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Biology of PRF: Fibrin Matrix, Growth Factor Release, and Cellular Activity
Contributors
Masako Fujioka-Kobayashi
Yufeng Zhang
Reinhard Gruber
Richard J. Miron
Chapter Highlights
What is PRF?
How does PRF differ from PRP at the biologic and cellular level?
What is the role of each cell type found in PRF?
What is the role of each GF found in PRF?
How does centrifugation speed and time affect PRF?
What advantages exist for horizontal centrifugation versus fixed-angle centrifugation?
Much can be discussed with respect to the biology of PRF and its ability to impact tissue regeneration. During the natural wound healing process, vascularization of tissues plays a pivotal role, facilitating the invasion of incoming cells, growth factors (GFs), cytokines, and other regenerative factors. The main aim of platelet concentrates, discovered over two decades ago, is to favor new blood flow (angiogenesis) to damaged tissues, thereby improving their healing potential by delivering a supraphysiologic concentration of blood-derived cells (namely platelets) and regenerative GFs. This chapter takes a deep look into the actual separation of blood layers during the centrifugation process to provide the clinician a general overview of the cell types and GFs found in PRF, including their roles, and also discusses the effects of centrifugation speed and time on cell layer separation. Furthermore, the advantages of producing an autologous fibrin scaffold are presented as it being a key regulator of wound healing because of its autologous source and its ability to promote the slow and gradual release of GFs over time. The advantages of horizontal centrifugation versus fixed-angle centrifugation are also discussed based on recent data from various laboratories from around the world.
The wound healing process is divided into three stages: the inflammatory phase, the proliferative phase, and the remodeling phase (Fig 2-1). The inflammatory phase starts at the time of injury and generally involves a wide array of cytokines and growth factors (GFs) that are released within the first 24 to 48 hours. Accordingly, a dynamic interaction occurs between endothelial cells, angiogenic cytokines, and the extracellular matrix (ECM) in an attempt to accelerate wound healing via an orchestrated delivery of multiple GFs in a well-controlled fashion.1
Fig 2-1 The three stages of wound healing: (a) Inflammatory phase. (b) Proliferative phase. (c) Remodeling phase.
In general, blood provides essential components to the healing process that comprise both cellular and protein products that essentially are the base components of wound healing. During the healing process, blood will undergo clotting within a few minutes to prevent further blood loss. This is an important step that will be later discussed in the PRF tube section, because in order for clotting to occur and even be improved in both speed and quality (in particular in patients taking anticoagulants), a proper understanding of the clotting cascade is required. In its simplest of forms, oxygen helps improve blood clotting, and for this reason, the simple removal of centrifugation tube lids following the spin process will lead to faster clotting of PRF and a superior fibrin mesh. One of the major roles of platelets is to assist during hemostasis through a fibrin clot formation.1,2 Not surprisingly, the additional use of PRF for wound healing (for example, following tooth removal and extraction socket healing) in patients taking anticoagulants can drastically improve the healing outcomes simply by improving clotting. Because PRF contains many platelets and a fibrin nucleus is already formed, bleeding has been shown to be significantly reduced postoperative when PRF is utilized in patients on anticoagulant therapy.3
Tips
Oxygen helps improve blood clotting, so the simple removal of centrifugation tube lids following the spin process will lead to faster clotting of PRF.
In patients undergoing anticoagulant therapy, the simple addition of PRF during surgery can help favor faster clotting, thereby reducing bleeding times postoperative.
Platelets also release various GFs and cytokines that further lead to tissue regeneration but also attract macrophages and neutrophils to the defect site. These cells are responsible for clearing debris, replacing necrotic tissue, and removing bacteria from the wound site.
The proliferative phase begins by day 3, where the blood clot within the wound is further supplied with a provisional matrix typically composed in part with fibrin, which facilitates cell migration, while the clot within the vessel lumen contributes to hemostasis.2 Fibroblast cells are recruited to the wound site and begin producing new collagen in a random and somewhat disorganized order. Simulta-neously, new blood vessel formation leads to new angiogenesis, and the wound gradually begins to gain initial stability.
During the third and final stage (the remodeling phase), disorganized collagen is replaced by newly organized collagen fibrils that provide enhanced stability and strength to the injured site, where tissue regeneration takes place4 (see Fig 2-1).
Whole blood is comprised of four main components: blood plasma, red blood cells (RBCs), white blood cells (WBC), and platelets. Initially, platelets were reported as the major responsible component for the activation and release of crucial GFs for wound healing, including PDGF, coagulation factors, adhesion molecules, cytokines, and angiogenic factors. Their role has been extremely well described in the literature, so typically the entire field has been referred to as platelet concentrates or platelet-rich plasma/fibrin. Interestingly, however, over the years more attention has been placed on leukocytes, which are not only responsible for host defense but also highly implicated in the wound healing and regenerative phases.
Table 2-1 highlights the various cell types found in blood, including their density, frequency, and surface area. Note that while platelets are the lightest of the group, WBCs and RBCs are very similar in density. For these reasons they are also harder to separate in a centrifuge based on density. Noteworthy is the fact that per µL, there are 5,000,000 RBCs when compared to only 5,000 WBCs. Therefore, RBCs outnumber WBCs in a 1,000:1 ratio, which make them difficult to separate, especially on a fixed-angle centrifugation device as discussed later in this chapter (Video