calcification and ii) determine an effective concentration of both substances for further testing in a mouseosteotomy-model. Concurrently, two absorbable bovine Col I scaffolds (ACSs) were tested for their suitability to be used as a delivery system of these substances into the osteotomy gap. Finally, both HIF-stabilizers were evaluated for their potential to accelerate fracture healing in a mouse-osteotomy-model. In vitro, a concentration-dependent inhibitory effect of the glucocorticoid dexamethasone was observed on osteogenic differentiation and calcification of hMSCs via a quantitative Alizarin red assay. This inhibition was counteracted by applying different concentrations DFO and MIF in combination. As suitable scaffolds for the in vivo application Lyostypt® (ACS-L) and Helistat® (ACS-H) were investigated in vitro for their structural components and impact on hMSC osteogenesis, cytotoxicity and immunogenicity. Proteomics analysis of both scaffolds yielded several proteins beside collagens that might be advantageous or disadvantageous regarding the fracture healing outcomes. Moreover, ACS-H induced a strong tumor necrosis factor (TNFα) release when applied to whole human blood and inhibited the calcification during the osteogenic differentiation of hMSCs. Consequently, ACS-L was examined in more detail in a mouse-osteotomy-model and revealed an inhibitory effect on mineralized callus formation, cellular recruitment to the osteotomy gap and vessel formation. The application of DFO and MIF in combination and DFO alone during the initial healing phase accelerated the vessel formation, the ACS degradation and the callus mineralization. The results support the fact that stabilization of HIF-1α enhances osteogenic differentiation in vitro and is capable to counteract e.g. glucocorticoid-induced inhibition. Both ACSs negatively influenced either the hMSC differentiation in vitro or the bone healing process in vivo, although being routinely used in research and clinic routine. These results display a delayed healing process that parallels observed compromised conditions in Rheumatoid arthritis patients or smokers – reduced vessel and bone formation. The combination of MIF and DFO was evaluated in this model of compromised condition to test their counteracting ability in this clinically relevant model. This study provides evidence for a promising therapeutic strategy to accelerate fracture healing capacities and to prevent disorders by applying potent HIF-stabilizers in a specific patient cohort with a higher risk of a bone healing delay.