Abstract
Bone tissue engineering is based on bone grafting to repair bone defects. Bone graft substitutes can contribute to the addition of mesenchymal stem cells (MSCs) in order to enhance the rate and the quality of defect regeneration. The stem cell secretome contains many growth factors and chemokines, which could affect cellular characteristics and behavior. Conditioned medium (CM) could be used in tissue regeneration avoiding several problems linked to the direct use of MSCs. In this study, we investigated the effect of human periodontal ligament stem cells (hPDLSCs) and their CM on bone regeneration using a commercially available membrane scaffold Evolution (EVO) implanted in rat calvarias. EVO alone or EVO + hPDLSCs with or without CM were implanted in Wistar male rats subjected to calvarial defects. The in vivo results revealed that EVO membrane enriched with hPDLSCs and CM showed a better osteogenic ability to repair the calvarial defect. These results were confirmed by acquired micro-computed tomography (CT) images and the increased osteopontin levels. Moreover, RT-PCR in vitro revealed the upregulation of three genes (Collagen (COL)5A1, COL16A1 and transforming growth factor (TGF)β1) and the down regulation of 26 genes involved in bone regeneration. These results suggest a promising potential application of CM from hPDLSCs and scaffolds for bone defect restoration and in particular for calvarial repair in case of trauma.
Highlights
Bone damage is difficult to repair, especially for calvaria defects, and bone tissue possesses a low ability of endogenous self-repair by regenerating new bone without forming a fibrotic scar [1]
The human periodontal ligament stem cells (hPDLSCs) were positive for Oct3/4, Sox-2, SSEA-4, CD29, CD44, CD73, CD90 and CD105 and negative for CD14, CD34 and CD45 (Figure 1A). hPDLSCs have been expanded to form confluent cultures of adherent cells with a fibroblastic morphology, as demonstrated by toluidine blue staining (Figure 1B)
We used hPDLSCs that express mesenchymal stem cells (MSCs) features such as the positivity for Oct3/4, Sox-2, SSEA-4, CD29, CD44, CD73, CD90 and CD105, the negativity for CD14, CD34, CD45, the ability to differentiate into osteogenic and adipogenic lineages other than the adherence to a plastic substrate, as previously demonstrated by our group [26]. hPDLSCs seem promising in bone regenerative medicine and may have advantages compared to other MSCs
Summary
Bone damage is difficult to repair, especially for calvaria defects, and bone tissue possesses a low ability of endogenous self-repair by regenerating new bone without forming a fibrotic scar [1]. It is mandatory to guarantee a fast and suitable regeneration of bone tissue and its properties, in order to ameliorate the healing process and to re-establish the lost functions. Bone tissue engineering is a research field that provides the right microenvironments to promote cell differentiation together with optimal scaffold development. The scaffolds need to be biocompatible, lead the progenitor cells to commit to an osteogenic lineage and avoid the possible host tissue inflammation or reaction. Scaffolds should provide an optimal microenvironment to support bone growth and development, the vascular network formation and cell recruitment [3]
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