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.     

Clinical effects of different types of red cell concentrates in patients with thalassemia and sickle cell disease

The treatment of thalassemia is still essentially based on continuous transfusion supporting using red cell concentrates (RCC) prepared in different ways. For patients with sickle-cell disorders, either urgent or chronic red blood cell transfusion therapy, is widely used in the management of sickle cell disease (SCD) because it reduces HbS level and generally prevents recurrent vaso-occlusive disease (VOD). Recently, the introduction of pre-storage filtration to remove leukocytes and the use of techniques for multicomponent donation have increased the types of blood components available for transfusion purposes. The clinical effects of different types of blood components in thalassaemic and sickle-cell patients have not been extensively studied so far. We evaluated the impact of the various different blood components currently available on transfusion needs, transfusion intervals and adverse reactions in order to determine which is the most advantageous for transfusion-dependent thalassaemic and sickle-cell patients followed in our centre. We believe that the optimal characteristics of the RCC are aged less than 10 days from time of collection; Hb content greater than 56 g per unit; Hct: 55-60%; volume (including additive) 300 mL+/-20%; leucodepleted to less than 200,000 leukocytes per unit; low cytokine content (achievable by pre-storage filtration carried out between two and 24 hours after the collection); lack of microaggregates (achievable by pre-storage filtration or filtration in the laboratory) and protein content less than 0.5 g per unit for patients allergic to plasma proteins (achievable with manual or automated washing). It is still recommended that the blood transfused should be as fresh as possible, compatible with the centre's product availability and the centre's organisation should be continuously adapted to this aim. We always transfuse blood within 10 days of its collection, respecting Rh and Kell system phenotypes. Pre-storage filtration is strongly recommended, both in order to prevent adverse reactions through the marked leucodepletion (less than 200,000 leukocytes per unit) and for a better standardisation of the final product, including the certainty that the product does not contain clots, an assurance that bed-side filtration cannot give. The RCC should be produced using a method causing as little as possible stress to the red cell membrane. The use of RCC with a high content of Hb (less than 56 g per unit) is strongly recommended, because our study clearly shows that this reduces the number of exposures to donors and the number of accesses to hospital, thus improving the patient's quality of life.     

The Quality of Red Blood Cells

The evolving practice of medicine has required a number of changes in red cell product manufacture to ensure that the final product is more specifically tailored to the needs of the individual patient. As a result of the increasing concern over the risks of transfusion pharmaceutical standards of manufacture are now applied to blood component preparation. Studies have been undertaken to define the optimum method of blood processing, and newer technologies are emerging to allow acquisition of a more consistent dose of red cells in a fashion which may minimize the lesion of collection. Use of high efficiency 3+ generation filter technologies reduces leukokine build up during storage and improves the quality and purity of the stored blood product. The combination of new plasticizers for packaging and improved red cell additive solutions should allow the blood center to supply a more functional red cell with longer storage shelf life. Overall these developments should result in the provision of a more consistent dose of fully functional red cells to the recipient who will be less exposed to the undesirable sequelae of transfusion than previously.     

Transfusion et drépanocytose : axes d’optimisation de la sécurité transfusionnelle

Transfusion remains a major treatment in sickle cell disease. In France, sickle cell disease patients are mainly from Sub-Saharan Africa and West Indies. The immuno-hematological characteristics of these patients of African ancestry induce a short supply of compatible packed red blood cells and an increased rate of haemolytic transfusion reactions, compared to the general transfused population. The optimization of transfusion safety relies on all steps of the transfusion chain. This article aims to describe the current situation in France and to determine the axes of optimization at all steps of the transfusion organization: promotion of donation, preparation of products, taking into account the sickle trait, qualification of packed red blood cells, supply of the blood banks concerned by transfusion of these patients, transfusion protocols and pre transfusion analysis. Research and formation play an important part.     

Rosetting in Plasmodium falciparum: A cytoadherence phenotype with multiple actors

The capacity of Plasmodium falciparum-infected red blood cells to bind uninfected red blood cells (“rosetting”) has been associated with high parasite density in numerous geographic areas and with severe malaria in African children. We summarize here the associations that have emerged from field studies and describe the various experimental models of rosetting that have been developed. A variety of erythrocyte receptors, several serum factors and a number of rosette-mediating PfEMP1 adhesins have been identified. Several var genes code for rosette-forming PfEMP1 adhesins in each P. falciparum genome, so that each clonal line has the capacity to generate distinct types of rosettes. To clarify their respective role in malaria pathogenesis, each of the multiple ligand/receptor interactions should be further studied for fine specificity, binding affinity and the impact of the large population polymorphism of the parasite variant repertoires should be assessed. Interestingly, some major human erythrocyte surface polymorphisms have been identified as affecting rosette formation, consistent with a role for rosetting in life-threatening falciparum malaria.     

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.     

Red blood cell alloimmunisation after platelet transfusion (excluding ABO blood group system)

Red blood cell alloimmunisation after transfusion of red blood cell concentrates carries a risk for every recipient. This risk is particularly high for patients with conditions such as sickle cell disease. However, red blood cell alloimmunisation can also occur after platelet concentrate transfusion. All blood group systems other than ABO are affected, and there are several mechanisms responsible for this alloimmunisation. The practical implications of this are a need to match red blood cell concentrates in all alloimmunised patients and, in pregnant women, recongnition of the risk of developing haemolytic disease of the foetus and newborn. Several measures can be taken to prevent alloimmunisation: in the case of the D antigen, for example, anti-RhD immunoglobulins can be infused before transfusing platelet concentrates from an RhD-positive donor in a RhD-negative recipient.     

The role and impact of the transfusion medicine consultation service in the management of patients in the hematopoietic transplant service: A retrospective analysis

BackgroundAdverse reactions secondary to transfusion of incorrect blood components can be fatal. We have established numerous processes to prevent these reactions in patients with cancer who continuously need blood component support, especially hematopoietic transplant recipients. The development of an active transfusion medicine consultation service at our institution to serve patients undergoing hematopoietic transplantation has led to more organized and simpler management of providing blood components to such patients.Study design and methodsSafety tools were employed to attain our goal of providing safe blood components to hematopoietic transplant recipients. These tools were consultation request forms, blood component selection stickers on the patients’ charts, and transfusion medicine physician consultation notes posted in the patients’ medical records. One hundred randomly selected hematopoietic transplant recipients were reviewed over 16 months. Fifty patients received blood components from ABO-compatible donors, whereas the other 50 patients received components from ABO-incompatible donors. Deviation reports regarding the issuance of blood components in these patients over the study period were reviewed retrospectively.ResultsWe identified eight reported deviations from the recommended blood components: red blood cells in one case, fresh frozen plasma in one case, single donor platelets in one case, and random donor platelets in five cases. Our transfusion service issued all eight components, but none of them were transfused. In all eight cases, the blood components were intended for transfusion to ABO-mismatched hematopoietic transplant recipients. Nurses identified the incorrect blood components by verifying the recommended blood groups on the patients’ chart stickers, returned the components to the transfusion service, and transfused the correct blood components.ConclusionUse of these safety tools has improved the safety culture regarding transfusion of blood components in hematopoietic transplant recipients at our institution.     

Evidence for indications of fresh frozen plasma

There continues to be a general but unfounded enthusiasm for fresh frozen plasma (FFP) usage across a range of clinical specialties in hospital practice. Clinical use of plasma has grown steadily over the last two decades in many countries. In England and Wales, there has not been a significant reduction in the use of FFP over the last few years, unlike red cells. There is also evidence of variation in usage among countries--use in England and Wales may be proportionately less per patient than current levels of usage in other European countries and the United States. Plasma for transfusion is most often used where there is abnormal coagulation screening tests, either therapeutically in the face of bleeding, or prophylactically in non-bleeding subjects prior to invasive procedures or surgery. Little evidence exists to inform best therapeutic plasma transfusion practice. Most studies have described plasma use in a prophylactic setting, in which laboratory abnormalities of coagulation tests are considered a predictive risk factor for bleeding prior to invasive procedures. The strongest randomised controlled trial (RCT) evidence indicates that prophylactic plasma for transfusion is not effective across a range of different clinical settings and this is supported by data from non-randomised studies in patients with mild to moderate abnormalities in coagulation tests. There are also uncertainties whether plasma consistently improves the laboratory results for patients with mild to moderate abnormalities in coagulation tests. There is a need to undertake new trials evaluating the efficacy and adverse effects of plasma, both in bleeding and non-bleeding patients, to understand whether the "presumed" benefits outweigh the "real risks". In addition, new haemostatic tests should be validated which better define risk of bleeding.     

Autologous blood predonation in cardiac surgery

The risk of transfusion of allogeneic blood products on outcome is well documented. Autologous blood donation prior to elective cardiac surgery has repeatedly shown to be an effective practice to reduce the exposure to allogeneic blood, but was criticized because of the alleged high costs. We analyzed the data of 4878 patients undergoing elective open-heart surgery, in whom 18% underwent autologous donation. Overall, autologous blood donation reduced the incidence of allogeneic blood transfusion from 48 to 13% during hospitalization. Additionally, it is shown that diagnosis adjusted autologous blood donation is cost-effective. The higher the probability of transfusion the better is efficacy and cost-effectiveness of predonation. Cardiac surgery is a high-transfusion area, thus, it offers ideal conditions for autologous donation. Autologous blood donation still remains a promising and cost-effective alternative to reduce allogeneic blood transfusion in elective cardiac surgery.     

Use of random versus apheresis platelet concentrates

The respective use of random (RPC) and apheresis (APC) platelet concentrates is highly heterogeneous among countries, ranging from 10 to 98% RPC in countries supposed to provide a similar transfusion service to patients. Moreover, when considering each country in the past 10 years, one can observe that some have changed their policy, switching from a majority of APC to RPC or vice versa. This presentation intends to analyse which factors may impact such decisions. For many years, the only available platelet component was a RPC obtained from whole blood donation by a two centrifugation steps process, the "platelet rich plasma" or PRP method. Since the beginning of the 1970s, APCs became available, with in fact many different techniques leading to many APCs that may not be equivalent. Since the end of the 1980s, a new method of RPC preparation was developed, using the buffy-coat (BC-PC), providing a blood component with highly preserved platelet functions as compared to RPCs prepared by the PRP technique. Finally, the use of each of these components either native, or leuco-reduced, or suspended in a storage solution, or processed with a pathogen inactivation technique adds new layers of complexity to compare them. Innumerable references can be found in the literature describing in vitro functional parameters of platelet concentrates. Although it is clear that BC-RPC retain much more their in vitro functions than PRP-RPC, indicating that no one should use the latter any more, it is much more difficult to distinguish differences between other PCs. Conversely, only a very few studies have been published related to a comparison of clinical efficacy of RPC versus APC, the endpoints being mainly CCI. Similarly to the in vitro studies, although RPC prepared with the PRP method show the lowest CCIs, no clear difference exists between "modern" RPC and APC. Another factor that may impact policy decision is the occurrence of adverse reactions in recipients. When considering only comparable data, for example leuco-reduced RPC versus leuco-reduced APC, there is now evidence that the latter is more associated with adverse reactions in recipients: data from hemovigilance in France show that, although no difference is noted for febrile non haemolytic transfusion reactions, nor for bacteria contamination, the incidence of allergic adverse reactions is about four times higher with APC as compared with RPC. Other aspects may impact the decision: the fact that using APC in place of RPC reduces the total donor exposure of patients was considered critical in some countries to reduce the risk of transmission of blood transmissible disease. Finally, the cost of the components, much higher for APC may be considered.     

Role of Lu/BCAM in abnormal adhesion of sickle red blood cells to vascular endothelium

Lutheran (Lu) blood group and Basal Cell Adhesion Molecule (BCAM) antigens are both carried by two glycoprotein (gp) isoforms of the immunoglobulin superfamily representing receptors for laminin alpha5 chain. They are expressed in red blood cells, in endothelial cells of vascular capillaries and in epithelial cells of several tissues. Lu/BCAM gps are overexpressed in sickle red blood cells (SS RBCs). Stimulation of SS RBCs by epinephrine activates the PKA depending signaling pathway and induces reinforced Lu/BCAM-mediated adhesion to laminin10/11. We have analyzed the phosphorylation state of Lu/BCAM long isoform cytoplasmic tail and showed that it is phosphorylated by CKII, GSK3b and PKA. Phosphorylation of this isoform in transfected K562 cells is stimulated by effectors of the PKA pathway and induces cell adhesion to laminin10/11. Lu/BCAM gps are highly expressed in endothelial cells and exhibit potential integrin binding motifs. We showed that they interact with integrin alpha4beta1, the unique integrin expressed on the surface of young reticulocytes. Adhesion assays under flow conditions showed that SS RBCs adhere to primary human endothelial cells (HUVEC) after selective activation of intergin alpha4beta1 and that this adhesion is mediated by endothelial Lu/BCAM gps. Our studies show that Lu/BCAM gps expressed either on erythroid or on endothelial cells are involved in SS RBC-endothelium interactions and could play a role in the abnormal adhesion of SS RBCs to vascular endothelium contributing to the vaso-occlusive crises reported for sickle cell disease patients.     

Cys shotgun labeling of macrophages adhering to and engulfing Ig-opsonized cells

A method is being developed to study cytoskeletal reorganization in cell adhesion processes. The initial model process is adhesion and phagocytosis of beads or red blood cells by macrophages. Live cell labeling with Cys reactive fluorophores is performed before and during phagocytosis with different color labeling dyes. Since Cys is a relatively hydrophobic amino acid, its differential exposure and labeling in principle reflects changes in tertiary or quaternary structure of specific proteins. Similar studies conducted on red blood cells under fluid shear conditions showed that specific domains in spectrin undergo extensible unfolding within sheared cells. The initial work here with macrophages also suggests some structural changes in phagocytosis although the proteins and specific sites have yet to be identified.     

Erythroid adhesion molecules in sickle cell disease: Effect of hydroxyurea

In sickle cell disease, the complex scenario of vaso-occlusive crisis (VOC) typical of this disease is clearly multifactorial and not fully understood. Cell-cell and cell-cell matrix interactions mediated by adhesive molecules present on blood cells and endothelial cells (ECs) are thought to play an important role. Early studies have shown that sickle red blood cells (RBCs) are abnormally adherent to ECs and some of the molecules involved in these interactions have been identified, such as the alpha4beta1 integrin and CD36, exclusively present on stress reticulocytes, and CD47 on mature RBCs. More recently, attention focused on Lu/BCAM, the unique RBC receptor for laminin, and on ICAM-4, a red cell-specific adhesion receptor, which is a ligand for a large repertoire of integrins (alphaLbeta2, alphaMbeta2, alphaxbeta2, alphaVbeta3). The counter-receptors on ECs and the role of plasma proteins forming bridges between blood cells and ECs have been clarified in part. It has also been shown that reticulocytes from SCD patients express higher levels of alpha4beta1 integrin and CD36, and that under hydroxyurea (HU) therapy, both cell adhesion to ECs or extracellular matrix proteins and the levels of these adhesion molecules are reduced. These findings are consistent with the view that enhanced adhesion of blood cells to ECs is largely determined by the membrane expression level of adhesion molecules and could be a crucial factor for triggering or aggravating vaso-occlusion. In SCD patients, membrane expression of Lu/BCAM (and perhaps ICAM-4) is enhanced on RBCs whose adherence to laminin or ECs is also increased. Interestingly, Lu/BCAM- and ICAM-4-mediated adhesion are enhanced by the stress mediator epinephrine through a PKA-dependent pathway initiated by a rise in intracellular cAMP and leading to receptor activation by phosphorylation according to the same signaling pathway. More recently, studies based on quantitative expression analysis of adhesion molecules on RBCs and during erythroid differentiation in patients undergoing HU therapy, surprisingly revealed that Lu/BCAM level was enhanced, although alpha4beta1, CD36 and ICAM-4 (to a lower extent) levels were indeed reduced. CD47 and CD147 expression were also enhanced in HU-treated patients. Based on these findings we suggest that the signalization cascade leading to receptor activation rather than the expression level only of adhesion molecules may be the critical factor regulating cell adhesion, although both mechanisms are not mutually exclusive.     

An overview of red blood cell and platelet alloimmunisation in transfusion

Post-transfusion alloimmunisation is the main complication of all those observed after one or more transfusion episodes. Alloimmunisation is observed after the transfusion of red blood cell concentrates but also of platelet concentrates. Besides alloimmunisation due to antigens carried almost exclusively by red blood cells such as those of the Rhesus-Kell system, alloimmunisation often raises against HLA antigens; the main responsibility for that, apart from platelet transfusions, lies with residual leukocytes in the products transfused, hence the central importance of effective leukoreduction right from the blood product preparation stage. Alloimmunization is not restricted to transfusion, but it is also observed during pregnancies, carrying out microtransfusions of blood from the fetus immunizing the mother through the placenta (in a retrograde way). Preexisting maternal-fetal immunization can complicate a transfusion program and intensify the creation of alloantibodies in several blood and tissue group systems. The occurrence of autoantibodies, created by several pathogenic reasons, can also interfere with the propensity of certain recipients of blood components to produce alloantibodies. The genetic condition of individuals is in fact strongly linked to the ability or not to recognize antigenic variants foreign to their own biological program and mount an alloimmune response. Some hemoglobin diseases, in carriers of which transfusions can be iterative and lifelong, are complicated by frequent alloimmunizations and amplification of the complications of these alloimmunizations, imposing even stricter transfusion rules. This review details the mechanisms favoring the occurrence of alloimmunization and the immunological principles for the production of molecular and cellular tools for alloimmunization. It concludes with the main preventive measures available to limit the occurrence of these frequent complications of varying severity but sometimes severe.     

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.     

L'adhérence des érythrocytes à l'endothélium vasculaire

Blood cells are in continuous contact with the vascular endothelium. Endothelial cell culture, intravital videomicroscopy allowed the investigation of blood cell-endothelium interactions in dynamic conditions.In the various diseases, diabetes mellitus, sickle cell anemia and malaria, erythrocytes have an increased adhesion to endothelial cells. The presence of advanced glycation end products (AGE) on erythrocytes of diabetics is responsible for their binding to the receptor RAGE present on the endothelium. The AGE-RAGE binding provokes an oxidant stress and induces the expression of the adhesion molecule. Furthermore, erythrocyte AGE induce an increase in vascular permeability. In sickle cell anemia, the increased adhesiveness and the sickling of red blood cells are responsible for thrombosis. Plasmodium falciparum infestation of erythrocytes induces knob formation at the cell surface and the P. falciparum protein binding to CD36, ICAM-1 and thrombospondin present on the endothelium, and facilitates the parasite dissemination. © 2999 Éditions scientifiques et médicales Elsevier SAS     

Sickle red cell adhesion: Many issues and some answers

Among multiple pathologies associated with sickle cell disease, sickle red cell-endothelial interaction has been implicated as a potential initiating mechanism in vaso-occlusive events that characterize this disease. Vast literature exists on various aspects of sickle red cell adhesion, but many issues remain unresolved, especially pertaining to the role of sickle red cell heterogeneity, the relative role of multiple adhesion mechanisms and targets of antiadhesive therapy. This review briefly analyzes these issues.     

Do multiple blood transfusions predispose for a higher rate of non-blood-related infection complications?

Allogeneic transfusions of red blood cell (RBC) concentrates have been related to an increase in postoperative infections. Leukocytes present in RBC units might have deleterious effects on the receptor immune system, provoking a state of immunosuppression that favors the development of postoperative infections (TRIM effect). The bioactive substances released by leukocytes in a time-dependent form, accumulating in blood components during storage, might be responsible for the TRIM effect. Multiple observational studies with logistic regression have demonstrated a direct relationship between transfusion and infection. However, several factors related to surgical difficulty and patient illness severity might act as strong confounding variables on the relationship studied. Randomized controlled trials designed to establish a causal relationship between transfusion and infection have yielded contradictory results. While we await new studies, allogeneic transfusions should be considered as a possible risk factor for postoperative infection.     

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.