Abstract
Liquid platelet-rich fibrin (PRF) can be prepared by high centrifugation forces separating the blood into a platelet-poor plasma (PPP) layer and a cell-rich buffy coat layer, termed concentrated PRF (C-PRF). Heating the liquid PPP was recently introduced to prepare an albumin gel (Alb-gel) that is later mixed back with the concentrated liquid C-PRF to generate Alb-PRF. PRF is a rich source of TGF-β activity; however, the overall TGF-β activity in the PPP and the impact of heating the upper plasma layer remains unknown. Here, we investigated for the first time the in vitro TGF-β activity of all fractions of Alb-PRF. We report that exposure of oral fibroblasts with lysates of PPP and the buffy coat layer, but not with heated PPP, provoked a robust increase in the TGF-β target genes interleukin 11 and NADPH oxidase 4 by RT-PCR, and for IL11 by immunoassay. Consistent with the activation of TGF-β signaling, expression changes were blocked in the presence of the TGF-β receptor type I kinase inhibitor SB431542. Immunofluorescence and Western blot further confirmed that lysates of PPP and the buffy coat layer, but not heated PPP, induced the nuclear translocation of Smad2/3 and increased phosphorylation of Smad3. The immunoassay further revealed that PPP and particularly BC are rich in active TGF-β compared to heated PPP. These results strengthen the evidence that not only the cell-rich C-PRF but also PPP comprise a TGF-β activity that is, however, heat sensitive. It thus seems relevant to mix the heated PPP with the buffy coat C-PRF layer to regain TGF-β activity, as proposed during the preparation of Alb-PRF.
Highlights
Platelet-rich fibrin (PRF), originally introduced in 2006 in an article series [1], has received increasing attention in clinical regenerative dentistry [2] and other fields, including aesthetic medicine [3], as an autologous source of cells and growth factors embedded in a fibrin-rich extracellular matrix that has handling properties similar to a graft or biomaterial
This research was prompted by the recently established PRF formulation consisting of heat-coagulated platelet-poor plasma (PPP) mixed with the liquid cell-rich buffy coat concentrated PRF (C-PRF) to serve as a long-term stable autologous and injectable matrix [13,20]
Previous reports clearly demonstrated that TGF-β is among the growth factors supporting the formation of a fibrous extracellular matrix [29,30,34]
Summary
Platelet-rich fibrin (PRF), originally introduced in 2006 in an article series [1], has received increasing attention in clinical regenerative dentistry [2] and other fields, including aesthetic medicine [3], as an autologous source of cells and growth factors embedded in a fibrin-rich extracellular matrix that has handling properties similar to a graft or biomaterial. Even though the preparation of solid PRF is simple, as it basically requires the spontaneous coagulation of centrifuged blood, the protocols have been refined towards gaining an ideal ratio of the PRF clot and its content of platelets, leucocytes, and growth factors. Solid PRF was introduced as L-PRF prepared with blood collection tubes containing clot activators at around 700 g for 12 min in a fixed angle centrifuge. PRF protocols prepared via horizontal centrifugation using plain glass tubes without clot activators to prepare H-PRF have been shown to lead to more favorable accumulation of platelets and leukocytes in the upper plasma layers [4,12].
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