In Vitro and Ex Vivo Kinetic Release Profile of Growth Factors and Cytokines from Leucocyte- and Platelet-Rich Fibrin (L-PRF) Preparations

Abstract

L-PRF is an autologous blood-derived biomaterial (ABDB) capable of releasing biologically active agents to promote healing. Little is known about its release profile of growth factors (GFs), cytokines, and MMPs. This study reported the in vitro and ex vivo release kinetics of GFs, cytokines, and MMPs from L-PRF at 6, 24, 72, and 168 h. The in vitro release rates of PDGF, TGF-β1, EGF, FGF-2, VEGF, and MMPs decreased over time with different rates, while those of IL-1β, IL-6, TNF-α, IL-8, and IL-10 were low at 6 h and then increased rapidly for up to 24 h and subsequently decreased. Of note, the release rates of the GFs followed first-order kinetics both in vitro and ex vivo. Higher rates of release were found ex vivo, suggesting that significant amounts of GFs were produced by the local cells within the wound. In addition, the half-life times of GFs locally produced in the wound, including PDGF-AA, PDGF-AB/BB, and VEGF, were significantly extended (p < 0.05). This work demonstrates that L-PRF can sustain the release of GFs and cytokines for up to 7 days, and it shows that the former can activate cells to produce additional mediators and amplify the communication network for optimizing the wound environment, thereby enhancing healing.

Increased local concentrations of growth factors from leucocyte‐ and platelet‐rich fibrin do not translate into improved alveolar ridge preservation: An intra‐individual mechanistic randomized controlled trial

“…GF concentrations were quantified using a multiplex immunoassay. A Milliplex® MAP transforming growth factor‐β1 (TGF‐β1) Magnetic Bead Single Plex Kit (Merck & Co., USA) and Milliplex® Customized 13‐Plex Human Growth Factor/Cytokine/MMP Mag Kit (Merck & Co) were used to detect and quantify 14 biomarkers, including TGF‐β1, platelet‐derived growth factor‐AA (PDGF‐AA), platelet‐derived growth factor‐AB/BB (PDGF‐AB/BB), epidermal growth factor (EGF), fibroblast growth factor‐2 (FGF‐2), vascular endothelial growth factor (VEGF), angiopoietin‐2 (ANG‐2), interleukin‐1β (IL‐1β), interleukin‐6 (IL‐6), interleukin‐8 (IL‐8), interleukin‐10 (IL‐10), tumor necrosis factor‐α (TNF‐α), matrix metalloproteinase‐1 (MMP‐1), matrix metalloproteinase‐2 (MMP‐2)) as previously described (Wang et al, 2022). Streptavidin/phycoerythrin conjugates were quantified using the Bio‐Plex® 200 system (Bio‐Rad Laboratories Inc., USA).…”

“…Thereafter, paper strips were placed into Eppendorf microcentrifuge tubes containing 100 μl of phosphate-buffered saline with an ethylenediaminetetraacetic acid (EDTA)-free Protease Inhibitor Cocktail (cOmplete™, Roche Applied Science, Indianapolis, IN) and vortexed for 10 s. Eluates were centrifuged at 10,000 rpm for 10 min at 4 C and stored at À80 C until analysis. were used to detect and quantify 14 biomarkers, including TGF-β1, platelet-derived growth factor-AA (PDGF-AA), platelet-derived growth factor-AB/BB (PDGF-AB/BB), epidermal growth factor (EGF), fibroblast growth factor-2 (FGF-2), vascular endothelial growth factor (VEGF), angiopoietin-2 (ANG-2), interleukin-1β (IL-1β), interleukin-6 (IL-6), interleukin-8 (IL-8), interleukin-10 (IL-10), tumor necrosis factor-α (TNF-α), matrix metalloproteinase-1 (MMP-1), matrix metalloproteinase-2 (MMP-2)) as previously described (Wang et al, 2022). Streptavidin/phycoerythrin conjugates were quantified using the Bio-Plex ® 200 system (Bio-Rad Laboratories Inc., USA).…”

Lyophilized Platelet-Rich Fibrin Exudate-Loaded Carboxymethyl Chitosan/GelMA Hydrogel for Efficient Bone Defect Repair

“…Further, the sustained release profiles of TGF-β1 and PDGF-AB were investigated to demonstrate the sustained release ability of CGP hydrogels for growth factors (Figure i,j). Since many growth factors, including TGF-β1 and PDGF-AB, have already been investigated in PRF using elution, TGF-β1 and PDGF-AB can reflect the levels of other growth factors in the experiment . The results showed a prolonged and sustainable release of TGF-β1 and PDGF-AB from CGP hydrogels in the observation period of 21 days.…”

“…Since many growth factors, including TGF-β1 and PDGF-AB, have already been investigated in PRF using elution, TGF-β1 and PDGF-AB can reflect the levels of other growth factors in the experiment. 66 The results showed a prolonged and sustainable release of TGF-β1 and PDGF-AB from CGP hydrogels in the observation period of 21 days. The CGP hydrogels released 7.31% ± 0.34% of TGF-β1 and 7.36% ± 0.29% of PDGF-AB after 21 days, demonstrating the efficiency of the CGP hydrogels for sustained delivery of growth factors.…”

Platelet-rich fibrin as an autologous biomaterial for bone regeneration: mechanisms, applications, optimization

“…In the meantime, a heatmap was generated using the mean of total amounts of release per hour to show the changes in the release rate of growth factors, cytokines, or MMPs over 168 h ( Figure 5 ). For growth factors and MMPs, the highest release was found at the first 6 h, followed by a reduction in concentrations, although with different rates of decrease, while the highest release rates for cytokines and the chemokine occurred from 6 to 24 h ( Wang X. et al, 2022 ). Besides, kinetic analysis of the released factors showed that, in contrast to the bimodal release of TGF-ß1 in L-PRP and clot, in L-PRF, TGF-ß1 showed a single-peak release at day 7; the release profile of IGF-1 in L-PRF showed a gradual release over the first 3 days, and at the same time, in L-PRF, the release of IL-1ß reached the peak of its release at day 1 ( Schär et al, 2015 ).…”