In vitro study of osteoclastogenesis under simulated bone augmentationThe effects of bone-conditioned medium and saliva on osteoclastogenesis

Resumen

The present PhD thesis is a compendium of four publications broadening the knowledge on osteoclastogenesis under simulated bone augmentation, more especially about the effects of saliva and bone-conditioned medium on osteoclastogenesis. Resorption of bone grafts and host bone, can be a challenge especially when a bonny defect has to be regenerated or there is a lack of host bone due to a trauma, pathology, aging or tooth extraction among others. In the oral cavity, saliva is present and can reach mineralized surfaces, however, the relationship between saliva and bone resorption is yet unknown. Herein, we examined whether saliva affects the process of osteoclastogenesis in vitro, possibly affecting bone healing and bone regeneration. Bone regeneration is a common procedure in traumatology, periodontology, oral and maxillofacial surgery that involves the use of bone fillers. Bone autograft is considered to be the gold standard bone substitute due to its trinity of properties: osteoinductivity, osteoconductivity and osteogenesis. Paracrine factors released from bone autografts might contribute to the overall process of graft consolidation, however the underlying mechanisms are unknown. Here, we determined the protein spectrum released from porcine bone chips into the conditioned medium (BCM) to mimic the paracrine environment of cortical bone grafts. Some of the factors released by bone autografts could maybe influence on the autograft resorption and therefore explain why osteoclasts rapidly form on the surface of bone chips at augmentation sites. The underlying molecular mechanism, however, is unclear. Soluble factors released from bone chips in vitro have a robust impact on mesenchymal cell differentiation. Here we determined whether these soluble factors change the differentiation of hematopoietic cells into osteoclasts, still unknown. Based on the in vitro results here presented, it can be observed that saliva suppresses osteoclastogenesis and leads to the development of a phagocytic cell phenotype, therefore affecting function of osteoclasts, the bone resorbing cells. Resorption of bone autografts could be attributed to some of the proteins detected on the secretions of bone autografts, termed bone conditioned medium (BCM). Proteomic analysis showed that BCM contains more than 150 proteins, among which, 43 were categorized into “secreted” and “extracellular matrix”. We discovered growth factors that are not only detectable in BCM, but potentially also target cellular processes involved in bone regeneration e.g. pleiotrophin, galectin-1, TGF-?-induced gene (TGFBI), latency-associated peptide forming a complex with TGF-?1, and TGF-?2. Results here presented on the influence of BCM on osteoclastogenesis demonstrated that activated BCM by heat is able to stimulate osteoclastogenesis in vitro. These in vitro results support the notion that the resorption of autografts may be supported by as yet less defined regulatory mechanisms. Moreover the presented protocols on the use of BCM should encourage to further reveal the paracrine effects of bone grafts during bone regeneration and open a path for translational research in the broad field of reconstructive surgery. Taking everything together, it can be concluded that saliva affects bone resorption towards the development of a phagocytic cell line, and that not only saliva affects bone resorption but also the secretions from autologous bone grafts. There is enough evidence to conclude that bone autografts not only have three properties, but one more: a regulation property, the fourth dimension of autologous bone grafts.