Red blood cell storage: the story so far.

Red blood cells are still the most widely transfused blood component worldwide and their story is intimately entwined with the history of transfusion medicine and the changes in the collection and storage of blood1,2. At present, the most widely used protocol for the storage of red blood cells (for up to 42 days) is the collection of blood into anticoagulant solutions (typically citrate-dextrose-phosphate); red cell concentrates are prepared by the removal of plasma and, in some cases, also leukoreduction. The product is stored at 4 ± 2° C in a slightly hypertonic additive solution, generally SAGM (sodium, adenine, glucose, mannitol, 376 mOsm/L)1. Despite this, a definitive protocol that reconciles long-term storage on the one hand and safety and efficacy of the transfusion therapy on the other is still the subject of intense debate and discussion. In fact, although the organisation of the blood system, through the achievement of self-sufficiency, currently enables ordinary requests of the transfusion ‘market’ to be met, in the case of a calamity, disaster, or emerging infections3, or in particular periods of the year, local reserves can sometimes reach a minimum. There is still an underlying concern about the real need to store blood components for as long as possible in order to obtain a gradual increase in the interval between the donation and the transfusion, and how much this elastic time span can be prolonged without definitively compromising the quality of the product and, in the final analysis, the recipients’ health2. Indeed, although the transfusion establishment initially pursued both objectives (product quality and prolongation of the storage period), recent retrospective studies (whose results are, therefore, weakened by all the statistical limitations of this type of analysis)5–8 have indicated the apparent irreconcilability of the two aims. These studies seem to suggest that the quality (in terms of safety and efficiency) of red blood cells decreases in proportion to the time the storage period is prolonged. Furthermore, there is extremely convincing molecular evidence9,10 which, together with the results of clinical studies11–33, appears to confirm the preliminary conclusions regarding the likely poorer quality of red blood cells stored for a long time. However, the statistical validity and methodological rigour, in terms of evidence-based medicine, of the clinical studies have recently been challenged, highlighting the need for prospective, double-blind, randomized studies, in like fashion to the one carried out by Walsh et al.34 in 2004, which led the authors to conclude “the data did not support the hypothesis that transfusing red blood cells stored for a long time has detrimental effects on tissue oxygenation in critically ill, anaemic, euvolumaeic patients without active bleeding”. The international scientific community now seems much more convinced of the need of prospective studies, since such studies, on large cohorts of subjects, are currently underway35,36. The key point of the problem is probably the lack of universally accepted standard criteria that closely reflect the dramatic molecular changes that occur during prolonged storage of red blood cells and which, simply put, would enable ‘good’ blood to be distinguished from ‘no longer sufficiently good’ blood. The current standard requirements for patenting new additive solutions in the USA, and also suggested in the recommendations of the European Council37, are essentially based on two parameters: the level of haemolysis (below the threshold of 0.8% at the end of the storage period, following the introduction of the “95/95” rule38) and a survival rate of the transfused cells of more than 75% at 24 hours after transfusion. This latter parameter can be assessed by measuring the half-life of red blood cells labelled with 51 chromium prior to transfusion. These parameters are, however, fairly general and easily affected by the considerable biological variability between donors, given that it is known that blood from some donors resists storage better than that from other donors39. Haemolysis is an easier parameter to monitor. Typically, between 0.2 and 0.4% of red blood cells stored in the presence of standard additive solutions are haemolysed after 5–6 weeks of storage, while pre-storage leukoreduction halves the incidence of this phenomenon40. These widely accepted and well-established parameters do not, however, reflect the profound molecular changes that affect red blood cells during their storage. A brief list of the elements of the so-called “red blood cell storage lesion” includes10: morphological changes, slowed metabolism with a decrease in the concentration of adenosine triphosphate (ATP), acidosis with a decrease in the concentration of 2,3-diphosphoglycerate (2,3-DPG), loss of function (usually transient) of cation pumps and consequent loss of intracellular potassium and accumulation of sodium within the cytoplasm, oxidative damage with changes to the structure of band 341 and lipid peroxidation, apoptotic changes with racemisation of membrane phospholipids and loss of parts of the membrane through vesiculation9. Some of these changes occur within the first few hours of storage, for example, the decrease in pH or the increases in potassium and lactate; others, however, take days or weeks10. Together, these events risk compromising the safety and efficacy of long-stored red blood cells, reducing their capacity to carry and release oxygen, promoting the release of potentially toxic intermediates (for example, free haemoglobin can act as a source of reactive oxygen species) and negatively influencing physiological rheology (through the increased capacity of the red blood cells to adhere to the endothelium42,43 or through their enhanced thrombogenic44 or pro-inflammatory45 potential). These observations at a molecular level were supported by the results of a series of clinical studies (albeit retrospective and not randomised). These studies appeared to show a relationship between the duration of storage and a proportional increase in adverse events in the transfused patients, although the data available are preliminary and the statistically more reliable studies that conform more closely with the gold standard criteria represented by evidence-based medicine are considered necessary by many4 and are, indeed, underway5.