A new assay to evaluate microvesicle plasmin generation capacity: validation in disease with fibrinolysis imbalance. Issue 1 (16th July 2018)

• Record Type: Journal Article
• Authors: Cointe, Sylvie; Harti Souab, Karim; Bouriche, Tarik; Vallier, Loris; Bonifay, Amandine; Judicone, Coralie; Robert, Stéphane; Armand, Romain; Poncelet, Philippe; Albanese, Jacques; Dignat‐George, Françoise; Lacroix, Romaric
• Is Part Of: Journal of extracellular vesicles. Volume 7:Issue 1(2018)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2018-07-16
• Subjects: Extracellular vesicles -- microvesicles -- fibrinolysis -- immunomagnetic separation -- septic shock; Cells -- Mechanical properties -- Periodicals; Transport Vesicles; Cells -- Mechanical properties; Periodicals; Fulltext; Internet Resources; Periodicals; Electronic journals; Periodicals; 571.63
• DOI: 10.1080/20013078.2018.1494482
• Languages: English
• ISSNs: 2001-3078

Abstract

ABSTRACT: Among extracellular vesicles, leukocyte‐derived microvesicles (LMVs) have emerged as complex vesicular structures. Primarily identified as procoagulant entities, they were more recently ascribed to plasmin generation capacity (MV‐PGC). The objectives of this work were (1) to develop a new hybrid bio‐assay combining the specific isolation of LMVs and measurement of their PGC, and compare its performance to the original method based on centrifugation, (2) to validate MV‐PGC in septic shock, combining increased levels of LMVs and fibrinolytic imbalance. Using plasma sample spiked with LMVs featuring different levels of PGC, we demonstrated that CD15‐beads specifically extracted LMVs. The MV dependency of the test was demonstrated using electron microscopy, high speed centrifugation, nanofiltration and detergent‐mediated solubilization and the MV‐PGC specificity using plasmin‐specific inhibitors, or antibodies blocking elastase or uPA. Thanks to a reaction booster (ε‐ACA), we showed that the assay was more sensitive and reproducible than the original method. Moreover, it exhibited a good repeatability, inter‐operator and inter‐experiment reproducibility. The new immunomagnetic bio‐assay was further validated in patients with septic shock. As a result, we showed that MV‐PGC values were significantly lower in septic shock patients who died compared to patients who survived, both at inclusion and 24 h later (1.4 [0.8–3.0] vs 3.1 [1.7–18] A 405 × 10 −3 /min, p = 0.02; 1.4 [1–1.6] vs 5.2 [2.2–16] A 405 × 10 −3 /min, p = 0.004). Interestingly, combining both MV‐PGC and PAI‐1 in a ratio significantly improved the predictive value of PAI‐1. This strategy, a hybrid capture bioassay to specifically measure LMV‐PGC using for the first time, opens new perspectives for measuring subcellular fibrinolytic potential in clinical settings with fibrinolytic imbalance.