Transfusion of red cell concentrates (RCCs) is associated with increased risk of adverse outcomes that may be affected by different blood manufacturing methods and the presence of extracellular vesicles (EVs). We investigated the effect of different manufacturing methods on hemolysis, residual cells, cell-derived EVs, and immunomodulatory effects on monocyte activity. Thirty-two RCC units produced using whole blood filtration (WBF), red cell filtration (RCF), apheresis-derived (AD), and whole blood-derived (WBD) methods were examined (n = 8 per method). Residual platelet and white blood cells (WBCs) and the concentration, cell of origin, and characterization of EVs in RCC supernatants were assessed in fresh and stored supernatants. Immunomodulatory activity of RCC supernatants was assessed by quantifying monocyte cytokine production capacity in an in vitro transfusion model. RCF units yielded the lowest number of platelet and WBC-derived EVs, whereas the highest number of platelet EVs was in AD (day 5) and in WBD (day 42). The number of small EVs (<200 nm) was greater than large EVs (≥200 nm) in all tested supernatants, and the highest level of small EVs were in AD units. Immunomodulatory activity was mixed, with evidence of both inflammatory and immunosuppressive effects. Monocytes produced more inflammatory interleukin-8 after exposure to fresh WBF or expired WBD supernatants. Exposure to supernatants from AD and WBD RCC suppressed monocyte lipopolysaccharide-induced cytokine production. Manufacturing methods significantly affect RCC unit EV characteristics and are associated with an immunomodulatory effect of RCC supernatants, which may affect the quality and safety of RCCs.
Exposure to spray-dried solvent/detergent plasma resulted in marked augmentation of monocyte inflammatory cytokine production. Solvent/detergent plasma exposure resulted in the lowest cytokine production, suggesting lower immunomodulatory potential. Further work is needed to determine how these in vitro findings may translate to the bedside.
Extracellular vesicles (EVs) are small, subcellular vesicles that are released from a variety of cells and play important roles in cell-to-cell communication.• Transfused blood products, including red blood cell, platelet, and plasma products contain EVs that are capable of interacting with and altering immune cell function.• The extent to which EVs may contribute to clinically meaningful immunomodulatory effects of transfusion remains unclear and deserving of further study.T he term extracellular vesicle (EV) describes a broad class of small, subcellular vesicular particles that are released from a variety of cell types. Once considered inert debris, emerging data suggest that EVs play pivotal roles in cell-to-cell communication. 1 EVs contain a myriad of biologic response elements, including proteins, nucleic acids, lipid mediators, and other metabolites. While exact mechanisms of EV packaging and release remain unclear, EV content often does not match that of parent cells suggesting differential packaging and release for the purposes of intercellular transport. 2 EV contents also differ based on the mechanism of EV release. There are three main EV categories based on size and mechanism of release. Exosomes are nanoparticles smaller than 0.15 μm released by fusion of the plasma membrane with the endosomal compartment. Microvesicles (also termed ectosomes or microparticles) can be up to 1 to 2 μm and are released by membrane shedding. Finally, apoptotic bodies are the largest particles, ranging approximately 1 to 5 μm, produced from apoptotic cells. While size is often used to differentiate among classes, there is a fair degree of overlap in sizes and at present it is difficult to specifically distinguish EV subpopulations. Once released, EVs are taken up by target cells by a variety of mechanisms, including endocytosis, membrane fusion, micropinocytosis, or phagocytosis. 2 There, EVs are able to regulate a variety of cellular processes. EVs ACCUMULATE IN BLOOD PRODUCTSMuch attention has focused on red blood cell (RBC)-derived EVs as potential mediators of the RBC storage lesion. Membrane vesiculation and EV release are part of the RBC aging process, which is accelerated during hypothermic storage. 3 Multiple reports document accumulation of RBC-derived EVs during storage. 3 It is important to note, however, that RBC products also contain EVs from a variety of cell types, including platelets (PLTs), white blood cells (WBCs), and endothelial cells. Blood collection and component preparation methods significantly impact quantities and composition of EVs released from RBCs and from other cell types. 4,5 There is also a high degree of donor-related variability in EV populations, likely due to differences in donor circulating EV populations and/or in susceptibility of cells to form additional vesicles during component preparation and storage. Thus, each given bag of RBC product may contain vastly different EV populations with accompanying differences in biologic activity.While much of the work on transfusion-related ...
Immunoparalysis in children with septic shock is associated with increased risk of nosocomial infections and death. Myeloid-derived suppressor cells (MDSCs) potently suppress T cell function and may perpetuate immunoparalysis. Our goal was to test the hypothesis that children with septic shock would demonstrate increased proportions of MDSCs and impaired immune function compared with healthy controls.DESIGN: Prospective observational study. SETTING: Fifty-four bed PICU in a quaternary-care children's hospital. PATIENTS:Eighteen children with septic shock and thirty age-matched healthy children. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS:Peripheral blood mononuclear cells (PBMCs) were isolated from whole blood and stained for cell surface markers to identify MDSCs by flow cytometric analysis, including granulocytic and monocytic subsets. Adaptive and innate immune function was measured by ex vivo stimulation of whole blood with phytohemagglutinin-induced interferon (IFN) γ production and lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF)-α production, respectively. Prolonged organ dysfunction (OD) was defined as greater than 7 days. Children with septic shock had a higher percentage of circulating MDSCs, along with lower LPS-induced TNFα and phytohemagglutinininduced IFNγ production capacities, compared with healthy controls. A cut-off of 25.2% MDSCs of total PBMCs in initial samples was optimal to discriminate children with septic shock who went on to have prolonged OD, area under the curve equal to 0.86. Children with prolonged OD also had decreased TNFα production capacity over time compared with those who recovered more quickly (p = 0.02). CONCLUSIONS:This article is the first to describe increased MDSCs in children with septic shock, along with an association between early increase in MDSCs and adverse OD outcomes in this population. It remains unclear if MDSCs play a causative role in sepsis-induced immune suppression in children. Additional studies are warranted to establish MDSC as a potential therapeutic target.
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