Vesiculation in irradiated and cation-leaky-stored red blood cells.

BACKGROUND: Extracellular vesicles (EVs) are released by red blood cells (RBCs) throughout their life-span and also during hypothermic storage when they accumulate in the blood bag. We queried whether stored RBCs with increased cation permeability, either from donors with familial pseudohyperkalaemia (FP) or caused by irradiation, vesiculate more readily. STUDY DESIGN AND METHODS: Recent technical advances have revealed at least two sub-populations of MVs in RBC storage units: macrovesicles (2-6 µm) and microvesicles (1-2 µm). Using nanoparticle tracking analysis, imaging flow cytometry, and protein quantification methods, we measured and characterized vesicles released by RBCs from control and FP individuals at three different storage time-points (day 4, day 17, and day 29). The RBCs had either been stored untreated or irradiated on either day 1 or day 14 of storage. RESULTS: We found no difference in the number or size of vesicles released between cation-leaky FP RBCs and non-FP controls. Similarly, irradiated and non-irradiated RBCs showed very similar patterns of vesicle release to during cold-storage. The only significant difference in vesicle release was the increase in accumulated vesicles with length of storage time which has been reported previously. DISCUSSION: EVs in stored blood are potential contributors to adverse transfusion reactions. The number of vesicles released during 35-day hypothermic storage varies between donors and increases with storage duration. However, increased cation permeability and irradiation do not appear to affect vesicle formation during RBC cold-storage.

International guidelines regarding the role of IVIG in the management of Rh- and ABO-mediated haemolytic disease of the newborn.

Haemolytic disease of the newborn (HDN) can be associated with significant morbidity. Prompt treatment with intensive phototherapy (PT) and exchange transfusions (ETs) can dramatically improve outcomes. ET is invasive and associated with risks. Intravenous immunoglobulin (IVIG) may be an alternative therapy to prevent use of ET. An international panel of experts was convened to develop evidence-based recommendations regarding the effectiveness and safety of IVIG to reduce the need for ETs, improve neurocognitive outcomes, reduce bilirubin level, reduce the frequency of red blood cell (RBC) transfusions and severity of anaemia, and/or reduce duration of hospitalization for neonates with Rh or ABO-mediated HDN. We used a systematic approach to search and review the literature and then develop recommendations from published data. These recommendations conclude that IVIG should not be routinely used to treat Rh or ABO antibody-mediated HDN. In situations where hyperbilirubinaemia is severe (and ET is imminent), or when ET is not readily available, the role of IVIG is unclear. High-quality studies are urgently needed to assess the optimal use of IVIG in patients with HDN.

Evaluating apheresis platelets at reduced dose as a contingency measure for extreme shortages.

BACKGROUND: The COVID19 pandemic highlights the need for contingency planning in the event of blood shortages. To increase platelet supply, we assessed the operational impact and effect on platelet quality of splitting units prior to storage. STUDY DESIGN AND METHODS: Using production figures, we modeled the impact on unit numbers, platelet counts, and volumes of splitting only apheresis double donations into three units (yielding ⅔ doses), or all standard dose units in half. To assess quality, eight pools of three ABO/Rh-matched apheresis (Trima Accel) double donations in plasma were split to ⅔ and ½ volumes in both Terumo and Fresenius storage bags. These were irradiated and subject to maximal permitted periods of nonagitation (3 × 8 h) before comparing platelet quality markers (including pH, CD62P expression) to Day 9 of storage. RESULTS: Splitting all double donations into three predicted inventory expansion of 23% overall whereas halving all standard dose units clearly doubles stock. In our study, ⅔ and ½ doses contained 153 ± 15 × 109 (~138 ml) and 113 ± 11 × 109 (~102 ml) platelets respectively. Following storage, higher pH was observed in ⅔ than in ½ doses and in Terumo compared to Fresenius bags. The higher pH was reflected in better quality markers, including lower CD62P expression. Despite the differences, on Day 8 (of pH monitoring at expiry) all ⅔ doses and most ½ doses were ≥pH 6.4. CONCLUSION: A strategy to split apheresis platelets in plasma to lower doses is feasible, maintains acceptable platelet quality, and should be considered by blood services in response to extreme shortages.

The compound effect of irradiation and familial pseudohyperkalemia on potassium leak from red blood cells.

BACKGROUND: Familial pseudohyperkalemia (FP) is a rare asymptomatic condition characterized by an increased rate of potassium leak from red blood cells (RBC) on refrigeration. Gamma irradiation compromises RBC membrane integrity and accelerates potassium leakage. Here, we compared the effect of irradiation, applied early or late in storage, on FP versus non-FP RBC. STUDY DESIGN: Five FP and 10 non-FP individuals from the National Institute for Health Research Cambridge BioResource, UK, and three FP and six non-FP individuals identified by Australian Red Cross Lifeblood consented to the study. Blood was collected according to standard practice in each center, held overnight at 18-24°C, leucocyte-depleted, and processed into red cell concentrates (RCC) in Saline Adenine Glucose Mannitol. On Day 1, RCC were split equally into six Red Cell Splits (RCS). Two RCS remained non-irradiated, two were irradiated on Day 1 and two were irradiated on Day 14. RBCs were tested over cold storage for quality parameters. RESULTS: As expected, non-irradiated FP RCS had significantly higher supernatant potassium levels than controls throughout 28 days of storage (p < .001). When irradiated early, FP RCS released potassium at similar rates to control. When irradiated late, FP RCS supernatants had higher initial post-irradiation potassium concentration than controls but were similar to controls by the end of storage (14 days post-irradiation). No other parameters studied showed a significant difference between FP and control. DISCUSSION: FP does not increase the rate of potassium leak from irradiated RBCs. Irradiation may cause a membrane defect similar to that in FP RBCs.

A comparison of the effect of X and gamma irradiation on red cell storage quality.

BACKGROUND AND OBJECTIVES: Irradiation of red cell components is indicated for recipients at risk of transfusion-associated graft vs. host disease. Current technologies available comprise of a gamma (γ) or an x source of radiation. The benefits of x vs. γ include non-radioactivity and hence no decay of the source. We aimed to compare the effect of the two technologies on red cell component storage quality post-irradiation. MATERIALS AND METHODS: Paired units of red cell concentrates (RCC), neonatal red cell splits (RCS), red cells for intra-uterine transfusion (IUT) or neonatal exchange transfusion (ExTx) were either γ- or x-irradiated. Units were sampled and tested for five storage parameters until the end of shelf life. Equivalence analysis of storage quality parameters was performed for pairs of the same components (RCC, RCS, IUT or ExTx) that were either γ- or x-irradiated. RESULTS: Nearly all component comparisons studied showed equivalence between γ and x irradiation for haemolysis, ATP, 2,3-DPG, potassium release and lactate production. The exceptions found that were deemed non-equivalent were higher haemolysis with x irradiation for ExTx, lower 2,3-DPG with x irradiation for RCS irradiated early and higher ATP with x irradiation for IUT. However, these differences were considered not clinically significant. CONCLUSION: This study has demonstrated that a range of red cell components for use in different age groups are of acceptable quality following x irradiation, with only small differences deemed clinically insignificant in a few of the measured parameters.

UK national survey of anticoagulation in extra-corporeal photopheresis-Is it time for a UK consensus statement?

BACKGROUND: Extra-corporeal photopheresis (ECP) requires anticoagulation to prevent circuit clotting. Unfractionated heparin (UFH) is currently the only anticoagulant licensed for the ECP system in use in the United Kingdom (UK). Acid citrate dextrose-A (ACD-A) is the preferred anticoagulant for most other apheresis procedures. Anecdotal evidence suggested variability in ECP practice across the UK with some providers using off-label ACD-A. AIMS: We developed a survey together with the UK Photopheresis Society to establish current practice. MATERIALS & METHODS: This was distributed to all 17 ECP providers covering 34 UK sites. RESULTS: Significant variability in practice was demonstrated with only 36% of responding providers (5/14) using UFH exclusively and 29% (4/14) using ACD-A as standard. CONCLUSION: This survey highlights the need for a UK consensus.

Familial pseudohyperkalemia induces significantly higher levels of extracellular potassium in early storage of red cell concentrates without affecting other standard measures of quality: A case control and allele frequency study.

BACKGROUND: Familial pseudohyperkalemia (FP) is characterized by an increased rate of potassium leakage in refrigerated red cells and is associated with the minor allele of the single nucleotide polymorphism rs148211042 (R723Q) in the ABCB6 gene. The study aims were to obtain the minor allele frequencies of ABCB6 variants and to measure supernatant potassium accumulation, and other red cell storage parameters, in red cell concentrates (RCC) from carriers of variant rs148211042 under standard blood bank conditions. STUDY DESIGN: Whole blood units were collected from 6 FP individuals and 11 controls and processed into RCC in additive solution. RCC were sampled and tested over cold storage for full blood count, extracellular potassium, glucose, lactate, microvesicle release, deformability, hemolysis, pH, adenosine triphosphate, and 2,3-diphosphoglycerate. RESULTS: Screening of genotyped cohorts identified that variant rs148211042 is present in 1 in 394 British citizens of European ancestry. FP RCC had significantly higher supernatant potassium at all time points from day 3 onwards (p < .001) and higher mean cell volume (p = .032) than controls. The initial rate of potassium release was higher in FP RCC; supernatant potassium reached 46.0 (23.8-57.6) mmol/L (mean [range]) by day 5, increasing to 68.9 (58.8-73.7) mmol/L by day 35. Other quality parameters were not significantly different between FP RCC and controls. CONCLUSION: These data suggest that if a blood donor has FP, reducing the RCC shelf-life to 5 days may be insufficient to reduce the risk of hyperkalemia in clinical scenarios such as neonatal large volume transfusion.

Fatigue, executive function and psychological effects in children with immune thrombocytopenia: a cross-sectional study.

Childhood ITP is often considered to be a relatively mild haematological disorder, with only a minority of patients requiring treatment for troublesome bleeding. Over recent years, wider effects of the condition have been identified in some adults, particularly relating to fatigue and cognitive impairment. In this study, we sought to investigate such effects in a group of children with ITP and further our understanding of their psychological profile. Children attending routine haematology outpatient clinics and their parents were asked to complete standardised questionnaires designed to assess a range of psychological and cognitive factors. Although the majority of children had some scores within the normal range, a significantly high proportion had difficulties with fatigue (70·6%), emotional and behavioural symptoms (25·7%) or executive functioning (19·4%). Quality of life and subjective evaluation of the illness (appraisal) correlated significantly with each of these domains, but bleeding severity and platelet count did not. Our findings provide valuable insight into the broader impact of childhood ITP, which could aid in providing holistic care, potentially contribute to decisions regarding medical treatment, and guide future research.

Washed red cells: theory and practice.

Washing of red cells is sometimes performed to reduce allergic reactions due to contaminating plasma proteins or to reduce the concentration of potassium accumulating in the supernatant of red cells during storage as an alternative to transfusion of fresher red cells in patients at risk of hyperkalaemia. There are a variety of methods for washing red cells, and the laboratory data suggest that variables such as age of red cell before washing, washing method and solution, storage medium and length of storage time after washing can all effect the final red cell quality. Studies suggest that washing removes 90-95% plasma, but the proportion of units below 0·05 mg/dl IgA (equivalent to IgA deficient) is variable dependent upon methods used. Although potassium levels are reduced immediately following washing, the rate of leakage following subsequent storage is method-dependent, requiring careful consideration of shelf life if potassium reduction is the goal. Other markers of red cell quality such as haemolysis are negatively impacted by washing so a reduced shelf life compared to standard red cells is appropriate, especially following irradiation. Data from animal models and clinical studies on possible additional benefits of washing, such as reduced lung or kidney injury, are mixed, ranging from some benefit to some harm, and further studies are warranted.

Improving blood-prescribing decisions: Evaluating the efficacy of a blood guidelines app.

OBJECTIVES: To assess the effect of an app providing national blood transfusion guidelines on prescribing decisions. BACKGROUND: National, regional and local audits in England consistently show inappropriate use of all blood components; around 15%-20% of red blood cells (RBC) and 20%-30% of platelets and fresh frozen plasma (FFP). Hospital transfusion guidelines may be difficult to locate and not agree with national guidelines. We developed and tested a dedicated app providing national evidence-based guidelines for use at the point of care to help clinicians make better decisions when authorising blood. METHODS/MATERIALS: We identified areas of blood authorisation with high frequency of component use and evidence of widespread unnecessary authorisation. We developed seven representative clinical scenarios where the transfusion of blood components may or may not benefit the adult patient. Responding doctors were invited to select their authorisation choice via an online questionnaire, initially without and then with access to the app. Adherence to guidelines was assessed with and without aid of the app. RESULTS: Using the app, doctors were much more likely to select the correct decision, in accordance with national guidance. Compared with baseline measurements, decisions improved by 67% for RBC, 58% for platelets and 73% for FFP. These improvements were statistically significant. CONCLUSION: Apps such as "Blood Components" can help doctors do "the right thing rather than the wrong thing". Further studies are required to assess the impact of using the app in clinical practice and the effect on blood component management and financial savings.

Rejuvenation of RBCs: validation of a manufacturing method suitable for clinical use.

BACKGROUND: Rejuvenation of stored red blood cells (RBCs) increases levels of adenosine 5'-triphosphate (ATP) and 2,3-diphosphoglycerate (2,3-DPG) to those of fresh cells. This study aimed to optimize and validate the US-approved process to a UK setting for manufacture and issue of rejuvenated RBCs for a multicenter randomized controlled clinical trial in cardiac surgery. STUDY DESIGN AND METHODS: Rejuvenation of leukoreduced RBC units involved adding a solution containing pyruvate, inosine, phosphate, and adenine (Rejuvesol, Zimmer Biomet), warming at 37°C for 60 minutes, then "manual" washing with saline adenine glucose mannitol solution. A laboratory study was conducted on six pools of ABO/D-matched units made the day after donation. On Days 7, 21, and 28 of 4 ± 2°C storage, one unit per pool was rejuvenated and measured over 96 hours for volume, hematocrit, hemolysis, ATP, 2,3-DPG, supernatant potassium, lactate, and purines added (inosine) or produced (hypoxanthine) by rejuvenation. Subsequently, an operational validation (two phases of 32 units each) was undertaken, with results from the first informing a trial component specification applied to the second. Rejuvenation effects were also tested on crossmatch reactivity and RBC antigen profiles. RESULTS: Rejuvenation raised 2,3-DPG to, and ATP above, levels of fresh cells. The final component had potassium and hemolysis values below those of standard storage Days 7 and 21, respectively, containing 1.2% exogenous inosine and 500 to 1900 µmoles/unit of hypoxanthine. The second operational validation met compliance to the trial component specification. Rejuvenation did not adversely affect crossmatch reactivity or RBC antigen profiles. CONCLUSION: The validated rejuvenation process operates within defined quality limits, preserving RBC immunophenotypes, enabling manufacture for clinical trials.

Patient Blood Management for Neonates and Children Undergoing Cardiac Surgery: 2019 NATA Guidelines.

Pediatric cardiac surgery is associated with a substantial risk of bleeding, frequently requiring the administration of allogeneic blood products. Efforts to optimize preoperative hemoglobin, limit blood sampling, improve hemostasis, reduce bleeding, correct coagulopathy, and incorporate blood sparing techniques (including restrictive transfusion practices) are key elements of patient blood management (PBM) programs, and should be applied to the pediatric cardiac surgical population as across other disciplines. Many guidelines for implementation of PBM in adults undergoing cardiac surgery are available, but evidence regarding the implementation of PBM in children is limited to systematic reviews and specific guidelines for the pediatric cardiac population are missing. The objective of the task force from the Network for the Advancement of Patient Blood Management, Haemostasis and Thrombosis (NATA, www.nataonline.com) is to provide evidence-based recommendations regarding anemia management and blood transfusion practices in the perioperative care of neonates and children undergoing cardiac surgery, and to highlight potential areas where additional research is urgently required.

Red blood cell transfusion practice in children: current status and areas for improvement? A study of the use of red blood cell transfusions in children and infants.

BACKGROUND: Patterns of red blood cell (RBC) transfusion are less well understood for children than adults. This study was undertaken to document current pediatric practice, to identify specific areas for improving patient care and safety. STUDY DESIGN AND METHODS: All UK hospitals were invited to participate. All children less than 18 years old admitted and receiving a RBC transfusion during a 3-month period in 2009 were eligible for inclusion. RESULTS: A total of 160 of 247 (65%) sites treating children or neonates responded; 119 provided data on 1302 pediatric patients transfused in nonneonatal wards. A total of 74% of patients received a single RBC transfusion during their admission. More than half (53%) of recipients had a hematologic or oncologic underlying diagnosis, and 33% were on general pediatric wards. The median pretransfusion hemoglobin (Hb) level was 7.9 g/dL (interquartile range [IQR], 6.9-9.4 g/dL), varying by location and diagnosis. The median volume prescribed was 15 mL/kg (IQR, 11.8-19.2 mL/kg). Prescribing by units instead of milliliters was recorded for 493 of 1264 (39%) of transfusions. For 734 of 1302 (56%) where Hb levels were available within 2 days between pre- and posttransfusion Hb, the median transfusion increment was 2.8 g/dL (IQR, 1.4-3.9 g/dL). CONCLUSION: This study of UK pediatric RBC transfusion practice has demonstrated significant variation in pretransfusion Hb, frequent prescribing in units rather than milliliters, and a high proportion of single transfusions during admissions. Future education and research should target transfusion triggers and prescription volumes for children in all clinical areas.

Familial pseudohyperkalemia in blood donors: a novel mutation with implications for transfusion practice.

BACKGROUND: Familial pseudohyperkalemia (FP) is a dominantly inherited condition in which red blood cells (RBCs) have an increased cold-induced permeability to monovalent cations. Potassium leaks into the supernatant of all stored blood with time, but FP RBCs leak potassium more rapidly. We investigated two unrelated blood donors whose RBC donations demonstrated unexpectedly high potassium after 5 and 6 days' storage. We matched the observed pattern of RBC cation leak to a previously recognized family with FP (FP-Cardiff) and investigated the likely cause with targeted DNA analysis. STUDY DESIGN AND METHODS: Cation leakage from the donor RBCs and from standard donor units was measured. DNA analysis of donors and family members with FP-Cardiff was performed. Allele frequencies were obtained from human variation databases. RESULTS: Both implicated donors were found to have increased cold-induced potassium leak identical in pattern to affected members of the family with FP-Cardiff. We found a heterozygous substitution Arg723Gln in the ATP-binding cassette, Subfamily B, Member 6 protein that segregated with FP in the Cardiff family and was also present in both blood donors. Arg723Gln is listed in human variation databases with an allele frequency of approximately 1:1000. CONCLUSIONS: We describe a novel FP mutation that may affect 1:500 European blood donors and causes rapid loss of potassium from stored RBCs. This finding has implications for neonates and infants receiving large-volume RBC transfusions. Genomic screening of donors could be used to identify donors with this mutation and potentially improve the quality and safety of donor units.