Vesicles Generated during Storage of Red Blood Cells Enhance the Generation of Radical Oxygen Speciesin Activated Neutrophils

Abstract

Erythrocytes are known to shed vesiclesin vivo, under various conditionsin vitro, and, with impact for transfusion medicine, during storage of red blood cell concentrates (Vstovesicles). Vstovesicles of blood transfusions have been shown to deliver glycosylphosphatidylinositol-linked proteins to recipient erythrocytes, to display prothrombotic activity, and to have an inhibitory effect on macrophages. The interaction of Vstovesicles with and their effect on neutrophilic granulocytes has not yet been studied in detail. Fluorescentlylabeled Vstoand calcium-induced vesicles were preparedin order to study the uptake of labeled vesicular components by neutrophils as compared to the process of phagocytosis of zymosan using flow cytometry and confocal microscopy. The activating effect of Vstovesicles on neutrophils was addressed by a luminometric assay for stimulated radical oxygen species (ROS) generation. Coincubation of vesicles and neutrophils results in a transfer of vesicular components to the cells. This uptake is different from a phagocytotic process and is enhanced upon interference with the cellular actin cytoskeleton. Preincubation of neutrophils with Vstovesicles results in an enhanced ROS generation by neutrophils, which is further increased upon fMLP stimulation and during zymosan phagocytosis. The activating effect of Vstovesicles on neutrophils might be due to the specific accumulation of lysophospholipidsin Vstovesicles and should be considered as a possible contributor to the pathogenesis of transfusion-related acute lung injury.