José F. Lana

Relevant Aspects of Centrifugation Step in the Preparation of Platelet-Rich Plasma

Introduction. Platelet-Rich Plasma (PRP) is rich in growth factors, playing important role in tissue healing. The wide variation of reported protocols for preparation of PRP leads to variable compositions, which induce different biological responses and prevent results comparison. This study aims to highlight relevant aspects of the centrifugation step to obtain reproducible results and overall quality. Material and Methods. Samples of blood were collected from 20 healthy donors that have signed free informed consent. Two centrifugation steps (spins) were analyzed for the influence of centrifugal acceleration, time, processed volume, and platelet gradient. The Pure Platelet-Rich Plasma (P-PRP) was characterized as platelet concentration, integrity, and viability (sP-selectin measurement). Results. Lower centrifugal accelerations favour platelet separation. The processing of 3.5 mL of blood at 100 ×g for 10 min (1st spin), 400 ×g for 10 min (2nd spin), withdrawing 2/3 of remnant plasma, promoted high platelet recovery (70–80%) and concentration (5x) maintaining platelet integrity and viability. The recovery of platelets was reduced for a larger WB volume (8.5 mL) processed. Conclusion. Centrifugal acceleration, time, WB processed volume, and minimization of the platelet gradient before sampling are relevant aspects to ensure reproducible compositions within the autologous nature of PRP.

Prediction and Modulation of Platelet Recovery by Discontinuous Centrifugation of Whole Blood for the Preparation of Pure Platelet-Rich Plasma

Abstract The aim of this study was to describe the behavior of the separation of red blood cells (RBCs) by discontinuous centrifugation (DC) of whole blood to modulate and control the platelet recovery in the preparation of pure platelet-rich plasma (P-PRP). P-PRP is a platelet-rich plasma (PRP) in which the white blood cell layer is not included. To achieve this goal, an analytical model was derived that takes into account the packing of RBCs and predicts the behavior of platelet and plasma recovery efficiencies (PtPlRE) based on the volume of whole blood, the hematocrit, and the volume of supernatant, as a function of the operating variables, centrifugal acceleration, and time. The model was derived from the basic equation of DC, which originates from the equilibrium balance of forces on a particle, and included the addition of one factor that corrected the terminal velocity of RBCs and was also correlated to the PtPlRE in the supernatant. This factor was the ratio between the fractional volume concentrations of plasma and RBCs in the centrifugation pellet after centrifugation. The model was validated and the variability of the data was determined using experimental data from 10 healthy donors in the age range of 25–35 years. The predicted behavior for the packing of RBCs and the PtPlRE was consistent with the behavior seen in the experimental data. Thus, the PtPlRE could be modulated and controlled through centrifugal acceleration, time, and hematocrit. Use of this model based on a physical description of events is the first step of a reliable standardization of PRP preparations.

Fibrin network architectures in pure platelet-rich plasma as characterized by fiber radius and correlated with clotting time.

Abstract Fibrin networks are obtained through activation of platelet-rich plasma (PRP) for use in tissue regeneration. The importance of fibrin networks relies on mediation of release of growth factors, proliferation of tissue cells and rheological properties of the fibrin gels. Activation of PRP usually involves the decomposition of fibrinogen by agonists, in a wide range of concentrations. Therefore fibrin networks with a large structural diversity are formed, making comparative evaluations difficult. In order to standardize the fibrin networks, we used the statistical techniques central composite rotatable design and response-surface analysis, to correlate the radius of the fibers with the ratios between the agonists (autologous serum/calcium chloride) and agonist/PRP. From an individual and interactive analysis of the variables, architectures characterized by thick, medium and thin fibers were delineated on the response-surface. Furthermore, the architectures were correlated with coagulation time. This approach is valuable for standardizing the PRP preparation for clinical applications.

Platelet Rich Plasma and Its Growth Factors: The State of the Art

This study aims to offer a general idea of the current progress and discussions about the aspects of technical preparation and biological foundation of PRP for clinical application. We seek to gather the best therapeutic indications that have a scientific foundation on the use of this new tool of Regenerative Medicine. The articles of this study were acquired from the leading data bases of medical literature.

Contributions for classification of platelet rich plasma – proposal of a new classification: MARSPILL

Platelet-rich plasma (PRP) has emerged as a significant therapy used in medical conditions with heterogeneous results. There are some important classifications to try to standardize the PRP procedure. The aim of this report is to describe PRP contents studying celular and molecular components, and also propose a new classification for PRP. The main focus is on mononuclear cells, which comprise progenitor cells and monocytes. In addition, there are important variables related to PRP application incorporated in this study, which are the harvest method, activation, red blood cells, number of spins, image guidance, leukocytes number and light activation. The other focus is the discussion about progenitor cells presence on peripherial blood which are interesting due to neovasculogenesis and proliferation. The function of monocytes (in tissue-macrophages) are discussed here and also its plasticity, a potential property for regenerative medicine treatments.

Platelet-Rich Plasma

This study aims to offer a general idea of the current progress and discussions about the aspects of technical preparation and biological foundation of PRP for clinical application. We seek to gather the best therapeutic indications that have a scientific foundation on the use of this new tool of Regenerative Medicine. The articles of this study were acquired from the leading data bases of medical literature.

Use of Platelet-Rich Plasma (PRP) in Treating Chronic Wounds

The healing process is dynamic and involves complex events that include hemostasis, inflammation, granulation tissue formation, epithelialization, neovascularization, collagen synthesis, and wound contraction. Several experimental clinical studies have demonstrated the reduction of growth factors of chronic wounds. Platelet aggregation has the leading role in the process of skin healing since it is responsible for releasing growth factors, adhesion molecules and lipids, which regulate migration, proliferation and function of keratinocytes, fibroblasts and endothelial cells. The platelet-leukocyte gel (L-PRP), besides releasing the growth factors that start tissue regeneration, can also strengthen the antimicrobial activity, which shows its potential as an infection prevention and treatment agent. PRP is a powerful weapon for treating chronic ulcers, providing healing, reducing infection rates, besides its preventive action, which reduces amputation rates.

Distribution, recovery and concentration of platelets and leukocytes in L-PRP prepared by centrifugation

Platelet-rich plasma (PRP) is an autologous product prepared from whole blood (WB) that is widely used in regenerative medicine. In clinical practice, discontinuous centrifugation is used for both hand- and machine-prepared PRP. However, separation of WB fractions via centrifugation is a complex process, and the lack of clear mechanisms limits the understanding and evaluation of PRP preparation methods This paper focuses on the distribution, recovery and concentration factor of platelets and leukocytes in L-PRP (leukocyte and platelet-rich plasma) to define a concentration pattern for these blood components due to centrifugation conditions. WB collected from three healthy donors was centrifuged for 10min at 50-800 xg in a first step and then at 400 xg in a second step. The results from the first centrifugation step showed most platelets to be distributed in the upper layer (UL) and the buffy coat (BC), with approximately 14.5±5.2% retained in the bottom layer (BL). Most leukocytes were present in the BL. The greatest platelet recoveries from L-PRP were obtained at up to 150 xg (88.5±16.9%). The cumulative concentration factors with respect to the WB from the second centrifugation step were 6 and 1.2 for platelets and leukocytes, respectively. Thus, the concentration patterns delineated three centrifugation ranges with platelet/leukocyte ratios of 205±18, 325±15 and 107±4 and lymphocyte/granulocyte ratios of 1.54±0.74, 0.90±0.08 and 0.42±0.07. These findings contribute to a scientifically based standardization of L-PRP preparations.

Platelet-Rich Plasma and Tissue Engineering

The focus in this chapter is an evaluation, based in data from the literature, of the use of Platelet-Rich Plasma (PRP) in cell culture for treatment of bone and cartilage defects. For the preparation of this chapter the data bank from PubMed, developed by the National Center for Biotechnology Information, U.S. National Library of Medicine was used as a tool. Key words used were: platelet-rich plasma, mesenchymal stem cells, biomaterials, tissue engineering; cartilage repair; bone healing. The role of PRP on tissue regeneration, cell proliferation and mesenchymal stem-cells is emphasized. In vitro and in vivo studies of PRP in bone and cartilage regeneration are described and referenced.

Challenges and a Feasible Strategy for Studies and Standardization of Platelet-Rich Plasma

Platelet-rich plasma (PRP) is currently used in regenerative medicine and has opened up an exciting field of study. In this chapter, we present the main challenges limiting the study and standardization of PRP, based on variables and interactions involved in its preparation. Furthermore, we present and discuss some approaches to basic science studies of PRP from the simplest case to the most complex or real scenario. A feasible strategy for studying P-PRP (Pure-PRP) preparation in a real situation is also illustrated. The key point of the strategy is the predictive description of the main steps of PRP preparation, based on the physicochemical events. Within the expected variability for an autologous product, the quality of PRP can be modulated by preparation conditions and also tailored for evaluation of biological responses. The interconnected characterization and standardization of the quality and biological properties of PRP are the basis for further clinical studies