COMPARATIVE METHODS FOR PROCESSING PLATELET LYSATE
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Keywords
Platelet lysate; Platelet releaseate; PRP
Abstract
Introduction: Platelets play an important role in numerous physiologic processes through the release of bioactive proteins and growth factors contained in the α granules. These proteins can be collected after physical and chemical processes stimulate their release from platelets. After filtration, this solution contains concentrated growth factors and is then referred to as platelet lysate (PL). Among the several methods used to produce PL, no method has been established as superior in producing the highest concentration of growth factors. This study evaluated six methods of producing PL including CaCl2, freeze-thaw methods at −80˚ C and at −40˚ C, ozone exposure, USb, and USp, along with an “un-activated” Platelet-rich plasma (PRP) sample served as a control. Our goal was to evaluate each sample to determine which method produced the
highest concentration of growth factors.
Methods: PRP was produced from whole blood and subsequently divided into seven samples. One sample served as a control while the other six were used to produce PL by different methods, including ultrasonic probe homogenization, ultrasonic bath homogenization, freeze-thaw at −80˚ C, freeze-thaw at −40˚ C, addition of calcium chloride, and ozone administration.
Outcome measures: The concentration of six growth factors were measured using digital enzyme-linked immunosorbent assay from the produced samples.
Results: PL produced by ultrasonic bath produced the highest concentrations of BDNF, EGF HB-EGF, PDGF-BB, and Vascular Endothelial Growth Factor (VEGF). PL produced by ultrasonic probe produced the highest concentration of IL-1 RA. The concentration of growth factors produced by the ultrasonic bath and probe methods did not differ significantly. The growth factors found in the freeze-thaw methods did not statistically differ from control. Ozone was the least effective at releasing measurable growth factors from PRP. The calcium chloride method resulted in a clotted sample which inhibited growth factor analysis.
Conclusion: The results of this study support the effectiveness of ultrasound homogenization in releasing growth factors from PRP over other current activation methods.
References
2. Mautner K, et al. A call for a standard classification system for future biologic research: the rationale for new PRP nomenclature. PM R, 2015;7(4 Suppl):S53-S59.
3. Hall MP, et al. Platelet-rich plasma: current concepts and application in sports medicine. J Am Acad Orthop Surg 2009;17(10):602-8.
4. Hsu WK, et al. Platelet-rich plasma in orthopaedic applications: evidence-based recommendations for treatment. J Am Acad Orthop Surg 2013;21(12):739-48.
5. Franklin SP, et al. Influence of Cellular Composition and Exogenous Activation on Growth Factor and Cytokine Concentrations in Canine Platelet-Rich Plasmas. Front Vet Sci 2017;4:40.
6. Zimmermann R, et al. Sample preparation technique and white cell content influence the detectable levels of growth factors in platelet concentrates. Vox Sang 2003; 85(4):283-9.
7. Eppley BL, Woodell JE, and Higgins J. Platelet quantification and growth factor analysis from platelet-rich plasma: implications for wound healing. Plast Reconstr Surg 2004;114(6):1502-8.
8. Barsotti MC, et al. Effect of platelet lysate on human cells involved in different phases of wound healing. PLoS One 2013;8(12):e84753.
9. Iudicone P, et al. Pathogen-free, plasma-poor platelet lysate and expansion of human mesenchymal stem cells. J Transl Med 2014;12:28.
10. Borghese C, et al. Clinical-grade quality platelet-rich plasma releasate (PRP-R/SRGF) from CaCl2 -activated platelet concentrates promoted expansion of mesenchymal stromal cells. Vox Sang, 2016;111(2):197-205.
11. Altaie, A., H. Owston, and E. Jones, Use of platelet lysate for bone regeneration - are we ready for clinical translation? World J Stem Cells 2016;8(2):47-55.
12. Alsousou, J., et al., The biology of platelet-rich plasma and its application in trauma and orthopaedic surgery: a review of the literature. J Bone Joint Surg Br 2009;91(8):987-96.
13. Rawson, B., Platelet-Rich Plasma and Epidural Platelet Lysate: Novel Treatment for Lumbar Disk Herniation. J Am Osteopath Assoc 2020;120(3):201-207.
14. Kruger, J.P., et al., Bioactive factors in platelet-rich plasma obtained by apheresis. Transfus Med Hemother 2013;40(6):432-40.
15. Ayhan E, Kesmezacar H and Akgun I. Intraarticular injections (corticosteroid, hyaluronic acid, platelet rich plasma) for the knee osteoarthritis. World J Orthop 2014;5(3):351-61.
16. Rawson, B., Ruptured ACL Repaired with Autologous Platelet-Rich Plasma and Bone Marrow Aspirate Concentrate. EC Orthopaedics 2021;30-34.
17. Qian X, et al., Efficacy and Safety of Autologous Blood Products Compared With Corticosteroid Injections in the Treatment of Lateral Epicondylitis: A Meta-Analysis of Randomized Controlled Trials. PM R 2016;8(8):780-91.
18. Lai LP, et al. Use of Platelet-Rich Plasma in Intra-Articular Knee Injections for Osteoarthritis: A Systematic Review. PM R, 2015;7(6):637-48.
19. Barsotti, M.C., et al., Effect of platelet lysate on human cells involved in different phases of wound healing. PLoS One, 2013;8(12):84753.
20. Amable PR, et al. Platelet-rich plasma preparation for regenerative medicine: optimization and quantification of cytokines and growth factors. Stem Cell Res Ther 2013;4(3):67.
21. Borghese C, et al. Clinical-grade quality platelet-rich plasma releasate (PRP-R/SRGF) from CaCl2 -activated platelet concentrates promoted expansion of mesenchymal stromal cells. Vox Sang 2016.
22. Roffi A, et al. Does platelet-rich plasma freeze-thawing influence growth factor release and their effects on chondrocytes and synoviocytes? Biomed Res Int 2014.
23. Arnoczky SP and Sheibani-Rad S. The basic science of platelet-rich plasma (PRP): what clinicians need to know. Sports Med Arthrosc Rev 2013;21(4):180-5.
24. Mautner K, et al. A call for a standard classification system for future biologic research: the rationale for new PRP nomenclature. PM R 2015;7(4 Suppl):S53-9.
25. Fortunato TM, et al. Platelet lysate gel and endothelial progenitors stimulate microvascular network formation in vitro: tissue engineering implications. Sci Rep 2016;6.
26. Bernardi M, et al. Production of human platelet lysate by use of ultrasound for ex vivo expansion of human bone marrow-derived mesenchymal stromal cells. Cytotherapy 2013;15(8):20-9.
27. Anitua, E., et al., Ozone dosing alters the biological potential and therapeutic outcomes of plasma rich in growth factors. J Periodontal Res 2015;50(2):240-7.
28. Renn TY, et al. Anti-inflammatory effects of platelet biomaterials in a macrophage cellular model. Vox Sang 2015;109(2):138-47.
29. Sonker A and Dubey A. Determining the Effect of Preparation and Storage: An Effort to Streamline Platelet Components as a Source of Growth Factors for Clinical Application. Transfus Med Hemother 2015;42(3):174-80.
30. Anitua E, et al. Autologous platelets as a source of proteins for healing and tissue regeneration. Thromb Haemost 2004;91(1):4-15.
31. do Amaral RJ, et al. Platelet-rich plasma releasate differently stimulates cellular commitment toward the chondrogenic lineage according to concentration. J Tissue Eng 2015;6:2041731415594127.
32. Arakawa T, et al. Factors affecting short-term and long-term stabilities of proteins. Adv Drug Deliv Rev 2001;46(1-3):307-26.
33. Bocci V. Ozone as Janus: this controversial gas can be either toxic or medically useful. Mediators Inflamm 2004;13(1):3-11.
34. Valacchi G and Bocci V. Studies on the biological effects of ozone: 10. Release of factors from ozonated human platelets. Mediators Inflamm 1999;8(4-5):205-9.
35. Pietrzak WS and Eppley BL. Platelet rich plasma: biology and new technology. J Craniofac Surg 2005;16(6):1043-54.
36. Magalon J, et al. DEPA classification: a proposal for standardising PRP use and a retrospective application of available devices. BMJ Open Sport Exerc Med 2016;2(1):e000060.
37. Weibrich G, et al. Growth factor levels in platelet-rich plasma and correlations with donor age, sex, and platelet count. J Craniomaxillofac Surg 2002;30(2):97-102.
38. Xiong G, et al. Men and Women Differ in the Biochemical Composition of Platelet-Rich Plasma. Am J Sports Med;2017:363546517740845.
39. Sheth U, et al. Efficacy of autologous platelet-rich plasma use for orthopaedic indications: a meta-analysis. J Bone Joint Surg Am 2012;94(4):298-307.
40. Miller LE, et al. Efficacy of platelet-rich plasma injections for symptomatic tendinopathy: systematic review and meta-analysis of randomised injection-controlled trials. BMJ Open Sport Exerc Med 2017;3(1):e000237.
41. Andia I and Maffulli N. Platelet-rich plasma for muscle injury and tendinopathy. Sports Med Arthrosc 2013;21(4):191-8.
42. Chahla J, et al. A Call for Standardization in Platelet-Rich Plasma Preparation Protocols and Composition Reporting: A Systematic Review of the Clinical Orthopaedic Literature. J Bone Joint Surg Am 2017;99(20):1769-1779.
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