Two Cases of "Quad Fever" in Spinal Cord Injury Department of Emergency and Critical Care Medicine Kurume University School of Medicine, United States
Introduction: Recently, platelet lysate(PL) has gained attention in the field of regenerative medicine. While platelet-derived microvesicles reportedly possess hemostatic abilities similar to platelets, there are only a few standardized protocols for their preparation from PL. Here, three different fractions were obtained from PL and their procoagulant and anticoagulant functions were investigated.
Methods: Sample preparation: Washed platelets were obtained from mouse whole blood and their lysate solutions were prepared using a freeze–thaw method. Three fractions with different properties (large extracellular vesicles [LEVs], small EVs [SEVs], and a protein fraction devoid of EVs [Plt-P]), were isolated from the lysate solutions using ultracentrifugation. Evaluation 1: The coagulation potential of LEVs, SEVs, and Plt-Pin the whole blood was assessed using thromboelastometry (ROTEM®). Diluted mouse whole blood with phosphate buffer saline was supplemented with each fraction as a coagulation trigger, and clotting time (CT), clot formation time (CFT), and maximum clot firmness (MCF) were evaluated. Evaluation 2: The thrombin-producing ability of LEVs, SEVs, and Plt-P in microparticle-free plasma (MpFP) was evaluated using thrombin generation assay (TGA). MpFP was obtained from the preparation of washed platelets, and a thrombin-specific fluorogenic substrate (Z-G-G-R-AMC) was used. Evaluation 3: The effect of Plt-P on thrombin-generation was investigated. The impact of adding Plt-P on thrombin generation was evaluated using TGA triggered by LEVs.
Results: 1. The CT values obtained for LEVs, SEVs, and Plt-P were 194 ± 17 sec, 254 ± 49 sec, 603 ± 59 sec; CFT values were 72 ± 12 sec, 81 ± 12 sec, 174 ± 43 sec; and MCF values were 57 ± 3 mm, 57 ± 3 mm, 49 ± 5 mm , respectively. LEVs exhibited the most significant procoagulant properties, whereas Plt-P showed potential anticoagulant properties. 2. Only LEVs demonstrated concentration-dependent thrombin-generating ability. 3. Plt-P suppressed both the thrombin-generating ability of LEVs and the time to thrombin generation.
Conclusions: Platelet-derived fractions exhibited distinct procoagulant or anticoagulant functions. In trauma cases, the administration of only LEVs fraction extracted from PL may alter future treatment strategies.
