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BACKGROUND: Transfusion of stored red blood cells (RBCs) can be associated with adverse side effects. Recent studies in mice transfused with stored RBCs showed that a strong proinflammatory cytokine storm was induced due to extravascular hemolysis already at 2 hours after transfusion. Therefore, we here investigated if transfusion of 2 units of cryopreserved autologous RBCs induced a proinflammatory response in healthy human volunteers.

STUDY DESIGN AND METHODS: Two units of autologous RBCs, cryopreserved for 16 weeks, were transfused into 10 healthy human volunteers. Serum and blood samples taken at 2 hours before and at 2 and 48 hours after transfusion were analyzed for signs of extravascular hemolysis and the presence of proinflammatory cytokines.

RESULTS: At 2 hours after transfusion, transferin-bound serum iron, as well as transferin saturation and total bilirubin, were already significantly increased. These measures all returned back toward that in pretransfusion samples at 48 hours after transfusion. No increases in the production of the proinflammatory cytokines interleukin (IL)-1β, IL-6, IL-8, monocyte chemotactic protein-1, macrophage inflammatory protein-1β, or tumor necrosis factor-α were detected at any time point after transfusion.

CONCLUSION: Although a significant level of extravascular hemolysis already occurred at 2 hours after transfusion of cryopreserved RBCs, there were no signs of proinflammatory cytokine production up to 48 hours after transfusion.

Normally senescent red blood cells (RBCs) have a rather fixed life-span after which they are eliminated from the circulation mainly by macrophages in the spleen and liver. However, the normal life-span may be significantly reduced when RBCs are stored under blood bank conditions, which ultimately results in a reduced 24 hour survival after transfusion. Although not completely understood, the damage occurring to some of the stored RBCs is probably multifactorial as stored RBCs shares features of both senescence and suicidal RBC death (eryptosis). One key cellular event associated with eryptosis is an increased intracellular Ca2+-concentration. We found that human RBCs stored for up to 42 days under blood bank conditions contained a small subset of cells with an increased intracellular Ca2+-concentration corresponding to that during eryptosis. Since little is known about the mechanisms mediating uptake and clearance of eryptotic RBCs in vivo, we further investigated this using a murine transfusion model of Ca2+ ionophore-treated RBCs (Ca2+-RBCs). Ca2+-RBCs were mainly trapped by MARCO+ splenic marginal zone macrophages and CD11c+ dendritic cells (DCs) already at 1 hour after transfusion. Among DCs, CD11c+ CD207+ DCs in the marginal zone were particularly efficient in mediating uptake of Ca2+-RBCs. Similar to that found in vitro, CD47 on the Ca2+-RBCs did not affect their clearance in vivo. Thus, RBCs with an increased intracellular Ca2+-concentration accumulates during RBC storage, and in a murine model such RBCs are recognized by splenic macrophages and DCs in ways similar to what has been reported for nucleated apoptotic cells.

BACKGROUND: Red blood cells (RBCs) undergo structural and metabolic changes with prolonged storage, which ultimately may decrease their survival after transfusion. Although the storage-induced damage to RBCs has been rather well described biochemically, little is known about the mechanisms underlying the recognition and rapid clearance of the damaged cells by macrophages.STUDY DESIGN AND METHODS: We here used a murine model for cold (+4°C) RBC storage and transfusion. Phagocytosis of human or murine RBCs, liquid stored for 6-8 weeks or 10-14 days respectively, was investigated in murine peritoneal macrophages.RESULTS: The effects of storage on murine RBCs resembled that described for stored human RBCs with regard to decreased adenosine triphosphate (ATP) levels, accumulation of microparticles during storage, and RBC recovery kinetics after transfusion. Under serum-free conditions, phagocytosis of stored human or murine RBCs was reduced by 70-75%, as compared with that in the presence of heat-inactivated fetal calf serum (FCS). Human serum promoted phagocytosis of stored human RBCs similar to that seen with FCS. By blocking macrophage class A scavenger receptors with fucoidan or dextran sulphate, phagocytosis of human or murine RBCs was reduced by more than 90%. Phagocytosis of stored human RBCs was also sensitive to inhibition by the phosphatidylinositol 3 kinase-inhibitor LY294002, the ERK1/2-inhibitor PD98059, or the p38 MAPK-inhibitor SB203580.CONCLUSIONS: RBCs damaged during liquid storage may be recognized by macrophage class A scavenger receptors and serum-dependent mechanisms. This species-independent recognition mechanism may help to further understand the rapid clearance of stored RBCs shortly after transfusion.