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Red blood cell (RBC) transfusion is common in critically ill, postsurgical, and posttrauma patients in whom both systemic inflammation and immune suppression are associated with adverse outcomes. RBC products contain a multitude of immunomodulatory mediators that interact with and alter immune cell function. These interactions can lead to both proinflammatory and immunosuppressive effects. Defining clinical outcomes related to immunomodulatory effects of RBCs in transfused patients remains a challenge, likely due to complex interactions between individual blood product characteristics and patient-specific risk factors. Unpacking these complexities requires an in-depth understanding of the mechanisms of immunomodulatory effects of RBC products. In this review, we outline and classify potential mediators of RBC transfusion-related immunomodulation and provide suggestions for future research directions.

BACKGROUND We have previously shown that critically ill children transfused with red blood cells (RBCs) of longer storage durations have more suppressed monocyte function after transfusion compared to children transfused with fresher RBCs and that older stored RBCs directly suppress monocyte function in vitro, through unknown mechanisms. We hypothesized that RBC-derived microvesicles (MVs) were responsible for monocyte suppression. STUDY DESIGN AND METHODS To determine the role of stored RBC unit–derived MVs, we cocultured monocytes with supernatants, isolated MVs, or supernatants that had been depleted of MVs from prestorage leukoreduced RBCs that had been stored for either 7 or 30 days. Isolated MVs were characterized by electron microscopy and flow cytometry. Monocyte function after coculture experiments was measured by cytokine production after stimulation with lipopolysaccharide (LPS). RESULTS Monocyte function was suppressed after exposure to supernatants from 30-day RBC units compared to monocytes cultured in medium alone (LPS-induced tumor necrosis factor-α production, 17,611 ± 3,426 vs. 37,486 ± 5,598 pg/mL; p = 0.02). Monocyte function was not suppressed after exposure to MV fractions. RBC supernatants that had been depleted of MVs remained immunosuppressive. Treating RBC supernatants with heat followed by RNase (to degrade protein-bound RNA) prevented RBC supernatant–induced monocyte suppression. CONCLUSION Our findings implicate soluble mediators of stored RBC-induced monocyte suppression outside of MV fractions and suggest that extracellular protein-bound RNAs (such as microRNA) may play a role in transfusion-related immunomodulation.

Red blood cell (RBC) transfusion is common in critically ill, postsurgical, and posttrauma patients in whom both systemic inflammation and immune suppression are associated with adverse outcomes. RBC products contain a multitude of immunomodulatory mediators that interact with and alter immune cell function. These interactions can lead to both proinflammatory and immunosuppressive effects. Defining clinical outcomes related to immunomodulatory effects of RBCs in transfused patients remains a challenge, likely due to complex interactions between individual blood product characteristics and patient-specific risk factors. Unpacking these complexities requires an in-depth understanding of the mechanisms of immunomodulatory effects of RBC products. In this review, we outline and classify potential mediators of RBC transfusion-related immunomodulation and provide suggestions for future research directions.

Background Our previous in vitro work showed that stored red blood cells (RBCs) increasingly suppress markers of innate immune function with increased storage time. This multicenter prospective observational study tests the hypothesis that a single RBC transfusion in critically ill children is associated with immune suppression as a function of storage time. Study Design and Methods Blood samples were taken immediately before and 24 (±6) hours after a single RBC transfusion ordered as part of routine care. Innate and adaptive immune function was assessed by ex vivo whole blood stimulation with lipopolysaccharide (LPS) and phytohemagglutinin, respectively. Monocyte HLA-DR expression, regulatory T cells, plasma interleukin (IL)-6, and IL-8 levels were also measured. Results Thirty-one transfused critically ill children and eight healthy controls were studied. Critically ill subjects had lower pretransfusion LPS-induced tumor necrosis factor-α production capacity compared to healthy controls, indicating innate immune suppression (p

BACKGROUND: Reduced monocyte function is associated with adverse outcomes from critical illness. Red blood cells (RBCs) are thought to impair monocyte function but relationships between RBC storage solution and monocyte suppression are unknown. This study was designed to test the hypothesis that immunosuppressive effects of RBCs on monocytes are related to both storage time and preservative solution. STUDY DESIGN AND METHODS: Monocytes from healthy adult donors were co-cultured with RBCs that had been stored in AS-1, AS-3, or CPD only for 7, 14, or 21 days. Cells were then stimulated with lipopolysaccharide (LPS) and their supernatants assayed for tumor necrosis factor (TNF)-α and interleukin (IL)-10. Transwell experiments were performed to evaluate the role of cell-to-cell contact. Monocyte mRNA expression was quantified by real-time–polymerase chain reaction. RESULTS: LPS-induced TNF-α production capacity was reduced compared to controls for all groups, but CPD-only RBCs suppressed monocyte function more than RBCs stored in AS-1 (p = 0.007) and AS-3 (p = 0.006). IL-10 production was preserved or augmented in all groups. A longer storage time was associated with reduced TNF-α production capacity for AS-1 and AS-3 groups but not CPD. Preventing cell-to-cell contact did not eliminate the inhibitory effect of RBCs on monocyte responsiveness. RBC exposure was associated with decreased LPS-induced TNFA mRNA expression (p

We have previously shown that critically ill children transfused with red blood cells (RBCs) of longer storage durations have more suppressed monocyte function after transfusion compared to children transfused with fresher RBCs and that older stored RBCs directly suppress monocyte function in vitro, through unknown mechanisms. We hypothesized that RBC-derived microvesicles (MVs) were responsible for monocyte suppression.To determine the role of stored RBC unit-derived MVs, we cocultured monocytes with supernatants, isolated MVs, or supernatants that had been depleted of MVs from prestorage leukoreduced RBCs that had been stored for either 7 or 30 days. Isolated MVs were characterized by electron microscopy and flow cytometry. Monocyte function after coculture experiments was measured by cytokine production after stimulation with lipopolysaccharide (LPS).Monocyte function was suppressed after exposure to supernatants from 30-day RBC units compared to monocytes cultured in medium alone (LPS-induced tumor necrosis factor-α production, 17,611 ± 3,426 vs. 37,486 ± 5,598 pg/mL; p = 0.02). Monocyte function was not suppressed after exposure to MV fractions. RBC supernatants that had been depleted of MVs remained immunosuppressive. Treating RBC supernatants with heat followed by RNase (to degrade protein-bound RNA) prevented RBC supernatant-induced monocyte suppression.Our findings implicate soluble mediators of stored RBC-induced monocyte suppression outside of MV fractions and suggest that extracellular protein-bound RNAs (such as microRNA) may play a role in transfusion-related immunomodulation.

Our previous in vitro work showed that stored red blood cells (RBCs) increasingly suppress markers of innate immune function with increased storage time. This multicenter prospective observational study tests the hypothesis that a single RBC transfusion in critically ill children is associated with immune suppression as a function of storage time.Blood samples were taken immediately before and 24 (±6) hours after a single RBC transfusion ordered as part of routine care. Innate and adaptive immune function was assessed by ex vivo whole blood stimulation with lipopolysaccharide (LPS) and phytohemagglutinin, respectively. Monocyte HLA-DR expression, regulatory T cells, plasma interleukin (IL)-6, and IL-8 levels were also measured.Thirty-one transfused critically ill children and eight healthy controls were studied. Critically ill subjects had lower pretransfusion LPS-induced tumor necrosis factor-α production capacity compared to healthy controls, indicating innate immune suppression (p 0.0002). Those who received RBCs stored for not more than 21 days demonstrated recovery of innate immune function (p = 0.02) and decreased plasma IL-6 levels (p = 0.002) over time compared to children transfused with older blood, who showed persistence of systemic inflammation and innate immune suppression. RBC storage time was not associated with changes in adaptive immune function.In this pilot cohort of critically ill children, transfusion with older prestorage leukoreduced RBCs was associated with persistence of innate immune suppression and systemic inflammation. This was not seen with fresher RBCs. RBC transfusion had no short-term association with adaptive immune function. Further studies are warranted to confirm these findings in a larger cohort of patients.