Regulation of gingival fibroblast gene expression by leukocyte and platelet rich fibrin

Objectives: Leukocyte and Platelet-Rich Fibrin (L-PRF), a new clinically used blood product, is rich in leukocytes and platelets embedded in a high-density fibrin network. L-PRF has shown some clinical promise to promote gingival wound healing. In gingival surgeries where L-PRF is being used, it will be in direct contact with human gingival fibroblasts (HGFs) of the surgical flap. However, little is known about the combined effects of the growth factors and cells present in L-PRF on HGFs. We hypothesized that L-PRF regulates expression of key wound healing genes in HGFs. Thus, our aim was to investigate the effects of soluble biological factors released from L-PRF on HGF gene expression. Methods: L-PRF membranes were produced from blood samples of six volunteer donors using standard clinical protocol. The membranes were placed into upper compartments of transwell plate inserts (0.4 µm pore size) with confluent layers of HGFs at the bottom of the wells. Thus, only soluble mediators were able to influence HGF gene expression. HGFs with the L-PRF membranes in the wells were incubated for 48h followed by isolation of total RNA for real-time quantitative PCR of wound healing related genes. The levels of selected proteins produced by the cells were analyzed using Western blotting. Results: Among the 86 genes studied, the expression of 35 genes (41%) was significantly regulated by the L-PRF membranes in HGFs. Replicate membranes from the same donor or between different donors produced similar responses, with membranes from different individuals varying somewhat in magnitude of the response. Among the most regulated genes, matrix metalloproteinase-1 (MMP-1) and MMP-3 showed several-fold increase by L-PRF-treatment, both at the mRNA and protein level. Angiogenesis related genes, VEGF-α and FGF-2 showed also significant up-regulation in HGFs by the L-PRF-treatment. Conclusions: The results demonstrate that the L-PRF membranes have a strong and a specific regulatory effect on HGFs that may modulate several aspects of wound healing.Dentistry, Faculty ofGraduat