Regulation of kynurenine metabolism by blood donor genetics and biology impacts red cell hemolysis in vitro and in vivo.

ABSTRACT: In the field of transfusion medicine, the clinical relevance of the metabolic markers of the red blood cell (RBC) storage lesion is incompletely understood. Here, we performed metabolomics of RBC units from 643 donors enrolled in the Recipient Epidemiology and Donor Evaluation Study, REDS RBC Omics. These units were tested on storage days 10, 23, and 42 for a total of 1929 samples and also characterized for end-of-storage hemolytic propensity after oxidative and osmotic insults. Our results indicate that the metabolic markers of the storage lesion poorly correlated with hemolytic propensity. In contrast, kynurenine was not affected by storage duration and was identified as the top predictor of osmotic fragility. RBC kynurenine levels were affected by donor age and body mass index and were reproducible within the same donor across multiple donations from 2 to 12 months apart. To delve into the genetic underpinnings of kynurenine levels in stored RBCs, we thus tested kynurenine levels in stored RBCs on day 42 from 13 091 donors from the REDS RBC Omics study, a population that was also genotyped for 879 000 single nucleotide polymorphisms. Through a metabolite quantitative trait loci analysis, we identified polymorphisms in SLC7A5, ATXN2, and a series of rate-limiting enzymes (eg, kynurenine monooxygenase, indoleamine 2,3-dioxygenase, and tryptophan dioxygenase) in the kynurenine pathway as critical factors affecting RBC kynurenine levels. By interrogating a donor-recipient linkage vein-to-vein database, we then report that SLC7A5 polymorphisms are also associated with changes in hemoglobin and bilirubin levels, suggestive of in vivo hemolysis in 4470 individuals who were critically ill and receiving single-unit transfusions.

Genetic regulation of carnitine metabolism controls lipid damage repair and aging RBC hemolysis in vivo and in vitro.

ABSTRACT: Recent large-scale multiomics studies suggest that genetic factors influence the chemical individuality of donated blood. To examine this concept, we performed metabolomics analyses of 643 blood units from volunteers who donated units of packed red blood cells (RBCs) on 2 separate occasions. These analyses identified carnitine metabolism as the most reproducible pathway across multiple donations from the same donor. We also measured l-carnitine and acyl-carnitines in 13 091 packed RBC units from donors in the Recipient Epidemiology and Donor Evaluation study. Genome-wide association studies against 879 000 polymorphisms identified critical genetic factors contributing to interdonor heterogeneity in end-of-storage carnitine levels, including common nonsynonymous polymorphisms in genes encoding carnitine transporters (SLC22A16, SLC22A5, and SLC16A9); carnitine synthesis (FLVCR1 and MTDH) and metabolism (CPT1A, CPT2, CRAT, and ACSS2), and carnitine-dependent repair of lipids oxidized by ALOX5. Significant associations between genetic polymorphisms on SLC22 transporters and carnitine pools in stored RBCs were validated in 525 Diversity Outbred mice. Donors carrying 2 alleles of the rs12210538 SLC22A16 single-nucleotide polymorphism exhibited the lowest l-carnitine levels, significant elevations of in vitro hemolysis, and the highest degree of vesiculation, accompanied by increases in lipid peroxidation markers. Separation of RBCs by age, via in vivo biotinylation in mice, and Percoll density gradients of human RBCs, showed age-dependent depletions of l-carnitine and acyl-carnitine pools, accompanied by progressive failure of the reacylation process after chemically induced membrane lipid damage. Supplementation of stored murine RBCs with l-carnitine boosted posttransfusion recovery, suggesting this could represent a viable strategy to improve RBC storage quality.

Clinical outcomes in hospitalized plasma and platelet transfusion recipients prior to and following widespread blood donor SARS-CoV-2 infection and vaccination.

BACKGROUND: The safety of transfusion of SARS-CoV-2 antibodies in high plasma volume blood components to recipients without COVID-19 is not established. We assessed whether transfusion of plasma or platelet products during periods of increasing prevalence of blood donor SARS-CoV-2 infection and vaccination was associated with changes in outcomes in hospitalized patients without COVID-19. METHODS: We conducted a retrospective cohort study of hospitalized adults who received plasma or platelet transfusions at 21 hospitals during pre-COVID-19 (3/1/2018-2/29/2020), COVID-19 pre-vaccine (3/1/2020-2/28/2021), and COVID-19 post-vaccine (3/1/2021-8/31/2022) study periods. We used multivariable logistic regression with generalized estimating equations to adjust for demographics and comorbidities to calculate odds ratios (ORs) and 95% confidence intervals (CIs). RESULTS: Among 21,750 hospitalizations of 18,584 transfusion recipients without COVID-19, there were 697 post-transfusion thrombotic events, and oxygen requirements were increased in 1751 hospitalizations. Intensive care unit length of stay (n = 11,683) was 3 days (interquartile range 1-5), hospital mortality occurred in 3223 (14.8%), and 30-day rehospitalization in 4144 (23.7%). Comparing the pre-COVID, pre-vaccine and post-vaccine study periods, there were no trends in thromboses (OR 0.9 [95% CI 0.8, 1.1]; p = .22) or oxygen requirements (OR 1.0 [95% CI 0.9, 1.1]; p = .41). In parallel, there were no trends across study periods for ICU length of stay (p = .83), adjusted hospital mortality (OR 1.0 [95% CI 0.9-1.0]; p = .36), or 30-day rehospitalization (p = .29). DISCUSSION: Transfusion of plasma and platelet blood components collected during the pre-vaccine and post-vaccine periods of the COVID-19 pandemic was not associated with increased adverse outcomes in transfusion recipients without COVID-19.

Testosterone supplementation increases red blood cell susceptibility to oxidative stress, decreases membrane deformability, and decreases survival after cold storage and transfusion.

BACKGROUND: Blood collection from donors on testosterone therapy (TT) is restricted to red blood cell (RBC) concentrates to avoid patient exposure to supraphysiological testosterone (T). The objective of this study was to identify TT-related changes in RBC characteristics relevant to transfusion effectiveness in patients. STUDY DESIGN: This was a two-part study with cohorts of patients and blood donors on TT. In part 1, we conducted longitudinal evaluation of RBCs collected before and at three time points after initiation of T. RBC assays included storage and oxidative hemolysis, membrane deformability (elongation index), and oximetry. In part 2, we evaluated the fate of transfused RBCs from TT donors in immunodeficient mice and by retrospective analyses of NIH's vein-to-vein databases. RESULTS: TT increased oxidative hemolysis (1.45-fold change) and decreased RBC membrane deformability. Plasma free testosterone was positively correlated with oxidative hemolysis (r = .552) and negatively correlated with the elongation index (r = -.472). Stored and gamma-irradiated RBCs from TT donors had lower posttransfusion recovery in mice compared to controls (41.6 ± 12 vs. 55.3 ± 20.5%). Recipients of RBCs from male donors taking T had 25% lower hemoglobin increments compared to recipients of RBCs from non-TT male donors, and had increased incidence (OR, 1.80) of requiring additional RBC transfusions within 48 h of the index transfusion event. CONCLUSIONS: TT is associated with altered RBC characteristics and transfusion effectiveness. These results suggest that clinical utilization of TT RBCs may be less effective in recipients who benefit from longer RBC survival, such as chronically transfused patients.

The prevalence of hepatitis B virus, human T-lymphotropic virus and human immunodeficiency virus in patients receiving blood transfusions in South Africa.

BACKGROUND AND OBJECTIVES: South Africa has a high prevalence of human immunodeficiency virus (HIV) and hepatitis B virus (HBV) and to a lesser extent human T-lymphotropic virus (HTLV). Each of these agents is transfusion-transmissible (TT) but deciding whether to implement preventive screening depends upon knowledge of background prevalence in transfused patients. We determined the prevalence of HIV, HBV and HTLV I/II among blood transfusion recipients in South African hospitals. MATERIALS AND METHODS: We obtained identity-unlinked samples used for blood cross-matching at 634 South African hospitals served by the South African National Blood Service (SANBS). The ABBOTT Alinity S® Immunochemiluminescent system measured HIV, HBV and HTLV I/II antibodies. Repeatedly reactive samples were confirmed using the Roche Cobas® 8000. Logistic regression was performed to investigate the determinants of associations for HIV, HBV and HTLV infections. RESULTS: The overall prevalences of HIV, HBV and HTLV were 37.8%, 7.4% and 0.6%, respectively. The HIV prevalence in blood recipients was twice as high as general population estimates. Public hospital patients had a significantly higher prevalence compared with private hospital patients for HIV and HBV. HIV prevalence was significantly higher in females, and HBV prevalence was significantly higher in males, excluding the unknown gender results. CONCLUSION: Patients receiving blood transfusions in South Africa have high rates of HIV and HBV infection that should be taken into consideration when determining donor screening strategies for other viral infections. Measurable prevalence of HTLV indicates endemicity of this infection in South Africa.

Genetic determinants of plasma testosterone in male blood donors are associated with altered red blood cell characteristics and survival in cold storage and after transfusion.

Genetic mutations in genes regulating plasma testosterone in men may interfere with effective erythropoiesis, and may result in red blood cell (RBC) dysfunction and hemolysis. The aim of this study was to identify genetic polymorphisms in male donors that regulate plasma testosterone and impact RBC survival in cold storage and after transfusion. We evaluated nine single nucleotide polymorphisms (SNPs) previously reported to be associated with circulating testosterone in male plasma. These SNPs were linked with donor-component-recipient databases (NIH REDS program) to determine SNP associations with donor RBC hematological indices, osmotic and oxidative hemolysis, and RBC transfusion effectiveness defined as adjusted hemoglobin increments (delta hemoglobin, ΔHb) following a single RBC unit transfusion. Four of the nine testosterone SNPs were located on the X chromosome, of which two (rs7057002, rs73629199) were significantly associated with reduced hemoglobin increments (0.2 and 0.3 g/dL, respectively) compared with reference alleles in transfused recipients. Seven of the nine testosterone SNPs were associated with significant changes in RBC susceptibility to osmotic hemolysis including a missense mutation in the major plasma carrier of testosterone (SHBG, rs6259), and four SNPs with changes in oxidative hemolysis. Four SNPs were associated with decreased RBC count, hemoglobin, and hematocrit. Ancestry/ethnicity-specific (African and Hispanic) associations were observed between two SNPs (rs7057002, rs7879462) and oxidative hemolysis. Genetic determinants of plasma testosterone in male donors significantly impact the quality and transfusion effectiveness of cold stored RBCs. Testosterone SNPs associated with decreased RBC transfusion effectiveness may have clinical implications and warrant further revaluation.

Associations of donor, component, and recipient factors on hemoglobin increments following red blood cell transfusion in very low birth weight infants.

BACKGROUND: Anemia in very low birth weight (VLBW) infants is common and frequently managed with red blood cell (RBC) transfusions. We utilized a linked vein-to-vein database to assess the role of blood donors and component factors on measures of RBC transfusion effectiveness in VLBW infants. STUDY DESIGN AND METHODS: We linked blood donor and component manufacturing data with VLBW infants transfused RBCs between January 1, 2013 and December 31, 2016 in the Recipient Epidemiology Donor Evaluation Study-III (REDS III) database. Using multivariable regression, hemoglobin increments and subsequent transfusion events following single-unit RBC transfusion episodes were examined with consideration of donor, component, and recipient factors. RESULTS: Data on VLBW infants (n = 254) who received one or more single-unit RBC transfusions (n = 567 units) were linked to donor demographic and component manufacturing characteristics for analysis. Reduced post-transfusion hemoglobin increments were associated with RBC units donated by female donors (-0.24 g/dL [95% confidence interval (CI) -0.57, -0.02]; p = .04) and donors <25 years old (-0.57 g/dL [95% CI -1.02, -0.11]; p = .02). For RBC units donated by male donors, reduced donor hemoglobin levels were associated with an increased need for subsequent recipient RBC transfusion (odds ratio 3.0 [95% CI 1.3, 6.7]; p < .01). In contrast, component characteristics, storage duration, and time from irradiation to transfusion were not associated with post-transfusion hemoglobin increments. CONCLUSION: Donor sex, age, and hemoglobin levels were associated with measures of RBC transfusion effectiveness in VLBW infants. Mechanistic studies are needed to better understand the role of these potential donor factors on other clinical outcomes in VLBW infants.

Blood donor, component, and recipient-specific factors associated with venous thromboembolism in transfused hospitalized adult patients: Data from the recipient epidemiology and donor evaluation Study-III (REDS-III).

OBJECTIVE: Growing evidence suggests multiple pathophysiological mechanisms linking red blood cells (RBC) transfusions to thrombosis. This study examined blood donor, component, and recipient factors which may be associated with thromboembolic outcomes following RBC transfusion. METHODS: We utilized the Recipient Epidemiology Donor Evaluation Study-III (REDS-III) database on patients transfused in 12 hospitals between 2013-2016. Stratified Cox proportional hazards regression models with time-dependent exposures were used to examine associations of donor and component modification characteristics on venous thromboembolism (VTE) in patients transfused RBC units. RESULTS: 59,603 patients were transfused 229,500 RBC units during 79,298 hospitalizations with post-transfusion VTE occurring in 1869 (2.4%) of patients. In adjusted regression analyses, a per RBC-unit risk of VTE was present for gamma irradiation (HR = 1.03; 95% CI: 1.02-1.03), female donor sex (HR = 1.01; 95% CI: 1.00-1.01), storage duration greater than 5 weeks (HR = 1.01; 95% CI: 1.01-1.02), AS-1 storage solution (HR = 1.01; 95% CI: 1.00-1.01), and apheresis-derived collections (HR = 1.01; 95% CI: 1.01-1.02). Among recipient factors, male sex (HR = 1.03; 95% CI: 1.02-1.04), pre-transfusion hemoglobin level (HR = 0.94; 95% CI: 0.94-0.94), body mass index strata (HR = 1.11; 95% CI: 1.08-1.14), and principal diagnoses including malignancy (HR = 1.13; 95% CI: 1.10-1.16), cardiac arrest (HR = 1.38; 95% CI:1.07-1.77) and hip fracture (HR = 1.59; 95% CI:1.53-1.66) were associated with VTE in adjusted analyses. DISCUSSION: We identified several donor, component, and recipient-specific factors associated with VTE in transfused hospitalized adult patients. In adjusted models, the dose-dependent associations of donor and component-specific factors with VTE were modest and unlikely to be clinically significant in the majority of transfused patients. Additional mechanistic and clinical studies linking blood donor and component factors with thrombotic outcomes are needed.

Associations between ABO non-identical platelet transfusions and patient outcomes-A multicenter retrospective analysis.

BACKGROUND: Due to platelet availability limitations, platelet units ABO mismatched to recipients are often transfused. However, since platelets express ABO antigens and are collected in plasma which may contain ABO isohemagglutinins, it remains controversial as to whether ABO non-identical platelet transfusions could potentially pose harm and/or have reduced efficacy. STUDY DESIGN AND METHODS: The large 4-year publicly available Recipient Epidemiology and Donor Evaluation Study-III (REDS-III) database was used to investigate patient outcomes associated with ABO non-identical platelet transfusions. Outcomes included mortality, sepsis, and subsequent platelet transfusion requirements. RESULTS: Following adjustment for possible confounding factors, no statistically significant association between ABO non-identical platelet transfusion and increased risk of mortality was observed in the overall cohort of 21,176 recipients. However, when analyzed by diagnostic category and recipient ABO group, associations with increased mortality for major mismatched transfusions were noted in two of eight subpopulations. Hematology/Oncology blood group A and B recipients (but not group O) showed a Hazard Ratio (HR) of 1.29 (95%CI: 1.03-1.62) and intracerebral hemorrhage group O recipients (but not groups A and B) showed a HR of 1.75 (95%CI: 1.10-2.80). Major mismatched transfusions were associated with increased odds of receiving additional platelet transfusion each post-transfusion day (through day 5) regardless of the recipient blood group. DISCUSSION: We suggest that prospective studies are needed to determine if specific patient populations would benefit from receiving ABO identical platelet units. Our findings indicate that ABO-identical platelet products minimize patient exposure to additional platelet doses.

Proceedings of the 2022 NHLBI and OASH state of the science in transfusion medicine symposium.

BACKGROUND: State of the Science (SoS) meetings are used to define and highlight important unanswered scientific questions. The National Heart, Lung, and Blood Institute (NHLBI), National Institutes of Health, and the Office of the Assistant Secretary for Health (OASH), Department of Health and Human Services held a virtual SoS in transfusion medicine (TM) symposium. STUDY DESIGN AND METHODS: In advance of the symposium, six multidisciplinary working groups (WG) convened to define research priorities in the areas of: blood donors and the supply, optimizing transfusion outcomes for recipients, emerging infections, mechanistic aspects of components and transfusion, new computational methods in transfusion science, and impact of health disparities on donors and recipients. The overall objective was to identify key basic, translational, and clinical research questions that will help to increase and diversify the volunteer donor pool, ensure safe and effective transfusion strategies for recipients, and identify which blood products from which donors best meet the clinical needs of specific recipient populations. RESULTS: On August 29-30, 2022, over 400 researchers, clinicians, industry experts, government officials, community members, and patient advocates discussed the research priorities presented by each WG. Dialogue focused on the five highest priority research areas identified by each WG and included the rationale, proposed methodological approaches, feasibility, and barriers for success. DISCUSSION: This report summarizes the key ideas and research priorities identified during the NHLBI/OASH SoS in TM symposium. The report highlights major gaps in our current knowledge and provides a road map for TM research.

Early Post-Hospitalization Hemoglobin Recovery and Clinical Outcomes in Survivors of Critical Illness: A Population-Based Cohort Study.

Anemia is common during critical illness, is associated with adverse clinical outcomes, and often persists after hospitalization. The goal of this investigation is to assess the relationships between post-hospitalization hemoglobin recovery and clinical outcomes after survival of critical illness. This is a population-based observational study of adults (≥18 years) surviving hospitalization for critical illness between January 1, 2010 and December 31, 2016 in Olmsted County, Minnesota, United States with hemoglobin concentrations and clinical outcomes assessed through one-year post-hospitalization. Multi-state proportional hazards models were utilized to assess the relationships between 1-month post-hospitalization hemoglobin recovery and hospital readmission or death through one-year after discharge. Among 6460 patients that survived hospitalization for critical illness during the study period, 2736 (42%) were alive, not hospitalized, and had available hemoglobin concentrations assessed at 1-month post-index hospitalization. Median (interquartile range) age was 69 (56, 80) years with 54% of male gender. Overall, 86% of patients had anemia at the time of hospital discharge, with median discharge hemoglobin concentrations of 10.2 (9.1, 11.6) g/dL. In adjusted analyses, each 1 g/dL increase in 1-month hemoglobin recovery was associated with decreased instantaneous hazard for hospital readmission (HR 0.87 [95% CI 0.84-0.90]; p < 0.001) and lower mortality (HR 0.82 [95% CI 0.75-0.89]; p < 0.001) through one-year post-hospitalization. The results were consistent in multiple pre-defined sensitivity analyses. Impaired early post-hospitalization hemoglobin recovery is associated with inferior clinical outcomes in the first year of survival after critical illness. Additional investigations are warranted to evaluate these relationships.

Effect of donor, component, and recipient characteristics on hemoglobin increments following red blood cell transfusion.

Significant research has focused individually on blood donors, product preparation and storage, and optimal transfusion practice. To better understand the interplay between these factors on measures of red blood cell (RBC) transfusion efficacy, we conducted a linked analysis of blood donor and component data with patients who received single-unit RBC transfusions between 2008 and 2016. Hemoglobin levels before and after RBC transfusions and at 24- and 48-hour intervals after transfusion were analyzed. Generalized estimating equation linear regression models were fit to examine hemoglobin increments after RBC transfusion adjusting for donor and recipient demographic characteristics, collection method, additive solution, gamma irradiation, and storage duration. We linked data on 23 194 transfusion recipients who received one or more single-unit RBC transfusions (n = 38 019 units) to donor demographic and component characteristics. Donor and recipient sex, Rh-D status, collection method, gamma irradiation, recipient age and body mass index, and pretransfusion hemoglobin levels were significant predictors of hemoglobin increments in univariate and multivariable analyses (P < .01). For hemoglobin increments 24 hours after transfusion, the coefficient of determination for the generalized estimating equation models was 0.25, with an estimated correlation between actual and predicted values of 0.5. Collectively, blood donor demographic characteristics, collection and processing methods, and recipient characteristics accounted for significant variation in hemoglobin increments related to RBC transfusion. Multivariable modeling allows the prediction of changes in hemoglobin using donor-, component-, and patient-level characteristics. Accounting for these factors will be critical for future analyses of donor and component factors, including genetic polymorphisms, on posttransfusion increments and other patient outcomes.

Blood donor obesity is associated with changes in red blood cell metabolism and susceptibility to hemolysis in cold storage and in response to osmotic and oxidative stress.

BACKGROUND: Obesity is a global pandemic characterized by multiple comorbidities, including cardiovascular and metabolic diseases. The aim of this study was to define the associations between blood donor body mass index (BMI) and RBC measurements of metabolic stress and hemolysis. STUDY DESIGN AND METHODS: The associations between donor BMI (<25 kg/m2 , normal weight; 25-29.9 kg/m2 , overweight; and ≥30 kg/m2 , obese) and hemolysis (storage, osmotic, and oxidative; n = 18 donors) or posttransfusion recovery (n = 14 donors) in immunodeficient mice were determined in stored leukocyte-reduced RBC units. Further evaluations were conducted using the National Heart, Lung, and Blood Institute RBC-Omics blood donor databases of hemolysis (n = 13 317) and metabolomics (n = 203). RESULTS: Evaluations in 18 donors revealed that BMI was significantly (P < 0.05) and positively associated with storage and osmotic hemolysis. A BMI of 30 kg/m2 or greater was also associated with lower posttransfusion recovery in mice 10 minutes after transfusion (P = 0.026). Multivariable linear regression analyses in RBC-Omics revealed that BMI was a significant modifier for all hemolysis measurements, explaining 4.5%, 4.2%, and 0.2% of the variance in osmotic, oxidative, and storage hemolysis, respectively. In this cohort, obesity was positively associated (P < 0.001) with plasma ferritin (inflammation marker). Metabolomic analyses on RBCs from obese donors (44.1 ± 5.1 kg/m2 ) had altered membrane lipid composition, dysregulation of antioxidant pathways (eg, increased oxidized lipids, methionine sulfoxide, and xanthine), and dysregulation of nitric oxide metabolism, as compared to RBCs from nonobese (20.5 ± 1.0 kg/m2 ) donors. CONCLUSIONS: Obesity is associated with significant changes in RBC metabolism and increased susceptibility to hemolysis under routine storage of RBC units. The impact on transfusion efficacy warrants further evaluation.

Transfusion thresholds for guiding red blood cell transfusion.

BACKGROUND: The optimal haemoglobin threshold for use of red blood cell (RBC) transfusions in anaemic patients remains an active field of research. Blood is a scarce resource, and in some countries, transfusions are less safe than in others because of inadequate testing for viral pathogens. If a liberal transfusion policy does not improve clinical outcomes, or if it is equivalent, then adopting a more restrictive approach could be recognised as the standard of care.  OBJECTIVES: The aim of this review update was to compare 30-day mortality and other clinical outcomes for participants randomised to restrictive versus liberal red blood cell (RBC) transfusion thresholds (triggers) for all clinical conditions. The restrictive transfusion threshold uses a lower haemoglobin concentration as a threshold for transfusion (most commonly, 7.0 g/dL to 8.0 g/dL), and the liberal transfusion threshold uses a higher haemoglobin concentration as a threshold for transfusion (most commonly, 9.0 g/dL to 10.0 g/dL). SEARCH METHODS: We identified trials through updated searches: CENTRAL (2020, Issue 11), MEDLINE (1946 to November 2020), Embase (1974 to November 2020), Transfusion Evidence Library (1950 to November 2020), Web of Science Conference Proceedings Citation Index (1990 to November 2020), and trial registries (November 2020). We  checked the reference lists of other published reviews and relevant papers to identify additional trials. We were aware of one trial identified in earlier searching that was in the process of being published (in February 2021), and we were able to include it before this review was finalised. SELECTION CRITERIA: We included randomised trials of surgical or medical participants that recruited adults or children, or both. We excluded studies that focused on neonates. Eligible trials assigned intervention groups on the basis of different transfusion schedules or thresholds or 'triggers'. These thresholds would be defined by a haemoglobin (Hb) or haematocrit (Hct) concentration below which an RBC transfusion would be administered; the haemoglobin concentration remains the most commonly applied marker of the need for RBC transfusion in clinical practice. We included trials in which investigators had allocated participants to higher thresholds or more liberal transfusion strategies compared to more restrictive ones, which might include no transfusion. As in previous versions of this review, we did not exclude unregistered trials published after 2010 (as per the policy of the Cochrane Injuries Group, 2015), however, we did conduct analyses to consider the differential impact of results of trials for which prospective registration could not be confirmed.   DATA COLLECTION AND ANALYSIS: We identified trials for inclusion and extracted data using Cochrane methods. We pooled risk ratios of clinical outcomes across trials using a random-effects model. Two review authors independently extracted data and assessed risk of bias. We conducted predefined analyses by clinical subgroups. We defined participants randomly allocated to the lower transfusion threshold as being in the 'restrictive transfusion' group and those randomly allocated to the higher transfusion threshold as being in the 'liberal transfusion' group. MAIN RESULTS: A total of 48 trials, involving data from 21,433 participants (at baseline), across a range of clinical contexts (e.g. orthopaedic, cardiac, or vascular surgery; critical care; acute blood loss (including gastrointestinal bleeding); acute coronary syndrome; cancer; leukaemia; haematological malignancies), met the eligibility criteria. The haemoglobin concentration used to define the restrictive transfusion group in most trials (36) was between 7.0 g/dL and 8.0 g/dL.  Most trials included only adults; three trials focused on children. The included studies were generally at low risk of bias for key domains including allocation concealment and incomplete outcome data. Restrictive transfusion strategies reduced the risk of receiving at least one RBC transfusion by 41% across a broad range of clinical contexts (risk ratio (RR) 0.59, 95% confidence interval (CI) 0.53 to 0.66; 42 studies, 20,057 participants; high-quality evidence), with a large amount of heterogeneity between trials (I² = 96%). Overall, restrictive transfusion strategies did not increase or decrease the risk of 30-day mortality compared with liberal transfusion strategies (RR 0.99, 95% CI 0.86 to 1.15; 31 studies, 16,729 participants; I² = 30%; moderate-quality evidence) or any of the other outcomes assessed (i.e. cardiac events (low-quality evidence), myocardial infarction, stroke, thromboembolism (all high-quality evidence)). High-quality evidence shows that the liberal transfusion threshold did not affect the risk of infection (pneumonia, wound infection, or bacteraemia). Transfusion-specific reactions are uncommon and were inconsistently reported within trials. We noted less certainty in the strength of evidence to support the safety of restrictive transfusion thresholds for the following predefined clinical subgroups: myocardial infarction, vascular surgery, haematological malignancies, and chronic bone-marrow disorders. AUTHORS' CONCLUSIONS: Transfusion at a restrictive haemoglobin concentration decreased the proportion of people exposed to RBC transfusion by 41% across a broad range of clinical contexts. Across all trials, no evidence suggests that a restrictive transfusion strategy impacted 30-day mortality, mortality at other time points, or morbidity (i.e. cardiac events, myocardial infarction, stroke, pneumonia, thromboembolism, infection) compared with a liberal transfusion strategy. Despite including 17 more randomised trials (and 8846 participants), data remain insufficient to inform the safety of transfusion policies in important and selected clinical contexts, such as myocardial infarction, chronic cardiovascular disease, neurological injury or traumatic brain injury, stroke, thrombocytopenia, and cancer or haematological malignancies, including chronic bone marrow failure.  Further work is needed to improve our understanding of outcomes other than mortality. Most trials compared only two separate thresholds for haemoglobin concentration, which may not identify the actual optimal threshold for transfusion in a particular patient. Haemoglobin concentration may not be the most informative marker of the need for transfusion in individual patients with different degrees of physiological adaptation to anaemia. Notwithstanding these issues, overall findings provide good evidence that transfusions with allogeneic RBCs can be avoided in most patients with haemoglobin thresholds between the range of 7.0 g/dL and 8.0 g/dL. Some patient subgroups might benefit from RBCs to maintain higher haemoglobin concentrations; research efforts should focus on these clinical contexts.

NT-proBNP levels in the identification and classification of pulmonary transfusion reactions.

BACKGROUND: Consensus definitions for transfusion-related acute lung injury (TRALI) and transfusion-associated circulatory overload (TACO) have recently been revised; however, pulmonary transfusion reactions remain difficult to diagnose. We hypothesized that N-terminal pro-brain natriuretic peptide (NT-proBNP) levels could have utility in the identification and classification of pulmonary transfusion reactions. STUDY DESIGN AND METHODS: We performed a secondary analysis of a case-control study of pulmonary transfusion reactions at four academic hospitals. We evaluated clinical data and measured NT-proBNP levels prior to and following transfusion in patients with TACO (n = 160), transfused acute respiratory distress syndrome (ARDS) [n = 51], TRALI [n = 12], TACO/TRALI [n = 7], and controls [n = 335]. We used Wilcoxon Rank-Sum tests to compare NT-proBNP levels, and classification and regression tree (CART) algorithms to produce a ranking of covariates in order of relative importance for differentiating TACO from transfused controls. RESULTS: Pre-transfusion NT-proBNP levels were elevated in cases of transfused ARDS and TACO (both P < .001) but not TRALI (P = .31) or TACO/TRALI (P = .23) compared to transfused controls. Pre-transfusion NT-proBNP levels were higher in cases of transfused ARDS or TRALI with a diagnosis of sepsis compared to those without (P < .05 for both). CART analyses resulted in similar differentiation of patients with TACO from transfused controls for models utilizing either NT-proBNP levels (AUC 0.83) or echocardiogram results (AUC 0.80). CONCLUSIONS: NT-proBNP levels may have utility in the classification of pulmonary transfusion reactions. Prospective studies are needed to test the predictive utility of pre-transfusion NT-proBNP in conjunction with other clinical factors in identifying patients at risk of pulmonary transfusion reactions.

Nicotine exposure increases markers of oxidant stress in stored red blood cells from healthy donor volunteers.

BACKGROUND: Cigarette smoking is a frequent habit across blood donors (approx. 13% of the donor population), that could compound biologic factors and exacerbate oxidant stress to stored red blood cells (RBCs). STUDY DESIGN AND METHODS: As part of the REDS-III RBC-Omics (Recipient Epidemiology Donor Evaluation Study III Red Blood Cell-Omics) study, a total of 599 samples were sterilely drawn from RBC units stored under blood bank conditions at Storage Days 10, 23, and 42 days, before testing for hemolysis parameters and metabolomics. Quantitative measurements of nicotine and its metabolites cotinine and cotinine oxide were performed against deuterium-labeled internal standards. RESULTS: Donors whose blood cotinine levels exceeded 10 ng/mL (14% of the tested donors) were characterized by higher levels of early glycolytic intermediates, pentose phosphate pathway metabolites, and pyruvate-to-lactate ratios, all markers of increased basal oxidant stress. Consistently, increased glutathionylation of oxidized triose sugars and lipid aldehydes was observed in RBCs donated by nicotine-exposed donors, which were also characterized by increased fatty acid desaturation, purine salvage, and methionine oxidation and consumption via pathways involved in oxidative stress-triggered protein damage-repair mechanisms. CONCLUSION: RBCs from donors with high levels of nicotine exposure are characterized by increases in basal oxidant stress and decreases in osmotic hemolysis. These findings indicate the need for future clinical studies aimed at addressing the impact of smoking and other sources of nicotine (e.g., nicotine patches, snuff, vaping, secondhand tobacco smoke) on RBC storage quality and transfusion efficacy.

Additive effects of blood donor smoking and gamma irradiation on outcome measures of red blood cell transfusion.

BACKGROUND: Recent publications have reported conflicting results regarding the role of blood donor tobacco use on hemoglobin (Hb) levels in patients after red blood cell (RBC) transfusion. We examined associations and interactions between donor, component, and recipient factors to better understand the impact of donor smoking on transfusion outcomes. STUDY DESIGN AND METHODS: We linked blood donor and component manufacturing data, including self-reported cigarette smoking, with a cohort of patients transfused RBCs between 2013 and 2016. Using multivariable regression, we examined Hb increments and subsequent transfusion requirements after single-unit RBC transfusion episodes, adjusting for donor, component, and recipient factors. RESULTS: We linked data on 4038 transfusion recipients who received one or more single-unit RBC transfusions (n = 5086 units) to donor demographic and component manufacturing characteristics. Among RBC units from smokers (n = 326), Hb increments were reduced after transfusion of gamma-irradiated units (0.76 g/dL; p = 0.033) but not unirradiated units (1.04 g/dL; p = 0.54) compared to those from nonsmokers (1.01 g/dL; n = 4760). In parallel with changes in Hb levels, donor smoking was associated with the receipt of additional RBC transfusions for irradiated (odds ratio [OR], 2.49; p = 0.01) but not unirradiated RBC units (OR, 1.10; p = 0.52). CONCLUSION: Donor smoking was associated with reduced Hb increments and the need for additional transfusions in recipients of gamma-irradiated RBC units. Additional research is needed to better understand interactions between donor, component, and recipient factors on efficacy measures of RBC transfusion.

Antenatal blood transfusion in South Africa: indications and practice in a high-HIV-prevalence setting.

BACKGROUND: Globally, data on antenatal blood transfusion practices are scarce. We sought to characterize the epidemiology of antenatal transfusion in South Africa. STUDY DESIGN AND METHODS: A cross-sectional study was conducted of women who were transfused during pregnancy (>48 hr before anticipated delivery) at two hospitals in Durban and Soweto in 2014 to 2015. Medical record data on demographics, obstetric history, anemia, HIV status, and indications for blood transfusion were abstracted. RESULTS: The records on a total of 560 transfused pregnant women were evaluated; mean age was 28 years, 98% were of black African ethnicity, and 28% were HIV positive. At time of transfusion, one-half were in the first trimester. Hemorrhage was noted in 76% of women, most of which was associated with abortion (67%) or ectopic pregnancy (27%). Most women were transfused with red blood cells (RBCs; median, 2 units); 14% of women were transfused with plasma and 2% with platelets. Median pre- and posttransfusion hemoglobin levels were 6.9 g/dL and 9.2 g/dL, respectively; the latter differed by hospital (8.7 g/dL vs. 9.5 g/dL; p < 0.01). Hemorrhage was associated with missing HIV status, lower gestational age, and transfusion of 3 or more RBC units (all p < 0.01). In contrast, diagnoses of anemia (Soweto only) were associated with HIV infection, later gestational age, and lower (<3 units) RBC dose (all p < 0.01). CONCLUSION: Abortion and ectopic pregnancy with associated hemorrhage were the leading indications for antenatal transfusion and were concentrated in early gestation. By contrast, anemia was associated with HIV infection and transfusion in the third trimester.

The evanescence and persistence of RBC alloantibodies in blood donors.

BACKGROUND: Blood donors represent a healthy population, whose red blood cell (RBC) alloantibody persistence or evanescence kinetics may differ from those of immunocompromised patients. A better understanding of the biologic factors impacting antibody persistence is warranted, as the presence of alloantibodies may impact donor health and the fate of the donated blood product. METHODS: Donor/donation data collected from four US blood centers from 2012 to 2016 as part of the Recipient Epidemiology and Donor Evaluation Study-III (REDS-III) were analyzed. Clinically significant antibodies from blood donors with more than one donation who underwent at least one follow-up antibody screen after the initial antibody identification were included. Of 632,378 blood donors, 481 (128 males and 353 females) fit inclusion criteria. RESULTS: Antibody screens detected 562 alloantibodies, with 368 of 562 (65%) of antibodies being persistently detected and with 194 of 562 (35%) becoming evanescent. Factors associated with antibody persistence included antibody specificity, detection at the first donation, reported history of transfusion, and detection of multiple antibodies concurrently. Anti-D, C, and Fya were most likely to persist, while anti-M, Jka , and S were most frequently evanescent. CONCLUSIONS: These data provide insight into variables impacting the duration of antibody detection, and they may also influence blood donor center policies regarding donor recruitment/acceptance.

Red blood cell transfusions for emergency department patients with gastrointestinal bleeding within an integrated health system.

STUDY OBJECTIVE: To assess trends over time in red blood cell (RBC) transfusion practice among emergency department (ED) patients with gastrointestinal (GI) bleeding within an integrated healthcare system, inclusive of 21 EDs. METHODS: Retrospective cohort of ED patients diagnosed with GI bleeding between July 1st, 2012 and September 30th, 2016. The primary outcome was receipt of an RBC transfusion in the ED. Secondary outcomes included 90-day rates of RBC transfusion, repeat ED visits, rehospitalization, and all-cause mortality. Logistic regression was used to obtain confounder-adjusted outcome rates. RESULTS: A total of 24,868 unique patient encounters were used for the primary analysis. The median hemoglobin level in the ED prior to RBC transfusion decreased from 7.5 g/dl to 6.9 g/dl in the first versus last twelve months of the study period (p < 0.0001). A small trend was observed in the overall adjusted rate of ED RBC transfusion (absolute quarterly change of -0.1%, R2 = 0.18, p = 0.0001) largely attributable to the subgroup of patients with hemoglobin nadirs between 7.0 and 9.9 g/dl (absolute quarterly change of -0.4%, R2 = 0.38, p < 0.0001). Rates of RBC transfusions through 90 days likewise decreased (absolute quarterly change of -0.4%, R2 = 0.85, p < 0.0001) with stable to decreased corresponding rates of repeat ED visits, rehospitalizations and mortality. CONCLUSION: Rates of ED RBC transfusion decreased over time among patients with GI bleeding, particularly in those with hemoglobin nadirs between 7.0 and 9.9 g/dl. These findings suggest that ED providers are willing to adopt evidence-based restrictive RBC transfusion recommendations for patients with GI bleeding.