Bone infections are one of the main reasons for impaired bone regeneration and non-union formation. In previous experimental animal studies we could already demonstrate that bone defects due to prior infections showed a markedly reduced healing capacity, which could effectively be enhanced via application of Wnt3a and Adipose-derived stromal cells (ASCs). For a more in-depth analysis, we investigated proliferation and mineralization of cultured osteoblasts infected with staph aureus and sought to investigate effects of Wnt3a and ASCs on infected osteoblasts.
Materials and methods
Primary murine osteoblasts were isolated from calvariae and infected with staph aureus. Infected osteoblasts received treatment via application of recombinant Wnt3a, ASC conditioned medium and were furthermore cocultured with ASCs. Osteoblasts were evaluated by Alamar blue assay for metabolic activity, TUNEL-assay for apoptosis, ALP and Alizarin Red staining for mineralization. In addition, immunoflourescent staining (IF) and qRT-PCR analyses were performed.
Infected osteoblasts showed a markedly reduced ability for mineralization and increased apoptosis, which could be restored to physiological levels by Wnt3a and ASC treatment. Interestingly, metabolic activity of osteoblasts seemed to be unaffected by staph aureus infection. Additional analyses of Wnt-pathway activity revealed effective enhancement of canonical Wnt-pathway activity in Wnt3a-treated osteoblasts.
In summary, we gained further osteoblast-related insights into pathomechanisms of reduced bone healing capacity upon infections.
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Adipose-derived stromal cell
Receptor activator nuclear factor-κB ligand
- GSK-3β :
Glycogen synthase kinase 3 beta
Secreted frizzled-related protein 1
Tumor necrosis factor-related apoptosis-inducing ligand
Bone marrow-derived stem cells
- Runx 2:
Runt-related transcription factor 2
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This work was supported by a grant of the DFG (Deutsche Forschungsgemeinschaft) (BE 4169/8-1).
Conflict of interest
The authors declare that they have no conflicts of interests.
Isolation of murine osteoblasts was performed according to the guidelines of the National Institute of Health for the use of experimental animals and after approval by the German legislation. The protocol was approved by the LANUV (NRW, Germany; Permit-Number: 84–02.04.2014.A044).
Informed consent is not applicable for this study.
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Wagner, J.M., Steubing, Y., Dadras, M. et al. Wnt3a and ASCs are capable of restoring mineralization in staph aureus-infected primary murine osteoblasts. J Bone Miner Metab (2021). https://doi.org/10.1007/s00774-021-01269-4