CD47 on erythrocytes inhibits phagocytosis through interaction with the inhibitory immunoreceptor SIRP␣ expressed by macrophages. Thus, the CD47-SIRP␣ interaction constitutes a negative signal for erythrocyte phagocytosis. However, we report here that CD47 does not only function as a "do not eat me" signal for uptake but can also act as an "eat me" signal. In particular, a subset of old erythrocytes present in whole blood was shown to bind and to be phagocytosed via CD47-SIRP␣ interactions. Furthermore, we provide evidence that experimental aging of erythrocytes induces a conformational change in CD47 that switches the molecule from an inhibitory signal into an activating one. Preincubation of experimentally aged erythrocytes with human serum before the binding assay was required for this activation. We also demonstrate that aged erythrocytes have the capacity to bind the CD47-binding partner thrombospondin-1 (TSP-1) and that treatment of aged erythrocytes with a TSP-1-derived peptide enabled their phagocytosis by human red pulp macrophages. Finally, CD47 on erythrocytes that had been stored for prolonged time was shown to undergo a conformational change and bind TSP-1. These findings reveal a more complex role for CD47-SIRP␣ interactions in erythrocyte phagocytosis, with CD47 acting as a molecular switch for controlling erythrocyte phagocytosis. (Blood. 2012;119(23):5512-5521) IntroductionErythrocyte clearance has been studied for many years in the context of normal turnover, 1,2 enhanced clearance resulting from defects in erythrocyte metabolism or changes in membrane composition, [3][4][5] and in blood transfusion. 6 A large number of factors that may influence erythrocyte clearance have been proposed, ranging from autoantibody binding to band 3 after conformational changes resulting from aging (the "senescent cell antigen"), 7,8 to the expression of phosphatidylserine on the outer leaflet of the erythrocyte membrane. 9 In general, it is thought that erythrocytes are cleared after accumulating so-called "eat me" signals and are then phagocytosed by macrophages of the reticuloendothelial system, predominantly in the spleen and liver. 1,10,11 In contrast, one of the membrane proteins expressed by erythrocytes, CD47, has been shown to inhibit phagocytosis of erythrocytes by macrophages of the reticuloendothelial system. 12-16 CD47 exerts its inhibitory effect through binding to SIRP␣ on the macrophage, which induces inhibitory signaling by the immunoreceptor tyrosinebased inhibition motifs (ITIMs) residing in the cytoplasmic tail of SIRP␣. 17,18 On ligation of SIRP␣ by CD47, the SIRP␣ ITIMs recruit and activate tyrosine phosphatases SHP-1 and SHP-2, and this regulates, generally in a negative fashion, downstream signaling pathways and effector functions. The inhibitory effect of CD47 on erythrocyte clearance can be illustrated by transfusion of CD47-deficient erythrocytes into a wild-type recipient, which leads to rapid phagocytosis of the CD47-deficient erythrocytes by red pulp macrophages in the spleen. 12,19 ...

The use of the diversion bag and, to a lesser extent, the use of double swabs with 70 percent isopropyl alcohol, led to a reduction of contamination. As expected, predominant contamination with resident skin bacteria was reduced. The combination of diversion bag and new disinfection led to a frequency of initial positive results for pooled five-donor PCs, which is similar to that of single-donor apheresis PCs. Furthermore, the bacterial detection system and associated product recall procedures have been shown to be effective in preventing transfusion of contaminated PCs and/or related red cells, especially for rapidly growing bacteria.

The effect of rapid cooling to 20-24 degrees C of whole blood immediately after collection, using 'cooling units' with butane-1,4-diol and prolonged storage up to 24 h at ambient temperature was investigated in the whole blood and the subsequently prepared plasma, buffy coat and buffy-coat-poor red cell concentrate (BC-poor RCC) in saline-adenine-glucose-mannitol (SAG M) solution. Factor VIII:C content of the plasma (n = 10), after 24 h storage was 80 +/- 3% of the initial value. In routine procedures factor VIII:C content in the plasma (n = 129 pools of 20 donor units plasma) was 0.77 +/- 0.078 IU/ml, after storage of the whole blood for 16-20 h. In whole blood (n = 10), the 2,3-diphosphoglycerate (2,3-DPG) content of the red cells decreased from 4.36 +/- 0.55 to 1.47 +/- 0.6 mumol/ml red cells after 24 h storage at 20-24 degrees C. After storage of the BC-poor RCC (n = 10) at 2-6 degrees C for 1 week, the 2,3-DPG had dropped to 0.76 +/- 0.46 mumol/ml red cells. During the first 24 h of storage of whole blood, the adenine triphosphate (ATP) levels of the red cells remained stable. A mean increase of 20% of the initial value was observed after addition of SAG M solution. In the BC-poor RCC the ATP slowly decreased to 81 +/- 5% after 5 weeks and to 68 +/- 6.6% of the initial value after 6 weeks storage.(ABSTRACT TRUNCATED AT 250 WORDS)

Key Points FCA is a novel flow cytometry–based platelet aggregation assay that allows single receptor analysis in small volume/thrombocytopenic samples FCA facilitates platelet studies in experimental animal models even during gestation and allows kinetic measurements in individual animals

This study shows that at low hematocrit, the oxygen-delivering capacity of human red blood cells stored 5-6 wks is reduced compared with fresh cells and red blood cells stored for an intermediate period. Although red blood cells stored for 2-3 wks are completely devoid of 2,3-diphosphoglycerate, their oxygen-delivering capacity to the intestines was the same as fresh red blood cells. Our study showed that red blood cell deformability was preserved during storage, suggesting that other mechanisms may account for the observed decrease in oxygen delivery by red blood cells stored 2-3 wks.

Resuspension of RBCs in our alkaline medium PAGGG-M resulted in a RBC unit of high quality during storage for up to at least 35 days, with 2,3-DPG levels of higher than 10 mumol per g Hb, hemolysis of less than 0.2 percent, and ATP levels of higher than 5 mumol per g Hb.

No abstract

Diversion of the first 10 ml of blood was shown to contribute significantly to a reduction in the prevalence of superficial skin bacteria in whole-blood units. In our opinion, blood collection systems should be adapted to use the first 10-30 ml of a whole-blood donation for testing purposes.