An in vitro study into three different PRF preparations for osteogenesis potential

Objective: To investigate the effect of Advanced Platelet-Rich Fibrin (A-PRF+), Leukocyte Platelet-Rich Fibrin (L-PRF), and injectable Platelet-Rich Fibrin (i-PRF) on osteogenesis of a human osteoblast-like cell line in vitro. Background: Different PRF protocols are used in clinical dentistry in the last years. Recent literature documented the positive impact of PRF derivatives in vivo and in vitro, on different types of cells. However, hardly any literature comparing the new protocols for PRF (the A-PRF+ and i-PRF) with the original protocol of PRF (L-PRF) is present for osteoblast-like cells. Materials and Methods: A-PRF+, L-PRF, and i-PRF were prepared from six male donors and pre-cultured with 10 mL culture medium for 6 days. 5 x 10³ cells/ml osteoblasts from the osteoblast cell line (U2OS) were seeded and cultured either with conditioned medium derived from the different PRF conditions or with regular culture medium. At five different time points (0, 7, 14, 21, 28 days), the osteogenic capacity of the cells was assessed with Alizarin Red S to visualize mineralization. Also in these cells, the calcium concentration and alkaline phosphatase activity were investigated. Using qPCR, the expression of alkaline phosphatase, osteocalcin, osteonectin, ICAM-1, RUNX-2, and collagen 1a was assessed. Results: In osteoblast-like cells cultured with conditioned medium, the A-PRF+ conditioned medium induced more mineralization and calcium production after 28 days of culturing compared with the control (p <.05). No significant differences were found in the extent of cell proliferation between the different conditions. RUNX-2 and osteonectin mRNA expression in the cells were lower in all PRF-stimulated cultures compared with control at different time points. The i-PRF-conditioned medium induced more ALP activity (p <.05) compared with control and osteoblasts-like cells differentiated more compared with osteoblasts cultured with L-PRF. Conclusions: The three PRF preparations seem to have the capacity to increase the osteogenic potential of osteoblast-like cells. A-PRF+ seems to have the highest potential for mineralization, while i-PRF seems to have the potential to enhance early cell differentiation.