The development of surgical adjuvants for the local stimulation of healing is an important field of research in biomaterial and pharmaceutical sciences. The first act of healing associates many actors, first of all platelets, leuko- cytes, fibrin matrix and many growth factors. All these actors work in synergy during the coagulation process, and many products logically tried to mimic these natural mechanisms in order to improve healing on a surgical site. This trend started many years ago with fibrin glues (autologous, or more frequently allogeneous) [1, 2], and evolved recently with platelet concentrates technologies [3]. The concept of these latter [4] is to use a centrifugation procedure (often in 2 steps) in order to concentrate and collect most platelets from a blood harvest (taken with anticoagulant), and to inject them into a wounded site in order to improve healing. The activa tion of the product (with thrombin for example) induces the platelet growth factors release and the polymerization of fibrinogen (released by platelets or free in the plasma) into fibrin [5], leading to a platelet gel.

Platelet concentrates and fibrin glues are thus in fact not so different, both being based on similar technologies of po- lymerization of blood fibrinogen into a fibrin gel [3, 6]. However the concept of platelet-rich product was often wrongly summarized by one magical concept: growth fac- tors. This craze for growth factors often led to forget the key roles of the other actors present in these blood-derived prod- ucts: between mystical belief, commercial interests and sci- entific truth, for many researchers in this field it required several years to accept the key role of fibrin [7, 8] and some- times leukocytes in these products. When considering the complexity of coagulation and healing, growth factors are only actors among many others. The platelet concentrates are more complex products than classical pharmaceutical prepa- rations, because their clinical effects are dependent on the intrinsic versatile and adaptive characteristics of the patient blood and on the numerous -and often not investigated- ac- tors contained in these products [9]. Platelet concentrates are in fact blood concentrates, and their biology is as complex as blood itself. These products are living biomaterials, and are more difficult to handle and evaluate than synthetic materials shaped on demand.

The terminologies used in the field thus significantly suffered from the initial confusions about these products and mystical belief in platelet growth factors. The craze for plate- let concentrates technologies started first in oral and maxillo- facial surgery after the first article of Marx et al. [10]. These authors used the term « Platelet-Rich Plasma (PRP) », in reference to the term used for platelet concentrates in hema- tology for the treatment of patients suffering from severe thrombopenia. The “platelet-rich plasma (PRP)” term was first used in 1954 by Kingsley to designate thrombocyte con- centrate during blood coagulation experiments [11], and is still used today in blood transfusion science.
However, the concept of platelet concentrate for topical use was in fact much older than the article of Marx and co- workers. A few years after the first publications of Matras about fibrin glues [1], autologous preparations called
« platelet-fibrinogen-thrombin mixtures » were tested in oph- thalmology [12, 13], general surgery [14] and neurosurgery [15]. Another author called it « gelatin platelet (gel foam) » [16]. These platelet-rich products were only used as fibrin tissue adhesives, and not as healing stimulators: the platelets were only supposed to support a stronger fibrin polymeriza- tion, and therefore a more efficient tissue sealing than basic fibrin glues. Growth factors and healing properties were then not considered.

A first clinical demonstration that platelet concentrates promote healing locally was reported by Knighton et al. In
1986 [17]. These authors used the term “platelet-derived wound healing factors” (PDWHF) for a preparation pro- duced using an empirical 2-step centrifugation procedure. Using this protocol, these authors treated 49 patients with chronic non-healing cutaneous ulcers and reported good out- comes. In other articles in 1988 and 1990 [18, 19], these authors used a slightly different term to define their tech- nique: “platelet-derived wound healing formula (PDWHF)”. These authors already used the term “platelet-rich plasma” as a general term of transfusion medicine, but it did not refer to the final product for clinical use itself. A few years later, Whitman et al. [20] presented their clinical results in oral and maxillofacial surgery, using a platelet concentrate that was produced by a gradient density cell separator from the hematology laboratory. It was named PRP during collection, but the authors understood that the final product was a fibrin gel, and therefore named it “platelet gel”. Unfortunately, in all these studies, the exact content of these products re- mained unknown, particularly concerning the leukocyte con- tent. Only platelets were quantified.
The use of the “PRP” term thus truly started with Marx et al. [10], in a first study about the use of these platelet-rich products during bone graft reconstruction in maxillofacial surgery. The clinical results for bone healing were very in- teresting and sustained the use of these technologies in this field. This PRP terminology was maybe too general, but it was adequate: the PRP from Marx et al. Was produced using a cell separator from the hematology laboratory, and there- fore was very similar to a professional PRP for transfusion. But this product was also used as a fibrin gel, since it was injected during activation with bovine thrombin. Moreover, the leukocyte content of the product used by these authors was not confirmed.

From this moment, the term “PRP” was used every- where, whatever the platelet concentrate technology was [3]. During the following years, most studies have employed various in-house protocols, where the basic two-step cen- trifugation process was modified with regard to centrifuga- tion forces (from 160g to 3000g) and time (from 3 to 20 min for the different centrifugation steps). The definition of these parameters frequently seems to be empirical, and cross- examination of these technical data is an impasse. In most articles, the various platelet concentrates were not correctly described or characterized, particularly concerning their cell contents. Most authors were somehow considering that all platelet concentrates were the same, whatever was inside.
This confusing situation created incredible inconsisten- cies in the literature. It is frequent to see articles where authors quantified the growth factor content of these prod- ucts, but without activating the platelets [21, 22]: obviously it cannot lead to relevant results and conclusions. The re- leasates are often tested, but without investigating the key role of the fibrin matrix in the release kinetics [21-24], even if this parameter was proved to be a considerable issue [25,
26]. It is also frequent to read in vitro cell studies where PRP and cell lineages in culture are coming from 2 different do- nors [21, 22], or even 2 different species [23, 24]: consider- ing that many platelet concentrates also contain leukocytes, cell cultures with platelet concentrates are often cocultures with leukocytes [27, 28], but many authors did not take this key issue into consideration. As a whole, the enormous lit- erature on the field is very difficult to review, and the pub- lished results are so contradictory that this literature could be called a blind library of knowledge.

2. IN SEARCH FOR A BETTER TERMINOLOGY: A SLOW EVOLUTION

The vagueness of the terminology and the associated lack of characterization of the various platelet concentrates were key issues, and several actors of this field tried to identify and define clearly some of these products.

First, some companies started to use specific commercial names for their products in order to give them a better visi- bility. In 1999, Anitua (Biotechnology Institute, BTI, Vito- ria, Spain, a dental implant company) described a manual version of the original PRP technology, using a one-step centrifugation process and several pipetting steps [29]. The final product was called PRGF (Plasma [29] or Preparation [30] Rich in Growth Factors), and contained no leukocyte and lower platelet and growth factor concentrations than other products [31]. It could be activated with calcium chlo- ride in order to produce a PRGF gel. Other companies also tried to use similar strategies, such as Vivostat PRF (Platelet- Rich Fibrin): this latter is in fact only another variation of the cell separator PRP technology [4]. Using the name of a commercial product to define a specific version of the plate- let concentrate technology is a smart solution and of great help for a better understanding of the data published in the literature. Unfortunately, it also raises serious suspicions of commercialism [32], since these names are not based on the

consensual classification of a scientific reality, but only shaped by the need of commercial communication.

Some techniques however were so different from the initial PRP concept, that the need for a different name be- came obvious and natural. When Choukroun et al. Described in 2001 the Platelet-Rich Fibrin (PRF) technique [33], it was considered as a second-generation platelet concentrate tech- nology [3, 34-38]. Indeed Choukroun’s PRF shows very dif- ferent characteristics than the other products: blood is col- lected without anticoagulant and centrifuged immediately, leading to the natural formation of a strong PRF clot in the middle of the tube (without the addition of any adjuvants or triggering factors). PRF is a solid biomaterial [39-42], an optimized natural blood clot [43, 44], and not an injectable liquid platelet suspension like the PRPs. The platelet activa- tion and the fibrin polymerization are not the final step of the process, they are the process [43]. With the invention of this technique, the key role of the fibrin matrix in these technolo- gies became an obvious issue [3], even if the confusions per- sisted. At least, it was obvious that platelet concentrate tech- nologies resulting in strong fibrin biomaterials could not be called PRP anymore. But another issue was raised by the studies on PRF: PRF also contains leukocytes [36, 43]. The immune parameters started to be investigated and the key role of leukocytes was hypothesized and pointed out [25, 27,
28].

Obviously, the fibrin architecture and leukocyte content could not be forgotten. An important series of opinions and original articles challenged the spineless consensus on the historical PRP term and gave the opportunity to create a bet- ter terminology.

In Bielecki et al.’s opinion in 2006 [45], the authors in- sisted on the different forms of PRP used in clinical practice: PRP could be injected without thrombin (for example in chronic severe elbow tendinosis)[46], but was more often used after activation with thrombin (or another activator) resulting in a gelatinous mass formation. These authors therefore proposed to define Platelet-Rich Plasma as an inac- tive substance, while “platelet-rich gel” (PRG) represented the biologically active fibrin matrix rich in platelets, leuko- cytes and related active molecules (growth factors, serotonin, catecholamines, von Willebrand factor, proaccelerin, os- teonectin and others released from the platelet granules). The presence of leukocytes was also pointed out, but no other specific terminology was proposed. In 2008, Everts et al. Completed this first opinion [47], insisting on the leukocyte content and the 2 forms (activated/non activated) of these products. A more accurate terminology was proposed: the inactivated product was named platelet-leukocyte rich plasma (P-LRP), and the activated gel was named platelet- leukocyte gel (PLG).

The proposals of these 2 groups were significant contri- butions for the clarification of these products, and these authors used these new terminologies in several following articles [48-57]. However, this terminology remained in- complete [32]: indeed, some PRPs do not contain leukocytes [31], and non activated PRPs cannot be considered as inac- tive products, their full activation is only delayed and more natural (after the contact with the tissues on the site of injec- tion)[46]. Moreover, even after activation, these gels (PRG,

PLG or whatever the name) never reach the degree of natural fibrin polymerization obtained in the PRF subfamily [43,
58].

The final step in this terminology history was the defini- tion of a full classification system for all platelet concen- trates [4]. Four families were simply designed, based on their leukocyte and fibrin content. Liquid platelet concentrate sus- pensions (before activation) were named PRP: “Pure Plate- let-Rich Plasma” (P-PRP) without leukocytes, “Leukocyte- and Platelet-Rich Plasma” (L-PRP) with leukocytes. On the contrary, solid platelet concentrate biomaterials, with a strong fibrin architecture (and therefore always activated), were called PRF: “Pure Platelet-Rich Fibrin” (P-PRF) with- out leukocytes, “Leukocyte- and Platelet-Rich Fibrin” (L-PRF) with leukocytes. The main commercially available technologies were classified following these principles (Table 1). This classification is probably the more simple and accurate [32], but a few clarifications are still necessary in order to find a consensus terminology.

3. FINDING A CONSENSUS TERMINOLOGY

A terminology must be simple, accurate and pragmatic. It also must avoid commercial interests and thus remain scien- tifically homogeneous [59]. For clarity and to be user- friendly, we should avoid the multiplication of pointless ac- ronyms that only create confusions. Following these princi- ples, a consensus on a simple terminology has been reached and summarized in the Table 1.
First of all, all the products of this category are « platelet concentrates ». This is the general term that defines all these products: activated or not, with or without leukocytes, all these products concentrate the platelets.
Second, the previous classification from 2009 [4] should be considered as a good and user-friendly system to classify products by their leukocyte content and fibrin architecture. Indeed, considering that most of the literature on the topic used the acronym PRP, and since this term is still valid for some of these products, this acronym should be completed but conserved in order to keep the coherence of the database. When searching « PRP » articles on Medline, « P-PRP » and
« L-PRP » are immediately identified. For this reason, the term « L-PRP » should be preferred to « P-LRP » for this subfamily, even if both are correct from a strict meaning standpoint.
« PRG » [45] and « PLG » [47] acronyms are too general and should therefore be avoided. Indeed from a strict defini- tion standpoint, P-PRF and activated P-PRP are both Plate- let-Rich Gels, and L-PRF and activated L-PRP are both Platelet-Leukocyte Gels, but the PRP and PRF families show two completely different fibrin matrix architectures [5, 43,
58]. Moreover PRG and PLG also contain plasma in the gel, as a colloidal solution trapped within the fibrin mesh. For all these reasons, the activated version of a P-PRP should sim- ply be a « P-PRP gel » (and not a PRG), and the activated version of a L-PRP should simply be a « L-PRP gel » (and not a PLG). The 2 PRF subfamilies only exist in the gel form, so they do not need a more accurate terminology.

FULL ARTICLE IS AVAILABLE DOWN

Authors
David M. Dohan Ehrenfest, Tomasz Bielecki, Allan Mishra, Piero Borzini, Francesco Inchingolo, Gilberto Sammartino, Lars Rasmusson and Peter A. Everts