Changes in neutrophil aggregation and chemotaxis response during 18- hour cold storage

Providing neutrophil transfusions to septic neonatal patients with depleted neutrophil reserves can be troublesome and require unscheduled blood donations. Buffy coats from stored whole blood are a potential source of neutrophils provided they remain viable during the interval between the whole blood collection and the buffy coat production. This study determined neutrophil function during storage of whole blood at 4 degrees C for 18 hours. Whole blood pH, hematocrit, platelet, and white cell counts remained unchanged. Chemotactic response to formyl methionyl leucyl penylalanine (FMLP) declined from 172.5 +/- 29 units (mean +/- SD) to 125 +/- 48 at 9 hours and 63.6 +/- 48 (p less than 0.05) at 18 hours. Aggregation response to FMLP remained normal for 9 hours but dropped to 15.5 percent of normal after 18 hours. Neutrophil response to cytaxins was maintained for at least 9 hours during storage of whole blood at 4 degrees C but seriously declined within 18 hours. Limiting 4 degrees C storage of whole blood to 9 hours prior to preparing buffy coats may provide flexibility needed for urgent provision of neutrophil transfusions.     

Mini buffy coat photopheresis for children and critically ill patients with extracorporeal photopheresis contraindications

BACKGROUND: Conventional extracorporeal photopheresis (ECP) has proven efficacy for the treatment of several diseases but is limited to patients with sufficient body weight. A novel simplified mini buffy coat ECP technique that allows treatment of small children and patients with apheresis contraindications has been developed.
STUDY DESIGN AND METHODS: White blood cell (WBC)-rich buffy coat fractions were prepared from 5 to 8 mL/kg whole blood in a closed system, diluted, and ultraviolet A (UVA)-irradiated after addition of 8-methoxypsoralen (8-MOP). Apoptosis and cell death were analyzed by annexin V and 7-aminoactinomycin staining. Lymphocyte proliferation was measured after CD3/CD28 and phytohemagglutinin (PHA) stimulation. Autologous residual blood and UVA-irradiated buffy coat were returned to the patients. Fifty-six mini buffy coat ECP procedures were applied to three children with acute steroid-refractory skin graft-versus-host disease and apheresis contraindications.
RESULTS: Mean whole blood and buffy coat volumes were 166 (±61.8) and 8 (±1.6) mL, respectively, and resulted in a hematocrit of 2.2% (±0.4) after saline dilution (median ± SD). UVA irradiation of 8-MOP buffy coat preparations resulted in significant induction of WBC apoptosis at 48-72 hours (p ≤ 0.006). WBC proliferation was significantly inhibited both after CD3/CD28 stimulation and after PHA stimulation when compared to controls (p ≤ 0.001). No clinical or laboratory side effects were observed during mini ECP procedures and the three patients responded to the therapy.
CONCLUSION: Mini buffy coat ECP induces apoptosis and lymphocyte proliferation inhibition, both of which occur after standard ECP. This study proposes that mini buffy coat ECP be used as a simple and inexpensive alternative to classical ECP in children and adult patients with apheresis contraindications.     

Storage of Blood Components Does Not Decrease Haemostatic Potential: In vitro Assessment of Fresh versus Stored Blood Components Using Thromboelastography

Summary

Background

Major surgery and severe trauma typically lead to massive blood loss requiring rapid transfusion of large amounts of blood products. It has been suggested that fresh, unrefrigerated whole blood provides a haemostatic advantage in this setting. The aim of the current study was to compare the clot formation parameters of fresh, unrefrigerated whole blood and whole blood reconstituted from components stored for varying periods of time, using rotational thromboelastography (ROTEM®).

Methods

Fresh whole blood and reconstituted whole blood using combinations of non-leucoreduced red cell units (stored for 7, 14, 21, 28, or 35 days), platelet concentrates (stored for 1, 3 or 5 days), and fresh frozen plasma (stored for 6 months) were analysed using ROTEM. Measurements of the clotting time (CT), clot formation time (CFT), and maximal clot firmness (MCF) were compared between units of fresh whole blood and reconstituted whole blood samples.

Results

There was no difference in the haemostatic parameters measured of fresh whole blood and reconstituted whole blood using red cell units stored for less than 21 days. ROTEM demonstrated that the CT and CFT were significantly shorter for reconstituted whole blood samples using red cells stored for longer than 21 days when compared to fresh whole blood and to reconstituted whole blood samples using red cell units stored for less than 21 days. The CT was inversely correlated to the duration of platelet storage. The MCF was unchanged regardless of duration of blood product storage.

Conclusion

Fresh unrefrigerated whole blood and blood products stored for short duration (less than 21 days) were not superior to those stored for longer durations.     

Storage of saline-adenine-glucose-mannitol suspended red cells in diethylhexyl phtalate and butyryl-n-trihexyl-citrate plasticized polyvinyl chloride containers. An in vitro comparative study

Units of whole blood collected into butyryl-n-trihexyl-citrate (BTHC) and diethylhexyl phtalate (DEHP) plasticized polyvinylchloride (PVC) blood storage containers and processed by means of an 'Optipress', which allows automated removal of the buffy coat, were compared. Units collected into standard PVC containers processed by the traditional method (no buffy coat removal) were used as a control group. The red cell concentrates were suspended in saline-adenine-glucose-mannitol (SAGM) and stored 42 days at 2-6 degrees C. Comparison of the buffy coat depleted red cell concentrates showed that red cell energy and oxygen delivery capacity, as evidenced by ATP and 2,3-DPG values, were slightly better preserved in the BTHC plasticized container, compared to the DEHP container. The red cell membrane, however, was slightly less well preserved, (the hemolysis at day 42 with BTHC ww 0.39%; with DEHP, 0.20%) in this container. The higher ATP levels might lead to a better in vivo recovery of stored red cells. In vivo studies comparing both plastic containers, therefore are indicated in order to determine if these differences have practical significance. A longer holding time of the whole blood at room temperature before processing reduced the hemolysis (42 days stored RCC as 0.26%). Slightly more fibrinopeptide A (FPA) generation and marginally lower pH and 2,3-DPG values were observed in this situation. This finding suggests an effect of higher plasticizer levels on the red cell membrane.     

White cells protect donor blood against bacterial contamination

The possible beneficial role of white cells (WBCs) in donor blood has been investigated with respect to their capacity to remove bacteria. Preparations of buffy coat and whole blood, containing as well as reduced of WBCs, were inoculated with Staphylococcus epidermidis, S. aureus, Escherichia coli, Pseudomonas aeruginosa, and Propionibacterium species. Upon storage at room temperature, the presence of WBCs resulted in a reduction of the bacterial content. Units inoculated with S. epidermidis and E. coli were completely cleared of bacteria within 5 to 24 hours. On the other hand, S. aureus, after an initial reduction in number, started to multiply. In WBC-reduced units, the initial bacterial content remained unchanged for 5 hours, but the bacteria then exhibited vigorous growth within 48 hours in buffy coat and slower growth in whole blood. Propionibacterium sp. did not grow with or without WBCs. P. aeruginosa did not grow in buffy coat but showed a growth pattern similar to that of S. aureus in whole blood. The presence of WBCs in the donor blood during the first hours after collection thus seems to rid the blood of at least some species of bacteria. These results indicate that it would be favorable not to perform WBC reduction during blood collection and that several hours of contact can be needed to obtain sterility.     

全血手工制备浓缩血小板后的血浆再制备冷沉淀的质量评价

目的评价室温新鲜全血白膜法制备浓缩血小板后的血浆再制备冷沉淀的质量。方法实验组为24例,新鲜全血(400 mL)置室温于<8 h用白膜法制备浓缩血小板后所得的血浆,冰冻保存。对照组1为12例,常规制备新鲜冰冻血浆,冰冻保存。对照组2为12例,新鲜冰冻单采血浆,血浆单采完毕分装为200 mL/袋并立即冰冻保存。3组血浆按常规制备冷沉淀,评价其质量:外观、凝血因子FⅧ及Fib的含量;血细胞残留量。结果 3组冷沉淀外观均正常;WBC含量在3组间无统计学意义。与对照组1比较:实验组凝血因子FⅧ(81.76±34.07)IU较低,Fib(202.63±48.58)mg及Plt(7.81±5.81)×109均较高。与对照组2比较:实验组凝血因子FⅧ含量相当,Fib(202.63±48.58)mg较高、Plt(7.81±5.81)×109较低。结论全血来源的制备浓缩血小板后的冰冻血浆还可以用于冷沉淀的制备,其质量符合国家标准。     

A new continuous-flow cell separation method based on cell density: principle, apparatus, and preliminary application to separation of human buffy coat

With the recent progress in transfusion medicine, separation and isolation of cells in a large quantity is becoming increasingly important. At present, the continuous cell separation method in a preparative scale is limited to apheresis and elutriation: the former is mainly used for collection of platelet and buffy coat from the whole blood, while the latter separates cells virtually according to their size. Here we introduce a continuous flow method that separates cells entirely based on cell density. The method is gentle and capable of processing a large number of cells. The potential capability of the method was demonstrated on separation of lymphocytes and granulocytes from human buffy coat. Lymphocytes were enriched to 90% in the fraction at density = 1.065 and granulocytes are isolated in fractions at density = 1.075-1.080 while red cells were completely retained at the periphery of the channel. CD34 cells were distributed around 1.065 and coeluted with lymphocytes, suggesting that further enrichment requires focusing the density gradient around 1.070. The method could process 10(9) nucleated cells in 2 hours. Our preliminary results suggest that the present method is an effective and efficient means to separate blood cells.     

Glycogen metabolism in stored granulocytes

Optimal function of transfused granulocytes (PMNs) requires adequate glycogen metabolism. Previous studies in our laboratory suggested that stored PMNs had decreased glycogen. We report here the glycogen content and chemotaxis of stored PMNs, and the ability of fresh and stored PMNs to use glycogen as the fuel source for chemotaxis. PMNs were prepared from 8 fresh units of blood drawn into citrate-phosphate-dextrose-adenine, suspended at 2 or 8 X 10(7) PMN per ml in autologous plasma with or without 15 mM sodium bicarbonate, and stored at 22 to 24 degrees C in transfer packs for 48 hours. Glycogen was measured on resting PMNs, and after challenge with opsonized zymosan and F-Met-Leu-Phe (FMLP). The chemotaxis of fresh and stored PMNs was measured in the presence or absence of extracellular glucose. Fresh PMNs contained 10.3 +/- 0.5 (mean +/- SEM) micrograms of glycogen per 10(6) PMN. Glycogen decreased by 4.2 +/- 0.9 micrograms per 10(6) PMN after challenge with opsonized zymosan and by 1.1 +/- 0.6 micrograms per 10(6) PMN after FMLP. After 48 hours of storage, neutrophil glycogen increased by 18 percent, except in units stored at a concentration of PMN of 8 X 10(7) per ml without sodium bicarbonate. In PMNs from these units stored without bicarbonate, glycogen decreased by 9 percent (p less than .05), and there was a 19 and 55 percent decrease in the ability of PMN from these units to metabolize glycogen after exposure to opsonized zymosan and FMLP, respectively (p less than 0.05). In addition, in PMNs from units stored at a concentration of PMN of 8 X 10(7) per ml without bicarbonate, there was a 47 and 70 percent decrease in chemotaxis at 24 and 48 hours, respectively (p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Physical Characteristics of Microaggregates in Stored Blood

The volume of microaggregates 13 to 80 μ in size which developed in blood components stored under varying conditions was measured with a particle size analyzer. The microaggregates were found to develop progressively during storage of ACD whole blood at 4 to 6 C. Coincident with this there was a drop in the platelet count during the first week of storage and a progressive reduction in the absolute granulocyte count. Microaggregate development after storage of various components of ACD blood was proportional to the concentration of platelets and leukocytes prior to storage. The microaggregates settled into the buffy coat after centrifugation and became larger. In vitro studies indicated that they were resistant to dissociation in vitro in comparison to platelet aggregates induced in fresh blood by adenosine diphosphate. Microaggregate formation was greater in CPD than in ACD anticoagulated blood stored at 4 to 6 C for 24 hours, but was not different after seven days of storage. A greater volume of microaggregates was formed in aliquots of ACD blood stored at 4 to 6 C than at room temperature, while no differences were noted after storage of blood in plastic bags or glass vacuum bottles.     

Monocyte chemoattractant protein production in red cell incompatibility

BACKGROUND: Mononuclear phagocytes play a central role in hemolytic transfusion reactions by erythrophagocytosis and production of inflammatory mediators. Factors that affect the number or function of monocyters would be expected to alter the clinical course of hemolytic transfusion reactions, and thus the production of monocyte chemoattractant protein-1 (MCP-1), a recently described chemotactic and activating cytokine specific for monocytes, was investigated in two distinct settings of red cell (RBC) incompatibility. STUDY DESIGN AND METHODS: Fresh heparinized whole blood was incubated with ABO- compatible or -incompatible RBCs. Isolated peripheral blood mononuclear cells were incubated with anti-D-coated or uncoated RBCs. MCP-1 was measured in the plasma or culture medium by enzyme-linked immunosorbent assay. MCP-1 gene expression was detected by Northern blot analysis of buffy coat or mononuclear cell total RNA. RESULTS: Significant levels of MCP-1 protein in plasma or medium were detected 24 hours after the addition of incompatible RBCs, but not in the first 6 hours. Nonimmune hemolysis of added RBCs did not stimulate MCP-1 production. The inactivation of complement by heat treatment of plasma prior to the addition of RBCs to whole blood did not prevent MCP-1 production. Nor did neutralizing antibodies to tumor necrosis factor prevent MCP-1 production in ABO incompatibility. MCP-1 production was associated with increased steady-state levels of white cell MCP-1 mRNA, which occurred more rapidly in ABO than Rh incompatibility. CONCLUSION: The monocyte- specific chemotactic cytokine MCP-1 is produced by peripheral blood leukocytes in response to RBC incompatibility. MCP-1 may act in a positive feedback loop to recruit and activate monocytes during hemolytic transfusion reactions, thus contributing to the maintenance of these reactions.     

Prestorage white cell reduction in saline-adenine-glucose-mannitol red cells by use of an integral filter: evaluation of storage values and invivo recovery

BACKGROUND : Prestorage white cell (WBC) reduction in blood components may decrease the incidence of adverse reactions and improve component quality. A bottom-and-top system with an integral third-generation WBC- reduction filter has been studied. STUDY DESIGN AND METHODS : Whole blood was collected from 30 healthy donors: from 20 by using a blood container system with an integral filter and from 10 controls by using a standard blood container system. Ten test units were buffy coat- depleted, stored for 72 hours at 4 degrees C, and then filtered, while an additional 10 test units were buffy coat-depleted and filtered at room temperature within 8 hours of collection. All units were stored at 4 degrees C for 42 days and sampled weekly. RESULTS : The mean WBC content of the 72-hour, 4 degrees C units was 0.33 × 10(6), that of the room-temperature units was 2.6 × 10(6), and that of the buffy coat- depleted controls was 460 × 10(6) (p < 0.0005). No significant differences were found among lactate, glucose, sodium, potassium, and plasma hemoglobin levels in the three groups. ATP and 2,3 DPG levels were significantly better preserved in control units than in 72-hour, 4 degrees C units (p = 0.016 and p = 0.032, respectively), but not better than in the room-temperature units. Significant differences were observed between pH values in filtered units and both groups of test units (p = 0.016). In biologic terms however, these differences were small. Red cells from an additional eight healthy volunteer donors were processed by an 8-hour room-temperature method and stored for 35 days. Studies in vivo 24-hour recovery of autologous red cells were performed by transfusing a radiolabeled (51Cr plus 131I-albumin) aliquot after 35 days' storage. Good recovery (mean > 80%) was found by both the single- and double-isotope-label methods. Recovery was significantly greater when calculated by the single-isotope method (p = 0.02). CONCLUSION : The combination of buffy coat removal and filtration in the blood container system with an integral filter achieved effective WBC reduction (> or = 3 log10 reduction from whole blood) without biologically significant detriment to in vitro or in vivo storage values.     

Hydrogen ion maintenance improves the chemotaxis of stored granulocytes

Through technological advances in granulocyte collection, it has become possible to collect neutrophils (PMNs) routinely in high concentration (greater than 5 X 10(7) PMN/ml) for transfusion. Previous studies in this laboratory suggested that storage of neutrophils for transfusion at high PMN concentrations resulted in impaired adenosine triphosphate (ATP) and hydrogen ion maintenance. The studies we report here were designed to assess the effect of PMN storage at concentrations which are usual (2 X 10(7) PMN/ml), intermediate (5 X 10(7) PMN/ml), and high (8 X 10(7) PMN/ml) on chemotactic responses, and to identify variables which are easily measured and might predict the chemotactic function of stored PMNs. Granulocyte concentrates were stored in plastic bags at 2,5, and 8 X 10(7) PMN per ml, with or without 15 mM bicarbonate (HCO3). The random migration (RM) chemotaxis (CTX), ATP, and relative cell size (VOL) of the fresh and stored cells and the pH, glucose, and lactate concentrations in the supernatant medium were measured in the freshly prepared units after 24 and 48 hours storage at room temperature. We found that RM, CTX, ATP, glucose, and pH decreased significantly (p less than .02) following storage for 24 and 48 hours, particularly in units stored at the higher cell concentrations. Cell volume and lactate increased significantly with storage for 24 and 48 hours, and these values were also greater in units stored at the higher cell concentration (p less than .02).(ABSTRACT TRUNCATED AT 250 WORDS)     

Remodelling whole blood processing through automation and pathogen reduction technology at the Luxembourg Red Cross

In 2014–2015, the Luxembourg Red Cross (LRC) implemented a fully automated system (FAS) able to process 4 whole blood units simultaneously, and a pathogen reduction technology (PRT) based on riboflavin and ultraviolet light to improve safety of platelet concentrates (PCs). In this observational study, the impact of both technologies to enable this centralised blood transfusion centre to provide safe and timely blood components supply for the whole country was analysed. Standard quality control parameters for blood components, productivity and safety were compared from data collected with the conventional semi- automated buffy coat method and with FAS/PRT. The FAS decreased processing time when compared with the buffy coat method and facilitated the daily routine at the LRC. Red blood cell concentrates, plasma units and PCs prepared with both methods were conform to the European Directorate for the Quality of Medicines & HealthCare specifications. PCs prepared by FAS showed high yields, with decreased variability when the device-related software (T-Pool Select) was used. PRT had minimal impact on platelet yields and product quality and induced no increase in transfusion reaction notifications. The FAS and PRT transformed the daily routine of blood component manufacture by allowing increased productivity and efficiency, notwithstanding resource containment and without impacting quality, yet promoting safety.     

Packed blood cells stored in AS-5 become proinflammatory during storage

BACKGROUND: Studies have shown that packed blood cells (PBCs) stored in AS-1 (Adsol, Baxter) and AS-3 (Nutricel, Medsep Corp.) accumulate proinflammatory substances, which may contribute to increased complications from allogeneic blood transfusion. This study assessed whether supernates from PBCs stored in AS-5 (Optisol, Terumo Corp.) prime neutrophils (PMNs), activate platelets (PLTs), and accumulate proinflammatory cytokines and PMN granule constituents.
STUDY DESIGN AND METHODS: PBC units were prepared in AS-5 from nonleukoreduced (NLR) and leukoreduced (LR) whole-blood units and stored at 4°C. Supernates from samples of PBCs collected at various storage times were analyzed by multiplex enzyme-linked immunosorbent assay for proinflammatory cytokines and myeloperoxidase (MPO) and were incubated with type-matched blood, which was assessed by flow cytometry for expression of CD11b on PMNs, CD62P on PLTs, and formation of PMN-PLT aggregates.
RESULTS: Supernates from NLR PBCs stored for at least 14 days elevated CD11b expression on PMNs and the number of PMN-PLT aggregates compared to supernates from collection day PBCs. The magnitude of these effects correlated with storage age. Supernates from LR PBCs did not elicit these responses. Expression of CD62P on PLTs was not affected by supernates from either NLR or LR PBCs. Levels of interleukin (IL)-1β, IL-6, IL-8, IL-18, NAP-2, MCP-1, RANTES, and MPO were elevated in supernates from 28- and 42-day NLR units. Tumor necrosis factor α and MIP-1α did not increase, and cytokine levels in LR PBC units did not increase.
CONCLUSION: Units of NLR PBCs stored in AS-5 become increasingly proinflammatory as a function of storage time. Leukoreduction prevents this change.     

Studies on Stored Whole Blood

Blood stored at 10 C displayed no more evidence of cell destruction in vitro than did blood stored at 4 C. Agitation for one hour at weekly intervals of storage was also not obviously damaging, nor was warming blood to 22 C for up to 16 hours. Exposure to 22 C for 24 hours led to erythrocyte deterioration that was most evident when units were more than 21 days old, and agitation plus this warming increased these signs of damage. Units stored as packed cells were moderately affected by exposure to 22 C for 24 hours, and agitation reinforced this effect. Posttransfusion survival of blood warmed to 22 C for 24 hours tended to be lower than nonwarmed controls; the warmed group average was below 70 per cent survival at 28 days of storage.
From these studies, the temperature variation commonly encountered in clinical blood banking (4 C to 10 C and short exposure to 22 C prior to transfusion) would not appear to contribute significantly to erythrocyte damage unless the units were in the oldest stages of storage or had been exposed to warm temperatures for longer than 24 hours. Mechanical stress had minimal adverse effect, but this became more evident when blood was stored as packed cells or when whole blood had been stored for 21 days.     

Stored red blood cells selectively activate human neutrophils to release IL-8 and secretory PLA2

Packed red blood cell (PRBC) transfusion has been invoked previously with immunosuppression and increased infections, but it has now been demonstrated that stored PRBCs (>14 days) can prime PMNs and provoke multiple organ failure. Recently, the role of PMNs in the genesis of MOF has been extended to their release of inflammatory cytokines, notably IL-1, IL-8, TNFalpha, and secretory phospholipase A2 (sPLA2). We hypothesize that stored PRBCs can act as a second event via stimulating the release of inflammatory cytokines from PMNs. Isolated human PMNs were incubated for 24 h in RPMI with either 20% fresh plasma or plasma from 42 day old PRBC (day of outdate) and release of IL-8, IL-1beta, TNFalpha, and sPLA2 were measured. Plasma from stored PRBCs contained small amounts of IL-8, sPLA2, and TNFalpha (102.1 +/-5.6 pg/ml, 87.6+/-6.0 pg/ml and 9.7+/-.7 pg/ml). Levels of IL-1beta were below detection (<1 pg/ml). Day 42 PRBC plasma stimulated significant PMN release of both IL-8 and sPLA2 as compared to both control and day 0 plasma (*P < .05), but PRBC plasma did not stimulate PMN release of either IL-1beta or TNFalpha. Transfused blood is emerging as an inflammatory agent that is capable of producing PMN priming. In this study we have demonstrated that PRBC plasma selectively activates PMNs to release both IL-8 and sPLA2. Thus, transfusion of PRBCs may represent a preventable inflammatory insult via modification of both blood banking and transfusion practices.     

Microarray kit analysis of cytokines in blood product units and segments

BACKGROUND: Cytokine concentrations in transfused blood components are of interest for some clinical trials. It is not always possible to process samples of transfused components quickly after their administration. Additionally, it is not practical to sample material in an acceptable manner from many bags of components before transfusion, and after transfusion, the only representative remaining fluid of the component may be that in the “segment,” because the bag may have been completely transfused. Multiplex array technology allows rapid simultaneous testing of multiple analytes in small‐volume samples. This technology was used to measure white blood cell (WBC) cytokine levels in blood products to determine 1) whether concentrations in segments correlate with those in the main bag and, thus, whether segments could be used for estimation of the concentrations in the transfused component and 2) whether concentrations after sample storage at 4°C for 24 hours do not differ from concentrations before storage, thus allowing for processing within 24 hours, rather than immediately after transfusion. STUDY DESIGN AND METHODS: WBC cytokines were measured in the supernatant from bags and segments of leukoreduced red blood cells (RBCs), nonleukoreduced whole blood, and leukoreduced plateletphereses using a human cytokine array kit (ProteoPlex, Novagen). RESULTS: Cytokine concentrations in RBCs and whole blood or plateletphereses stored at 4°C did not differ between bag and segment samples (all p > 0.05). There was no evidence of systematic differences between segment and bag concentrations. Cytokine concentrations in samples from plateletphereses did not change within 24 hours storage at 4°C. CONCLUSION: Samples from either bag or segment can be used to study cytokine concentrations in groups of blood products. Cytokine concentrations in plateletphereses appear to be stable for at least 24 hours of storage at 4°C and, thus, samples stored with those conditions may be used to estimate the cytokine concentrations of the component at the time of transfusion.     

Effects of filtration and gamma radiation on the accumulation of RANTES and transforming growth factor-β1 in apheresis platelet concentrates during storage

BACKGROUND: Platelet-derived biologic response modifiers (BRMs) including RANTES and transforming growth factor (TGF)-beta1 accumulate in platelet components during storage because of platelet activation, and they may play a causative role in nonhemolytic febrile transfusion reactions. The majority of PCs with high unit values are provided by single donor apheresis in Japan. STUDY DESIGN AND METHODS: RANTES and TGF-beta1 levels in platelet units prepared from single-donor apheresis platelet concentrates (apheresis PCs) and units from whole blood (buffy coat PCs) were investigated. The effects of prestorage and poststorage filtration and gamma radiation on the levels of RANTES and TGF-beta1 in the supernatant of apheresis PCs during storage were also examined. RESULTS: The levels of RANTES and TGF-beta1 increased during storage from Day 0 to Day 5. The levels of RANTES and of TGF-beta1 correlated with the platelet concentration (p<0.01), but not with the residual white cell concentration in apheresis PCs that were not white cell reduced by filtration (p>0.05). In addition, there was a correlation between RANTES and TGF-beta1 levels (p<0.01). In white cell-reduced apheresis PCs using negatively charged filters as well as in gamma-radiated apheresis PCs, the levels of these two BRMs-did not differ at any storage time from those of untreated apheresis PCs. Filtration of apheresis PCs with negatively charged filters after 3 days of storage significantly (p<0.05) reduced the levels of RANTES, but not of TGF-beta1. There was no reduction in the levels of RANTES and TGF-beta1 levels by positively charged filters. The RANTES levels in buffy coat PCs were slightly higher than but not significantly different from those of apheresis PCs during storage, except for the level on Day 1. There were no differences in the TGF-beta1 levels in apheresis and buffy coat PCs during storage. CONCLUSION: Prestorage filtration and gamma radiation had neither preventive effects on the accumulation of RANTES and TGF-beta1 nor adverse effects on platelet activation. Negatively charged filters might be useful for the reducing the levels of RANTES in stored apheresis PCs.     

Paired comparison of platelet concentrates prepared from platelet-rich plasma and buffy coats using a new technique with 111In and 51Cr

Two techniques for the preparation of platelet concentrate (PC), the standard platelet-rich plasma (PRP) and buffy coat (BC) methods, were compared in nine paired studies with regard to platelet harvest, white cell (WBC) contamination, and PC quality after 5 days of 22 degrees C storage. Platelet harvest using the BC method averaged approximately 56 percent of the whole blood level (6.2 x 10(10)/concentrate), which was less than the 76 percent achieved with the PRP-PC method (8.7 x 10(10)/concentrate). An additional 5 units collected into an experimental siphon bag for BC-PC processing showed improved platelet harvest (6.7 x 10(10)/concentrate, or approx. 70% of whole blood). WBCs remaining in the BC-PC averaged 0.19 x 10(8) per unit compared to 3.6 x 10(8) per unit for PRP-PC. Buffy coat processing produced red cell (RBC) units with 50 percent of the WBC contamination of conventionally prepared units (9.8 +/- 6.2 x 10(8)/unit vs. 18.9 +/- 7.1 x 10(8)/unit). The siphon bag further reduced WBC levels in the AS-3 RBC units (6.4 +/- 3.7 x 10(8)/unit). In vitro studies performed on Days 1 and 5 after collection showed no significant differences in platelet metabolic and biologic function or cell integrity. Beta-thromboglobulin and surface glycoprotein levels, indicators of platelet activation and membrane alteration, respectively, did not differ significantly in the PRP-PC and BC-PC; nor was lactate production higher in PRP-PC, despite the substantially higher WBC counts. Autologous in vivo platelet viability determinations were performed by using concurrent transfusion of 111In-labeled freshly drawn platelets and 51Cr-labeled stored platelets.(ABSTRACT TRUNCATED AT 250 WORDS)     

Warm storage of whole blood for 72 hours

BACKGROUND: In field emergency medicine, fresh whole-blood units are stored at room temperature up to 24 hours or occasionally longer. Few data exist on the integrity and in vitro functional properties of whole blood stored warm beyond 24 hours. STUDY DESIGN AND METHODS: Ten citrate phosphate dextrose solution whole-blood units were collected and divided into two equal volumes. One-half of each unit was stored at 19 degrees C and the other half was stored at 25 degrees C, encompassing the accepted range for room temperature storage. At 6, 24, 48, and 72 hours, aliquots were collected from each unit and whole blood analyzed for cell counts, gases, and clotting function with thromboelastography, red cells for intracellular analytes, platelet (PLT)-rich plasma for aggregometry, and the supernatant for hemoglobin, potassium, glucose, lactate, and plasma clotting studies. RESULTS: Whole-blood units stored at room temperature maintained cellular counts and coagulation activity for up to 72 hours. Units stored at 19 degrees C demonstrated greater RBC adenosine triphosphate and 2,3-diphosphoglycerate (DPG) content and stronger responses in PLT aggregation studies when compared with 25 degrees C storage. No significant hemolysis was observed, and no bacterial growth was detected. CONCLUSION: Storage of whole blood at room temperature for 72 hours leads to marked reductions in pH and DPG, but the observed reduction in PLT function and plasma coagulation factor activity was surprisingly modest compared to literature values. These findings should prompt additional investigation, given their potential importance for whole blood processing and field-expedient transfusion.