Introduction to Platelet Rich Fibrin (PRF) and its uses in Dentistry �

Introduction to Platelet Rich Fibrin (PRF) and its uses in Dentistry

1. Introduction to Platelet Rich Fibrin ( PRF) and its uses in Dentistry Dr. Hamza Jawed Resident OMFS

2. Introduction The conception of healing process is still partial and is a keen subject for research, but it is well known that platelets play a significant role in both hemostasis and wound healing activity . There is rationale on the fact that platelets play a crucial role in tissue healing and inflammation .

3. The presence of growth factors and cytokines are important guidelines to regenerate the wound area. Upon the activation of platelets there is proof of release of not only cytokines, enzymes, proteins but also fibrinolytic and anti-fibrinolytic proteins , which act as a matrix during the pathway of tissue repair. This has led to the notion in the use of platelets as a remedial tool to improve tissue healing

4. Cytokines are also released from the platelets, being responsible in modulating platelet activation and the proliferation and differentiation of leukocytes, playing an important role in immunology, specifically, in inflammation mechanism.

5. What are the differences between PRP , PRGF and PRF ?

6. Platelet Rich Plasma (PRP): The Platelet-Rich Plasma is a product derived from blood, its characteristic is due to the fact that the platelets present in the PRP release numerous substances that promote tissue repair and affect the behavior of other cells by modulating the inflammation and the formation of new blood vessels .

7. Platelet Rich Plasma (PRP): Despite the growing success and use of PRP in the initial years following its launch, there were several reported limitations that prevented its full potential. The technique itself was lengthy and therefore required the additional use of anti-coagulant factors to prevent clotting using bovine thrombin or CaCl2, both known inhibitors of wound healing . These drawbacks in combination with the lengthy harvesting/centrifugation preparation times were then frequently being utilized in large maxillofacial surgeries, whereas the typical dental or medical practitioner was resistant to its use due to lengthy preparation times

8. Platelet Rich Plasma (PRP): One of the other drawbacks of PRP was the fact that it was liquid by nature , and therefore required its combination with other biomaterials including bone grafts derived from human cadavers (allografts) or animal products (xenografts), thereby further combining its use with other “ unnatural ” products.

9. Platelet Rich in Growth Factors ( PRGF ) is a type of plasma enriched of proteins and circulating growth factors able to aid the bone and soft tissue regeneration. PRGF contains many different cells and cell-types highly concentrated in a gelatinous form which can be placed into the site of the injury these cells stimulate and accelerate the healing process by forming blood clots and releasing growth factors into the wound. PRGF does not need bovine or human thrombin for coagulation PRGF includes plasma proteins and coagulative factors and is then more advantageous compared to PRP.

10. PRGF preparation was carried out by following a previously described protocol. Blood sample was obtained from basilic vein using a large needle to avoid platelet rupture.

11. Sampled blood was combined with anticoagulant ( 1 ml of 3.8% sodium citrate for 10 ml blood ) and centrifuged at 460 G in 8 minutes : after the centrifugation, PRGF was taken from the bottom of the tube Calcium Chloride was then added to PRGF ( 0.05 ml per ml ): this action promotes the coagulation , usually obtained within at most 10 minutes. At the end of the procedure, we obtained a gelatinous PRGF, to be immediately placed in the surgical site.

12. PRGF preparation was carried out by following a previously described protocol. Blood sample was obtained from basilica vein using a large needle to avoid platelet rupture. Sampled blood was combined with anticoagulant (1 ml of 3.8% sodium citrate for 10 ml blood) and centrifuged at 460 G in 8 minutes: after the centrifugation, PRGF was taken from the bottom of the tube. 3 . Calcium Chloride was then added to PRGF (0.05 ml per ml): this action promotes the coagulation, usually obtained within at most 10 minutes. At the end of the procedure, we obtained a gelatinous PRGF, to be immediately placed in the surgical site.

13. WHAT IS PRF ?

14. The world of dentistry was first familiarized with the regenerative capacity of platelets in the 70s . It is a preparation of platelets present in a small volume of plasma containing a large amount of growth factors ( GFs ), which is essential for bone growth and regeneration . Platelet-rich fibrin (PRF) is frequently named as Choukroun’s PRF after its inventor, and was described as a second- generation platelet concentrate which contains platelets and growth factors in the form of fibrin membranes prepared from the patient’s own blood free of any anticoagulant or other artificial biochemical modifications

15. Platelet rich fibrin (PRF) is a fibrin matrix in which Platelet cytokines, Growth factors and Cells are trapped and may be released after a certain time and that can serve as a resorbable membrane . It is obtained from blood with the help of a simple process. PRF is basically a concentrate of growth factors that promote wound healing and regeneration which is used in various disciplines of dentistry to repair various lesions and regenerate dental and oral tissues. PRF was developed in France by Joseph Choukroun et al. in 2001 .

16. Growth factors are released after activation from the platelets trapped within fibrin matrix, and have been shown to stimulate the mitogenic response in the periosteum for bone repair during normal wound healing. It is an autogenous osteo-inductive material that enhances osteogenesis in the extraction tooth socket in comparison to the physiological healing process. It is an optimized blood clot. It also provides a significant postoperative protection of the surgical site and seems to accelerate the integration and remodeling of the grafted biomaterial.

17. The various cytokines [8] that are involved in PRF are: • Transforming growth factor-β • Platelet-derived growth factor • Vascular endothelial growth factor • Insulin growth factor-1 • Fibroblast growth factor • Epidermal growth factor.

18. Though the classical method for preparation of PRF was given by Dr. Choukroun , the present technique for PRF preparation is legitimized by the French Health Ministry. Currently, PRF is prepared without any use of anticoagulant or bovine thrombin.

19. A PRF with standard quality and quantity of the fibrin matrix, leukocytes, platelets, and growth factors demands a standard protocol for preparation

20. The armamentarium for preparation of PRF includes a PC-02 table centrifuge and a blood collection kit that contains in it a 24G butterfly needle and 9-ml blood collection tubes . The collected patient's blood sample in 10-ml tubes that is devoid of any anticoagulant is centrifuged immediately at a rate of 3000 rpm for 10 min

21. Following the centrifugation, the clot of PRF obtained contains; The highest values of platelets The highest values of the growth factors PDGF, VEGF and TGF A share extremely representative of fibrin, fibronectin and vitronectin About 65% of leukocytes

22. Important studies have found that the PRF can be a node on the local immune regulation, with ability to show a feedback control of the local inflammation. This notion may explain the reduction of postoperative infections when the PRF is used as an additive surgery

23. Current data show that there is a DIFFERENTIAL DISTRIBUTION of red blood cells, platelets, and leukocytes in the PRF clot depending on the CENTRIFUGAL FORCE used. ( Ghanaati S et al 2014) In vitro studies showed that a LONGER CENTRIFUGATION PROTOCOL (2700rpm) produces a DENSER (STRONGER) FIBRIN CLOT with less inter-fibrous space containing less cells compared to the SHORTER CENTRIFUGATION PROTOCOL OF APRF (1300rpm) that produced a LESS DENSE FIBRIN CLOT with a looser inter-fibrous structure containing more cells. ( Ghanaati S et al 2014)

24. Dohan ehrenfest and coworkers found in their in vitro studies that the original L-PRF protocol produces larger clots and membranes, and a more intense release of growth factors than the modified A-PRF protocol. ( dohan ehrenfest dm 2014)

25. In-vivo study showed that choukroun’s new formulation of PRF ( A-PRF ) had a more gradual release of growth factors, up to a 10-day period, and stimulated significantly higher growth factor release over time when compared to choukroun’s standard prf. ( kobayashi e 2016

26. The latter investigators concluded that A-PRF may prove CLINICALLY BENEFICIAL for future regenerative procedures.

27. How to prepare a PRF Membrane? Each fibrin clot concentrates most platelets (97%) and more than half of the leukocytes from a 10-ml blood harvest ( dohan ehrenfest dm) STEP 1 : the prf clot is removed from the tube with a sterile tweezer. STEP 2 : the fibrin clot is separated from the red blood cell fragment, approximately 2mm below the dividing line , using a scissor. The section of the blood clot attached to the fibrin clot contains the stem cells

28. The PRF clots are placed in the PRF BOX and covered with the lid. The PRF membranes are ready for use after 2 minutes A PRF membrane remains USABLE MANY HOURS after preparation, as long as the PRF is prepared correctly and conserved in physiologic condition

29. The use of the prf box is a user-friendly and inexpensive tool, allows for standardized preparation of homogeneous PRF membranes with a higher growth factor content, avoids the dehydration of the leukocytes living in the PRF clot, and also prevents the shrinkage of the fibrin matrix architecture ( dohan ehrenfest dm 2006)

30. Different Ways PRF can be used are

31. The purpose of PRF is to activate and facilitate the healing and regenerative capacity of the host tissue, by providing a strong fibrin scaffold, major growth factors and Allowing space for tissue regeneration. Using PRF as a protective barriers on bone graft sites helps to avoid perforations of the damaged gingival tissues and to prevent associated contamination of the bone graft below.

32. PRF membranes are not comparable to heterologous resorbable collagen or non-resorbable membranes. PRF membranes belong to a completely different category of membrane, namely natural autologous membrane. ( gassling v et al 2010) A PRF membrane is as natural as the host tissue, while heterologous membranes are considered as foreign bodies by the host tissues and interfere with the natural tissue healing process.

33. A PRF MEMBRANE CAN BE USED FOR THREE PURPOSES BIOACTIVE BARRIER A PRF membrane is a blood clot prepared in an optimized form that is rich in cells and growth factors, and acts as a NATURAL BIOACTIVE BARRIER, allowing interaction with the tissues below and above it. This interaction with tissues facilitates NATURAL TISSUE REGENERATION (NTR) and healing. (Del Corso M 2009) PRF will undergo a quicker remodeling (biodegradation) in situ than a resorbable collagen membrane, but will also promote a strong induction on the periosteum/gingival tissue due to the slow release of growth factors and other matrix proteins

34. COMPETITIVE INTERPOSITION BARRIER GTR MEMBRANES are CELL-PROOF BARRIERS against soft tissue invagination, whereas PRF MEMBRANES allow cells to migrate through it, thus allowing new blood vessel formation that will facilitate regenerative and healing interactions between the tissues BELOW AND ABOVE the PRF membrane

35. The PRF matrix becomes the interface between the tissues and therefore avoids the migration of the soft tissues deeper within grafted defect or augmented site. This biological characteristic is referred to as a COMPETITIVE BARRIER. (Del Corso M 2009)

36. However, it is important to recognize that using PRF as a competitive barrier does not have the graft stability or space maintenance characteristics of a normal collagen membrane, and therefore CANNOT BE RECOMMENDED TO USE as such.

37. PROTECTIVE BARRIER AND HEALING BOOSTER PRF membranes are frequently used for the protection of the grafted area and as a healing booster for the soft tissues above the grafted defects or augmented sites (promote the induction of a strong and thick periosteum and gingiva) (Del Corso M 2009) This boosted periosteum functions as a TRUE BARRIER between the soft tissue and bone compartments, and constitutes probably the BEST PROTECTION AND REGENERATIVE BARRIER for the intrabony defects. (Del Corso M 2009)

38. BENEFITS of PRF Natural (autologous) biomaterial while other membranes are considered as foreign bodies by the host tissues and interfere with the natural tissue healing process, a prf membrane is as natural as the host tissue with virtually no risk of infection, immune or a rejection reaction (foreign body response). ( dohan dm et al 2006)

39. EASY AND EFFICIENT TO USE Preparing PRF is EASY, FAST AND USER-FRIENDLY within the daily clinical routine. ( Mazor Z et al 2009)

40. SAFETY AND LOW RISK Blood is drawn from the patient and therefore reduced donor site morbidity. PRF rarely causes complications such as membrane exposure, an unwanted outcome that has been observed in cases using biodegradable barrier membranes. ( Hitti RA 2011) A further advantage of PRF is the extremely low risk of infection. Moreover, no in vitro cytotoxicity effects were detected whatever the quantity of PRF used.(Dohan Ehrenfest DM 2009)

41. INCREASED HEALING POTENTIAL PRF increases the predictability of wound healing and regeneration potential of tissues. (Hauser F et al 2013)

42. REDUCED MORBIDITY A clinical advantage of PRF as a graft material is related to avoidance of a donor site and risk of morbidity, thus resulting in a decrease in patient discomfort, post-surgical pain and bleeding after operation. (Del Corso M 2012) PRF is not only a platelet concentrate but also an ‘IMMUNE NODE’ that is able to stimulate defence mechanisms. (Dohan DM et al 2006)

43. Furthermore, evidence suggest that the content of platelet alpha granules might have a bactericidal effect , mediated by molecules called THROMBOCIDINES that may have an important contribution toward reducing postoperative infections. ( Rozman P 2007) The antihemorrhagic properties (helps in the clotting of blood and prevents hemorrhage) of PRF are also advantageous and convenient during surgical procedures. (Del Corso M 2012)

44. COST-BENEFIT PRF has a potential outstanding cost to benefit RATIO. A platelet-rich fibrin (PRF) membrane is a readily available and inexpensive biomaterial. the ease of preparation and cost-effectiveness of PRF membrane offers a huge advantage over other commercially available membranes. ( Choukroun J 2006) PRF is currently the safest and most economical choice for patients and clinicians for improving healing and regeneration outcomes

45. NO CONTRAINDICATIONS PRF has no contraindications, they can be used in all kinds of patients, especially in patients with systemic conditions where healing is compromised (i.e. diabetics and smokers) or in surgically compromised situations (damaged flap). In these situations PRF will promote soft tissue healing and stimulate the healing of a damaged flap and reduce the risks of flap necrosis after a surgery. All fibrin-based products (platelet concentrates), are frequently used for the stimulation of angiogenesis and to reduce the risk of flap necrosis in many general surgery applications. (Clark RA 2001) (Van Hinsbergh , V.W et al 2001)

46. IMPLNTOLOGY and PRF PRF technology is currently focused in the fields of improving clinical outcomes with: SINUS FLOOR ELEVATIONS USING PRF AS SOLE GRAFTING MATERIAL (Ali S et al 2016) SINUS FLOOR ELEVATION USING A COMBINATION OF PRF AND BONE GRAFT ( Choukroun J et al 2006) ALVEOLAR RIDGE PRESERVATION (SOCKET AUGMENTATION) (Hauser F et al 2013) PERI-IMPLANT TISSUE HEALING (Boora P et al 2015) IMPROVING IMPLANT STABILITY ( Öncu E 2015)

47. In-vitro studies have shown prf -induces gene expression of the early and late markers of osteogenesis thus stimulates bone and soft tissue healing. ( Clipet F et al 2012)

48. SINUS FLOOR ELEVATION USING PRF AS A SOLE OR COMBINATION WITH BONE GRAFTS: A systematic review showed that PRF used as a sole filling material in sinus floor elevation with simultaneous implant placement is a simple technique with promising results. ( kanayama t et al 2016) various clinical case reports describe the lateral approach for sinus floor elevation using only PRF as the grafting material. ( mazor z et al 2009) ( simonpieri a et al 2011)

49. C ase studies have demonstrated that PRF membranes can be used successfully as a protective barrier to cover the sinus membrane during grafting procedures. ( tatullo m et al 2012) ( mazor z et al 2009) PRF membranes also represent an easy and successful method to cover sinus membrane or osteotomy window to protect the SCHNEIDERIAN MEMBRANE , facilitate wound closure and to enhance healing. ( tatullo m et al 2012) cases have also been reported showing that a- prf membrane can be used as a healing barrier when perforations or tears of the schneiderian membrane occur. (diss a et al 2008) ( toffler m et al 2010)

50. ALVEOLAR RIDGE PRESERVATION: use of prf membranes to fill the socket after tooth extraction has shown to improve alveolar bone healing and preservation of the alveolar crest width ( hauser f et al 2013) PRF PLUGS OR MEMBRANES can also be used with compromised extraction sockets (peck mt et al 2011) severe cystic destructions or after cyst enucleations ( choukroun j et al 2006) to allow early bone and gingival regeneration required for implant placement. Clinical and histological findings suggest that filling a fresh extraction socket with PRF provides a viable therapeutic alternative for implant site preparation. ( zhao jh et al 2011)

51. IMMEDIATE POSTEXTRACTION IMPLANT PLACEMENT: PRF can be considered as a healing biomaterial with potential beneficial effect on peri-implant tissue and can be used as a therapeutic adjuvant with immediate implant placement in the clinical scenario of one stage, single tooth implant placement procedure in maxillary anterior region. ( boora p et al 2015) with immediate implant placement the peri-implant jump gap can be augmented with PRF CLOT (A-PRF OR L-PRF) OR SOLUTION (I-PRF) mixed with a bone substitute.( rao sg et al 2013

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