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An optimised protocol for platelet-rich plasma preparation to improve its angiogenic and regenerative properties.

ABSTRACT: Although platelet-rich plasma (PRP) is used as a source of growth factors in regenerative medicine, its effectiveness remains controversial, partially due to the absence of PRP preparation protocols based on the regenerative role of platelets. Here, we aimed to optimise the protocol by analysing PRP angiogenic and regenerative properties. Three optimising strategies were evaluated: dilution, 4?°C pre-incubation, and plasma cryoprecipitate supplementation. Following coagulation, PRP releasates (PRPr) were used to induce angiogenesis in vitro (HMEC-1 proliferation, migration, and tubule formation) and in vivo (chorioallantoic membrane), as well as regeneration of excisional wounds on mouse skin. Washed platelet releasates induced greater angiogenesis than PRPr due to the anti-angiogenic effect of plasma, which was decreased by diluting PRPr with saline. Angiogenesis was also improved by both PRP pre-incubation at 4?°C and cryoprecipitate supplementation. A combination of optimising variables exerted an additive effect, thereby increasing the angiogenic activity of PRPr from healthy donors and diabetic patients. Optimised PRPr induced faster and more efficient mouse skin wound repair compared to that induced by non-optimised PRPr. Acetylsalicylic acid inhibited angiogenesis and tissue regeneration mediated by PRPr; this inhibition was reversed following optimisation. Our findings indicate that PRP pre-incubation at 4?°C, PRPr dilution, and cryoprecipitate supplementation improve the angiogenic and regenerative properties of PRP compared to the obtained by current methods.

SUBMITTER: Etulain J

PROVIDER: S-EPMC5784112 | biostudies-literature | 2018 Jan

REPOSITORIES: biostudies-literature

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An optimised protocol for platelet-rich plasma preparation to improve its angiogenic and regenerative properties.

Etulain Julia J Mena Hebe A HA Meiss Roberto P RP Frechtel Gustavo G Gutt Susana S Negrotto Soledad S Schattner Mirta M

Scientific reports 20180124 1

Although platelet-rich plasma (PRP) is used as a source of growth factors in regenerative medicine, its effectiveness remains controversial, partially due to the absence of PRP preparation protocols based on the regenerative role of platelets. Here, we aimed to optimise the protocol by analysing PRP angiogenic and regenerative properties. Three optimising strategies were evaluated: dilution, 4 °C pre-incubation, and plasma cryoprecipitate supplementation. Following coagulation, PRP releasates (P ...[more]

PMID: 29367608

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Project description:BackgroundThe role of platelet-rich plasma (PRP) in the treatment of sport-related injuries is unclear, largely due to the heterogeneity of clinical results. This may relate to compositional differences in PRP from different separation systems.Questions/purposesThis study aims to compare the composition of PRP produced with five different commercially available systems, focusing on cellular concentrations and pH.MethodsSeven donors (41 ± 12 years) provided blood for PRP preparation using five systems (Arthrex Angel, Emcyte Genesis CS, Arteriocyte Magellan, Harvest SmartPrep, and Biomet GPS III). Post processing, cellular composition was measured including platelets (PLT), white blood cells (WBC), neutrophils (NE), and red blood cells (RBC), as well as pH.ResultsPlatelet concentration and capture efficiency were similar between systems, except the Angel 7% preparation had a greater concentration than Genesis CS (2310 ± 524 vs. 1129 ± 264 k/μL). WBC concentration was variable between systems; however, significant differences were only found between the Angel 2% and GPS III preparations (11.0 ± 4.5, 27.3 ± 7.1 k/μL). NE concentration was significantly lower in the Angel 2% and 7% preparations compared with GPS III (0.6 ± 0.6 and 1.8 ± 1.3 k/μL vs. 9.4 ± 7.0 k/μL). RBC concentration was highest in SmartPrep (3.2 ± 0.6 M/μL) and Genesis CS systems (3.1 ± 0.6 M/μL) compared with all other systems (≤1.1 ± 1.2 M/μL). Finally, pH was significantly lower with the SmartPrep system (6.95 ± 0.06) compared with all others (≥7.26 ± 0.06).ConclusionAside from platelet concentration and capture efficiency, significant compositional differences were identified between preparation systems. Caution should be employed when interpreting clinical results of studies utilizing PRP, as the role of compositional differences and their effect on outcome are unknown. Further study is necessary to determine the clinical significance of these differences.

Project description:The number of studies evaluating platelet-rich plasma (PRP) concentration has substantially grown in the last fifteen years. A systematic review on this field has been realized by evaluating in the identified studies the in vitro PRP concentration-also analyzing the platelet amount-and the in vivo PRP effects in tissue regeneration compared to any control. The protocol has been developed in agreement with the Preferred Reporting for Items for Systematic Reviews and Meta-Analyses-Protocols (PRISMA-P) guidelines. Multistep research of the PubMed, MEDLINE, Embase, PreMEDLINE, Ebase, CINAHL, PsycINFO, Clinicaltrials.gov, Scopus database and Cochrane databases has permitted to identify articles on different concentrations of PRP in vitro and related in vivo impact for tissue repair. Of the 965 articles initially identified, 30 articles focusing on PRP concentration have been selected and, consequently, only 15 articles have been analyzed. In total, 40% (n = 6) of the studies were related to the fixed PRP Concentration Group used a fixed PRP concentration and altered the platelet concentration by adding the different volumes of the PRP (lysate) to the culture. This technique led to a substantial decrease in nutrition available at higher concentrations. Sixty percent (n = 9) of the studies were related to the fixed PRP Volume Group that used a fixed PRP-to-media ratio (Vol/Vol) throughout the experiment and altered the concentration within the PRP volume. For both groups, when the volume of medium (nutrition) decreases, a lower rate of cell proliferation is observed. A PRP concentration of 1.0 × 106 plt/?L, appears to be optimal thanks to the constant and plentiful capillary nutrition supply and rapid diffusion of growth factors that happen in vivo and it also respects the blood decree-law. The PRP/media ratio should provide a sufficient nutrition supply to prevent cellular starvation, that is, PRP ? 10% (Vol/Vol) and thus best mimic the conditions in vivo.

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An optimised protocol for platelet-rich plasma preparation to improve its angiogenic and regenerative properties.

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An optimised protocol for platelet-rich plasma preparation to improve its angiogenic and regenerative properties.

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