Open AccessTunable activation of therapeutic platelet-rich plasma by pulse electric field: Differential effects on clot formation, growth factor release, and platelet morphology
Author(s) -
Andrew L. Frelinger,
Anja J. Gerrits,
V.B. Neculaes,
Thomas Gremmel,
Andrew S. Torres,
Anthony Caiafa,
Sabrina L. Carmichael,
Alan D. Michelson
Publication year - 2018
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0203557
Subject(s) - platelet , platelet rich plasma , platelet activation , thrombin , chemistry , thromboelastography , growth factor , platelet derived growth factor receptor , granule (geology) , biophysics , immunology , materials science , biochemistry , medicine , biology , receptor , composite material
Background Activation of platelet-rich plasma (PRP) by pulse electric field (PEF) releases growth factors which promote wound healing ( e . g ., PDGF, VEGF for granulation, EGF for epithelialization). Aims To determine after PEF activation of therapeutic PRP: 1) platelet gel strength; 2) profile of released growth factors; 3) alpha- and T-granule release; 4) platelet morphology. Methods Concentrated normal donor PRP was activated by 5 μsec (long) monopolar pulse, ~4000 V/cm (PEF A) or 150 nsec (short) bipolar pulse, ~3000 V/cm (PEF B) in the presence of 2.5 mM (low) or 20 mM (high) added CaCl 2 . Clot formation was evaluated by thromboelastography (TEG). Surface exposure of alpha granule (P-selectin) and T-granule (TLR9 and protein disulfide isomerase [PDI]) markers were assessed by flow cytometry. Factors in supernatants of activated PRP were measured by ELISA. Platelet morphology was evaluated by transmission electron microscopy (TEM). Results Time to initial clot formation was shorter with thrombin (<1 min) than with PEF A and B (4.4–8.7 min) but clot strength (elastic modulus, derived from TEG maximum amplitude) was greater with PEF B than with either thrombin or PEF A (p<0.05). Supernatants of PRP activated with PEF A had higher EGF levels than supernatants from all other conditions. In contrast, levels of PF4, PDGF, and VEGF in supernatants were not significantly different after PEF A, PEF B, and thrombin activation. T-granule markers (TLR9 and PDI) were higher after thrombin than after PEF A or B with low or high CaCl 2 . By TEM, platelets in PEF-treated samples retained a subset of granules suggesting regulated granule release. Conclusion Pulse length and polarity can be modulated to produce therapeutic platelet gels as strong or stronger than those produced by thrombin, and this is tunable to produce growth factor profiles enhanced in specific factors important for different stages of wound healing.