NO对不同天龄红细胞保存质量影响的初步研究

目的探讨一氧化氮(NO)对采用percoll密度梯度离心法分离的不同天龄红细胞(RBC)功能状态的影响作用。方法 1)将Percoll细胞分离液配制成5个不连续密度梯度(1.091,1.098 5,1.106,1.113 5,1.121 g/ml),将RBC按平均密度大小分层,依次吸取每层RBC分别做RBC内丙酮酸激酶(PK)含量测定并计算天龄、RBC内ATP含量检测、RBC变形性评价,以及RBC膜蛋白Western Blot。2)从同1份滤白全血中取出2等份,1份为实验组:加适宜浓度的NO溶液,另1份为对照组:加等体积红细胞保存液Ⅲ,分别于保存初期、中期、末期检测2组血液及不同天龄RBC的功能。结果 1)同1份样品经密度梯度离心分离后,RBC随着密度的增大,PK活性逐渐减小(6.192±1.25、5.165±0.84、4.538±0.76,P0.05)、天龄逐渐增加(10.409、34.957、49.945)、ATP含量逐渐减少(4.755±0.037、3.242±0.445、2.929±0.153,P0.01)、RBC变形性逐渐降低(200-1切变率下,0.381±0.005、0.340±0.033、0.281±0.028、P0.05)、RBC膜带3蛋白量减小、聚簇化的带3蛋白量增多、膜上结合的IgG含量增多。2)NO组与对照组相比,全血各指标无明显差异,但NO组保存中、末期老年RBC变形性明显高于对照组(200-1切变率下,中期:0.290±0.021 vs 0.276±0.021、0.229±0.024 vs 0.211±0.027;末期:0.277±0.017 vs0.263±0.019、0.213±0.038 vs 0.193±0.039,P0.01)。结论 RBC功能随密度的增大逐渐降低;NO可明显改善老年RBC的变形性,但对全血无明显影响。     

Transfusion of stored red blood cells adhere in the rat microvasculature

BACKGROUND: Ex vivo storage of red blood cells (RBCS) for transfusions is associated with a “storage lesion,” which decreases RBC deformability and increases RBC adhesiveness to vascular endothelium. This may impair microcirculatory flow with deleterious effects on oxygen delivery after transfusion. Previous studies have shown that human RBCs adhere to endothelial monolayers in vitro with prolonged storage and is reduced by prestorage leukoreduction (LR). The objective of this study was to determine whether duration of RBC storage and LR influence RBC adhesion in vivo in capillaries. STUDY DESIGN AND METHODS: Rat RBCs were collected and stored in CPDA‐1 under standard blood bank conditions. Three RBC products were compared: 1) fresh RBCs, less than 24 hours of storage (n = 6); 2) nonleukoreduced (NLR) RBCs stored for 7 days (n = 6); and 3) prestorage LR RBCs stored for 7 days (n = 6). RBCs were labeled with fluorescein isothiocyanate (FITC) 24 hours before transfusion and reinjected in an isovolemic manner into healthy rats. The FITC‐labeled RBCs were visualized in the extensor digitorum longus muscle using intravital video microscopy (20× magnification). The number of RBCs adherent in capillaries was counted 1 hour after transfusion in 10 random fields and the median values were compared with one‐way analysis of variance. RESULTS: Stored RBCs showed increased levels of adherence in capillaries compared to their fresh counterparts (p < 0.05). Prestorage LR decreased RBC adherence to levels equivalent to those of fresh RBCs (p < 0.05 for stored LR vs. stored NLR). CONCLUSION: Rat RBCs stored under conditions that closely mimicked clinical transfusion adhere in capillaries. The decreased RBC adherence with LR suggest a direct effect of white blood cells or their byproducts on RBC deformability and/or adhesiveness to microvascular endothelium. Further study will examine the mechanism of adherence and the impact it has on microcirculatory flow and oxygen delivery in the critically ill host.     

Unit-to-unit variability in the deformability of red blood cells

BackgroundIn blood banking practice, the storage duration is used as the primary criterion for inventory management, and usually, the packed red blood cells (PRBC) units are supplied primarily according to first-in-first-out (FIFO) principle. However, the actual functionality of individual PRBC units is mostly ignored. One of the main features of the RBCs not accounted for under this approach is the deformability of the red cells, i.e., their ability to affect the recipients' blood flow. The objective of the study was to analyze unit-to-unit variability in the deformability of PRBCs during their cold storage.MethodsRBC samples were obtained from twenty leukoreduced PRBC units, stored in SAGM. The deformability of cells was monitored from the day of donation throughout 42 days. RBC deformability was determined using the computerized cell flow-properties analyzer (CFA) based on cell elongation under a shear stress of 3.0 Pa, expressed by the elongation-ratio (ER). The image analysis determines the ER for each cell and provides the ER distribution in the population of 3000–6000 cells.ResultsThe deformability of freshly-collected RBCs exhibited marked variability already on the day of donation. We also found that the aging curve of PRBC deformability varies significantly among donors.SignificanceThe present study has demonstrated that storage duration is only one of the factors, and seemingly not even the major one, affecting the PRBCs functionality. Therefore, the FIFO approach is not sufficient for assessing the potential transfusion outcome, and the PRBC functionality should be determined explicitly for each unit.     

A comparison of biochemical and functional alterations of rat and human erythrocytes stored in CPDA-1 for 29 days: implications for animal models of transfusion

Animal models of transfusion are employed in many research areas yet little is known about the storage-related changes occurring in the blood used in these studies. This study assessed storage-related changes in red blood cell (RBC) biochemistry, function and membrane deformability in rat and human packed RBCs when stored in CPDA-1 at 4 degrees C over a 4-week period. Human blood from five volunteers and five bags of rat RBC concentrates (five donor rats per bag) were collected and stored at 4 degrees C. RBC function was assessed by post-transfusion viability and the ability to regenerate adenosine triphosphate (ATP) and 2,3-diphosphoglycerate (DPG) when treated with a rejuvenation solution. Membrane deformability was determined by a micropipette aspiration technique. ATP in rat RBCs declined more rapidly than human RBCs; after 1 week rat ATP fell to the same level as human cells after 4 weeks of storage (rat, 2.2 +/- 0.2 micromol g(-1) Hb; human, 2.5 +/- 0.3 micromol g(-1) Hb). Baseline DPG concentrations were similar in rat and human RBCs (16.2 +/- 2.3 micromol g(-1) Hb and 13.7 +/- 2.4 micromol g(-1) Hb) and declined very rapidly in both species. Human RBCs fully regenerated ATP and DPG when treated with a rejuvenation solution after 4 weeks of storage. Rat RBCs regenerated ATP but not DPG. Post-transfusion viability in rat cells was 79%, 26% and 5% after 1, 2 and 4 weeks of storage, respectively. In rats, decreased membrane deformability became significant (- 54%) after 7 days. Human RBC deformability decreased significantly by 34% after 4 weeks of storage. The rejuvenation solution restored RBC deformability to control levels in both species. Our results indicate that rat RBCs stored for 1 week in CPDA-1 develop a storage lesion similar to that of human RBCs stored for 4 weeks and underscores significant species-specific differences in the structure and metabolism of these cells.     

Loss of red cells from the use of blood warmers during apheresis procedures

In-line blood warmers increase the extracorporeal volume of apheresis procedures. A saline rinse of the blood warmer may be employed to minimize the loss of red blood cells (RBCs) from this extra volume. We measured the amount of RBCs remaining in three different brands of blood warmers at the end of a COBE Spectra rinseback to determine the clinical significance of blood warmer saline rinses. The volume of RBCs removed by a warmer rinse ranged from 15 to 24 ml per procedure and was statistically different among the three brands of warmers. The COBE warmer contained significantly less RBCs than either the Fenwal or Pharmaseal warmers. Patient hematocrit was not highly correlated with the residual RBC volume (r = 0.30). We estimate that 15–20 procedures are required before the equivalent of one unit of packed RBCs is lost to blood warmers. We conclude that the blood warmer saline rinse may be safely omitted for most patients. J. Clin. Apheresis 12:122–125, 1997. © 1997 Wiley-Liss, Inc.     

Extracellular Vesicles in Red Blood Cell Concentrates: An Overview

Red blood cell (RBC) concentrates may be stored for up to 42 days before transfusion to a patient. During storage extracellular vesicles (EVs) develop and can be detected in significant amounts in RBC concentrates. The concentration of EVs is affected by component preparation methods, storage solutions, and inter-donor variation. Laboratory investigations have focused on the effect of EVs on in vitro assays of thrombin generation and immune responses. Assays for EVs in RBC concentrates are not standardized. The aims of this review are to describe the factors that determine the presence of erythrocyte-EVs in RBC concentrates, the current techniques used to characterize them, and the potential role of EV analysis as a quality control maker for RBC storage.     

The effects of cryopreservation on red blood cell rheologic properties

BACKGROUND: In transfusion medicine, frozen red blood cells (RBCs) are an alternative for liquid‐stored RBCs. Little is known about the rheologic properties (i.e., aggregability and deformability) of thawed RBCs. In this study the rheologic properties of high‐glycerol frozen RBCs and postthaw stored in saline‐adenine‐glucose‐mannitol medium were compared to those of conventionally liquid‐stored and fresh RBCs. STUDY DESIGN AND METHODS: Fresh RBCs were obtained from healthy volunteers. Leukoreduced liquid‐stored and thawed deglycerolized RBC units were obtained from the Sanquin Blood Bank. RBCs were tested for aggregability (aggregation index [AI]), deformability (elongation index [EI]), and various hematologic variables. RESULTS: The AI of thawed RBCs was reduced, compared to fresh and liquid‐stored RBCs (p < 0.05). The EI of stored RBCs was significantly enhanced over a shear stress range of 2.0 to 50 Pa compared to fresh RBCs (p < 0.05). No significant differences in EI between thawed and 21‐ or 35‐day liquid‐stored RBCs were observed. The osmotic fragility, hemolysis, mean cell volume, and mean cell hemoglobin concentration of thawed RBCs were markedly altered, compared to fresh and liquid‐stored RBCs (p < 0.05). The adenosine triphosphate (ATP) content of thawed RBCs was similar to 3‐ or 21‐day liquid‐stored and fresh RBCs. CONCLUSIONS: Thawed RBCs are more fragile than conventionally liquid‐stored and fresh RBC. The freeze‐thaw‐wash process, however, did not adversely affect the aggregability and deformability or the ATP content of thawed RBCs. Based on the rheologic properties, cryopreserved RBCs are a valuable alternative to liquid‐stored RBCs.     

Blood banking-induced alteration of red blood cell flow properties.

BACKGROUND: Blood banking procedures are associated with damage to red blood cell (RBC) membranes, which can impair their flow properties, namely, their deformability, aggregability, and adherence to endothelial cells (ECs) and thus possibly introducing a circulatory risk to recipients. This study was undertaken to comprehensively explore the effect of cold storage and gamma irradiation on RBC flow properties. STUDY DESIGN AND METHODS: RBC flow properties were monitored as a function of shear stress with a computerized cell flow properties analyzer. Because we had previously studied storage effect on RBC aggregability (Transfusion 1999;39:277-81), here we determined the storage effect on RBC adherence and deformability, by measuring them before (control) and during storage. Gamma irradiation effect on RBC aggregability, adherence, and deformability was determined before (control) and after irradiation. RESULTS: Cold storage significantly elevated the number of adherent RBCs and the strength of their interaction with ECs, and was marked by decreased RBC deformability as early as 2 weeks into the storage period. The elevation of RBC-EC interaction was well correlated with translocation of phosphatidylserine to the RBC surface. Gamma irradiation induced an immediate and marked increase in the number of rigid cells, but did not affect RBC adherence and aggregability. CONCLUSION: RBC flow properties appear to be especially sensitive to cold storage and gamma irradiation because they are impaired long before the expiration date. Because impaired RBC flow properties facilitate circulatory disorders, the potential circulatory risk of transfusion RBC with blood banking-impaired rheology should be considered.     

Blood banking–induced alteration of red blood cell flow properties

BACKGROUND: Blood banking procedures are associated with damage to red blood cell (RBC) membranes, which can impair their flow properties, namely, their deformability, aggregability, and adherence to endothelial cells (ECs) and thus possibly introducing a circulatory risk to recipients. This study was undertaken to comprehensively explore the effect of cold storage and gamma irradiation on RBC flow properties.
STUDY DESIGN AND METHODS: RBC flow properties were monitored as a function of shear stress with a computerized cell flow properties analyzer. Because we had previously studied storage effect on RBC aggregability (Transfusion 1999;39:277-81), here we determined the storage effect on RBC adherence and deformability, by measuring them before (control) and during storage. Gamma irradiation effect on RBC aggregability, adherence, and deformability was determined before (control) and after irradiation.
RESULTS: Cold storage significantly elevated the number of adherent RBCs and the strength of their interaction with ECs, and was marked by decreased RBC deformability as early as 2 weeks into the storage period. The elevation of RBC–EC interaction was well correlated with translocation of phosphatidylserine to the RBC surface. Gamma irradiation induced an immediate and marked increase in the number of rigid cells, but did not affect RBC adherence and aggregability.
CONCLUSION: RBC flow properties appear to be especially sensitive to cold storage and gamma irradiation because they are impaired long before the expiration date. Because impaired RBC flow properties facilitate circulatory disorders, the potential circulatory risk of transfusion RBC with blood banking–impaired rheology should be considered.     

The viability of autologous human red cells stored in additive solution 5 and exposed to 25°C for 24 hours

BACKGROUND: No data exist on the viability of red cells (RBCs) stored in modern additive solution systems and allowed to warm above 10 degrees C. STUDY DESIGN AND METHODS: In a randomized crossover study, 3 units of blood were collected at least 8 weeks apart from 11 volunteer donors and stored in additive solution 5 (AS-5). Of 3 units from each volunteer, 1 was stored for 6 weeks at 4 degrees C, 1 for 5 weeks at 4 degrees C except for 24 hours at 25 degrees C on Day 14, and 1 for 5 weeks at 4 degrees C except for 24 hours at 25 degrees C on Day 28. Units were sampled periodically during storage; at the end of storage, viability was measured by the 99mTc/51 Cr double-label method. RESULTS: RBC viability was not significantly different in the storage protocols. Less than 1 percent of stored cells hemolyzed. RBC ATP concentrations at the end of storage correlated with viability and were approximately equal in the warmed units after 30 days' storage and the conventionally stored units after 42 days. CONCLUSIONS: The data suggest that RBCs stored in AS-5 and allowed to warm to 25 degrees C for 24 hours lose about 12 days of their shelf life.     

Abnormal membrane physical properties of red cells in McLeod syndrome

McLeod red cells (RBCs) lack Kx antigens and have weak expression of the Kell antigens. Individuals who carry the McLeod phenotype have acanthocytic RBCs and a compensated hemolytic state. To elucidate the role of the protein on which the Kx antigens reside in maintaining membrane deformability, the rheologic properties of McLeod RBCs were determined by ektacytometry. RBCs were obtained from normal individuals and from four patients with McLeod syndrome. Osmotic gradient deformability profiles of McLeod RBCs showed decreased whole cell deformability. Resealed ghosts from McLeod RBCs also showed decreased deformability, partly because of the decreased cell surface area and partly because of an intrinsic membrane stiffness in this syndrome. For the measurement of membrane mechanical stability, resealed ghosts were subjected to constant high shear stress in the ektacytomer, and deformability was recorded continuously as the deformable ghosts fragmented into rigid spherical vesicles. Membranes from McLeod RBCs showed a noticeable increase in mechanical stability. Acquired causes of acanthocytosis, such as liver disease, did not cause the rheologic abnormalities observed in McLeod cells. Other abnormalities noted in McLeod RBCs were decreased RBC potassium content and an increased number of dense RBCs, as determined by centrifugation on a discontinuous density gradient. The data indicate that McLeod RBCs are rigid and have decreased surface area and that their membranes are intrinsically rigid with increased mechanical stability. These abnormalities may account for the reduced RBC survival observed in McLeod syndrome. The protein that carries the Kx surface antigen seems to be required for the maintenance of the normal physical function of RBC skeletal proteins.     

Alteration of Deformability of the Erythrocyte Membrane in Stored Blood

The deformability of stored erythrocytes decreases progressively with duration of storage. This change can be measured in terms of mean pressure required to deform the cells sufficiently to cause them to enter and traverse a micropipette analogue of the microcirculation. The disc shape, with relatively higher surface area to volume ratio, is more deformable than the spherical form; and mean deformability at specific time intervals correlates with reported posttransfusion survival values. Only 76 per cent of red blood cells stored for three weeks were able to pass through a pipette with a minimal dimension of 2.85 μ, a dimension through which all fresh cells pass. This dimension is of magnitude similar to that previously reported for the microcirculation of the spleen. Regeneration of cellular ATP by incubation in adenosine is associated with restoration of mean deformability toward the normal range; and the majority of cells, having regained the biconcave disc shape, has values for individual cell deformability similar to fresh cells. The remaining spherical cells retain their relatively decreased deformability.     

Leukoreduction makes a difference: A pair proteomics study of extracellular vesicles in red blood cell units

Prestorage filtration of blood to remove contaminating donor leukocytes and platelets has substantially increased the safety level of transfusion therapy. We have previously shown that leukoreduction has a mitigating effect on the storage lesion profile by lowering the extent of hemolysis and of RBC aging and removal phenotypes, including surface signaling and microvesiculation. Even though protein composition may determine the fate of EVs in the recipient, the probable effect of leukoreduction on the EV proteome has been scarcely investigated. In the present paired study, we characterized the proteome of EVs released in prestorage leukoreduced (L) and nonleukoreduced (N) RBC units prepared from the same donors, by immunoblotting and qualitative proteomics analyses at two storage intervals. Apart from common proteofrms typically associated with the established EV biogenesis mechanisms, the comparative proteomics analyses revealed that both leukoreduction and storage duration affect the complexity of the EV proteome. Membrane and cytoskeleton-related proteins and regulators, metabolic enzymes and plasma proteins exhibited storage duration dependent variation in L- and N-EVs. Specific proteoforms prevailed in each EV group, such as transferrin in L-units or platelet glycoproteins, leukocyte surface molecules, MHC HLA, histones and tetraspanin CD9 in N-units. Of note, several unique proteins have been associated with immunomodulatory, vasoregulatory, coagulatory and anti-bacterial activities or cell adhesion events. The substantial differences between EV composition under the two RBC preparation methods shed light in the underlying EV biogenesis mechanisms and stimuli and may lead to different EV interactions and effects to target cells post transfusion.     

Is red blood cell rheology preserved during routine blood bank storage?

BACKGROUND: Red blood cell (RBC) units stored for more than 2 weeks at 4°C are currently considered of impaired quality. This opinion has primarily been based on altered RBC rheologic properties (i.e., enhanced aggregability, reduced deformability, and elevated endothelial cell interaction), during prolonged storage of nonleukoreduced RBC units. In this study, the rheologic properties and cell variables of leukoreduced RBC units, during routine blood bank storage in saline‐adenine‐glucose‐mannitol, were investigated. STUDY DESIGN AND METHODS: Ten leukoreduced RBC units were stored at the blood bank for 7 weeks at 4°C. RBCs were tested weekly for aggregability, deformability, and other relevant variables. RESULTS: RBC aggregability was significantly reduced after the first week of storage but recovered during the following weeks. After 7 weeks aggregability was slightly, but significantly, reduced (46.9 ± 2.4‐44.3 ± 2.2 aggregation index). During storage the osmotic fragility was not significantly enhanced (0.47 ± 0.01% phosphate‐buffered saline) and the deformability at shear stress of 3.9 Pa was not significantly reduced (0.36 ± 0.01 elongation index [EI]). The deformability at 50 Pa was reduced (0.58 ± 0.01‐0.54 ± 0.01 EI) but remained within reference values (0.53 ± 0.04). During 5 weeks of storage, adenosine triphosphate was reduced by 54% whereas mean cell volume, pH, and mean cell hemoglobin concentration were minimally affected. CONCLUSIONS: RBC biochemical and physical alterations during storage minimally affected the RBC ability to aggregate and deform, even after prolonged storage. The rheologic properties of leukoreduced RBC units were well preserved during 7 weeks of routine blood bank storage.     

人血浆对红细胞保存的影响

为了研究人血浆对保存的红细胞结构和功能的影响，对悬浮红细胞、洗涤红细胞以及添加不同量血浆的红细胞成分血在不同保存时间内的红细胞的功能指标，包括pH值、K^＋浓度、Na^＋浓度、渗透脆性、血浆游离血红蛋白、乙酰胆碱酯酶活性、ATP含量、2．3-DPG、P50含量进行了比较。结果证明，血浆有利于保存血液维持其高pH值、低K^＋浓度和高Na^＋浓度，有利于保存红细胞维持其ATP含量、携氧能力和红细胞变形性，但对保存血液维持低渗透脆性，对保存血液的血浆游离血红蛋白、乙酰胆碱酯酶活性、ATP含量和2．3-DPG含量无影响。结论：在红细胞成分血中加入一定量的血浆可能有利于红细胞的长期保存。     

Preparation of packed red blood cell units in the blood bank: Alteration in red blood cell deformability

BackgroundDonated blood is stored in the blood bank as packed red blood cell units. In the process of packed cells preparation, the red blood cells (RBCs) are subjectedto high level of shear stress, which can induce alterations in their properties.In the present study, we examined the effect of packed RBCs preparation (which included leuko-filtration) on red cell deformability.MethodsBlood samples were collected from 25 healthy donors and from corresponding units of packed RBCs. The portion of undeformable cells (%UDFC) was determined for each sample.ResultsThe median value of %UDFC was equal to 6.75 ± 0.70 %, for freshly-donated RBCs, and to 6.36 ± 0.51 %, for packed cells. Wherein, %UDFC may increase or decrease following packed cells preparation, depending upon the initial portion of undeformable cells.ConclusionLikely, exposure of RBCs to high shear stress, during packed cells preparation, induces opposing effects: (a) removal/destruction of rigid (undeformable) cells, thereby reducing their total amount (i.e., decreasing the %UDFC) on the one hand, and (b) mechanical damage to the cell membrane and subsequent reduction of the cell deformability (thereby increasing the %UDFC) on the other. As a consequence, the final impact of packed cells preparation is primarily determined by the initial state of erythrocytes in the blood of the donor.     

Extracellular vesicle characteristics in stored red blood cell concentrates are influenced by the method of detection

Extracellular vesicles (EVs), including microvesicles and exosomes, are small phospholipid vesicles (≤1 μm in diameter) that are present in blood products, accumulate during storage, and have a potential transfusion-related immunomodulatory role. Knowledge of EVs in stored blood products is limited due to the challenges and difficulties in detecting these heterogeneous submicron-sized vesicles. The aim of this study was to assess the impact of different approaches to characterize EVs in stored RBC products. Quantification and size-profiling of EVs in leukoreduced red cell concentrates (RCCs) were examined on day 3, 7, 21, and 42 of storage using tunable resistive plus sensing (TRPS), flow cytometer (FC), and dynamic light scatting (DLS) methods. Using the TRPS method, the concentration of EVs < 200 nm significantly increased throughout storage (p < 0.05). This change in exosome concentration was not detectable with FC or DLS due to limitations in their ability to resolve particles <200 nm and/or accurately determine EV concentration. Both the TRPS and FC demonstrate that the concentration of EVs ≥ 200 nm significantly increases in RCCs by day 42/43 compared to EVs present on day 3 (p < 0.001). As the DLS measures the average size of particles in suspension, only an increase in the zeta-average size was observed during storage. EV size and concentration in RBC products is significantly influenced by the length of storage. Overall, this study shows that combining technologies may be important to improve the characterization and study of EVs in stored RCCs.     

Harmful effects of transfusion of older stored red blood cells: iron and inflammation

Retrospective studies suggest that the transfusion of older, stored red blood cells (RBCs) may be associated with increases in mortality, serious infections, multiorgan failure, thrombosis, and hospital length of stay. Our research is based on the overarching hypothesis that the adverse effects associated with transfusion of older, stored RBCs result from the acute delivery of hemoglobin iron to the monocyte-macrophage system. To test this "iron hypothesis," we are recruiting healthy human volunteers to donate double, leukoreduced, RBC units. We then transfuse them with one autologous fresh unit (i.e., after 3-7 days of storage) and one older, stored unit (i.e., at 40-42 days of storage). The primary study outcome will compare laboratory iron measures and proinflammatory cytokines after transfusion of fresh or older, stored RBCs. Similar studies using allogeneic RBC transfusions will be performed in chronically transfused patients with either sickle cell disease or β-thalassemia. Although prospective, randomized studies will ultimately determine the existence of adverse effects from transfusing older, stored RBCs, our goal is to determine the mechanism(s) for this potential effect.     

A novel mouse model of red blood cell storage and posttransfusion in vivo survival

BACKGROUND: Storage of red blood cells (RBCs) is necessary for an adequate blood supply. However, reports have identified potential negative sequelae of transfusing stored RBCs. An animal model would be useful to investigate the pathophysiology of transfusing stored RBCs. However, it has been reported that storage of rat RBCs in CPDA-1 resulted in an unexpected sudden decline in posttransfusion survival. A mouse model of RBC storage and transfusion was developed to assess survival kinetics of mouse RBCs.
STUDY DESIGN AND METHODS: RBCs expressing green fluorescent protein were collected in CPDA-1, filter leukoreduced, adjusted to a 75% hematocrit, and stored at 4°C. At weekly intervals, stored RBCs were transfused into C57BL/6 recipients. RBC survival was measured by flow cytometry and chromium-51 labeling. Phosphatidylserine externalization and CD47 expression was also evaluated.
RESULTS: Mean 24-hour survivals of transfused RBCs were 99, 91, 64, 54, 30, and 18% after 0, 7, 14, 21, 28, and 35 days of storage, respectively. Stored RBCs showed an initial rapid clearance with subsequent extended survival. Increased surface phosphatidylserine and decreased CD47 expression were also observed.
CONCLUSIONS: Mouse RBCs showed a progressive decline in survival, as a function of storage time, unlike the precipitous loss of viability reported for rat RBCs. Moreover, changes in the measured surface markers were analogous to trends reported for human RBCs. Together, these findings provide an initial characterization of a novel mouse model of RBC storage with the potential to serve as an experimental platform for studying the pathophysiologic consequences of transfusing stored RBCs.     

The role of electrolytes and pH in RBC ASs

BACKGROUND: Experimental additive solutions (EASs) containing saline, adenine, glucose, mannitol and disodium phosphate can support RBCs for 9 or 10 weeks if used in 200- or 300-mL volumes. The effects of variations in the electrolyte composition and volume of EASs were explored. STUDY DESIGN AND METHODS: In three four-arm studies, 24 RBC units were pooled in groups of 4 and realiquoted as test units to ensure that all donors were equally represented in each study arm. In Study 1, units were stored for 11 weeks in EAS containing 0, 10, 20, or 30 mmol per L of sodium bicarbonate. In Study 2, units were stored for 9 weeks in EAS containing 26, 50, 100, or 150 mmol per L of sodium chloride. In Study 3, units were stored in 100 or 200 mL of AS-3 or EAS-61. RBC ATP concentrations and hemolysis were measured weekly. RESULTS: Increasing the sodium bicarbonate content of EASs increased the pH throughout storage and increased RBC ATP concentrations in the later phases of storage, but it had no effect on hemolysis. Increased sodium chloride content of EASs led to lower RBC ATP concentrations and increased hemolysis. In EAS-61, RBC ATP concentrations were increased throughout storage, and hemolysis was lower than that of RBCs stored in AS-3. CONCLUSION: RBC ATP synthesis is highly dependent on the pH of the AS. Hemolysis is affected by the salt content and volume of the AS.