Purpose: Shear-mediated platelet activation (SMPA) results in frequent, serious thrombotic events in patients supported on ventricular assist device (VADs). Despite recognition of supra-physiologic shear stress as an initiator of SMPA details of its mechanism and associated events is only partially understood. Further, current assays of platelet activation typically measure biochemical markers, aggregation, or thrombin generation - which are not necessarily reflective of early processes of SMPA. We hypothesized that SMPA, driven by VADassociated supra-physiologic shear stress, can result in membrane-associated morphologic changes prior to assayed biochemical events. Methods: Fresh human gel filtered-platelets (GFP) were subjected to dynamic shear stress in novel microfluidic platforms fabricated to emulate VADs. Shear exposure included repetitive peaks of 7 Pa x 5 ms followed by shear stress of 1 Pa x 10 ms between consecutive peaks, at a constant flow rate of 15 ul/min, for 100, 200 and 300 peaks of stress exposure - emulating VADs . Platelet activation was assessed for morphologic change via scanning electron microscopy (SEM); and for biochemical activation via thrombin generation via platelet activity state (PAS) assay. Results: Platelets exposed to VAD-associated supra-physiologic shear stresses demonstrated early and progressive membrane-associated morphologic alteration, with increasing pseudopod formation and length (Fig 1B,C). In contrast, despite early morphologic changes platelet activation remained undetectable as measured by the PAS assay (thrombin generation, Fig 1A). Conclusion: SMPA advances via a progression of initial morphological, followed by biochemical signaling and processes. As such a negative PAS assay does not necessarily indicate the absence of mechanical PA or the presence of “resting” platelets. SEM appears to be a sensitive assay revealing the earliest changes in the natural history of SMPA.
Platelet Membrane-Related Morphologic Alterations: An Early Marker of Supra-Physiologic Shear-Mediated Platelet Activation Associated with VADs
CONSOLO, FILIPPO;
2017-01-01
Abstract
Purpose: Shear-mediated platelet activation (SMPA) results in frequent, serious thrombotic events in patients supported on ventricular assist device (VADs). Despite recognition of supra-physiologic shear stress as an initiator of SMPA details of its mechanism and associated events is only partially understood. Further, current assays of platelet activation typically measure biochemical markers, aggregation, or thrombin generation - which are not necessarily reflective of early processes of SMPA. We hypothesized that SMPA, driven by VADassociated supra-physiologic shear stress, can result in membrane-associated morphologic changes prior to assayed biochemical events. Methods: Fresh human gel filtered-platelets (GFP) were subjected to dynamic shear stress in novel microfluidic platforms fabricated to emulate VADs. Shear exposure included repetitive peaks of 7 Pa x 5 ms followed by shear stress of 1 Pa x 10 ms between consecutive peaks, at a constant flow rate of 15 ul/min, for 100, 200 and 300 peaks of stress exposure - emulating VADs . Platelet activation was assessed for morphologic change via scanning electron microscopy (SEM); and for biochemical activation via thrombin generation via platelet activity state (PAS) assay. Results: Platelets exposed to VAD-associated supra-physiologic shear stresses demonstrated early and progressive membrane-associated morphologic alteration, with increasing pseudopod formation and length (Fig 1B,C). In contrast, despite early morphologic changes platelet activation remained undetectable as measured by the PAS assay (thrombin generation, Fig 1A). Conclusion: SMPA advances via a progression of initial morphological, followed by biochemical signaling and processes. As such a negative PAS assay does not necessarily indicate the absence of mechanical PA or the presence of “resting” platelets. SEM appears to be a sensitive assay revealing the earliest changes in the natural history of SMPA.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
