Advancing biomaterials of human origin for tissue engineering

The utility of platelet-rich plasma (PRP) has spanned various fields of dermatology from chronic ulcer management to trichology and aesthetics, due to its role in wound healing. Though PRP is being used over a long time, there is still confusion over proper terminology to define, classify and describe the different variations of platelet concentrates. There is also a wide variation in the reported protocols for standardization and preparation of PRP, in addition to lack of accurate characterization of the tested products in most articles on the topic. Additionally, the high cost of commercially available PRP kits, precludes its use over a larger population. In this article, we review the principles and preparation methods of PRP based on available literature and place our perspective in standardizing a safe, simple protocol that can be followed to obtain an optimal consistent platelet yield.

Principles and Methods of Preparation of Platelet-Rich Plasma: A Review and Author's Perspective.

Platelet concentrates for surgical topical applications are nowadays often used, but quantification of the long-term growth factor release from these preparations in most cases is impossible. Indeed, in most protocols, platelets are massively activated and there is no significant fibrin matrix to support growth factor release and cell migration. Choukroun's platelet-rich fibrin (PRF), a second generation platelet concentrate, is a leucocyte- and platelet-rich fibrin biomaterial. Here, we show that this dense fibrin membrane releases high quantities of three main growth factors (Transforming Growth Factor b-1 (TGFbeta-1), platelet derived growth factor AB, PDGF-AB; vascular endothelial growth factor, VEGF) and an important coagulation matricellular glycoprotein (thrombospondin-1, TSP-1) during 7 days. Moreover, the comparison between the final released amounts and the initial content of the membrane (after forcible extraction) allows us to consider that the leucocytes trapped in the fibrin matrix continue to produce high quantities of TGFbeta-1 and VEGF during the whole experimental time.

Slow release of growth factors and thrombospondin-1 in Choukroun's platelet-rich fibrin (PRF): a gold standard to achieve for all surgical platelet concentrates technologies.

The use of platelet concentrates has gained increasing awareness in recent years for regenerative procedures in modern dentistry. The aim of the present study was to compare growth factor release over time from platelet-rich plasma (PRP), platelet-rich fibrin (PRF), and a modernized protocol for PRF, advanced-PRF (A-PRF). Eighteen blood samples were collected from six donors (3 samples each for PRP, PRF, and A-PRF). Following preparation, samples were incubated in a plate shaker and assessed for growth factor release at 15 min, 60 min, 8 h, 1 day, 3 days, and 10 days. Thereafter, growth factor release of PDGF-AA, PDGF-AB, PDGF-BB, TGFB1, VEGF, EGF, and IGF was quantified using ELISA. The highest reported growth factor released from platelet concentrates was PDGF-AA followed by PDGF-BB, TGFB1, VEGF, and PDGF-AB. In general, following 15–60 min incubation, PRP released significantly higher growth factors when compared to PRF and A-PRF. At later time points up to 10 days, it was routinely found that A-PRF released the highest total growth factors. Furthermore, A-PRF released significantly higher total protein accumulated over a 10-day period when compared to PRP or PRF. The results from the present study indicate that the various platelet concentrates have quite different release kinetics. The advantage of PRP is the release of significantly higher proteins at earlier time points whereas PRF displayed a continual and steady release of growth factors over a 10-day period. Furthermore, in general, it was observed that the new formulation of PRF (A-PRF) released significantly higher total quantities of growth factors when compared to traditional PRF. Based on these findings, PRP can be recommended for fast delivery of growth factors whereas A-PRF is better-suited for long-term release.

Comparative release of growth factors from PRP, PRF, and advanced-PRF

Objectives The objective of this study was to analyze the effects of Choukroun's PRF (platelet-rich fibrin), a leucocyte and platelet concentrate clinically usable as fibrin membrane or clot, on human primary cultures of gingival fibroblasts, dermal prekeratinocytes, preadipocytes, and maxillofacial osteoblasts. Study design For the proliferation study, these cells were cultivated with or without a PRF membrane originating from the same donor as for the cells. For osteoblasts and fibroblasts, dose-dependent effect was assessed (using 2 membranes). Cell counts and cytotoxicity tests were performed at 3, 7, 14, and 21 days, and even 28 days for osteoblasts. More osteoblast cultures were prepared in differentiation conditions according to 3 modalities (without PRF, with PRF, with PRF the first day and differentiation medium applied only after the first week of culture). Osteoblast differentiation was analyzed using Von Kossa staining and alkaline phosphatase, DNA and total cell proteins dosage. Results PRF induced a significant and continuous stimulation of proliferation in all cell types. It was dose dependent during all the experiment with osteoblasts, but only on day 14 with fibroblasts. Moreover, PRF induced a strong differentiation in the osteoblasts, whatever the culture conditions. The analysis of osteoblast cultures in differentiation conditions with PRF, using light and scanning electron microscopy, revealed a starting mineralization process in the PRF membrane itself after 14 days. Moreover, PRF leucocytes seemed to proliferate and interact with osteoblasts. Conclusions Cultures with PRF are always cocultures with leucocytes. These “chaperone leucocytes” could be the source of differential geographic regulation within the culture and explain the double contradictory effect proliferation/differentiation observed on osteoblasts.

In vitro effects of Choukroun's PRF (platelet-rich fibrin) on human gingival fibroblasts, dermal prekeratinocytes, preadipocytes, and maxillofacial osteoblasts in primary cultures

Platelet-rich plasma (PRP) is a platelet concentrate that has been used widely to accelerate soft-tissue and hard-tissue healing. The preparation of PRP has been described by several authors. Platelet-rich fibrin (PRF) was first described by Choukroun et al. in France. It has been referred to as a second-generation platelet concentrate, which has been shown to have several advantages over traditionally prepared PRP. Its chief advantages include ease of preparation and lack of biochemical handling of blood, which makes this preparation strictly autologous. This article describes the evolution of this novel platelet concentrate, referred to as PRF.

Platelet-rich fibrin: Evolution of a second-generation platelet concentrate

Purpose: To evaluate and compare the changes in crestal bone height around implants placed with flapless surgery and with-flap surgery. Materials and Methods: Ten implants were placed in six patients - five using flapless and five using with-flap techniques. Single-piece root-form implants and a one-stage approach with immediate nonfunctional loading protocol were used. The change in heights of crestal bone was measured on standardized digital periapical radiographs taken at 0, 1, and 3 months. Results: On mesial side, the mean change from months 0-1, months 1-3, and months 0-3 for flapless method was significantly lower than with-flap method [0.01-0.06 mm for flapless and 0.13-0.40 mm for with-flap ( P = 0.01)]. On the distal side, the mean change from months 0-1, months 1-3, and months 0-3 for flapless method was significantly lower than with-flap method [0.02-0.05 mm for flapless and 0.09-0.30 mm for with-flap ( P = 0.01)]. Conclusions: During the three-month period, reduction of crestal bone height around the implants placed with flapless surgery (0.06 mm) was not statistically significant, while the reduction of crestal bone height around the implants placed using with-flap surgery (0.4 mm) was statistically significant. Comparitively, flapless approach showed lesser crestal bone height reduction, which was statistically significant.

In vivo evaluation of crestal bone heights following implant placement with 'flapless' and 'with-flap' techniques in sites of immediately loaded implants.

E Munirathnam Naidu

Papers

Platelet-rich fibrin: Evolution of a second-generation platelet concentrate

In vivo evaluation of crestal bone heights following implant placement with 'flapless' and 'with-flap' techniques in sites of immediately loaded implants.