Platelet vesicles are potent inflammatory mediators in red blood cell products and washing reduces the inflammatory phenotype.

<bold>Background: </bold>Studies suggest that washing red cell concentrates (RCCs) to remove soluble mediators and/or inflammatory components, such as extracellular vesicles (EVs), may lead to better clinical outcomes. This study tested the hypothesis that non-red blood cell (RBC) generated vesicles in RCC are potent inflammatory mediators in vitro and washing RCCs can reduce these vesicles and subsequently decrease the inflammatory activity of RCCs.<bold>Study Design and Methods: </bold>Sixteen RCCs were pooled and split into four groups based on pre-wash storage time (Day 2 or 14; n = 4/group). Each group was tested 24 hours and 7 days post-wash. Characteristics of RBCs and EVs, cytokines released by monocytes, and expression of human umbilical vein endothelial cells (HUVECs) adhesion molecules were assessed.<bold>Results: </bold>All RCCs meet quality standards for hemolysis, hematocrit, and hemoglobin. Washing did not remove residual platelets from RCCs but led to a significant reduction in platelet-EV count regardless of the group. Supernatant of RCCs washed on Day 14 and stored for 24 hours had significantly lower concentrations of RBC-EVs and white blood cell EVs compared to unwashed controls. Supernatant of unwashed RCCs showed higher production of inflammatory cytokines/chemokines MCP-1, IL-8, and TNF-α, and heightened expression of HUVEC VCAM-1, which were significantly reduced by washing. Spiking washed RCC supernatants with platelet-EVs showed significant increase in IL-8, MCP-1, VCAM-1, and E-selection in groups washed on Day 14.<bold>Conclusions: </bold>Platelet-EVs in RCCs are associated with pro-inflammatory activity. As washing significantly reduced RCC immunomodulatory activity, implementation of this process may improve transfusion outcomes. [ABSTRACT FROM AUTHOR]