Deciphering the protein secretome of leukocyte-platelet rich fibrin

Leukocyte-platelet rich fibrin (L-PRF) is extensively used in the dentistry field and other clinical scenarios due to its regeneration properties. We obtained L-PRF membranes in absence of anticoagulants and cultured in DMEM. The secretomewas collected at days 3, 7 and 21 in order to identify the proteins secreted and the differences over time. Protein secretomeat day 3 was analysed by LC-MS/MS and differences in the proteome were analysed by Sequential Window Acquisition of all Theoretical Mass Spectra (SWATH). Overall, 705 proteins were identified in the secretome of L-PRF membranes after 3 days of culture, including growth factors (EGF, PDGFA) and proteins related to platelet and neutrophil degranulation. A total of 202 differentially secreted proteins were quantified by SWATH when comparing secretomes at days 3, 7 and 21. The majority of them were enriched at day 3 such as MMP9, TSP1 and CO3. On the contrary, fibrinogen and CATS were found down-regulated at day 3. Growth factor and western blotting analysis corroborated the proteomic results. This is the most detailed proteome analysis of the L-PRF secretome to date. Proteins and growth factors identified, and their kinetics, provide novel information to further understand the wound healing properties of L-PRF.