Red cell transfusion in medicine: Future challenges

Many side-effects of red blood cell transfusion have been described. They include iron-overload, as well as allo- and autoantibody formation against red cells. During storage, erythrocytes undergo complex structural and biochemical changes. It has been suggested that accelerated and/or aberrant forms of the physiological erythrocyte aging process underlie the red cell storage lesion. This storage lesion may contribute to side-effects of transfusion as endothelial damage by release of internal erythrocyte constituents, (pro)inflammatory consequences, hampered microcirculation and oxygen delivery. Understanding the process that determines the fate of red blood cells after transfusion may contribute to the prevention of side-effects after red blood cell transfusion. This should be the focus of research on red blood cell transfusion in clinical transfusion medicine.     

Red blood cell transfusion strategies.

Although the hemoglobin level of 100 g/L has been used for many years as the allogeneic red blood cell (RBC) transfusion trigger, current evidence indicates that for most patients a more restrictive transfusion strategy is at least as effective as and possibly superior to a liberal transfusion strategy. Moreover, the available data indicate that the use of smaller volumes of allogeneic RBCs may be associated with decreased risk of morbidity and mortality. Thus several recent studies indicate that the use of more restrictive triggers than 100 g/L does not appear to adversely affect patient outcomes. Indeed, the majority of recently published RBC transfusion guidelines recommend a more conservative and cautious approach to allogeneic RBC transfusion practice, primarily to reduce the risk of transfusion-related adverse effects. However, the available transfusion trigger studies do not provide sufficient data to allow the claim that the improved outcomes observed are the sole result of the transfusion strategy used. It is possible that the results are the consequence of effects yet to be defined clearly. Additional studies will be necessary to determine the effects of RBC storage time and the presence of allogeneic leukocytes in allogeneic RBC transfusion practice. Nonetheless, the available data, together with detailed information about alternatives to blood product transfusions, will enable physicians to improve outcomes in transfused patients.     

Stored red blood cell transfusions: Iron,inflammation, immunity,and infection

The potential adverse effects of transfusion of red blood cells after prolonged storage have been hotly debated. During refrigerated storage, red blood cells are damaged, a process known as the red blood cell “storage lesion.” We hypothesized that the delivery of a bolus of iron derived from these rapidly cleared, damaged, red blood cells is responsible for some of the adverse effects of transfusion. Iron may play a role in producing a pro-inflammatory response to transfused red blood cells, potentially through the effects of reactive oxygen species on stress pathways and inflammasome activation. Furthermore, the excess iron may impair the host's ability to combat infection by its innate iron-withholding pathways. This symposium paper summarizes the background for the “iron hypothesis” as it relates to transfusion of red blood cells after prolonged refrigerated storage. It also includes a summary of the data from recent murine and human studies, and concludes with a discussion of several unresolved questions arising from these published studies.     

Laboratory hemostatic abnormalities in massively transfused patients given red blood cells and crystalloid.

Most of the literature on massive transfusion concerns whole blood replacement, whereas clinically, packed red blood cells are commonly given. To determine when hemostatic abnormalities occur in patients resuscitated primarily with packed red blood cells and crystalloid, the cases of 39 consecutive patients who were transfused with 10 or more red blood cell units of any kind within 24 hours were reviewed. After transfusion with 20 or more units of red blood cell products of any kind (packed red blood cells, cell-saver units, or whole blood), 75% (3 of 4) of patients had platelet counts less than 50 x 10(9)/L, compared to 0 of 29 patients given less than 20 units (P less than 0.001). After transfusion of 12 units of relatively plasma-free red blood cell products (packed red blood cells or cell-saver units), 100% (8 of 8) of patients had prothrombin time prolonged by more than 1.5 times mid-range of normal, compared to 36% (5 of 14) of patients given less than 12 units (P = 0.012). These data confirm that patients massively transfused with red blood cells of any kind develop significant thrombocytopenia after 20 units. Importantly, probably clinically significant prothrombin time and partial thromboplastin time prolongations occurred consistently after transfusion of 12 units of relatively plasma-free red blood cells in unselected patients at an urban trauma hospital. These data suggest that coagulation factor replacement is necessary in patients who receive 12 or more units of packed red blood cells or cell-saver blood, and platelet replacement is necessary in patients who receive 20 or more units of any red blood cell product. A prospective study is needed to determine whether the expected abnormal clinical bleeding indeed occurs in patients with such laboratory coagulation abnormalities and to determine when plasma transfusion is indicated in patients massively transfused with red blood cells.     

Storage of red blood cell concentrates: Clinical impact

The storage of red blood cells for transfusion purposes induces modifications of biochemical and biological properties. Moreover, these modifications are modulated by the donors’ characteristics and the cell processing. These ex vivo alterations were suspected to decrease the transfusion efficiency and even to induce adverse events. This short article will review the red blood cells storage lesions and the clinical data related to them. In particular, the questions regarding the donors and recipients sex will be discussed.     

Platelet transfusion in adults: An update

Many patients worldwide receive platelet components (PCs) through the transfusion of diverse types of blood components. PC transfusions are essential for the treatment of central thrombocytopenia of diverse causes, and such treatment is beneficial in patients at risk of severe bleeding. PC transfusions account for almost 10% of all the blood components supplied by blood services, but they are associated with about 3.25 times as many severe reactions (attributable to transfusion) than red blood cell transfusions after stringent in-process leukoreduction to less than 10⁶ residual cells per blood component. PCs are not homogeneous, due to the considerable differences between donors. Furthermore, the modes of PC collection and preparation, the safety precautions taken to limit either the most common (allergic-type reactions and febrile non-hemolytic reactions) or the most severe (bacterial contamination, pulmonary lesions) adverse reactions, and storage and conservation methods can all result in so-called PC “storage lesions”. Some storage lesions affect PC quality, with implications for patient outcome. Good transfusion practices should result in higher levels of platelet recovery and efficacy, and lower complication rates. These practices include a matching of tissue ABH antigens whenever possible, and of platelet HLA (and, to a lesser extent, HPA) antigens in immunization situations. This review provides an overview of all the available information relating to platelet transfusion, from donor and donation to bedside transfusion, and considers the impact of the measures applied to increase transfusion efficacy while improving safety and preventing transfusion inefficacy and refractoriness. It also considers alternatives to platelet component (PC) transfusion.     

Proteomics of Blood-Based Therapeutics

Blood-based therapeutics are cellular or plasma components derived from human blood. Their production requires appropriate selection and treatment of the donor and processing of cells or plasma proteins. In contrast to clearly defined, chemically synthesized drugs, blood-derived therapeutics are highly complex mixtures of plasma proteins or even more complex cells. Pathogen transmission by the product as well as changes in the integrity of blood constituents resulting in loss of function or immune modulation are currently important issues in transfusion medicine. Protein modifications can occur during various steps of the production process, such as acquisition, enrichment of separate components (e.g. coagulation factors, cell populations), virus inactivation, conservation, and storage. Contemporary proteomic strategies allow a comprehensive assessment of protein modifications with high coverage, offer capabilities for qualitative and even quantitative analysis, and for high-throughput protein identification. Traditionally, proteomics approaches predominantly relied on two-dimensional gel electrophoresis (2-DE). Even if 2-DE is still state of the art, it has inherent limitations that are mainly based on the physicochemical properties of the proteins analyzed; for example, proteins with extremes in molecular mass and hydrophobicity (most membrane proteins) are difficult to assess by 2-DE. These limitations have fostered the development of mass spectrometry centered on non-gel-based separation approaches, which have proven to be highly successful and are thus complementing and even partially replacing 2-DE-based approaches. Although blood constituents have been extensively analyzed by proteomics, this technology has not been widely applied to assess or even improve blood-derived therapeutics, or to monitor the production processes. As proteomic technologies have the capacity to provide comprehensive information about changes occurring during processing and storage of blood products, proteomics can potentially guide improvement of pathogen inactivation procedures and engineering of stem cells, and may also allow a better understanding of factors influencing the immunogenicity of blood-derived therapeutics. An important development in proteomics is the reduction of inter-assay variability. This now allows the screening of samples taken from the same product over time or before and after processing. Optimized preparation procedures and storage conditions will reduce the risk of protein alterations, which in turn may contribute to better recovery, reduced exposure to allogeneic proteins, and increased transfusion safety.     

What quality of red blood cells shall we offer the transfused patient?

Individual units of red blood cells (RBCs) can vary widely in their benefit to recipients depending on how the unit is prepared and the duration and quality of pretransfusion storage. The haemoglobin (Hb) concentration of the donor blood, the volume of the collection, leucocyte depletion procedures and the composition of the additive solution are all factors that will influence the final product. It may not be generally appreciated that, as red cell preparation is currently practiced, the haemoglobin content of viable RBCs in an individual blood unit collected within current standards may range from as little as 30 g to as much as 90 g, a variation of which the transfusing physician is generally unaware and which should no longer be considered acceptable. The transfusion community is currently in the untenable position of attempting to balance the quality of transfused blood against perceived cost, a balance that needs to be re‐examined in the light of recent developments. A number of measures are currently available that can readily eliminate many of these shortcomings. There are four obvious ways by which the quality and uniformity of RBC units can be improved: (1) The volume of blood collected from the donor should be adjusted depending on the donor's haemoglobin concentration. (2) Red cell losses during preparation should be minimized and accounted for in the collection (3) The subsequent preservation procedure should both maintain RBC viability during storage and minimize the loss of 2,3‐DPG and of membrane flexibility. (4) The haemoglobin content of a unit of red cells should be standardized. We recommend 50 g ± 5 g of haemoglobin as the amount of blood that should be collected from a donor. For all of these measures the technology is currently available and may be readily introduced, especially through the use of automated collection systems. It is the responsibility of the transfusion organisations to redefine their goals and, in collaboration with industry, develop routine procedures that do not unnecessarily sacrifice to economics the best interests of blood recipients and their prescribing physicians.     

Transfusion-Associated Graft-Versus-Host Disease and the Irradiation of Blood Components

Transfusion-associated graft-versus-host disease (TA-GVHD) is a rare but lethal disorder caused when viable donor lymphocytes engraft and proliferate in a susceptible transfusion recipient. Patients with immune deficiency disorders, hematologic malignancies and bone marrow transplants are at risk to TA-GVHD, as are premature newborns and transfusion recipients who are HLA heterozygous for an HLA-haplotype that is shared with an HLA homozygous donor. Irradiation of blood components with 2500 cGy will inactivate donor lymphocytes and prevent TA-GVHD. Platelets and granulocytes are not functionally impaired by this radiation dose, but red cells sustain detectible damage. Red cell units irradiated and stored for 42 days have significantly higher supernatant recovery of chromium-51 labeled cells is sub-optimal. Based on these data, the maximum permissible storage time for irradiated red cells has been reduced to 28 days.     

Les technologies de biologie moléculaire en immunohématologie

The molecular basis of almost all antigens of the 33 blood group systems are known. These knowledge and the advent of the PCR technology have allowed the DNA-based genotyping in order to predict the presence or absence of a blood group antigen on the cell membrane of red blood cells. DNA genotyping is required in cases where red blood cells patient cannot be used for serological typing either after a recent transfusion or because of the presence of autoantibodies on the red blood cells. Numerous DNA assays are available to detect any nucleotide polymorphism on the genes encoding blood group antigens. The technologies have improved to answer quickly to any case of transfusion emergency and to limit the risk of DNA contamination in a molecular diagnostic laboratory. Some technologies are ready for high-throughput blood group genotyping. They will be used in the future to obtain a fully typed blood group card of each donor but also to detect blood donors with rare phenotypes to register them to the Banque Nationale de Sang de Phénotype Rare (BNSPR).     

Phosphatidylserine-expressing cell by-products in transfusion: A pro-inflammatory or an anti-inflammatory effect?

Labile blood products contain phosphatidylserine-expressing cell dusts, including apoptotic cells and microparticles. These cell by-products are produced during blood product process or storage and derived from the cells of interest that exert a therapeutic effect (red blood cells or platelets). Alternatively, phosphatidylserine-expressing cell dusts may also derived from contaminating cells, such as leukocytes, or may be already present in plasma, such as platelet-derived microparticles. These cell by-products present in labile blood products can be responsible for transfusion-induced immunomodulation leading to either transfusion-related acute lung injury (TRALI) or increased occurrence of post-transfusion infections or cancer relapse. In this review, we report data from the literature and our laboratory dealing with interactions between antigen-presenting cells and phosphatidylserine-expressing cell dusts, including apoptotic leukocytes and blood cell-derived microparticles. Then, we discuss how these phosphatidylserine-expressing cell by-products may influence transfusion.     

Concise review: stem cell-derived erythrocytes as upcoming players in blood transfusion

Blood transfusions have become indispensable to treat the anemia associated with a variety of medical conditions ranging from genetic disorders and cancer to extensive surgical procedures. In developed countries, the blood supply is generally adequate. However, the projected decline in blood donor availability due to population ageing and the difficulty in finding rare blood types for alloimmunized patients indicate a need for alternative red blood cell (RBC) transfusion products. Increasing knowledge of processes that govern erythropoiesis has been translated into efficient procedures to produce RBC ex vivo using primary hematopoietic stem cells, embryonic stem cells, or induced pluripotent stem cells. Although in vitro-generated RBCs have recently entered clinical evaluation, several issues related to ex vivo RBC production are still under intense scrutiny: among those are the identification of stem cell sources more suitable for ex vivo RBC generation, the translation of RBC culture methods into clinical grade production processes, and the development of protocols to achieve maximal RBC quality, quantity, and maturation. Data on size, hemoglobin, and blood group antigen expression and phosphoproteomic profiling obtained on erythroid cells expanded ex vivo from a limited number of donors are presented as examples of the type of measurements that should be performed as part of the quality control to assess the suitability of these cells for transfusion. New technologies for ex vivo erythroid cell generation will hopefully provide alternative transfusion products to meet present and future clinical requirements.     

Yersinia enterocolitica septicaemia from transfusion of red cell concentrate stored for 16 days.

Two cases of transfusion transmitted Yersinia enterocolitica biotype 3, serotype 09 infection occurred in south east Scotland within four months of each other. In one case, a 79 year old man died the day after receiving a unit of red cell concentrate that had been stored for 29 days after donation. In the second case a 78 year old man died three days after transfusion of a unit of red cell concentrate that had been collected 16 days before transfusion. The donors of both units had no symptoms attributed to gastrointestinal infection. Early outdating of blood for transfusion after three weeks of storage is unlikely to eradicate Y enterocolitica associated fatalities from blood transfusion, and alternative methods should be considered.     

Utilization patterns of frozen autologous red blood cells. Experience in a referral center and a community hospital

The risks of homologous blood transfusion have motivated some blood centers and private industry to consider providing long-term storage of frozen, autologous red blood cells as a service. The usefulness of this practice is unknown. We performed a retrospective analysis of frozen autologous red blood cell use in two hospitals. Records were available for 21- and 9-year intervals, respectively. A total of 104 autologous units were cryopreserved for 41 patients. Fifteen (37%) of 41 patients received one or more of their stored units of red blood cells. Twenty-two patients had autologous units frozen in anticipation of elective surgery; 11 (50%) of these 22 patients received some or all of their stored units. Sixteen patients had autologous units stored because of potential transfusion problems related to rare blood types or to the presence of multiple blood cell alloantibodies, and another 3 patients had units frozen simply at their personal request. Only 4 (21%) of these latter 19 patients who donated without a specific planned use eventually received their frozen autologous red blood cells. Long-term autologous frozen red blood cell storage can improve medical management of some patients with anticipated surgical procedures or unusual requirements for transfusion. However, our study suggests that most autologous units frozen without specific planned use will not be transfused.     

Impact clinique de la durée de conservation des globules rouges avant transfusion

Presently, red blood cell units are stored up to 42 days in France and Canada. Length of storage of red blood cell units is not based on clinical outcomes: it is rather based on a decision made by some experts in the 1940s that red blood cell units can be stored as long as the average hemolysis is lower than 1% and the proportion of red blood cells still alive 24 hours post-transfusion is higher than 70%. Data reported recently suggest that transfusion with older red blood cell units may jeopardize the outcome of severely ill patients. In this paper, we comment the data already published on this question, and we summarize the randomized clinical trials presently on-going that were undertaken to address the relationship between length of storage of red blood cell units and outcomes of transfused patients.     

Preservation of erythrocytes of heterozygous SC sickle cell patients]

In order to evaluate the feasibility of the autologous transfusion in an alloimmunized sickle cell patient, changes in the hematologic and biochemical characteristics of erythrocytes stored for 42 days from two patients with sickle cell SC anemia were compared with control subjects' (Hb A) red blood cells. Erythrocytes were stored in Saline Adenosine Dextrose Mannitol at +4 degrees C. The cryopreservation storage was made and 51Cr red cell survival was measured in one patient. No significant difference in the hematologic and biochemical parameters of the SC red blood cells and the control subjects was observed during the storage at +4 degrees C. Red cell survivals determined in fresh cells, cells stored for 42 days at +4 degrees C and thawed cells from one patient demonstrate much shorter half-life values than those of normal red blood cells. Before application, our results need to be confirmed by the same protocol with another patient with sickle cell SC.     

红细胞冰冻前的最佳保存期探讨

背景：既能适当延长冰冻红细胞在冰冻前的保存期,又能保证冰冻红细胞的质量,是十分有意义的研究课题。 目的：适当延长稀有血型红细胞冰冻前在(4±2) ℃的保存期,满足急症稀有血型患者的输血,降低血液成本,减轻工作强度。 方法：将冰冻前(4±2) ℃保存1~6 d和冰冻前(4±2) ℃保存7~12 d制备的冰冻红细胞进行质量比较;根据血液保养液的红细胞保护原理、美国AABB相关标准及质检结果将含添加剂的红细胞在冰冻前的保存期设定为自采血之日起14 d内,对采取此措施前后2001/2009本中心的Rh(D)阴性供血情况进行统计分析。 结果与结论：质量检测结果达到国家标准,两组质量检测数据比较差异无显著性意义;将含添加剂的红细胞在冰冻前的保存期设定为自采血之日起14 d内,本中心Rh(D)阴性血年供血量虽逐年增加,但冰冻红细胞的供血量却未增加,其占供血的比例还在逐年下降,(4±2) ℃保存血的供血比例则逐渐增加。冰冻前在(4±2) ℃保存14 d再制备的冰冻红细胞发往临床均未发现因此而产生的血液质量问题。结果可见增加Rh(D)阴性(4±2) ℃保存血供血量,减少冰冻红细胞供血量,能及时满足急症Rh(D)阴性患者的输血,降低制备冰冻红细胞导致的高血液成本,减轻员工制备冰冻红细胞的工作强度。     

Transitioning from ‘blood’ safety to ‘transfusion’ safety: addressing the single biggest risk of transfusion

Transfusion errors occur at all points in the transfusion chain, often occurring at multiple points in the transfusion process for the same patient. Such events have been reported to national haemovigilance programs in almost all countries, over and over again. An incredible number of safety changes have been implemented to improve blood safety, including but not limited to: nucleic acid testing for HIV/HBV/HCV, bacterial culture for platelet concentrates, use of male-only plasma, and the introduction of pathogen reduction strategies. By contrast, very little momentum has developed behind transfusion safety, in hope of improving the safe delivery of blood to patients. This article will review the interventions that have been studied by transfusion medicine services in attempt to improve transfusion safety at every link in the transfusion chain. The most important and indispensable safety step is the introduction of an error tracking system. Such a system should capture all deviations from standard operating procedures, including near-misses that are captured before the blood product is issued. Near-misses are 300-fold more common and represent latent safety concerns requiring urgent attention. The system should be anonymous to ensure that there is no barrier to reporting and no-fault to recognize that the vast majority of errors are due to latent system errors. The errors should be coded by type and location to allow for the ability to query the error database for the purposes of benchmarking and tracking and trending after system changes. Such a system will allow hospital transfusion services to focus their initiatives at the steps in the transfusion chain most in need of repair at their institution. The system changes that have been studied include: confirmatory group testing, computerized physician order entry, prospective screening of transfusion orders before/after issue, controlled patient registration, regional blood bank information systems, positive patient identification at time of sample collection and the start of transfusion (using barcode or RFID technology), controlled release refrigeration devices, patient involvement in the transfusion process, and healthcare professional education. For each area, the specific technologies or examples will be detailed, the reports from the literature will be reviewed, and the obstacles to implementation will be discussed. Now that blood safety has been assured, we need to re-focus our attentions on the single biggest threat to patients: errors in the transfusion chain at the hospital level. We need to ensure that patients get blood only when required, that they get the correct product of the correct blood group, at the right dose, at the appropriate infusion rate, to the correct patient, at the right time. We need to take a rigorous scientific approach to solving transfusion safety to ensure that each process change is properly tested and validated to verify that each newly introduced process is safe and effective.     

Oxidative stress and antioxidant defenses during blood processing and storage of erythrocyte concentrates

Oxidative lesions start accumulating in cells when the oxidant attacks overwhelm the antioxidant defenses. This review will briefly describe red blood cell storage lesions with emphasis on the consequences of oxidation and the cellular defense mechanisms, as well as the methods that can be used to monitor them. The sources of variability in red blood cell storage capacity depend on the donor characteristics, the product processing and the storage conditions. Suggestions to improve the product quality in order to ensure the best efficacy and safety for the transfused patient are also discussed.     

The direct effects of stored blood products may worsen prognosis of cancer patients; shall we transfuse or not? An explanation of the adverse oncological consequences of blood product transfusion with a testable hypothesis driven experimental research protocol

Francis and colleagues reported an association between blood transfusion and worsened cancer prognosis. Since then there has been much debate over whether there is in fact such an association. We propose a possible mechanism which could explain much of the conflicting clinical and experimental evidence, and which can be readily tested experimentally. It is suggested that the extracellular accumulation of bioactive factors in blood transfusion products can directly and indirectly cause tumour growth and hence a worsening of prognosis. This theory can be applied both in vitro and in vivo. Two separate UK studies have shown that perioperative blood transfusion is associated with worsened prognosis in head and neck squamous cell cancer patients. Furthermore, pilot experiments have shown that as blood ages, endothelial growth factors are leached from the metabolically compromised red cell. We believe that we have provided a rationale to explain the conflicting findings of research to date in this area. That red cells should store endothelial reparative growth factors would seem logical, as would the release of any factors as the metabolic processes of the anucleate red cell decline over time. As a result, leuco-depletion should be promoted and blood transfusion should be avoided if possible.