Development of new blood banking strategies for processing and storage of red blood cell components

Abstract

The field of red blood cell (RBC) components has been quite uneventful during the last few decades, save for the experimental exploration of alternative additive solution compositions. The reason is likely that the RBC quality has been good enough and the shelf-life of conventional RBC concentrates (RCC) has been long enough to not really motivate the workload that any dramatic changes would implicate.

However, the concern for future blood supply shortage is now growing in Europe. It is incited by multiple factors; perhaps foremost, the oncoming ban of the blood bag plasticizer di(2-ethylhexyl) phthalate (DEHP). DEHP prolongs the RBC’s lifespan during blood bank storage by stabilising the RBC membrane, and removal of DEHP has been linked to unacceptable haemolysis levels. For a long time, it has been a challenge to find a replacement to DEHP that does not compromise the RBC quality or RCC shelf-life. Concomitantly with the imminent non-DEHP transition, new regulatory frameworks call for better blood supply contingency and preparedness for emergency situations, and the healthcare community encourages more individualised treatment therapies. A contra-indication to all of these proceedings is that the donor population is decreasing due to smaller birth cohorts, simultaneously as the number of patients needing transfusion therapy is increasing due to longer life expectancy and more successful treatment of diseases.

The four studies of this theses have focused on different ways to approach the same aim of preserving or even improving the RBC quality, limit the outdating frequency and, ultimately, ensure a safe and sufficient blood supply.

Whole blood was processed into RCCs, which were further treated to fit the objectives of each respective study. In Paper I, a method was developed to cryopreserve split RCCs, by combining the ACP 215 automated cell processor with a subsequent manual centrifugation step. In Papers II and III, a four-armed study compared storage or RCCs in DEHP to a suggested substitute plasticizer, di(2-ethylhexyl) terephthalate (DEHT), combined with either of the two additive solutions saline-adenine-glucose-mannitol (SAGM) or phosphate-adenine-glucose-guanosine-saline-mannitol (PAGGSM). Paper II addressed ideal storage conditions whereas Paper III exposed the RBCs to extreme oxidative stress in the form of X-ray irradiation. Irradiated and ACP 215-washed RCCs were then compared to RCCs pathogen reduced with the Intercept blood system in Paper IV. All studies assessed the RBC quality after intervention and during subsequent storage by utilising a battery of analyses that together determined the RBC storage lesion, i.e. the RBCs metabolic, morphologic and oxidative status as well as the direct preservation of its membrane.

Paper I demonstrated that it was possible to successfully cryopreserve split RCCs without negatively impacting the quality of the final component compared to traditional, non-split cryopreserved RCCs. In fact, the levels of haemolysis and extracellular potassium ions (K+) were both lower than in non-split cryopreserved RCCs, which emphasised the compatibility of split cryopreserved RCCs in a paediatric transfusion setting. The quality of all the split RCCs was very even, demonstrating the robustness and reproducibility of the protocol, which is essential for everyday blood banking. Papers II and III verified DEHT as a strong potential substitute plasticizer to DEHP, even though the RBC membrane integrity was slightly impaired, foremost during SAGM storage. Irradiation expectedly introduced additional membrane damage, but satisfactory, the haemolysis was still well within the allowed margin. Independent of the level of stress exposure, the results suggested that if a transition to PAGGSM is adopted at the same time as a new plasticizer is implemented, any deleterious effects of the DEHP removal can be strongly mitigated. The two papers also confirmed that the RBC metabolism was unaffected by the switch of plasticizer. Paper IV demonstrated that, in addition to increased safety in terms of transfusion-transmitted infectious diseases, pathogen reduction is a promising future option to both irradiation and washing of blood components. Pathogen reduced RCCs exhibited membrane preservation similar to conventional RCCs and far superior to irradiated and automated-washed RCCs, where shelf-life reduction is a necessary adverse measure. With pathogen reduction, the shelf-life of conventional RCCs in Sweden, 42 days, would still be feasible. In addition, pathogen reduced RCCs implicated better ATP preservation, which may be beneficial for the RBC in vivo survival; however, at the expense of 2,3-DPG. Considering the washing-specific parameters, implying efficacy of plasma reduction, minor adjustments of the centrifugation protocol would still be desirable.

In conclusion, this thesis explores the multiple pathways of RBC processing in order to develop or refine RBC components or RBC storage. The purpose behind this is to propose a way to adapt to the growing urge of ensuring availability of a blood supply fit for all essential transfusions to all categories of patients. In four individual papers, this thesis shows the deleterious effects of RBC cell stress, but also proposes a joint common approach to mitigate it or, in some cases, even improve the components further compared to the previous standard. Hopefully, this innovative view on classical RBC components, along with the presentation of a satisfactory plasticizer option, may inspire to the introduction of a number of measures that can contribute to decreased RCC wastage or prevention of shelf-life reduction. These are two key attributes in increasing patient safety.