Platelet-rich plasma stimulated by pulse electric fields: Platelet activation, procoagulant markers, growth factor release and cell proliferation

Abstract

Therapeutic use of activated platelet-rich plasma (PRP) has been explored for wound healing, hemostasis and antimicrobial wound applications. Pulse electric field (PEF) stimulation may provide more consistent platelet activation and avoid complications associated with the addition of bovine thrombin, the current state of the art ex vivo activator of therapeutic PRP. The aim of this study was to compare the ability of PEF, bovine thrombin and thrombin receptor activating peptide (TRAP) to activate human PRP, release growth factors and induce cell proliferation in vitro. Human PRP was prepared in the Harvest SmartPreP2 System and treated with vehicle, PEF, bovine thrombin, TRAP or Triton X-100. Platelet activation and procoagulant markers and microparticle generation were measured by flow cytometry. Released growth factors were measured by ELISA. The releasates were tested for their ability to stimulate proliferation of human epithelial cells in culture. PEF produced more platelet-derived microparticles, P-selectin-positive particles and procoagulant annexin V-positive particles than bovine thrombin or TRAP. These differences were associated with higher levels of released epidermal growth factor after PEF than after bovine thrombin or TRAP but similar levels of platelet-derived, vascular-endothelial, and basic fibroblast growth factors, and platelet factor 4. Supernatant from PEF-treated platelets significantly increased cell proliferation compared to plasma. In conclusion, PEF treatment of fresh PRP results in generation of microparticles, exposure of prothrombotic platelet surfaces, differential release of growth factors compared to bovine thrombin and TRAP and significant cell proliferation. These results, together with PEF's inherent advantages, suggest that PEF may be a superior alternative to bovine thrombin activation of PRP for therapeutic applications.

Platelet-rich plasma stimulates angiogenesis in mice which may promote hair growth

“…The alpha granules of platelet began to secrete growth factors immediately once activated, and secreted almost all growth factors within an hour [18, 19]. Most PRP protocols in vitro experiments used bovine or autologous thrombin with or without calcium chloride to activate the platelets [8, 20], which was consistent with our study that PRP was activated by exogenous thrombin.…”

Platelet-Rich Plasma for the Treatment of Tissue Infection: Preparation and Clinical Evaluation

“…PEF was more effective than bovine thrombin or TRAP to yield platelet-derived microparticles, EGF, procoagulant particles, and P-selectin-positive particles. 46 In a rabbit model, PRP activated by nanosecond-pulsed electric field-enhanced blood reperfusion in areas of large surgical skin flaps and ischemic hindlimb wounds. 47 The utility of PEF to activate PRP may reduce variability and eliminate immune-mediated complications associated with the use of bovine thrombin, and thus has great clinical potential.…”

Autologous platelet‐rich plasma for healing chronic venous leg ulcers: Clinical efficacy and potential mechanisms

“…There is also evidence that profiles of growth factors can vary depending on the agonist or process used to manufacture APG. For example, pulsed electric field activation of PRP at bipolar low versus monopolar high field strength results in differential platelet‐derived microparticle production and activation of platelet surface pro‐coagulant markers . It is apparent that molecular contents of platelet releasates vary depending on the method used to induce platelet activation and degranulation (summarised in Table ), indicating the type of variation that could occur depending on how APG is produced.…”

“…For example, pulsed electric field activation of PRP at bipolar low versus monopolar high field strength results in differential platelet-derived microparticle production and activation of platelet surface pro-coagulant markers. 32,33 It is apparent that molecular contents of platelet releasates vary depending on the method used to induce platelet activation and degranulation (summarised in Table 1), indicating the type of variation that could occur depending on how APG is produced. It will be important to gain further insights that could also apply to the analysis of other measurable components of PRP in the future.…”