Abstract
Platelet-rich fibrin (PRF) contains a broad spectrum of bioactive molecules that can trigger several cellular responses. However, these molecules along with their upstream responses remain mostly uninvestigated. By means of proteomics we revealed that PRF lysates contain more than 650 proteins, being TGF-β one of the few growth factors found. To uncover the major target genes regulated by PRF lysates, gingival fibroblasts were exposed to lysates obtained from PRF membranes followed by a whole genome array. We identified 51 genes strongly regulated by PRF including IL11, NOX4 and PRG4 which are characteristic TGF-β target genes. RT-PCR and immunoassay analysis confirmed the TGF-β receptor I kinase-dependent increased expression of IL11, NOX4 and PRG4. The PRF-derived TGF-β activity was verified by the translocation of Smad2/3 into the nucleus along with the increased phosphorylation of Smad3. Considering that PRF is clinically used in combination with dental implants and collagen membranes, we showed here that PRF-derived TGF-β activity adsorbs to titanium implants and collagen membranes indicated by the changes in gene expression and immunoassay analysis. Our study points towards TGF-β as major target of PRF and suggest that TGF-β activity released by PRF adsorbs to titanium surface and collagen membranes
Highlights
Platelet-rich fibrin (PRF) contains a broad spectrum of bioactive molecules that can trigger several cellular responses
In the present study we show that (i) TGF-β is found in PRF lysates based on proteomic analysis; (ii) PRF lysates provoke a robust activation of TGF-β signalling in oral gingival fibroblasts based on genomic screening and a series of specific downstream analysis; (iii) PRF-derived TGF-β activity adsorbs to titanium implants and collagen membranes
We could confirm the presence of typical platelets proteins, such as platelet factor 4 (PF4), platelet basic protein (PPBP), platelets glycoproteins (GP1BA, GP1BB CD36, GP5, GP6, GP9), platelet endothelial cell adhesion molecule (PECAM1), and von Willebrand factor (VWF)
Summary
Platelet-rich fibrin (PRF) contains a broad spectrum of bioactive molecules that can trigger several cellular responses. Cell proliferation or osteogenic differentiation in response to PRF can be measured via changes in gene expression[12,13] This screening approach can be further refined by means of whole genome arrays. The second aim of the present study was to combine the proteomic approach with the findings from the gene array to understand which PRF-derived growth factors cause the major changes in gene expression. In order to mimic a clinical situation in an in vitro setting, we examined whether the growth factors released by PRF activates mesenchymal cells bound to the respective biomaterials This is clinically relevant as local application of recombinant TGF-β in a collagen sponge can enhance bone regeneration of rabbit skull defects[24]. The final aim of this research was to investigate if the growth factors, which cause the most robust gene expression changes, adsorb to titanium and collagen membranes
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